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  • <h1>SNAPSHOT</h1>
  • <h4>Created at: 2017-12-04 05:24</h4>
  • <h4>Created at: 2019-06-20 05:42</h4>
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  • <!-- Title Section, includes id, name and short name -->
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  • <h2>AOP ID and Title:</h2>
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  • AOP 149: Peptide Oxidation Leading to Hypertension
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  • <strong>Short Title: Hypertension</strong>
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  • <h2>Graphical Representation</h2>
  • <img src="https://aopwiki.org/system/dragonfly/production/2017/08/17/62rd12uxde_AOP_149_JPG.jpg" , height="500" , width="700"> </img>
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  • <hr>
  • <!-- Author section, includes text of author names as they have been entered by the user -->
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  • <h2>Authors</h2>
  • <hr>
  • <p><em>British American Tobacco:</em> Frazer Lowe (Frazer_Lowe@bat.com); Linsey Haswell; Marianna Gaca</p>
  • <p><em>Philip Morris International:</em> Karsta Luettich; Marja Talikka&nbsp;; Julia Hoeng</p>
  • <p><em>Selventa:</em> Vy Hoang (vhoang@selventa.com)</p>
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  • <!-- Status Section, lists status of aop -->
  • <div id="status">
  • <h2>Status</h2>
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  • <table class="table table-bordered table-striped">
  • <thead>
  • <th>Author status</th>
  • <th>OECD status</th>
  • <th>OECD project</th>
  • <th>SAAOP status</th>
  • </thead>
  • <tr>
  • <td>Under development: Not open for comment. Do not cite</td>
  • <td>Under Development</td>
  • <td>1.50</td>
  • <td>Included in OECD Work Plan</td>
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  • <!-- Abstract Section, text as generated by author -->
  • <div id="abstract">
  • <h2>Abstract</h2>
  • <hr>
  • <p style="text-align:justify">Hypertension is a cardiovascular risk factor that has a profound influence on cardiovascular morbidity and mortality.&nbsp; While good progress has been made in terms of identifying and managing this risk factor for patient care, methods to assess the potential of chemical compounds to induce hypertension, or to assess the efficacy of consumer products (e.g. e-cigarettes, tobacco heating products) targeted at reducing disease burden remain largely limited to epidemiological associations and <em>in vivo</em> studies.&nbsp;</p>
  • <p style="text-align:justify">Here we present the supporting information on an AOP describing how vascular endothelial peptide oxidation leads to hypertension via perturbation of endothelial nitric oxide (NO) bioavailability. &nbsp;The molecular initiating event is oxidation of amino acid (AA) residues on critical peptides of the NO pathway, notably protein kinase B (AKT), guanosine triphosphate cyclohydrolase-1 (GTPCH-1), endothelial nitric oxide synthase (eNOS), and also the cellular ROS scavenger; glutathione. &nbsp;Oxidation of the enzymic components of the pathway lead to reduced expression of the&nbsp;phosphorylated proteins, and protein loss via proteasomal degradation.&nbsp;Oxidation of reduced glutathione to GSSG promotes bonding of GSSG to critical AA residues on eNOS, and the reduced expression of GTPCH-1 reduces bioavailability of tetrahydrobiopterin (BH4), both of which lead&nbsp;to uncoupling of eNOS (reduced NO production, increased superoxide production). &nbsp;The combination of these molecular events lead to reduced bioavailabilty of NO, which in turn reduces the potential for vasodilation and shifts the balance of vascular tone towards vasoconstriction. &nbsp;Repeated perturbation of this pathway via chronic exposure to toxicants, ultimately increases vascular resistance and contributes towards the development of hypertension.</p>
  • <p style="text-align:justify">The evidence supporting the AOP is&nbsp;strong from the molecular level up to impaired vasodilation,&nbsp;however there is a knowledge gap concerning the magnitude of&nbsp;contribution of this mechanism towards the development of hypertension, given the complexity of the disorder. &nbsp;Further AOPs are likely required to characterise the contribution of competing/promoting mechanisms which alter vascular tone towards chronic vasoconstriction.</p>
  • <p style="text-align:justify">With respect to the regulatory context, the AOP is of relevance to the risk assessment of airborne pollutants and other chronically inhaled toxicants which affect vascular endothelial cells. Additionally, acute and chronic exposure to aerosols generated by alternative tobacco products and nicotine delivery devices are poorly understood with respect to their impact on cardiovascular disease risk. Reliable <em>in vitro</em> models and human clinical biomarkers would help to inform regulatory decision-making with respect to understanding the potential cardiovascular health impacts of these novel products.</p>
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  • <!-- Background Section, text as generated by author -->
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  • <h3>Background</h3>
  • <hr>
  • <p>The motivation for the AOP development, a deeper explaination of the underlying biology, and KE/KER assessments can be found in Lowe et al. 2017.</p>
  • <p>Originally, &quot;oxidative stress&quot; was proposed as the MIE for this AOP.&nbsp; Upon discussion with EAGMST, it was suggested to use a term that is more representative of the mechanism of action, hence &quot;peptide oxidation&quot; is proposed instead.&nbsp; All references to &quot;oxidative stress&quot; within this&nbsp;wiki are made within the context that localised conditions conducive to oxidative damage within the vascular endothelium would trigger the revised MIE, and are limited to the peptide targets named within the AOP; namely GSH, GTPCH1, AKT and eNOS&nbsp;(although other redox sensitive peptides&nbsp;are undoubtedly affected also).</p>
  • <p>The mechanisms of potentially deleterious peptide oxidation by ROS/RNS are discussed&nbsp;by Berlett and Stadtman (1997) in the context of health effects.&nbsp; References to &quot;oxidative stress&quot; within this wiki are made with this context in mind.</p>
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  • <!-- AOP summary, includes summary of each of the events associated with this aop -->
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  • <h2>Summary of the AOP</h2>
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  • <!-- stressor table -->
  • <h3>Stressors</h3>
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  • <!-- event table -->
  • <h3>Events</h3>
  • <h3>Molecular Initiating Events (MIE), Key Events (KE), Adverse Outcomes (AO)</h3>
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  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Name</th>
  • <th>Evidence</th>
  • <th>Sequence</th>
  • <th>Type</th>
  • <th>Event ID</th>
  • <th>Title</th>
  • <th>Short name</th>
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  • </thead>
  • <tbody>
  • <tr>
  • <td>Reactive oxygen species</td>
  • <td>Strong</td>
  • <td>1</td>
  • <td>MIE</td>
  • <td>209</td>
  • <td><a href="/events/209">Peptide Oxidation</a></td>
  • <td>Peptide Oxidation</td>
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  • </tbody>
  • </table>
  • </div>
  • <h4>Reactive oxygen species</h4>
  • <p>Compounds or environmental conditions,&nbsp;which generate endothelium-localised ROS <em>in vivo</em> are the primary source of the MIE. &nbsp;Notable examples include:&nbsp;</p>
  • <p>ROS/ROS donors : (Song et al. 2008, van Gorp et al. 1999, van Gorp et al. 2002, Park et al. 2013, Montecinos et al. 2007, Schuppe et al. 1992,&nbsp;</p>
  • <p>Hypoxia/ischaemia : Nozik-Grayck et al. 2014, Zhang et al. 2014, De Pascali et al. 2014,&nbsp;</p>
  • <p>SIN-1 (CAS № 16142-27-1) :&nbsp;&nbsp;Das et al. 2014&nbsp;&nbsp;</p>
  • <p>Heavy Metals (Lead, Cadmium, Mercury)&nbsp;: Vaziri&nbsp;et al. 2001, Wolf et al. 2007</p>
  • <p>Carbonyls (including methylglyoxal,&nbsp;<em>N</em>,<em>N</em>&prime;-bis(2-chloroethyl)-<em>N</em>-nitroso-urea), acrolein)&nbsp;: Morgan et al. 2014, Dhar et al. 2010, Su et al. 2013, Chen et al. 2010, Chen et al. 2011,&nbsp;Michaud et al. 2006, Qin et al. 2016, Zhang et al. 2011</p>
  • <p>Glucose : Zou et al. 2002, Song&nbsp;et al. 2007, Du et al. 2013, Du et al. 2001, Dhar et al. 2010, Su et al. 2008</p>
  • <p>Ultra-fine particulates : Du et al. 2013, Tseng et al. 2016</p>
  • <p>Cigarette smoke (known to contain carbonyls, metals and ROS) :&nbsp;Michaud et al. 2006, Zhang et al. 2006, Talukder et al. 2011</p>
  • <tr><td></td><td></td><td></td><td></td><td></td></tr>
  • <tr>
  • <td>2</td>
  • <td>KE</td>
  • <td>927</td>
  • <td><a href="/events/927">KE1 : S-Glutathionylation, eNOS</a></td>
  • <td>S-Glutathionylation, eNOS</td>
  • </tr>
  • <tr>
  • <td>3</td>
  • <td>KE</td>
  • <td>935</td>
  • <td><a href="/events/935">KE2 : Decrease, GTPCH-1</a></td>
  • <td>Decrease, GTPCH-1</td>
  • </tr>
  • <tr>
  • <td>4</td>
  • <td>KE</td>
  • <td>934</td>
  • <td><a href="/events/934">KE3 : Decrease, Tetrahydrobiopterin</a></td>
  • <td>Decrease, Tetrahydrobiopterin</td>
  • </tr>
  • <tr>
  • <td>5</td>
  • <td>KE</td>
  • <td>932</td>
  • <td><a href="/events/932">KE4 : Uncoupling, eNOS</a></td>
  • <td>Uncoupling, eNOS</td>
  • </tr>
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  • <td>6</td>
  • <td>KE</td>
  • <td>933</td>
  • <td><a href="/events/933">KE6 : Depletion, Nitric Oxide</a></td>
  • <td>Depletion, Nitric Oxide</td>
  • </tr>
  • <tr>
  • <td>7</td>
  • <td>KE</td>
  • <td>937</td>
  • <td><a href="/events/937">KE7 : Impaired, Vasodilation</a></td>
  • <td>Impaired, Vasodilation</td>
  • </tr>
  • <tr>
  • <td>8</td>
  • <td>KE</td>
  • <td>951</td>
  • <td><a href="/events/951">KE8 : Increase, Vascular Resistance</a></td>
  • <td>Increase, Vascular Resistance</td>
  • </tr>
  • <tr>
  • <td>9</td>
  • <td>KE</td>
  • <td>973</td>
  • <td><a href="/events/973">KE5 : Decrease, AKT/eNOS activity</a></td>
  • <td>Decrease, AKT/eNOS activity</td>
  • </tr>
  • <tr><td></td><td></td><td></td><td></td><td></td></tr>
  • <!-- mie table -->
  • <h3>Molecular Initiating Event</h3>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Title</th>
  • <th>Short name</th>
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  • <td><a href="/events/209">Peptide Oxidation</a></td>
  • <td>Peptide Oxidation</td>
  • <td>10</td>
  • <td>AO</td>
  • <td>952</td>
  • <td><a href="/events/952">Hypertension</a></td>
  • <td>Hypertension</td>
  • </tr>
  • </tbody>
  • </table>
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  • <div>
  • <h4><a href="/events/209">209: Peptide Oxidation</a><br></h4>
  • <h5>Short Name: Peptide Oxidation</h5>
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  • <h4>Key Event Component</h4>
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  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
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  • <td>oxidative stress</td>
  • <td></td>
  • <td>increased</td>
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  • </tbody>
  • </table>
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  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <!-- rel table -->
  • <h3>Key Event Relationships</h3>
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  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • <th>Upstream Event</th>
  • <th>Relationship Type</th>
  • <th>Downstream Event</th>
  • <th>Evidence</th>
  • <th>Quantitative Understanding</th>
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  • <td><a href="/aops/27">27: Cholestatic Liver Injury induced by Inhibition of the Bile Salt Export Pump (ABCB11)</a></td>
  • <td>KeyEvent</td>
  • <td><a href="/relationships/980">Peptide Oxidation</a></td>
  • <td>adjacent</td>
  • <td>KE2 : Decrease, GTPCH-1</td>
  • <td>Moderate</td>
  • <td>Low</td>
  • </tr>
  • <tr>
  • <td><a href="/aops/108">108: Inhibition of pyruvate dehydrogenase kinase leading to hepatocellular adenomas and carcinomas (in mouse and rat)</a></td>
  • <td>KeyEvent</td>
  • <td><a href="/relationships/955">KE1 : S-Glutathionylation, eNOS</a></td>
  • <td>adjacent</td>
  • <td>KE4 : Uncoupling, eNOS</td>
  • <td>High</td>
  • <td>Moderate</td>
  • </tr>
  • <tr>
  • <td><a href="/aops/144">144: Lysosomal damage leading to liver inflammation</a></td>
  • <td>KeyEvent</td>
  • <td><a href="/relationships/952">KE2 : Decrease, GTPCH-1</a></td>
  • <td>adjacent</td>
  • <td>KE3 : Decrease, Tetrahydrobiopterin</td>
  • <td>High</td>
  • <td>High</td>
  • </tr>
  • <tr>
  • <td><a href="/aops/149">149: Peptide Oxidation Leading to Hypertension</a></td>
  • <td>MolecularInitiatingEvent</td>
  • <td><a href="/relationships/953">KE3 : Decrease, Tetrahydrobiopterin</a></td>
  • <td>adjacent</td>
  • <td>KE4 : Uncoupling, eNOS</td>
  • <td>High</td>
  • <td>High</td>
  • </tr>
  • <tr>
  • <td><a href="/aops/210">210: Activation of c-Jun N-terminal kinase (JNK) and Forkhead box O (FOXO) and reduction of WNT pathways leading to reproductive failure: Integrated multi-OMICS approach for AOP building</a></td>
  • <td>KeyEvent</td>
  • <td><a href="/relationships/956">KE4 : Uncoupling, eNOS</a></td>
  • <td>adjacent</td>
  • <td>KE6 : Depletion, Nitric Oxide</td>
  • <td>High</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
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  • <!-- loop to find stressors under event -->
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  • <!-- biological organization -->
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  • <h4>Biological Organization</h4>
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  • <tr>
  • <th>Level of Biological Organization</th>
  • <td><a href="/relationships/958">KE6 : Depletion, Nitric Oxide</a></td>
  • <td>adjacent</td>
  • <td>KE7 : Impaired, Vasodilation</td>
  • <td>High</td>
  • <td>Moderate</td>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Molecular</td>
  • <td><a href="/relationships/982">KE7 : Impaired, Vasodilation</a></td>
  • <td>adjacent</td>
  • <td>KE8 : Increase, Vascular Resistance</td>
  • <td>Moderate</td>
  • <td>Low</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
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  • <h4>Cell term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
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  • <tr>
  • <th>Cell term</th>
  • <td><a href="/relationships/983">KE8 : Increase, Vascular Resistance</a></td>
  • <td>adjacent</td>
  • <td>Hypertension</td>
  • <td>Moderate</td>
  • <td>Low</td>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/relationships/988">KE5 : Decrease, AKT/eNOS activity</a></td>
  • <td>adjacent</td>
  • <td>KE6 : Depletion, Nitric Oxide</td>
  • <td>High</td>
  • <td>High</td>
  • </tr>
  • <tr>
  • <td><a href="/relationships/992">Peptide Oxidation</a></td>
  • <td>adjacent</td>
  • <td>KE5 : Decrease, AKT/eNOS activity</td>
  • <td>High</td>
  • <td>Moderate</td>
  • </tr>
  • <tr>
  • <td><a href="/relationships/1698">Peptide Oxidation</a></td>
  • <td>adjacent</td>
  • <td>KE1 : S-Glutathionylation, eNOS</td>
  • <td>Moderate</td>
  • <td>Low</td>
  • </tr>
  • <tr><td></td><td></td><td></td><td></td><td></td></tr>
  • </tbody>
  • </table>
  • </div>
  • <!-- stressor table -->
  • <h3>Stressors</h3>
  • <br>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Name</th>
  • <th>Evidence</th>
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  • <tbody>
  • <tr>
  • <td>eukaryotic cell</td>
  • <td>Reactive oxygen species</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of cell term -->
  • <h4>Reactive oxygen species</h4>
  • <p>Compounds or environmental conditions,&nbsp;which generate endothelium-localised ROS <em>in vivo</em> are the primary source of the MIE. &nbsp;Notable examples include:&nbsp;</p>
  • <p>ROS/ROS donors : (Song et al. 2008, van Gorp et al. 1999, van Gorp et al. 2002, Park et al. 2013, Montecinos et al. 2007, Schuppe et al. 1992,&nbsp;</p>
  • <p>Hypoxia/ischaemia : Nozik-Grayck et al. 2014, Zhang et al. 2014, De Pascali et al. 2014,&nbsp;</p>
  • <p>SIN-1 (CAS № 16142-27-1) :&nbsp;&nbsp;Das et al. 2014&nbsp;&nbsp;</p>
  • <p>Heavy Metals (Lead, Cadmium, Mercury)&nbsp;: Vaziri&nbsp;et al. 2001, Wolf et al. 2007</p>
  • <p>Carbonyls (including methylglyoxal,&nbsp;<em>N</em>,<em>N</em>&prime;-bis(2-chloroethyl)-<em>N</em>-nitroso-urea), acrolein)&nbsp;: Morgan et al. 2014, Dhar et al. 2010, Su et al. 2013, Chen et al. 2010, Chen et al. 2011,&nbsp;Michaud et al. 2006, Qin et al. 2016, Zhang et al. 2011</p>
  • <p>Glucose : Zou et al. 2002, Song&nbsp;et al. 2007, Du et al. 2013, Du et al. 2001, Dhar et al. 2010, Su et al. 2008</p>
  • <p>Ultra-fine particulates : Du et al. 2013, Tseng et al. 2016</p>
  • <p>Cigarette smoke (known to contain carbonyls, metals and ROS) :&nbsp;Michaud et al. 2006, Zhang et al. 2006, Talukder et al. 2011</p>
  • <!-- organ term -->
  • <div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • </div>
  • <!-- end summary -->
  • <h4>Evidence Supporting Applicability of this Event</h4>
  • <br>
  • <!-- Overall assessment section, *** what is included here? *** -->
  • <div id="overall_assessment">
  • <h2>Overall Assessment of the AOP</h2>
  • <hr>
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <h3>Domain of Applicability</h3>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>human</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>rodents</td>
  • <td>rodents</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=0" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>human and other cells in culture</td>
  • <td>human and other cells in culture</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=0" , target="_blank">NCBI</a>
  • </td>
  • <td>All life stages</td>
  • <td></td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>human</td>
  • <td>Homo sapiens</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>mouse</td>
  • <td>Mus musculus</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>rat</td>
  • <td>Rattus norvegicus</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>cow</td>
  • <td>Bos taurus</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td></td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <div>
  • <p>The concentrations of GSH and GSSG have been shown in tissues of human and laboratory animals, including rats, mice and cows (Chen et al., 2010; Giustarini et al., 2013).</p>
  • <br>
  • </div>
  • <!-- event text -->
  • <h4>How this Key Event Works</h4>
  • <p>Oxidative stress corresponds to an imbalance between the rate of oxidant production and that of their degradation. The term oxidative stress indicates the outcome of oxidative damage to biologically relevant macromolecules such as nucleic acids, proteins, lipids and carbohydrates. This occurs when oxidative stress-related molecules, generated in the extracellular environment or within the cell, exceed cellular antioxidant defenses. Major reactive oxygen species (ROS), such as hydrogen peroxide (H2O2) and superoxide anion, as well as 4-hydroxy- 2,3-nonenal (HNE) and related 4-hydroxy-2,3-alkenals (HAKs), major aldehydic end-products of lipid peroxidation, can act as potential mediators able to affect signal transduction pathways as well as the proliferative and functional response of target cells. H2O2 and superoxide anion may be also generated as molecular messengers within the cell as part of the cellular response to defined growth factors, cytokines and other mediators. The final consequence at tissue, cellular and molecular level is primarily affected by the steady state concentration of oxidative stress-related molecules. The main biological targets of free radicals are proteins, lipids and DNA.</p>
  • <p>Major consequences of reaction of ROS, HAKs and NO with biologically relevant macromolecules that can mediate pathophysiological effects:</p>
  • <p>ROS: DNA: oxidation, strand breaks, genotoxicity Proteins: oxidation, fragmentation, formation of carbonyls Lipids: lipid peroxidation and degradation</p>
  • <p><a href="#Life_Stage_Applicability">Life Stage Applicability</a>, <a href="#Taxonomic_Applicability"> Taxonomic Applicability</a>, <a href="#Sex_Applicability"> Sex Applicability</a><br />
  • <em>Elaborate on the domains of applicability listed in the summary section above. Specifically, provide the literature supporting, or excluding, certain domains. </em></p>
  • <p>HAKs: DNA: adducts (low doses), strand breaks, genotoxicity (high doses) Proteins: adducts (Michael type reactions on Lys, Cys and His residues)</p>
  • <p>This proposed AOP is applicable to males and females, however a single study did suggest that KE responses may vary with sex (Scotland et al. 2005).&nbsp;</p>
  • <p>NO: DNA: oxidation, strand breaks Proteins: oxidation, nitrosation, nitration (nytrosylation of tyrosine) Lipids: lipid peroxidation and degradation</p>
  • <p>The AOP is relevant for all life stages, as vascular oxidative stress (a common cause of the MIE)&nbsp;can occur at all life stages. &nbsp;However, the AO via this mechanism is likely limited to adults, due to a loss of vaso-protective mechanisms with age (a major&nbsp;risk factor) and cardiovascular re-modelling over time. &nbsp;Hypertension is known to occur in adolescents and children, however the condition can be due to a variety of causes, such as renal disease and cardiovascular complications in early life.&nbsp; For example, persistant pulmonary hypertension is&nbsp;a consequence of failed pulmonary vascular transition at birth and leads to pulmonary&nbsp;hypertension&nbsp;with shunting of deoxygenated blood across the ductus arteriosus&nbsp;and foramen ovale,&nbsp;resulting in severe hypoxemia. &nbsp;The condition is improved by administration of nitric oxide by inhalation (Lai et al. 2017). &nbsp;While the initial cause of the hypertension is not vascular oxidative stress, ROS-mediated perturbations of the vascular endothelium, which arise as a result of the&nbsp;hypoxic conditions, contribute to the development of hypertension, and may have health implications for the neonate later in&nbsp;life (de Wijs-Meijler et al. 2017). &nbsp;Therefore, the physiology described in the AOP is relevant to the condition, but not strictly causative.</p>
  • <p>Continued oxidative stress can lead to chronic inflammation. Oxidative stress can activate a variety of transcription factors including NF-&kappa;B, AP-1, p53, HIF-1&alpha;, PPAR-&gamma;, &beta;-catenin/Wnt, and Nrf2. Activation of these transcription factors can lead to the expression of over 500 different genes, including those for growth factors, inflammatory cytokines and chemokines, which can activate inflammatory pathways. <sup><a href="#cite_note-1">[1]</a></sup> <sup><a href="#cite_note-2">[2]</a></sup> <sup><a href="#cite_note-3">[3]</a></sup></p>
  • <p>The KEs in this AOP are well-documented and well-studied in humans, cows and rodents. &nbsp;Similar outcomes are observed across these species following chemical exposure and other phenomena which cause vascular oxidative stress (e.g. hypoxia, ischaemia/reperfusion).</p>
  • <p>Glutathione (GSH) oxidation refers to the conversion of reduced glutathione to its oxidized form glutathione disulfide (GSSG) in the presence of oxidative species. GSH plays an important role as an anti-oxidant in regulating cellular redox homeostasis, and is mainly present in the cell as the reduced form (98%). Deficiency in GSH or a decrease in GSH/GSSG ratio results in decreased anti-oxidant function and increased susceptibility to oxidative stress, thus making it a marker of cellular redox status. An imbalance in GSH/GSSG ratio has been implicated in the onset and progression of human diseases, such as neurodegenerative diseases, cancers, pulmonary diseases and cardiovascular diseases (Ballatori et al., 2009; Kalinina et al., 2014)</p>
  • <p>The specific amino-acid residues involved in post-translational modification of eNOS do differ across species, however the functional effect is similar. &nbsp;For example, human eNOS is phosphorylated by AKT at serine residue 1177&nbsp;(Reviewed by Heiss et al. 2014), whereas bovine eNOS is phosphoryated by AKT at serine 1179 (Chen et al. 2017).</p>
  • <br>
  • <h4>How it is Measured or Detected</h4>
  • <p>&nbsp;</p>
  • <p><em><strong>measuring oxidative stress</strong> </em></p>
  • <p><em>Agents for <strong>ROS detection</strong> are primarily fluorescence based, but recently luminescent based detections have been introduced. The biggest difficulty reported with much of the cellular ROS research has been with the lack of reporter agents specific for discrete molecules. ROS moieties by their nature are reactive with a number of different molecules; as such designing reporter agents has been difficult. With more specific chemistries, particularly for hydrogen peroxide, the specific mechanisms for regulation will be elucidated. </em></p>
  • <p><em><strong>Reduced glutathione</strong> (GSH) is regenerated from its oxidized form (GSSH) by the action of an NADPH dependent reductase GSSH + NADPH + H+ &agrave; 2 GSH + NADP+ Due to the rapid nature of the reduction of GSSH relative to its synthesis or secretion, the ratio of GSH to GSSH is a good indicator of oxidative stress within cells. GSH and GSSH levels can be determined by HPLC, capillary electrophoresis, or biochemically in microplates. Several different assays have been designed to measure glutathione in samples. By using a luciferin derivative in conjunction with glutathione S-transferase enzyme the amount of GSH would be proportional to the luminescent signal generated when luciferase is added in a subsequent step. Total glutathione can be determined colorimetrically by reacting GSH with DTNB (Ellman&rsquo;s reagent) in the presence of glutathione reductase. Glutathione reductase reduces GSSH to GSH, which then reacts with DTNB to produce a yellow colored 5-thio-2-nitrobenzoic acid (TNB), which absorbs at 412 nm. </em></p>
  • <p><em><strong>Lipid peroxidation</strong> is one of the most widely used indicators of free radical formation, a key indicator of oxidative stress. Measurement of lipid peroxidation has historically relied on the detection of thiobarbituric acid (TBA) reactive compounds such as malondialdehyde generated from the decomposition of lipid peroxidation products. While this method is controversial in that it is quite sensitive, but not necessarily specific to MDA, it remains the most widely used means to determine lipid peroxidation. This reaction, which takes place under acidic conditions at 90-100&ordm;C, results in an adduct that can be measured colorimetrically at 532 nm or by fluorescence using a 530 nm excitation wavelength and a 550 nm emission wavelength. A number of commercial assay kits are available for this assay using absorbance or fluorescence detection technologies. The formation of F2-like prostanoid derivatives of arachidonic acid, termed F2-isoprostanes (IsoP) has been shown to be specific for lipid peroxidation. Unlike the TBA assay, measurement of IsoP appears to be specific to lipid peroxides, they are stable and are not produced by any enzymatic pathway making interpretation easier. There have been a number of commercial ELISA kits developed for IsoPs, but interfering agents in samples requires partial purification of samples prior to running the assay. The only reliable means for detection is through the use of GC/MS, which makes it expensive and limits throughput. </em></p>
  • <h3>Essentiality of the Key Events</h3>
  • <p><a href="#Molecular_Initiating_Event">Molecular Initiating Event Summary</a>, <a href="#Key_Events"> Key Event Summary</a><br />
  • <em>Provide an overall assessment of the essentiality for the key events in the AOP. Support calls for individual key events can be included in the molecular initiating event, key event, and adverse outcome tables above. </em></p>
  • <p><em><strong>Superoxide detection</strong> is based on the interaction of superoxide with some other compound to create a measurable result. The reduction of ferricytochrome c to ferrocytochrome c has been used in a number of situations to assess the rate of superoxide formation. While not completely specific for superoxide this reaction can be monitored colorimetrically at 550 nm. Chemiluminescent reactions have been used for their potential increase in sensitivity over absorbance-based detection methods. The most widely used chemiluminescent substrate is Lucigenin, but this compound has a propensity for redox cycling, which has raised doubts about its use in determining quantitative rates of superoxide production. Coelenterazine has also been used as a chemiluminescent substrate. Hydrocyanine dyes are fluorogenic sensors for superoxide and hydroxyl radical. These dyes are synthesized by reducing the iminium cation of the cyanine (Cy) dyes with sodium borohydride. While weakly fluorescent, upon oxidation their fluorescence intensity increases 100 fold. In addition to being fluorescent, oxidation also converts the molecule from being membrane permeable to an ionic impermeable moiety. The most characterized of these probes are Hydro-Cy3 and Hydro-Cy5. </em></p>
  • <p>The essentiality for this AOP is strong since there is direct evidence from multiple experimental studies showing that downstream key events can be prevented or inhibited if an upstream key event is blocked. An increase in intracellular glutathione, rather than the oxidized form, prevented S-glutathionylation (De Pascali et al., 2014), and inhibiting S-glutathionylation attenuated ultrafine particle-induced reduction in NO production (Du et al., 2013). In addition, increasing GTPCH-1, BH4 and Akt/eNOS activity increased eNOS activity and NO production but decreased superoxide generation (Fulton et al., 1999; Alp et al., 2003; Carnicer et al., 2012; Antoniades et al., 2011; Chen et al., 2011; De Pascali et al., 2014; Landmessar et al., 2003; Shinozaki et al., 2000; Ozaki et al., 2002). Infusion of NO donor sodium nitroprusside reduced vascular resistance, while sodium nitrite increased vasodilation (Eugene, 2016; Sindlier et al., 2014).</p>
  • <p><em><strong>Hydrogen peroxide</strong> (H2O2) is the most important ROS in regards to mitogenic stimulation or cell cycle regulation. There are a number of fluorogenic substrates, which serve as hydrogen donors that have been used in conjunction with horseradish peroxidase (HRP) enzyme to produce intensely fluorescent products. The more commonly used substrates include diacetyldichloro-fluorescein, homovanillic acid, and Amplex&reg; Red. In these examples, increasing amounts of H2O2 form increasing amounts of fluorescent product. </em></p>
  • <p>The essentiality of increased vascular resistance to hypertension is moderate due to the involvement of the heart in regulating blood pressure. &nbsp;While increased vascular resistance is a hallmark of essential hypertension, Mayet and Hughes (2003) discussed the roles of the heart and vascular resistance and concluded that both are of importance, and should not be viewed in isolation. &nbsp;Their conclusion is drawn from observations in younger adults with elevated blood pressure, who have a normal vascular resistance, but higher cardiac index. &nbsp;Over time, such adults often go on to change to a phenotype with a lower cardiac index and high vascular resistance and a diagnosis of chronic hypertension. &nbsp;Such changes are likely due to vascular remodelling processes with advancing age, however the specifics of this process are not well understood.</p>
  • <p><em><strong>Nitric Oxide</strong> The free radical nitric oxide (&bull;NO) is produced by a number of different cell types with a variety of biological functions. Regardless of the source or role, the free radical &bull;NO has a very short half life (t&frac12;= 4 seconds), reacting with several different molecules normally present to form either nitrate (NO3-) or nitrite (NO2-) A commonly used method for the indirect determination of &bull;NO is the determination of its composition products nitrate and nitrite colorimetrically. This reaction requires that nitrate (NO3) first be reduced to nitrite (NO2), typically by the action of nitrate reductase. Subsequent determination of nitrite by a two step process provides information on the &ldquo;total&rdquo; of nitrate and nitrite. In the presence of hydrogen ions nitrite forms nitrous acid, which reacts with sulfanilamide to produce a diazonium ion. This then coupled to N-(1-napthyl) ethylenediamine to form the chromophore which absorbs at 543 nm. Nitrite only determinations can then be made in a parallel assay where the samples were not reduced prior to the colorimetric assay. Actual nitrate levels are then calculated by the subtraction of nitrite levels from the total. <sup><a href="#cite_note-4">[4]</a></sup> </em></p>
  • <p>&nbsp;</p>
  • <p>&nbsp;</p>
  • <br>
  • <h4>References</h4>
  • <p>Ballatori, N., Krance, S.M., Notenboom, S., Shi, S., Tieu, K., and Hammond, C.L. (2009). Glutathione dysregulation and the etiology and progression of human diseases. Biol. Chem. 390, 191&ndash;214.</p>
  • <p>Chen, C.-A., Wang, T.-Y., Varadharaj, S., Reyes, L.A., Hemann, C., Talukder, M.A.H., Chen, Y.-R., Druhan, L.J., and Zweier, J.L. (2010). S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature 468, 1115&ndash;1118.</p>
  • <p>&nbsp;</p>
  • <p>Giustarini, D., Dalle-Donne, I., Milzani, A., Fanti, P., and Rossi, R. (2013). Analysis of GSH and GSSG after derivatization with N-ethylmaleimide. Nat. Protoc. 8, 1660&ndash;1669.</p>
  • <p>&nbsp;</p>
  • <p>Held P., 2010 Biotek, Measurement of ROS in Cells, <a class="external free" href="http://www.biotek.com/assets/tech_resources/ROS%20Application%20Guide.pdf" rel="nofollow" target="_blank">http://www.biotek.com/assets/tech_resources/ROS%20Application%20Guide.pdf</a></p>
  • <table border="0" cellpadding="0" cellspacing="0" style="width:1322px">
  • <tbody>
  • </tbody>
  • </table>
  • <p>Kalinina, E.V., Chernov, N.N., and Novichkova, M.D. (2014). Role of glutathione, glutathione transferase, and glutaredoxin in regulation of redox-dependent processes. Biochem. Biokhimii︠a︡ 79, 1562&ndash;1583.</p>
  • <table border="0" cellpadding="0" cellspacing="0" style="width:1132px">
  • <tbody>
  • <tr>
  • <td rowspan="2" style="height:140px; text-align:center; width:128px"><strong>Support for Essentiality of KEs</strong></td>
  • <td rowspan="2" style="text-align:center; width:203px"><strong>KE Description</strong></td>
  • <td style="text-align:center; width:148px"><strong>Defining Question</strong></td>
  • <td style="text-align:center; width:205px"><strong>High</strong></td>
  • <td style="text-align:center; width:203px"><strong>Moderate</strong></td>
  • <td style="text-align:center; width:245px"><strong>Low</strong></td>
  • </tr>
  • <tr>
  • <td style="height:108px; text-align:center; width:148px"><strong>Are downstream KEs and/or the AO prevented if an upstream KE is blocked?</strong></td>
  • <td style="text-align:center; width:205px"><strong>Direct evidence from experimental studies illustrating essentiality for at least one of the important KEs.</strong></td>
  • <td style="text-align:center; width:203px"><strong>Indirect evidence that sufficient modification of an expected modulating factor attenuates or augments a KE.</strong></td>
  • <td style="text-align:center; width:245px"><strong>No or contradictory experimental evidence of the essentiality of any of the KEs.</strong></td>
  • </tr>
  • <tr>
  • <td style="height:104px; width:128px">Peptide Oxidation (MIE)</td>
  • <td style="width:203px">Generation of localised ROS and other oxidative species in the vascular endothelium leads to oxidation of amino acid residues of peptides.&nbsp;&nbsp;</td>
  • <td style="width:148px">&nbsp;</td>
  • <td colspan="3" style="width:653px">&nbsp;</td>
  • </tr>
  • <tr>
  • <td style="height:125px; width:128px">S-glutathionylation of eNOS (KE1)</td>
  • <td style="width:203px">The oxidation of GSH into GSSG, elevates levels of GSSG, which then covalently bind to critical serine residues on the eNOS enzyme&nbsp;</td>
  • <td style="text-align:center; width:148px">High</td>
  • <td colspan="3" style="width:653px"><span style="font-size:14px">Protein S-Glutathionylation can only occur following covalent binding of GSSG.&nbsp; GSSG commonly binds to protein thiol residues, which can affect protein function&nbsp;(Bendall et al 2014).&nbsp;S-glutathionylation of eNOS is observed under conditions of oxidative stress, at cysteine residues 689 and 908 leading to eNOS uncoupling&nbsp;(Chen et al. 2010).&nbsp; Application of Glutaredoxin-1 or dithiothreitol removes S-glutathionylation and restores eNOS function&nbsp;(Chen et al. 2013, Jayaram et al. 2015) &nbsp;Essentiality rating is high due to direct experimental evidence showing removal of S-glutathionylation of eNOS restores NO production.</span></td>
  • </tr>
  • <tr>
  • <td style="height:105px; width:128px">Loss of GTPCH-1 (KE2)</td>
  • <td style="width:203px">Oxidative damage to GTPCH-1 has been shown to lead to reduced expression/activity of the enzyme</td>
  • <td style="text-align:center; width:148px">High</td>
  • <td colspan="3" style="width:653px">
  • <p>&nbsp;</p>
  • <p>Kamencic, H., Lyon, A., Paterson, P.G., and Juurlink, B.H. (2000). Monochlorobimane fluorometric method to measure tissue glutathione. Anal. Biochem. 286, 35&ndash;37.</p>
  • <p><span style="font-size:14px">Numerous studies have demonstrated that deletion of GTPCH-1 led to the deficiency of BH4 in bovine and murine endothelial cells (Tatham et al. 2009, Crabtree et al. 2009, Adlam et al. 2012)&nbsp;and in knockout mice&nbsp;(Chen et al. 2011).&nbsp; Gene silencing and GTPCH-1 inhibition lead to elevated blood pressure in experimental animals (Wang et al. 2008, Mitchell et al. 2003).&nbsp; Essentiality rating is high due to sunstantial experimental evidence of GTPCH-1 blocking on downstream KEs.</span></p>
  • </td>
  • </tr>
  • <tr>
  • <td style="height:100px; width:128px">Decreased BH4 (KE3)</td>
  • <td style="width:203px">Oxidation of BH4 itself to BH2, and/or reduced bioavailability of BH4 by a reduction in de novo synthesis by GTPCH-1.</td>
  • <td style="text-align:center; width:148px">High</td>
  • <td colspan="3" style="width:653px">
  • <p>&nbsp;</p>
  • <p>Parola, M. and Robino, G. (2001). Oxidative stress-related molecules and liver fibrosis. J Hepatol. 35, 297-306</p>
  • <p><span style="font-size:14px">Depletion of BH4 has been extensively reported to uncouple eNOS, decrease NO bioavailability and increase superoxide production.&nbsp; Application of exogenous BH4 has been reported to reverse this phenomenon (De Pascali et al. 2014, Zweier et al. 2011, Dumitrescu et al. 2007, Tiefenbacher et al. 1996,&nbsp;V&aacute;squez-Vivar et al. 2002)&nbsp;&nbsp; Essentiality rating is high due to extensive direct experimental evidence.</span></p>
  • </td>
  • </tr>
  • <tr>
  • <td style="height:112px; width:128px">eNOS Uncoupling (KE4)</td>
  • <td style="width:203px">eNOS uncoupling is characterised by a loss of NO production and superoxide production as eNOS dimerisation is lost</td>
  • <td style="text-align:center; width:148px">High</td>
  • <td colspan="3" style="width:653px"><span style="font-size:14px">eNOS uncoupling leads to reduction in NO bioavailability.&nbsp; Reversal of eNOS uncoupling is achieved by supplementation with BH4 or removal of eNOS S-glutathionylation (as described earlier), which restores NO production (Du et al. 2013b,&nbsp;Nozik-Grayck et al. 2014,&nbsp;Talukder&nbsp;et al. 2011, Su et al. 2013).&nbsp; Essentiality rating is high due to direct experimental evidence.</span></td>
  • </tr>
  • <tr>
  • <td style="height:142px; width:128px">Loss of AKT/eNOS (KE5)</td>
  • <td style="width:203px">In humans, AKT phosphorylates eNOS at serine residue 1177, leading to eNOS activation and NO production.&nbsp; Loss of AKT/eNOS function depletes NO bioavailability.</td>
  • <td style="text-align:center">High</td>
  • <td colspan="3" style="width:653px">
  • <p><span style="font-size:14px">eNOS knockout mice are routinely used as models of hypertension&nbsp;(Huang&nbsp;2000).&nbsp; Aortic rings from eNOS knockout mice do not relax following application of acetylcholine, but do relax upon application of NO donor sodium nitroprusside (Huang et al. 1995).&nbsp; Many animal studies demonstrated that inhibition of NO via eNOS inhibitors impaired endothelium-dependent vasodilation (Li et al. 2007, Paulis et al. 2008, Luo et al. 2000,&nbsp;S&eacute;lley&nbsp;et al. 2014).&nbsp; Inhibition of Akt or mutant eNOS attenuated eNOS phosphorylation in human and bovine cells, resulting in decreased NO bioavailability (Michaud et al. 2006, Dhar et al. 2010, Dimmeler et al. 1999, Fulton et al. 1999, Das et al. 2014, Uruno et al. 2005).&nbsp; Essentiality rating is high due to extensive direct experimental evidence.</span></p>
  • </td>
  • </tr>
  • <tr>
  • <td style="height:187px; width:128px">NO Depletion (KE6)</td>
  • <td style="width:203px">Nitric oxide is a potent vasodilator radical released by endothelial (cell) eNOS.&nbsp; NO signalling leads to potassium ion efflux and hyperpolarization of vascular smooth muscle cells and blood vessel relaxation.&nbsp; Reduction in NO bioavailability blunts this response</td>
  • <td style="text-align:center">Moderate</td>
  • <td colspan="3" style="width:653px">
  • <p>&nbsp;</p>
  • <p>Reuter S. et al., (2010) Oxidative stress, inflammation, and cancer: how are they linked? Free Radic Biol Med.49, 1603-1616</p>
  • <p><span style="font-size:14px">Depletion of NO bioavailability by pharmacological blockade of AKT and eNOS is widely reported to shift vascular tone towards a more vasoconstrictive phenotype, leading to hypertension (Li et al. 2007, Paulis et al. 2008, Scotland et al. 2005, Luo et al. 2000,&nbsp;S&eacute;lley&nbsp;et al. 2014, Haynes et al. 1993).&nbsp; However gender differences in experimental animals were reported to influence vasodilation by different mechanisms&nbsp;(Scotland et al. 2005).&nbsp; Furthermore, compensatory mechanisms affecting vascular tone are evident when bioavailability of NO is decreased (Brandes 2014, Durand et al. 2013).&nbsp;&nbsp; Essentiality rating is moderate due to extensive evidence from pharmacological blocking studies in humans and animals which show impairment of vasodilation and elevated BP.&nbsp; However, compensatory effects of other biological mechanisms on downstream KEs are evident as are gender differences in experimental animals.</span></p>
  • </td>
  • </tr>
  • <tr>
  • <td style="height:167px; width:128px">Impaired vasodilation (KE7)</td>
  • <td style="width:203px">
  • <p>&nbsp;</p>
  • <p>S&aacute;nchez-Valle V. et al., (2012) Role of oxidative stress and molecular changes in liver fibrosis: a review. Curr Med Chem. 19, 4850-4860</p>
  • <p>Vasodilation is mediated by vascular smooth muscle cells and characterized by potassium ion efflux and hyperpolarization of the tissue in response to signalling from the nervous system and chemical mediators released by the vascular endothelium</p>
  • </td>
  • <td style="text-align:center; width:148px">Moderate</td>
  • <td colspan="3" style="width:653px"><span style="font-size:14px">It is well accepted that vasodilation and systemic vascular resistance (SVR) are negatively correlated.&nbsp; Blood flow volume is increased when blood vessels dilate due to decreased vascular resistance (Siddiqui 2011). When vasodilation is impaired as a result of NO depletion or blockade of potassium channels, vascular stiffness and SVR increase (Berg et al. 2011, Brett et al. 1998, Dessey et al. 2004, Li et al. 2007, McVeigh et al. 2001, Paulis et al. 2008, Wilkinson et al. 2002).&nbsp; Essentiality rating is high due to extensive direct experimental evidence.</span></td>
  • </tr>
  • <tr>
  • <td style="height:116px; width:128px">Increased vascular resistance (KE8)</td>
  • <td style="width:203px">Increases in vascular tone as a result of impaired vasodilation.&nbsp; Commonly referred to as systemic vascular resistance (SVR) or total peripheral resistance (TPR).</td>
  • <td style="text-align:center; width:148px">Moderate</td>
  • <td colspan="3" style="width:653px"><span style="font-size:14px">It is well established that increased SVR (TPR), increased vascular stiffness and increased vascular reactivity contribute to hypertension (Brandes 2014,&nbsp;Fo&euml;x&nbsp;and Sear 2004, Mayet and Hughes 2003). In patients with hypertension, SVR was elevated in approximately 66% of enrolled patients (Chan et al. 2016).&nbsp; However, due to the critical role of cardiac output in blood pressure regulation (Brandes 2014, Mayet and Hughes 2003), and other compensatory mechanisms described earlier, the essentiality for the AO is moderate.</span></td>
  • </tr>
  • <tr>
  • <td style="height:89px; width:128px">Hypertension (AO)</td>
  • <td style="width:203px">Chronic elevation of systolic and/or diastolic blood pressure in systemic circulation or localised organs</td>
  • <td style="width:148px">&nbsp;</td>
  • <td colspan="3" style="width:653px">&nbsp;</td>
  • </tr>
  • </tbody>
  • </table>
  • <p>Tipple, T.E., and Rogers, L.K. (2012). Methods for the Determination of Plasma or Tissue Glutathione Levels. Methods Mol. Biol. Clifton NJ 889, 315&ndash;324</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <p>&nbsp;</p>
  • <!-- ke table -->
  • <h3>Key Events</h3>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Title</th>
  • <th>Short name</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/events/927">KE1 : S-Glutathionylation, eNOS</a></td>
  • <td>S-Glutathionylation, eNOS</td>
  • </tr>
  • <tr>
  • <td><a href="/events/935">KE2 : Decrease, GTPCH-1</a></td>
  • <td>Decrease, GTPCH-1</td>
  • </tr>
  • <tr>
  • <td><a href="/events/934">KE3 : Decrease, Tetrahydrobiopterin</a></td>
  • <td>Decrease, Tetrahydrobiopterin</td>
  • </tr>
  • <tr>
  • <td><a href="/events/932">KE4 : Uncoupling, eNOS</a></td>
  • <td>Uncoupling, eNOS</td>
  • </tr>
  • <tr>
  • <td><a href="/events/933">KE6 : Depletion, Nitric Oxide</a></td>
  • <td>Depletion, Nitric Oxide</td>
  • </tr>
  • <tr>
  • <td><a href="/events/937">KE7 : Impaired, Vasodilation</a></td>
  • <td>Impaired, Vasodilation</td>
  • </tr>
  • <tr>
  • <td><a href="/events/951">KE8 : Increase, Vascular Resistance</a></td>
  • <td>Increase, Vascular Resistance</td>
  • </tr>
  • <tr>
  • <td><a href="/events/973">KE5 : Decrease, AKT/eNOS activity</a></td>
  • <td>Decrease, AKT/eNOS activity</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <div>
  • <h4><a href="/events/927">927: KE1 : S-Glutathionylation, eNOS</a><br></h4>
  • <h5>Short Name: S-Glutathionylation, eNOS</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>protein glutathionylation</td>
  • <td>nitric oxide synthase, endothelial</td>
  • <td>increased</td>
  • </tr>
  • <tr>
  • <td>protein glutathionylation</td>
  • <td>cysteine residue</td>
  • <td>increased</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">149: Peptide Oxidation Leading to Hypertension</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • <h4>Stressors</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Name</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Reactive oxygen species</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Organization</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Molecular</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • <h4>Cell term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Cell term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>endothelial cell of vascular tree</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h4>Evidence Supporting Applicability of this Event</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Weak</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Weak</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <div>
  • <p>S-glutathionylation of eNOS has been demonstrated in humans, cows, mice and rats (Chen et al., 2010; De Pascali et al., 2014; Du et al., 2013).</p>
  • <br>
  • </div>
  • <!-- event text -->
  • <h4>How this Key Event Works</h4>
  • <p>S-glutathionylation is a redox-dependent, reversible post-translational modification that is involved in the regulation of various regulatory, structural, and metabolic proteins (Pastore and Piemonte, 2013). Under oxidative stress, S-glutathionylation targets cysteine residues of a protein and adds glutathione through thiol-disulfide exchange with oxidized glutathione (GSSG) or reaction of oxidant-induced protein thiyl radicals with reduced glutathione (Chen et al., 2010, 2011, Schuppe et al. 1992). Endothelial nitric oxide synthase (eNOS) regulates vascular function by generating nitric oxide which is involved in endothelium-dependent relaxation, and control of blood pressure and vascular tone. It has been shown that cysteine residues are important for the maintenance of normal eNOS function. Under oxidative stress, S-glutathionylation of eNOS was induced by GSSG at residue sites Cys 689 and Cys 908, resulting in a decrease in eNOS activity and an increase in superoxide generation, also known as eNOS uncoupling. Furthermore, eNOS S-glutathionylation was shown to be abundant in the vessel walls of spontaneously hypertension rats (SHRs), in contrast to non-hypertensive rats. &nbsp;&nbsp;SHRs demonstrated impaired endothelium-dependent vasodilation, which was reversible upon administration of the reducing agent,&nbsp;dithiothreitol (Chen et al. 2010). &nbsp;Similarly in human aortic endothial cells, exposure to ultrafine particles caused a decrease in NO production in a dose-depedent manner. &nbsp;This was shown to be prevented upon over-expression of glutaredoxin-1, which inhibits eNOS S-glutathionylation (Du et al. 2013).</p>
  • <br>
  • <h4>How it is Measured or Detected</h4>
  • <p>There are four general approaches to detect protein S-glutathionylation (Pastore and Piemonte, 2013).</p>
  • <ol>
  • <li><strong>Quantification of Total S-Glutathionylated Proteins</strong>: Use sample lysis or homogenization in non-reducing buffer containing N-ethylmaleimide to eliminate thiols, followed by protein precipitation, reduction of gluthionyl-protein adducts, and derivatization of protein thiols or free glutathione with fluorescence probes. Fluorescence can be measured by fluorometric analysis with or without prior HPLC separation. This method allows for quantification of glutathionylated proteins but cannot detect glutathione adducts on specific proteins.</li>
  • <li><strong>Labeling of Glutathione</strong>: Use <sup>35</sup>S-cysteine radiolabeling or biotin labeling to detect glutathione adducts on S-thiolated proteins.</li>
  • <li><strong>Use of Anti-Glutathione Antibodies</strong>: Use commercially available anti-glutathione to detect glutathionylated proteins by Western blots, immunoprecipitation or immunocytolocalization. This method is useful for analysis of individual proteins like eNOS but not for large-scale detection of glutathionylated proteins.</li>
  • <li><strong>Top-Down Proteomic Approach</strong>: Use liquid chromatography-coupled mass spectrometry to identify S-glutathionylated proteins on whole protein extract from cells without using labeling or anti-glutathione antibody.</li>
  • </ol>
  • <br>
  • <h4>References</h4>
  • <p>Chen, C.-A., Wang, T.-Y., Varadharaj, S., Reyes, L.A., Hemann, C., Talukder, M.A.H., Chen, Y.-R., Druhan, L.J., and Zweier, J.L. (2010). S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature 468, 1115&ndash;1118.</p>
  • <p>Chen CA,&nbsp;Lin CH,&nbsp;Druhan LJ,&nbsp;Wang TY,&nbsp;Chen YR,&nbsp;Zweier JL. &nbsp;Superoxide induces endothelial nitric-oxide synthase protein thiyl radical formation, a novel mechanism regulating&nbsp;eNOS&nbsp;function and coupling. &nbsp;J Biol Chem.&nbsp;2011 286(33):29098-107.</p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p>Du, Y., Navab, M., Shen, M., Hill, J., Pakbin, P., Sioutas, C., Hsiai, T.K., and Li, R. (2013). Ambient ultrafine particles reduce endothelial nitric oxide production via S-glutathionylation of eNOS. Biochem. Biophys. Res. Commun. 436, 462&ndash;466.</p>
  • <p>Pastore, A., and Piemonte, F. (2013). Protein glutathionylation in cardiovascular diseases. Int. J. Mol. Sci. 14, 20845&ndash;20876.</p>
  • <p>Schuppe I, Mold&eacute;us P, and Cotgreave IA. Protein-specific S-thiolation in human endothelial cells during oxidative stress. (1992) Biochem. Pharmacol. 44: 1757&ndash;1764.</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <div>
  • <div>
  • <h4><a href="/events/935">935: KE2 : Decrease, GTPCH-1</a><br></h4>
  • <h5>Short Name: Decrease, GTPCH-1</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>proteasome complex disassembly</td>
  • <td>GTP cyclohydrolase 1</td>
  • <td>decreased</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">149: Peptide Oxidation Leading to Hypertension</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Organization</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Cellular</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • <h4>Cell term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Cell term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>endothelial cell of vascular tree</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h4>Evidence Supporting Applicability of this Event</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Weak</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <div>
  • <p>Several studies showed decreased GTPCH-1 activity and/or protein expression in cardiac reperfusion patients, bovine endothelial cells, a mouse model of diabetes and a rat model of hypertension (Cervantes-P&eacute;rez et al., 2012; Abdelghany et al., 2017; Jayaram et al., 2015; Zhao et al., 2013).</p>
  • <p>Furthermore, mice deficient in GTPCH-1&nbsp;demonstrate decreased BH4 bioavailability, increased eNOS uncoupling, pulmonary vascular resistance and pulmonary hypertension (Belik et al. 2011, Nandi et al. 2005, Khoo et al. 2005).</p>
  • <br>
  • </div>
  • <!-- event text -->
  • <h4>How this Key Event Works</h4>
  • <p>Guanosine triphosphate cyclohydrolase-1 (GTPCH-1)&nbsp;is the rate-limiting enzyme in the de novo biosynthesis of tetrahydrobiopterin (BH4), an essential cofactor for endothelial nitric oxide synthase (eNOS) and nitric oxide generation (Wang et al., 2008). GTPCH-1 catalyzes the rearrangement of GTP to 7-dihydroneopterin triphosphate, which is converted to BH4 through sequential actions of pyruvoyl tetrahydrobiopterin synthase and sepiapterin reductase. GTPCH-1 activity is regulated in a negative feedback by levels of BH4 which promotes binding of GTPCH-1 with its inhibitor GTPCH feedback regulatory protein (GFRP), but phosphorylation of GTPCH-1 reduces its binding to GFRP and prevents this negative feedback (Chen et al., 2011). &nbsp;Loss or inactivation of GTPCH-1 results in decreased BH4 levels, which causes eNOS uncoupling.</p>
  • <br>
  • <h4>How it is Measured or Detected</h4>
  • <p>The activity of GTPCH-1 can be detected through the quantification of neopterin by high-performance liquid chromatography (HPLC) after the conversion of enzymatically formed dihydroneopterin triphosphate into neopterin by sequential iodine oxidation and dephosphorylation.</p>
  • <br>
  • <h4>References</h4>
  • <p>AbdelGhany, T., Ismail, R., Elmahdy, M., Mansoor F, Zweier J, Lowe, F., and Zweier, JL. (2017). Cigarette Smoke Constituents Cause Endothelial Nitric Oxide Synthase Dysfunction and Uncoupling due to Depletion of Tetrahydrobiopterin with Degradation of GTP Cyclohydrolase.&nbsp;&nbsp;Nitric Oxide&nbsp;(Under review).</p>
  • <p>Belik J,&nbsp;McIntyre BA,&nbsp;Enomoto M,&nbsp;Pan J,&nbsp;Grasemann H,&nbsp;Vasquez-Vivar J. &nbsp;Pulmonary hypertension in the newborn GTP cyclohydrolase I-deficient mouse. &nbsp;Free Radic Biol Med.&nbsp;2011, 51(12):2227-33.</p>
  • <p>Cervantes-P&eacute;rez, L.G., Ibarra-Lara, M. de la L., Escalante, B., Del Valle-Mondrag&oacute;n, L., Vargas-Robles, H., P&eacute;rez-Severiano, F., Pastel&iacute;n, G., and S&aacute;nchez-Mendoza, M.A. (2012). Endothelial nitric oxide synthase impairment is restored by clofibrate treatment in an animal model of hypertension. Eur. J. Pharmacol. 685, 108&ndash;115.</p>
  • <p>Chen, W., Li, L., Brod, T., Saeed, O., Thabet, S., Jansen, T., Dikalov, S., Weyand, C., Goronzy, J., and Harrison, D.G. (2011). Role of increased guanosine triphosphate cyclohydrolase-1 expression and tetrahydrobiopterin levels upon T cell activation. J. Biol. Chem. 286, 13846&ndash;13851.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <h3>Weight of Evidence Summary</h3>
  • <p><a href="#Relationships_Among_Key_Events_and_the_Adverse_Outcome">Summary Table</a><br />
  • <em>Provide an overall summary of the weight of evidence based on the evaluations of the individual linkages from the Key Event Relationship pages. </em></p>
  • <p>Khoo JP, Zhao L, Alp NJ, Bendall JK, Nicoli T, Rockett K, et al. Pivotal role for endothelial tetrahydrobiopterin in pulmonary hypertension.&nbsp;Circulation.&nbsp;2005;111:2126&ndash;33</p>
  • <p>Quantitative measurements with dose and time response data from published studies cited below can be found here: <a href="/wiki/index.php/File:Hypertension_Empirical_Support_Concordance_Table.pdf" title="File:Hypertension Empirical Support Concordance Table.pdf">File:Hypertension Empirical Support Concordance Table.pdf</a>.</p>
  • <p>Nandi M, Miller A, Stidwill R, Jacques TS, Lam AA, Haworth S, et al. Pulmonary hypertension in a GTP-cyclohydrolase 1-deficient mouse.&nbsp;Circulation.&nbsp;2005;111:2086&ndash;90</p>
  • <table border="1" class="table">
  • <tbody>
  • <tr>
  • <th rowspan="2">Support for Biological Plausibility of KERs</th>
  • <th>Defining Question</th>
  • <th>High (Strong)</th>
  • <th>Moderate</th>
  • <th>Low (Weak)</th>
  • </tr>
  • <tr>
  • <td>Is there a mechanistic relationship between KEup and KEdown consistent with established biological knowledge?</td>
  • <td>Extensive understanding of the KER based on previous documentation and broad acceptance.</td>
  • <td>KER is plausible based on analogy to,accepted biological relationships, but scientific understanding is incomplete.</td>
  • <td>Empirical support for association between KEs, but the structural or functional relationship between them is not understood.</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to Glutathione, Oxidation:</td>
  • <td>Strong</td>
  • <td colspan="3">Multiple studies demonstrated that oxidative stress leads to the oxidation of glutathione (GSH) in the vascular endothelium. Exposure to a number of oxidants, including tert-butyl hydroperoxide, hydrogen peroxide, diamide, methylglyoxal, glucose, ischemia, and ultrafine particles caused a decrease in levels of GSH, which is indicative of its oxidation in human, bovine, and rat endothelial cells (De Pascali et al., 2014; Dhar et al., 2010; Du et al., 2013b; Montecinos et al., 2007; Park, 2013; Schuppe et al., 1992; van Gorp et al., 1999, 2002).</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to AKT/eNOS activity, Decrease</td>
  • <td>Strong</td>
  • <td colspan="3">Multiple experimental studies reported decreased Akt and eNOS phosphorylation/activity following oxidative stress as a consequence of exposure to peroxynitrite, high glucose, methylglyoxal, high fat, cigarette smoke extract (CSE) and ischemia in humans, bovine, mouse and rat endothelial cells (Das et al., 2014; Dhar et al., 2010; Du et al., 2001; Du et al., 2013; Michaud et al., 2006; Song et al., 2007, 2008; Su et al., 2013; Zhang et al., 2014; Zou et al., 2002).</td>
  • </tr>
  • <tr>
  • <td>AKT/eNOS activity, Decrease Directly Leads to Nitric Oxide, Depletion</td>
  • <td>Strong</td>
  • <td colspan="3">Several studies demonstrated that Akt can directly phosphorylate eNOS, leading to increased eNOS enzymatic activity and subsequent NO production (Dimmeler et al., 1999; Fulton et al., 1999). Inhibition of Akt or mutant eNOS attenuated eNOS phosphorylation in human and bovine cells, resulting in decreased NO bioavailability (Das et al., 2014; Dhar et al. 2010; Dimmeler et al., 1999; Fulton et al., 1999; Michaud et al., 2006; Uruno et al., 2005).</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to GTPCH-1, Decrease</td>
  • <td>Moderate</td>
  • <td colspan="3">Several studies demonstrated that GTPCH-1, the rate-limiting enzyme for BH4 synthesis, is affected by oxidative stress. GTPCH-1 expression or activity was inhibited by peroxynitrite and CSE (Abdelghany et al., 2017; Zhao et al., 2003). Cardiac reperfusion patients who experienced oxidative stress had reduced GTPCH-1 activity (Jayaram et al., 2015).</td>
  • </tr>
  • <tr>
  • <td>GTPCH-1, Decrease Directly Leads to Tetrahydrobiopterin, Decrease</td>
  • <td>Strong</td>
  • <td colspan="3">Many studies demonstrated that GTPCH-1 deletion led to reduced bioavailability of BH4 in bovine and murine endothelial cells (Adlam et al., 2012; Chen et al., 2011; Chuaiphichai et al., 2014; Crabtree et al., 2009; Tatham et al., 2009; Wang et al., 2008). Several studies also showed that overexpression of GTPCH-1 in human and mouse endothelium increased BH4 levels and eNOS activity (Alp et al., 2003; Antoniades et al., 2011; Carnicer et al., 2012).</td>
  • </tr>
  • <tr>
  • <td>Tetrahydrobiopterin, Decrease Directly Leads to eNOS, Uncoupling</td>
  • <td>Strong</td>
  • <td colspan="3">The depletion of BH4 leading to eNOS uncoupling is well-studied. Several studies showed reduced levels of BH4 induced eNOS uncoupling by reducing eNOS activity, decreasing NO and increasing superoxide levels in bovine, murine, and rat endothelium&nbsp;(Chuaiphichai et al., 2014; Crabtree et al., 2009; De Pascali et al., 2014; Dumitrescu et al., 2007; Whitsett et al., 2007). Many studies demonstrated that BH4 treatment improved endothelial function by reducing eNOS-mediated superoxide generation and increasing NO formation in human, bovine, mouse, and rat endothelium (Chen et al., 2011; De Pascali et al., 2014; Landmesser et al., 2003; Ozaki et al., 2002; Shinozaki et al., 2000; Wang et al., 2014).</td>
  • </tr>
  • <tr>
  • <td>Glutathione, Oxidation Directly Leads to eNOS, S-Glutathionylation</td>
  • <td>Strong</td>
  • <td colspan="3">Glutathione oxidation as determined by increased oxidized GSSG or decreased GSH levels caused S-glutathionylation of eNOS in bovine and human aortic endothelial cells, and in hypertensive rats and mice (Chen et al., 2010; De Pascali et al., 2014; Du et al., 2013).</td>
  • </tr>
  • <tr>
  • <td>eNOS, S-Glutathionylation Directly Leads to eNOS, Uncoupling</td>
  • <td>Strong</td>
  • <td colspan="3"><em>In vitro</em> experiments showed that S-glutathionylation of eNOS significantly decreased NO activity and greatly increased superoxide generation (Chen et al., 2010). These results were observed in bovine and human aortic endothelial cells as well as in vessels of spontaneously hypertensive rats and cardiac reperfusion patients (De Pascali et al., 2014; Du et al., 2013; Jayaram et al., 2015).</td>
  • </tr>
  • <tr>
  • <td>eNOS, Uncoupling Directly Leads to Nitric Oxide, Depletion</td>
  • <td>Strong</td>
  • <td colspan="3">It is well-established that uncoupling of eNOS causes eNOS to switch from producing NO to generating superoxides (F&ouml;rstermann and M&uuml;nzel, 2006). Studies reporting eNOS uncoupling as a result of BH4 depletion or S-glutathionylation measured levels of NO and superoxide which are indicative of eNOS uncoupling (Chen et al. 2010; De Pascali et al., 2014, Du et al., 2013; Whitsett et al., 2007).</td>
  • </tr>
  • <tr>
  • <td>Nitric Oxide, Depletion Directly Leads to Vasodilation, impaired</td>
  • <td>Strong</td>
  • <td colspan="3">Vasodilation is caused by the relaxation of vascular smooth muscle cells within the walls of blood vessels, and is regulated through a number of mechanisms, including cyclic GMP-dependent hyperpolarization and relaxation via NO. Thus, alterations to NO levels have an influence on vasodilation (Silva et al., 2012). Many animal studies demonstrated that inhibition of NO via eNOS inhibitors impaired endothelium-dependent vasodilation (Li et al., 2007; Luo et al., 2000; Paulis et al., 2008; S&eacute;lley et al., 2014).</td>
  • </tr>
  • <tr>
  • <td>Vasodilation, impaired Directly Leads to Vascular resistance, Increase</td>
  • <td>Strong</td>
  • <td colspan="3">It is well-accepted that vasodilation and systemic vascular resistance (SVR) are negatively correlated; blood flow is increased when blood vessels dilate due to decreased vascular resistance (Siddiqui, 2011). When vasodilation is impaired as a result of NO depletion or changes in potassium channels, vascular tone and SVR increase (Berg and Jensen, 2011; Brett et al., 1998; Dessy et al., 2004; Li et al., 2007; McVeigh et al., 2001; Paulis et al., 2008; Wilkinson et al., 2002).</td>
  • </tr>
  • <tr>
  • <td>Vascular resistance, Increase Directly Leads to Hypertension, N/A</td>
  • <td>Strong</td>
  • <td colspan="3">It is well-established that increased systemic vascular resistance (SVR), increased vascular stiffness and increased vascular reactivity contribute to hypertension (Brandes, 2014; Fo&euml;x and Sear, 2004; Mayet and Hughes, 2003). In patients with hypertension, SVR was elevated in about 66% of enrolled patients (Chan et al., 2016).</td>
  • </tr>
  • </tbody>
  • </table>
  • <p>Wang, S., Xu, J., Song, P., Wu, Y., Zhang, J., Chul Choi, H., and Zou, M.-H. (2008). Acute inhibition of guanosine triphosphate cyclohydrolase 1 uncouples endothelial nitric oxide synthase and elevates blood pressure. Hypertension 52, 484&ndash;490.</p>
  • <p>&nbsp;</p>
  • <p>Zhao, Y., Wu, J., Zhu, H., Song, P., and Zou, M.-H. (2013). Peroxynitrite-dependent zinc release and inactivation of guanosine 5&rsquo;-triphosphate cyclohydrolase 1 instigate its ubiquitination in diabetes. Diabetes 62, 4247&ndash;4256.</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <div>
  • <div>
  • <h4><a href="/events/934">934: KE3 : Decrease, Tetrahydrobiopterin</a><br></h4>
  • <h5>Short Name: Decrease, Tetrahydrobiopterin</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>biosynthetic process</td>
  • <td>5,6,7,8-tetrahydrobiopterin</td>
  • <td>decreased</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">149: Peptide Oxidation Leading to Hypertension</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • <h4>Stressors</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Name</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Reactive oxygen species</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Organization</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Cellular</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • <h4>Cell term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Cell term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>endothelial cell of vascular tree</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • </div>
  • <!-- end of organ term -->
  • <table border="1" class="table">
  • <tbody>
  • <tr>
  • <th rowspan="2">Empirical Support for KERs</th>
  • <th>Defining Question</th>
  • <th>High (Strong)</th>
  • <th>Moderate</th>
  • <th>Low (Weak)</th>
  • </tr>
  • <tr>
  • <td>Does empirical evidence support that a change in KEup leads to an appropriate change in KEdown? Does KEup occur at lower doses, earlier time points, and higher in incidence than KEdown&nbsp;? Inconsistencies?</td>
  • <td>Multiple studies showing dependent change in both events following exposure to a wide range of specific stressors. No or few critical data gaps or conflicting data.</td>
  • <td>Demonstrated dependent change in both events following exposure to a small number of stressors. Some inconsistencies with expected pattern that can be explained by various factors.</td>
  • <td>Limited or no studies reporting dependent change in both events following exposure to a specific stressor; and/or significant inconsistencies in empirical support across taxa and species</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to Glutathione, Oxidation:</td>
  • <td>Moderate</td>
  • <td colspan="3">Many studies demonstrated a dose-dependent relationship between known inducers of oxidative stress (tert-butyl hydroperoxide, hydrogen peroxide, methylglyoxal, high glucose, and ultrafine particles) and reduced GSH levels in human and rat studies (Dhar et al., 2010; Du et al., 2013; Montecinos et al., 2007; Park et al., 2013; van Gorp et al., 1999).</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to AKT/eNOS activity, Decrease</td>
  • <td>Moderate</td>
  • <td colspan="3">eNOS activity and reactive oxygen species (ROS) were modulated in the opposite manner by several stressors (methylglyoxal, high glucose, SIN-1, hydrogen peroxide) in human and bovine endothelial cells, resulting in increased ROS and decreased eNOS activity (Das et al., 2014; Dhar et al., 2010; Chen et al., 2010).</td>
  • </tr>
  • <tr>
  • <td>AKT/eNOS activity, Decrease Directly Leads to Nitric Oxide, Depletion</td>
  • <td>Strong</td>
  • <td colspan="3">Various stress inducers (ischemia, peroxynitrite, SIN-1, insulin plus orotic acidura, etc.) showed that a decrease in AKT and/or eNOS activity led to increased eNOS uncoupling and decreased NO (Choi et al., 2014, 2015; Das et al., 2014; Dhar et al., 2010; Dumitrescu et al., 2007 Uruno et al., 2005).</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to GTPCH-1, Decrease</td>
  • <td>Weak</td>
  • <td colspan="3">One study in a rat model of aortic coarctation-associated hypertension provides evidence that there is a interdependence between oxidative stress and GTPCH-1 with increased ROS and decreased GTPCH-1 expression following a perturbation, but there is no dose-response or temporal data (Cervantes-P&eacute;rez et al., 2012).</td>
  • </tr>
  • <tr>
  • <td>GTPCH-1, Decrease Directly Leads to Tetrahydrobiopterin, Decrease</td>
  • <td>Strong</td>
  • <td colspan="3">Exposure to a wide range of perturbations (e.g. CSE, 4-hydroxy-2-nonenal, cytokines) led to a decrease in both GTPCH-1 activity/expression and BH4 levels in human, cows and rats (Antoniades et al., 2011; Cervantes-P&eacute;rez et al., 2012; Chen et al., 2011; Ismail et al., 2015; Jayaram et al., 2015; Whitsett et al., 2007).</td>
  • </tr>
  • <tr>
  • <td>Tetrahydrobiopterin, Decrease Directly Leads to eNOS, Uncoupling</td>
  • <td>Strong</td>
  • <td colspan="3">Multiple studies demonstrated strong dependency between BH4 and eNOS uncoupling; decreased BH4 along with decreased eNOS activity, decreased NO production or increased superoxide generation were observed after various perturbations (Cervantes-P&eacute;rez et al., 2012; De Pascali et al., 2014; Dumitrescu et al., 2007; Jayaram et al., 2015; Whitsett et al., 2007; Wang et al., 2008).</td>
  • </tr>
  • <tr>
  • <td>Glutathione, Oxidation Directly Leads to eNOS, S-Glutathionylation</td>
  • <td>Moderate</td>
  • <td colspan="3">Treatment with GSSG induced a dose-dependent increase in human eNOS S-glutathionylation <em>in vitro</em> whereas 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) treatment also resulted in increased eNOS S-glutathionylation in a dose-dependent manner in bovine aortic endothelial cells (Chen et al., 2010). Exposure to ultrafine particles or hypoxia/reoxygenation in human or bovine aortic endothelial cells respectively, demonstrated a response-response relationship between reduced glutathione (GSH) and eNOS S-glutathionylation &nbsp;(Du et al., 2013; De Pascali et al., 2014).</td>
  • </tr>
  • <tr>
  • <td>eNOS, S-Glutathionylation Directly Leads to eNOS, Uncoupling</td>
  • <td>Moderate</td>
  • <td colspan="3">Treatment with BCNU resulted in increased eNOS S-glutathionylation, increased superoxide generation and decreased NO production in a dose-dependent manner in bovine aortic endothelial cells (Chen et al., 2010). Exposure to hypoxia/reoxygenation and treatment with angiotensin II demonstrated a response-response relationship between eNOS S-glutathionylation and superoxide generation in human and bovine endothelial cells (De Pascali et al., 2014; Galougahi et al., 2014).</td>
  • </tr>
  • <tr>
  • <td>eNOS, Uncoupling Directly Leads to Nitric Oxide, Depletion</td>
  • <td>Strong</td>
  • <td colspan="3">Multiple experiments demonstrated that eNOS uncoupling results in increased superoxide formation and decreased NO production (Chen et al., 2010; De Pascali et al., 2014; Dumitrescu et al., 2007; Wang et al., 2008; Whitsett et al., 2007; Zou et al., 2002).</td>
  • </tr>
  • <tr>
  • <td>Nitric Oxide, Depletion Directly Leads to Vasodilation, impaired</td>
  • <td>Moderate</td>
  • <td colspan="3">Treatment with eNOS inhibitors and two other stressors, BCNU and diaminohydroxypyrimidine (DAHP), caused a decrease in both NO production and vasodilation (Chen et al., 2010; Paulis et al, 2008; Wang et al., 2008).</td>
  • </tr>
  • <tr>
  • <td>Vasodilation, impaired Directly Leads to Vascular resistance, Increase</td>
  • <td>Weak</td>
  • <td colspan="3">No direct evidence was found for this KER, but there is indirect support. Treatment with eNOS inhibitor L-NG-monomethyl arginine citrate (L-NMMA) caused an increase in SVR and a reduction in NO (Stamler et al., 1994), while L-NG-nitroarginine methyl ester (L-NAME) decreased NO-dependent relaxation and increased blood pressure (Paulis et al, 2008). Infusion of NO donor sodium nitroprusside led to dose-dependent reductions in SVR (Eugene, 2016).</td>
  • </tr>
  • <tr>
  • <td>Vascular resistance, Increase Directly Leads to Hypertension, N/A</td>
  • <td>Moderate</td>
  • <td colspan="3">Several human studies showed a dose-dependent change in SVR and hypertension following treatment with eNOS inhibitors (Brett et al., 1998; Haynes et al., 1993; McVeigh et al., 2001; Stamler et al., 1994; Wilkinson et al., 2002).</td>
  • </tr>
  • </tbody>
  • </table>
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h3>Quantitative Consideration</h3>
  • <p><a href="#Relationships_Among_Key_Events_and_the_Adverse_Outcome">Summary Table</a><br />
  • <em>Provide an overall discussion of the quantitative information available for this AOP. Support calls for the individual relationships can be included in the Key Event Relationship table above. </em></p>
  • <p>Overall, there is a good amount of quantitative data available for this AOP as demonstrated by the weight of evidence tables, which shows that empirical support ranges from weak to strong. Four key event relationships (KERs) have strong quantitative support: decreased AKT/eNOS activity =&gt; NO depletion, decreased GTPCH-1 =&gt; decreased BH4, decreased BH4 =&gt; eNOS uncoupling and eNOS uncoupling =&gt; NO depletion. The relationships between these key events are well described in the literature as GTPCH-1 is the rate-limiting enzyme for BH4 synthesis (Wang et al., 2008), BH4 is an essential cofactor for eNOS and function (Wang et al., 2014), and the uncoupling of eNOS causes it to generate superoxide instead of NO (Carnicer et al., 2012). As they are functionally interconnected, many studies measure these key events together; thus providing strong support for their dependency.</p>
  • <p>Two key events have limited quantitative support (oxidative stress =&gt; decreased GTPCH-1, impaired vasodilation =&gt; increased vascular resistance). Generally, the oxidation of BH4 rather than decreased GTPCH-1 is measured when cells are under oxidative stress, and ROS are assumed to be increased so no quantitative measures are taken. For vasodilation and vascular resistance, there appears to be a correlative relationship, where increased vasodilation would mean decreased vascular resistance and vice versa, so studies do not measure both key events. Several studies showed that treatment with eNOS inhibitors led to increased vascular resistance, suggesting impaired vasodilation (Li et al., 2007; McVeigh et al., 2001; Paulis et al., 2008; Wilkinson et al., 2002).</p>
  • <p>The other KERs have moderate quantitative support, meaning there were studies showing a dependent change in both key events following treatment with stressors. Several studies measured decreased NO and vasodilation following perturbations to eNOS inhibitors, BCNU and DAHP (Chen et al., 2010; Paulis et al, 2008; Wang et al., 2008).</p>
  • <p>In general, experiments with both dose-dependent and temporal response data following a stressor are not readily available for all KERs as most measurements are generally taken at one time point after a perturbation, or the measurements are for one key event, not both. However, an exhaustive literature search was not performed. An ideal experiment would be to treat cells with three to four stressors at increasing concentrations and measure the key events at different time intervals; thus providing a greater understanding of the temporal and dose-dependent responses between the key events.</p>
  • <h4>Evidence Supporting Applicability of this Event</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Weak</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end life stages -->
  • <!-- potential consierations, text as entered by author -->
  • <div id="considerations_for_potential_applicaitons">
  • <h2>Considerations for Potential Applications of the AOP (optional)</h2>
  • <hr>
  • <p>One of the most widely reported hypertension risk factors is tobacco smoking.&nbsp; While smoking cessation remains the best way to reduce the harmful effects of tobacco smoking, tobacco harm reduction is being considered by some regulators (e.g. US Food and Drug Administration; FDA) as a complementary strategy to reduce smoking-related disease burden.&nbsp; The US FDA has published guidance on assessing a &ldquo;modified risk tobacco product&rdquo; (MRTP), either through demonstration of reduced toxicant exposure or reduction in health risks<span style="font-size:10.8333px">&nbsp;<span style="font-family:arial,helvetica,sans-serif">(FDA 2012)</span></span>. By gaining an understanding of how, and to what extent tobacco smoke initiates biological mechanisms of hypertension, such knowledge could be utilised as a baseline for comparison purposes in toxicological assessments of the risk reduction potential of e-cigarettes.&nbsp;</p>
  • <p>Given the wide variety of data requirements to support such risk assessments e.g. human exposure studies, behavioural studies, efficacy studies, <em>in vitro</em> studies, clinical biomarker studies, pharmacokinetic studies, quality of life studies etc., a data integration framework is required to organise these data types into a comprehensive story that (i) characterises the toxicological problem, (ii) demonstrates the likely outcome of an intervention, and (iii) can be utilised to monitor the performance of any intervention over time.&nbsp; Adverse outcome pathways offer scientists and regulators a way to inform future Integrated Approaches to Testing and Assessment (IATA) for the risk assessment/harm reduction potential of electronic cigarettes and heated tobacco products.</p>
  • <p>&nbsp;</p>
  • <p>Other applications could include informing risk assessments for long term exposure to airborne pollution, in the context of helping to set acceptable exposure limits in urban environments for specific pollutants e.g. diesel exhuast particles.&nbsp; Furthermore, health supplements which purport to benefit the cardiovascular system in the context of&nbsp;hypertension risk, could also be assessed for efficacy.</p>
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <div>
  • <p>Decreased BH4 is observed in humans (Jayaram et al., 2015), cows (Abdelghany et al., 2017; Whitsett et al., 2007, Wang et al., 2008), mice (Adlam et al., 2012; Chuaiphichai et al., 2014; Crabtree et al., 2009; Tatham et al., 2009; Wang et al., 2008) and rats (Cervantes-P&eacute;rez et al., 2012).</p>
  • <br>
  • </div>
  • <!-- event text -->
  • <h4>How this Key Event Works</h4>
  • <p>Tetrahydrobiopterin (BH4) is an essential cofactor for a group of enzymes including aromatic acid hydroxylases, nitric oxide synthase (NOS) isoforms, and alkylglycerol monooxygenase (Wang et al., 2014). BH4 is synthesized from guanosine triphosphate through sequential reactions catalyzed by enzymes <a href="/wiki/index.php/Event:935" title="Event:935">GTPCH-1</a>, pyruvoyl tetrahydropterin synthase, and sepiapterin reductase (Tatham et al., 2009). During NOS catalysis, BH4 donates electrons to the ferrous-dioxygen complex in the oxygenase domain, leading to oxidation of L-arginine to N-hydroxy-Larginine and eventually conversion to citrulline and nitric oxide production (Chen et al., 2011; Crabtree et al., 2009). BH4 also stabilizes dimers of NOS isoforms, which is required for their enzymatic activity. When BH4 levels are decreased or limited, for example under oxidative stress conditions, BH4 can be oxidized to dihydrobiopterin (BH2) and then converted to biopterin. This reduction in BH4 availability results in NOS uncoupling where NOS is uncoupled from L-arginine oxidation and superoxide (or other reactive species) is produced rather than nitric oxide (Carnicer et al., 2012). Decreased BH4 have been demonstrated in a variety of vascular diseases such as hypertension, diabetes and atherosclerosis where endothelial dysfunction occurs.</p>
  • <br>
  • <h4>How it is Measured or Detected</h4>
  • <p>Levels of BH4, BH2 and biopterin levels can be determined by reverse-phase high-performance liquid chromatography (HPLC) followed by electrochemical detection (for BH4) and fluorescence detection (for BH2 and biopterin) (Howells et al., 1986).</p>
  • <!-- reference section, text as of right now but should be changed to be handled as table -->
  • <div id="references">
  • <h2>References</h2>
  • <hr>
  • <p>AbdelGhany, T., Ismail, R., Elmahdy, M., Mansoor F, Zweier J, Lowe, F., and Zweier, JL. (2017). Cigarette Smoke Constituents Cause Endothelial Nitric Oxide Synthase Dysfunction and Uncoupling due to Depletion of Tetrahydrobiopterin with Degradation of GTP Cyclohydrolase.&nbsp;&nbsp;Nitric Oxide&nbsp;(Under review).</p>
  • <p>Adlam, D., Herring, N., Douglas, G., De Bono, J.P., Li, D., Danson, E.J., Tatham, A., Lu, C.-J., Jennings, K.A., Cragg, S.J., et al. (2012). Regulation of &beta;-adrenergic control of heart rate by GTP-cyclohydrolase 1 (GCH1) and tetrahydrobiopterin. Cardiovasc. Res. 93, 694&ndash;701.</p>
  • <p>Alp, N.J., Mussa, S., Khoo, J., Cai, S., Guzik, T., Jefferson, A., Goh, N., Rockett, K.A., and Channon, K.M. (2003). Tetrahydrobiopterin-dependent preservation of nitric oxide-mediated endothelial function in diabetes by targeted transgenic GTP-cyclohydrolase I overexpression. J. Clin. Invest. 112, 725&ndash;735.</p>
  • <p>Antoniades, C., Cunnington, C., Antonopoulos, A., Neville, M., Margaritis, M., Demosthenous, M., Bendall, J., Hale, A., Cerrato, R., Tousoulis, D., et al. (2011). Induction of vascular GTP-cyclohydrolase I and endogenous tetrahydrobiopterin synthesis protect against inflammation-induced endothelial dysfunction in human atherosclerosis. Circulation 124, 1860&ndash;1870.</p>
  • <p>Bendall JK, Douglas G, McNeill E <em>et al.</em>&nbsp; Tetrahydrobiopterin in cardiovascular health and disease.&nbsp; Antioxid Redox Signal. (2014) 20(18):3040-77.</p>
  • <p>Berg, T., and Jensen, J. (2011). Simultaneous parasympathetic and sympathetic activation reveals altered autonomic control of heart rate, vascular tension, and epinephrine release in anesthetized hypertensive rats. Front. Neurol. 2, 71.</p>
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  • <p>ELISA kits for BH4 are also commercially available.</p>
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  • <p>In each case, care must be taken to protect the sample from oxidation, and BH4 is highly redox sensitive. &nbsp;Dithioerythritol is commonly used as a preservation agent.</p>
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  • <p>Whitsett, J., Picklo, M.J., and Vasquez-Vivar, J. (2007). 4-Hydroxy-2-nonenal increases superoxide anion radical in endothelial cells via stimulated GTP cyclohydrolase proteasomal degradation. Arterioscler. Thromb. Vasc. Biol. 27, 2340&ndash;2347.</p>
  • <p>Zhao Y,&nbsp;Cao J,&nbsp;Chen YS,&nbsp;Zhu Y,&nbsp;Patrick C,&nbsp;Chien B,&nbsp;Cheng A,&nbsp;Foehr ED. &nbsp;Detection of tetrahydrobiopterin by LC-MS/MS in plasma from multiple species. &nbsp;Bioanalysis.&nbsp;2009;1(5):895-903.</p>
  • <p>de Wijs-Meijler DP,&nbsp;Duncker DJ,&nbsp;Tibboel D,&nbsp;Schermuly RT,&nbsp;Weissmann N,&nbsp;Merkus D,&nbsp;Reiss IKM. &nbsp;Oxidative&nbsp;injury of the&nbsp;pulmonary&nbsp;circulation in the perinatal period: Short- and long-term consequences for the human cardiopulmonary system. &nbsp;Pulm Circ.&nbsp;2017 &nbsp;1;7(1):55-66. doi: 10.1086/689748. eCollection 2017 Mar.</p>
  • <p>&nbsp;</p>
  • <p>Wilkinson, I.B., MacCallum, H., Cockcroft, J.R., and Webb, D.J. (2002). Inhibition of basal nitric oxide synthesis increases aortic augmentation index and pulse wave velocity in vivo. Br. J. Clin. Pharmacol. 53, 189&ndash;192.</p>
  • <p>&nbsp;</p>
  • <p>Zhang, W., Han, Y., Meng, G., Bai, W., Xie, L., Lu, H., Shao, Y., Wei, L., Pan, S., Zhou, S., et al. (2014). Direct renin inhibition with aliskiren protects against myocardial ischemia/reperfusion injury by activating nitric oxide synthase signaling in spontaneously hypertensive rats. J. Am. Heart Assoc. 3, e000606.</p>
  • <p>Zhao, H., Kalivendi, S., Zhang, H., Joseph, J., Nithipatikom, K., V&aacute;squez-Vivar, J., and Kalyanaraman, B. (2003). Superoxide reacts with hydroethidine but forms a fluorescent product that is distinctly different from ethidium: potential implications in intracellular fluorescence detection of superoxide. Free Radic. Biol. Med. 34, 1359&ndash;1368.</p>
  • <p>Zou, M.-H., Hou, X.-Y., Shi, C.-M., Nagata, D., Walsh, K., and Cohen, R.A. (2002). Modulation by peroxynitrite of Akt- and AMP-activated kinase-dependent Ser1179 phosphorylation of endothelial nitric oxide synthase. J. Biol. Chem. 277, 32552&ndash;32557.</p>
  • <p>Zweier JL, Chen CA, Druhan LJ. &nbsp;S-glutathionylation reshapes our understanding of endothelial nitric oxide synthase uncoupling and nitric oxide/reactive oxygen species-mediated signaling.&nbsp; (2011) Antioxid Redox Signal. 14(10):1769-75.</p>
  • <br>
  • <!-- end event text -->
  • <br>
  • </div>
  • <div>
  • <div id="appendicies">
  • <h2>Appendix 1</h2>
  • <h3>List of MIEs in this AOP</h3>
  • <div>
  • <div>
  • <h4><a href="/events/932">932: KE4 : Uncoupling, eNOS</a><br></h4>
  • <h5>Short Name: Uncoupling, eNOS</h5>
  • <h4><a href="/events/209">Event: 209: Peptide Oxidation</a><br></h4>
  • <h5>Short Name: Peptide Oxidation</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>oxidative stress</td>
  • <td></td>
  • <td>nitric oxide synthase, endothelial</td>
  • <td>functional change</td>
  • <td>increased</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">149: Peptide Oxidation Leading to Hypertension</a></td>
  • <td><a href="/aops/27">Aop:27 - Cholestatic Liver Injury induced by Inhibition of the Bile Salt Export Pump (ABCB11)</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • <tr>
  • <td><a href="/aops/108">Aop:108 - Inhibition of pyruvate dehydrogenase kinase leading to hepatocellular adenomas and carcinomas (in mouse and rat)</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • <tr>
  • <td><a href="/aops/149">Aop:149 - Peptide Oxidation Leading to Hypertension</a></td>
  • <td>MolecularInitiatingEvent</td>
  • </tr>
  • <tr>
  • <td><a href="/aops/210">Aop:210 - Activation of c-Jun N-terminal kinase (JNK) and Forkhead box O (FOXO) and reduction of WNT pathways leading to reproductive failure: Integrated multi-OMICS approach for AOP building</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Organization</h4>
  • <h4>Biological Context</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Cellular</td>
  • <td>Molecular</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • <h4>Cell term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Cell term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>endothelial cell</td>
  • <td>eukaryotic cell</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h4>Evidence Supporting Applicability of this Event</h4>
  • <h4>Domain of Applicability</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>human</td>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Strong</td>
  • <td>rodents</td>
  • <td>rodents</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=0" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Strong</td>
  • <td>human and other cells in culture</td>
  • <td>human and other cells in culture</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=0" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Not Specified</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Not Specified</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <div>
  • <p>eNOS uncoupling has been demonstrated in humans, cows, mice and rats (Chen et al., 2010; Crabtree et al., 2009; De Pascali et al., 2014; Du et al., 2013; Jayaram et al., 2015).</p>
  • <p>The concentrations of GSH and GSSG have been shown in tissues of human and laboratory animals, including rats, mice and cows (Chen et al., 2010; Giustarini et al., 2013).</p>
  • <br>
  • </div>
  • <!-- event text -->
  • <h4>How this Key Event Works</h4>
  • <p>Endothelial nitric oxide synthase (eNOS) is responsible for the generation of vascular nitric oxide (NO), a protective molecule that is involved in the regulation of endothelium-dependent vasodilation, vascular tone, and blood pressure (F&ouml;rstermann and M&uuml;nzel, 2006). To generate NO, eNOS hydroxylates L-arginine to N-hydroxy-L-arginine and then oxidizes N-hydroxy-L-arginine to L-citrulline and NO. This enzymatic process requires NADPH, Ca2+/calmondulin, flavin mononucleotide, flavin adenine dinucleotide and its cofactor <a href="/wiki/index.php/Event:934" title="Event:934">tetrahydrobiopterin (BH4)</a>. Limiting BH4 levels or <a href="/wiki/index.php/Event:927" title="Event:927">S-glutathionylation of eNOS</a> can lead to eNOS uncoupling in which eNOS produces superoxide (or other reactive oxygen species) and less NO. The uncoupling of eNOS has been demonstrated to cause endothelial dysfunction, and is implicated in a number of cardiovascular diseases such as hypertension, atherosclerosis, hypercholesterolemia, and diabetes mellitus (Dumitrescu et al., 2007).</p>
  • <h4>Key Event Description</h4>
  • <p>Oxidative stress corresponds to an imbalance between the rate of oxidant production and that of their degradation. The term oxidative stress indicates the outcome of oxidative damage to biologically relevant macromolecules such as nucleic acids, proteins, lipids and carbohydrates. This occurs when oxidative stress-related molecules, generated in the extracellular environment or within the cell, exceed cellular antioxidant defenses. Major reactive oxygen species (ROS), such as hydrogen peroxide (H2O2) and superoxide anion, as well as 4-hydroxy- 2,3-nonenal (HNE) and related 4-hydroxy-2,3-alkenals (HAKs), major aldehydic end-products of lipid peroxidation, can act as potential mediators able to affect signal transduction pathways as well as the proliferative and functional response of target cells. H2O2 and superoxide anion may be also generated as molecular messengers within the cell as part of the cellular response to defined growth factors, cytokines and other mediators. The final consequence at tissue, cellular and molecular level is primarily affected by the steady state concentration of oxidative stress-related molecules. The main biological targets of free radicals are proteins, lipids and DNA.</p>
  • <p>Major consequences of reaction of ROS, HAKs and NO with biologically relevant macromolecules that can mediate pathophysiological effects:</p>
  • <p>ROS: DNA: oxidation, strand breaks, genotoxicity Proteins: oxidation, fragmentation, formation of carbonyls Lipids: lipid peroxidation and degradation</p>
  • <p>HAKs: DNA: adducts (low doses), strand breaks, genotoxicity (high doses) Proteins: adducts (Michael type reactions on Lys, Cys and His residues)</p>
  • <p>NO: DNA: oxidation, strand breaks Proteins: oxidation, nitrosation, nitration (nytrosylation of tyrosine) Lipids: lipid peroxidation and degradation</p>
  • <p>Continued oxidative stress can lead to chronic inflammation. Oxidative stress can activate a variety of transcription factors including NF-&kappa;B, AP-1, p53, HIF-1&alpha;, PPAR-&gamma;, &beta;-catenin/Wnt, and Nrf2. Activation of these transcription factors can lead to the expression of over 500 different genes, including those for growth factors, inflammatory cytokines and chemokines, which can activate inflammatory pathways. <sup><a href="#cite_note-1">[1]</a></sup> <sup><a href="#cite_note-2">[2]</a></sup> <sup><a href="#cite_note-3">[3]</a></sup></p>
  • <p>Glutathione (GSH) oxidation refers to the conversion of reduced glutathione to its oxidized form glutathione disulfide (GSSG) in the presence of oxidative species. GSH plays an important role as an anti-oxidant in regulating cellular redox homeostasis, and is mainly present in the cell as the reduced form (98%). Deficiency in GSH or a decrease in GSH/GSSG ratio results in decreased anti-oxidant function and increased susceptibility to oxidative stress, thus making it a marker of cellular redox status. An imbalance in GSH/GSSG ratio has been implicated in the onset and progression of human diseases, such as neurodegenerative diseases, cancers, pulmonary diseases and cardiovascular diseases (Ballatori et al., 2009; Kalinina et al., 2014)</p>
  • <br>
  • <h4>How it is Measured or Detected</h4>
  • <p>The activity of eNOS can be measured indirectly through superoxide and NO production. Superoxides can be detected using several standard methods including lucigenin-enhanced chemiluminescence (M&uuml;nzel et al., 2002; Tarpey et al., 1999), electron paramagnetic resonance (EPR) spin-trapping (Roubaud et al., 1997), and HPLC/fluorescence detector-based assay using dihydroethidium (Fink et al., 2004; Zhao et al., 2003). NO production can be measured through the conversion of L-arginine to L-citrulline (de Bono et al., 2007) , in situ fluorescent signal detection with fluorescent indicator DAF-2 DA (Itoh et al., 2000; Nagata et al., 1999; Qiu et al., 2001), EPR spin-trapping (Xia et al., 2000), and the determination of total nitrate and nitrite concentration (Crabtree et al., 2009; Du et al., 2013).</p>
  • <br>
  • <h4>References</h4>
  • <p>de Bono, J.P., Warrick, N., Bendall, J.K., Channon, K.M., and Alp, N.J. (2007). Radiochemical HPLC detection of arginine metabolism: Measurement of nitric oxide synthesis and arginase activity in vascular tissue. Nitric Oxide 16, 1&ndash;9.</p>
  • <p>&nbsp;</p>
  • <p>Chen, X., Xu, J., Feng, Z., Fan, M., Han, J., and Yang, Z. (2010). Simvastatin combined with nifedipine enhances endothelial cell protection by inhibiting ROS generation and activating Akt phosphorylation. Acta Pharmacol. Sin. 31, 813&ndash;820.</p>
  • <p><em><strong>measuring oxidative stress</strong> </em></p>
  • <p>Crabtree, M.J., Tatham, A.L., Al-Wakeel, Y., Warrick, N., Hale, A.B., Cai, S., Channon, K.M., and Alp, N.J. (2009). Quantitative regulation of intracellular endothelial nitric-oxide synthase (eNOS) coupling by both tetrahydrobiopterin-eNOS stoichiometry and biopterin redox status: insights from cells with tet-regulated GTP cyclohydrolase I expression. J. Biol. Chem. 284, 1136&ndash;1144.</p>
  • <p><em>Agents for <strong>ROS detection</strong> are primarily fluorescence based, but recently luminescent based detections have been introduced. The biggest difficulty reported with much of the cellular ROS research has been with the lack of reporter agents specific for discrete molecules. ROS moieties by their nature are reactive with a number of different molecules; as such designing reporter agents has been difficult. With more specific chemistries, particularly for hydrogen peroxide, the specific mechanisms for regulation will be elucidated. </em></p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p><em><strong>Reduced glutathione</strong> (GSH) is regenerated from its oxidized form (GSSH) by the action of an NADPH dependent reductase GSSH + NADPH + H+ &agrave; 2 GSH + NADP+ Due to the rapid nature of the reduction of GSSH relative to its synthesis or secretion, the ratio of GSH to GSSH is a good indicator of oxidative stress within cells. GSH and GSSH levels can be determined by HPLC, capillary electrophoresis, or biochemically in microplates. Several different assays have been designed to measure glutathione in samples. By using a luciferin derivative in conjunction with glutathione S-transferase enzyme the amount of GSH would be proportional to the luminescent signal generated when luciferase is added in a subsequent step. Total glutathione can be determined colorimetrically by reacting GSH with DTNB (Ellman&rsquo;s reagent) in the presence of glutathione reductase. Glutathione reductase reduces GSSH to GSH, which then reacts with DTNB to produce a yellow colored 5-thio-2-nitrobenzoic acid (TNB), which absorbs at 412 nm. </em></p>
  • <p>Du, Y., Navab, M., Shen, M., Hill, J., Pakbin, P., Sioutas, C., Hsiai, T.K., and Li, R. (2013). Ambient ultrafine particles reduce endothelial nitric oxide production via S-glutathionylation of eNOS. Biochem. Biophys. Res. Commun. 436, 462&ndash;466.</p>
  • <p><em><strong>Lipid peroxidation</strong> is one of the most widely used indicators of free radical formation, a key indicator of oxidative stress. Measurement of lipid peroxidation has historically relied on the detection of thiobarbituric acid (TBA) reactive compounds such as malondialdehyde generated from the decomposition of lipid peroxidation products. While this method is controversial in that it is quite sensitive, but not necessarily specific to MDA, it remains the most widely used means to determine lipid peroxidation. This reaction, which takes place under acidic conditions at 90-100&ordm;C, results in an adduct that can be measured colorimetrically at 532 nm or by fluorescence using a 530 nm excitation wavelength and a 550 nm emission wavelength. A number of commercial assay kits are available for this assay using absorbance or fluorescence detection technologies. The formation of F2-like prostanoid derivatives of arachidonic acid, termed F2-isoprostanes (IsoP) has been shown to be specific for lipid peroxidation. Unlike the TBA assay, measurement of IsoP appears to be specific to lipid peroxides, they are stable and are not produced by any enzymatic pathway making interpretation easier. There have been a number of commercial ELISA kits developed for IsoPs, but interfering agents in samples requires partial purification of samples prior to running the assay. The only reliable means for detection is through the use of GC/MS, which makes it expensive and limits throughput. </em></p>
  • <p>Dumitrescu, C., Biondi, R., Xia, Y., Cardounel, A.J., Druhan, L.J., Ambrosio, G., and Zweier, J.L. (2007). Myocardial ischemia results in tetrahydrobiopterin (BH4) oxidation with impaired endothelial function ameliorated by BH4. Proc. Natl. Acad. Sci. U. S. A. 104, 15081&ndash;15086.</p>
  • <p><em><strong>Superoxide detection</strong> is based on the interaction of superoxide with some other compound to create a measurable result. The reduction of ferricytochrome c to ferrocytochrome c has been used in a number of situations to assess the rate of superoxide formation. While not completely specific for superoxide this reaction can be monitored colorimetrically at 550 nm. Chemiluminescent reactions have been used for their potential increase in sensitivity over absorbance-based detection methods. The most widely used chemiluminescent substrate is Lucigenin, but this compound has a propensity for redox cycling, which has raised doubts about its use in determining quantitative rates of superoxide production. Coelenterazine has also been used as a chemiluminescent substrate. Hydrocyanine dyes are fluorogenic sensors for superoxide and hydroxyl radical. These dyes are synthesized by reducing the iminium cation of the cyanine (Cy) dyes with sodium borohydride. While weakly fluorescent, upon oxidation their fluorescence intensity increases 100 fold. In addition to being fluorescent, oxidation also converts the molecule from being membrane permeable to an ionic impermeable moiety. The most characterized of these probes are Hydro-Cy3 and Hydro-Cy5. </em></p>
  • <p>Fink, B., Laude, K., McCann, L., Doughan, A., Harrison, D.G., and Dikalov, S. (2004). Detection of intracellular superoxide formation in endothelial cells and intact tissues using dihydroethidium and an HPLC-based assay. Am. J. Physiol. Cell Physiol. 287, C895&ndash;C902.</p>
  • <p><em><strong>Hydrogen peroxide</strong> (H2O2) is the most important ROS in regards to mitogenic stimulation or cell cycle regulation. There are a number of fluorogenic substrates, which serve as hydrogen donors that have been used in conjunction with horseradish peroxidase (HRP) enzyme to produce intensely fluorescent products. The more commonly used substrates include diacetyldichloro-fluorescein, homovanillic acid, and Amplex&reg; Red. In these examples, increasing amounts of H2O2 form increasing amounts of fluorescent product. </em></p>
  • <p>F&ouml;rstermann, U., and M&uuml;nzel, T. (2006). Endothelial nitric oxide synthase in vascular disease: from marvel to menace. Circulation 113, 1708&ndash;1714.</p>
  • <p><em><strong>Nitric Oxide</strong> The free radical nitric oxide (&bull;NO) is produced by a number of different cell types with a variety of biological functions. Regardless of the source or role, the free radical &bull;NO has a very short half life (t&frac12;= 4 seconds), reacting with several different molecules normally present to form either nitrate (NO3-) or nitrite (NO2-) A commonly used method for the indirect determination of &bull;NO is the determination of its composition products nitrate and nitrite colorimetrically. This reaction requires that nitrate (NO3) first be reduced to nitrite (NO2), typically by the action of nitrate reductase. Subsequent determination of nitrite by a two step process provides information on the &ldquo;total&rdquo; of nitrate and nitrite. In the presence of hydrogen ions nitrite forms nitrous acid, which reacts with sulfanilamide to produce a diazonium ion. This then coupled to N-(1-napthyl) ethylenediamine to form the chromophore which absorbs at 543 nm. Nitrite only determinations can then be made in a parallel assay where the samples were not reduced prior to the colorimetric assay. Actual nitrate levels are then calculated by the subtraction of nitrite levels from the total. <sup><a href="#cite_note-4">[4]</a></sup> </em></p>
  • <p>Itoh, Y., Ma, F.H., Hoshi, H., Oka, M., Noda, K., Ukai, Y., Kojima, H., Nagano, T., and Toda, N. (2000). Determination and bioimaging method for nitric oxide in biological specimens by diaminofluorescein fluorometry. Anal. Biochem. 287, 203&ndash;209.</p>
  • <p>&nbsp;</p>
  • <br>
  • <h4>References</h4>
  • <p>Ballatori, N., Krance, S.M., Notenboom, S., Shi, S., Tieu, K., and Hammond, C.L. (2009). Glutathione dysregulation and the etiology and progression of human diseases. Biol. Chem. 390, 191&ndash;214.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Chen, C.-A., Wang, T.-Y., Varadharaj, S., Reyes, L.A., Hemann, C., Talukder, M.A.H., Chen, Y.-R., Druhan, L.J., and Zweier, J.L. (2010). S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature 468, 1115&ndash;1118.</p>
  • <p>M&uuml;nzel, T., Afanas&rsquo;ev, I.B., Kleschyov, A.L., and Harrison, D.G. (2002). Detection of Superoxide in Vascular Tissue. Arterioscler. Thromb. Vasc. Biol. 22, 1761&ndash;1768.</p>
  • <p>Giustarini, D., Dalle-Donne, I., Milzani, A., Fanti, P., and Rossi, R. (2013). Analysis of GSH and GSSG after derivatization with N-ethylmaleimide. Nat. Protoc. 8, 1660&ndash;1669.</p>
  • <p>Nagata, N., Momose, K., and Ishida, Y. (1999). Inhibitory effects of catecholamines and anti-oxidants on the fluorescence reaction of 4,5-diaminofluorescein, DAF-2, a novel indicator of nitric oxide. J. Biochem. (Tokyo) 125, 658&ndash;661.</p>
  • <p>Held P., 2010 Biotek, Measurement of ROS in Cells, <a class="external free" href="http://www.biotek.com/assets/tech_resources/ROS%20Application%20Guide.pdf" rel="nofollow" target="_blank">http://www.biotek.com/assets/tech_resources/ROS%20Application%20Guide.pdf</a></p>
  • <p>Qiu, W., Kass, D.A., Hu, Q., and Ziegelstein, R.C. (2001). Determinants of shear stress-stimulated endothelial nitric oxide production assessed in real-time by 4,5-diaminofluorescein fluorescence. Biochem. Biophys. Res. Commun. 286, 328&ndash;335.</p>
  • <p>Kalinina, E.V., Chernov, N.N., and Novichkova, M.D. (2014). Role of glutathione, glutathione transferase, and glutaredoxin in regulation of redox-dependent processes. Biochem. Biokhimii︠a︡ 79, 1562&ndash;1583.</p>
  • <p>Roubaud, V., Sankarapandi, S., Kuppusamy, P., Tordo, P., and Zweier, J.L. (1997). Quantitative measurement of superoxide generation using the spin trap 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide. Anal. Biochem. 247, 404&ndash;411.</p>
  • <p>Kamencic, H., Lyon, A., Paterson, P.G., and Juurlink, B.H. (2000). Monochlorobimane fluorometric method to measure tissue glutathione. Anal. Biochem. 286, 35&ndash;37.</p>
  • <p>Tarpey, M.M., White, C.R., Suarez, E., Richardson, G., Radi, R., and Freeman, B.A. (1999). Chemiluminescent Detection of Oxidants in Vascular Tissue Lucigenin But Not Coelenterazine Enhances Superoxide Formation. Circ. Res. 84, 1203&ndash;1211.</p>
  • <p>Parola, M. and Robino, G. (2001). Oxidative stress-related molecules and liver fibrosis. J Hepatol. 35, 297-306</p>
  • <p>Xia, Y., Cardounel, A.J., Vanin, A.F., and Zweier, J.L. (2000). Electron paramagnetic resonance spectroscopy with N-methyl-D-glucamine dithiocarbamate iron complexes distinguishes nitric oxide and nitroxyl anion in a redox-dependent manner: applications in identifying nitrogen monoxide products from nitric oxide synthase. Free Radic. Biol. Med. 29, 793&ndash;797.</p>
  • <p>Reuter S. et al., (2010) Oxidative stress, inflammation, and cancer: how are they linked? Free Radic Biol Med.49, 1603-1616</p>
  • <p>Zhao, H., Kalivendi, S., Zhang, H., Joseph, J., Nithipatikom, K., V&aacute;squez-Vivar, J., and Kalyanaraman, B. (2003). Superoxide reacts with hydroethidine but forms a fluorescent product that is distinctly different from ethidium: potential implications in intracellular fluorescence detection of superoxide. Free Radic. Biol. Med. 34, 1359&ndash;1368.</p>
  • <p>S&aacute;nchez-Valle V. et al., (2012) Role of oxidative stress and molecular changes in liver fibrosis: a review. Curr Med Chem. 19, 4850-4860</p>
  • <p>Tipple, T.E., and Rogers, L.K. (2012). Methods for the Determination of Plasma or Tissue Glutathione Levels. Methods Mol. Biol. Clifton NJ 889, 315&ndash;324</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <h3>List of Key Events in the AOP</h3>
  • <div>
  • <div>
  • <h4><a href="/events/933">933: KE6 : Depletion, Nitric Oxide</a><br></h4>
  • <h5>Short Name: Depletion, Nitric Oxide</h5>
  • <h4><a href="/events/927">Event: 927: KE1 : S-Glutathionylation, eNOS</a><br></h4>
  • <h5>Short Name: S-Glutathionylation, eNOS</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>nitric oxide biosynthetic process</td>
  • <td>nitric oxide</td>
  • <td>decreased</td>
  • <td>protein glutathionylation</td>
  • <td>nitric oxide synthase, endothelial</td>
  • <td>increased</td>
  • </tr>
  • <tr>
  • <td>protein glutathionylation</td>
  • <td>cysteine residue</td>
  • <td>increased</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">149: Peptide Oxidation Leading to Hypertension</a></td>
  • <td><a href="/aops/149">Aop:149 - Peptide Oxidation Leading to Hypertension</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • <h4>Stressors</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Name</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Reactive oxygen species</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Organization</h4>
  • <h4>Biological Context</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Cellular</td>
  • <td>Molecular</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • <h4>Organ term</h4>
  • <h4>Cell term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Organ term</th>
  • <th>Cell term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>blood</td>
  • <td>endothelial cell of vascular tree</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h4>Evidence Supporting Applicability of this Event</h4>
  • <h4>Domain of Applicability</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Strong</td>
  • <td>Low</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Strong</td>
  • <td>Low</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <div>
  • <p>NO depletion was observed in humans, cows, mice and rats (Chen et al., 2010; Crabtree et al., 2009; De Pascali et al., 2014; Du et al., 2013; Jayaram et al., 2015).</p>
  • <p>S-glutathionylation of eNOS has been demonstrated in humans, cows, mice and rats (Chen et al., 2010; De Pascali et al., 2014; Du et al., 2013).</p>
  • <br>
  • </div>
  • <!-- event text -->
  • <h4>How this Key Event Works</h4>
  • <p>Nitric oxide (NO), constitutively produced by endothelial nitric oxide synthase (eNOS), is an important regulator of vascular homeostasis. Endothelial-derived NO promotes vasodilation and protects against atherogenesis through the inhibition of vascular smooth muscle cell proliferation and migration, platelet aggregation and adhesion, and leukocyte adherence. Its effects have an influence on vascular resistance, blood pressure, vascular remodeling and angiogenesis (Luo et al., 2000). Dysfunctional eNOS as a result of <a href="/wiki/index.php/Event:932" title="Event:932">eNOS uncoupling</a> leads to a decrease or loss of NO bioavailability and an elevation of superoxide production (Crabtree et al., 2009). The imbalance of NO and superoxide is associated with many disorders,&nbsp;such as hypertension, atherosclerosis, hypercholesterolemia, and diabetes mellitus.</p>
  • <h4>Key Event Description</h4>
  • <p>S-glutathionylation is a redox-dependent, reversible post-translational modification that is involved in the regulation of various regulatory, structural, and metabolic proteins (Pastore and Piemonte, 2013). Under oxidative stress, S-glutathionylation targets cysteine residues of a protein and adds glutathione through thiol-disulfide exchange with oxidized glutathione (GSSG) or reaction of oxidant-induced protein thiyl radicals with reduced glutathione (Chen et al., 2010, 2011, Schuppe et al. 1992). Endothelial nitric oxide synthase (eNOS) regulates vascular function by generating nitric oxide which is involved in endothelium-dependent relaxation, and control of blood pressure and vascular tone. It has been shown that cysteine residues are important for the maintenance of normal eNOS function. Under oxidative stress, S-glutathionylation of eNOS was induced by GSSG at residue sites Cys 689 and Cys 908, resulting in a decrease in eNOS activity and an increase in superoxide generation, also known as eNOS uncoupling. Furthermore, eNOS S-glutathionylation was shown to be abundant in the vessel walls of spontaneously hypertension rats (SHRs), in contrast to non-hypertensive rats. &nbsp;&nbsp;SHRs demonstrated impaired endothelium-dependent vasodilation, which was reversible upon administration of the reducing agent,&nbsp;dithiothreitol (Chen et al. 2010). &nbsp;Similarly in human aortic endothial cells, exposure to ultrafine particles caused a decrease in NO production in a dose-depedent manner. &nbsp;This was shown to be prevented upon over-expression of glutaredoxin-1, which inhibits eNOS S-glutathionylation (Du et al. 2013).</p>
  • <br>
  • <h4>How it is Measured or Detected</h4>
  • <p>NO production can be measured through the conversion of L-arginine to L-citrulline (de Bono et al., 2007) , in situ fluorescent signal detection with fluorescent indicator DAF-2 DA (Itoh et al., 2000; Nagata et al., 1999; Qiu et al., 2001), EPR spin-trapping (Xia et al., 2000), and the determination of total nitrate and nitrite concentration (Crabtree et al., 2009; Du et al., 2013).</p>
  • <p>There are four general approaches to detect protein S-glutathionylation (Pastore and Piemonte, 2013).</p>
  • <ol>
  • <li><strong>Quantification of Total S-Glutathionylated Proteins</strong>: Use sample lysis or homogenization in non-reducing buffer containing N-ethylmaleimide to eliminate thiols, followed by protein precipitation, reduction of gluthionyl-protein adducts, and derivatization of protein thiols or free glutathione with fluorescence probes. Fluorescence can be measured by fluorometric analysis with or without prior HPLC separation. This method allows for quantification of glutathionylated proteins but cannot detect glutathione adducts on specific proteins.</li>
  • <li><strong>Labeling of Glutathione</strong>: Use <sup>35</sup>S-cysteine radiolabeling or biotin labeling to detect glutathione adducts on S-thiolated proteins.</li>
  • <li><strong>Use of Anti-Glutathione Antibodies</strong>: Use commercially available anti-glutathione to detect glutathionylated proteins by Western blots, immunoprecipitation or immunocytolocalization. This method is useful for analysis of individual proteins like eNOS but not for large-scale detection of glutathionylated proteins.</li>
  • <li><strong>Top-Down Proteomic Approach</strong>: Use liquid chromatography-coupled mass spectrometry to identify S-glutathionylated proteins on whole protein extract from cells without using labeling or anti-glutathione antibody.</li>
  • </ol>
  • <br>
  • <h4>References</h4>
  • <p>de Bono, J.P., Warrick, N., Bendall, J.K., Channon, K.M., and Alp, N.J. (2007). Radiochemical HPLC detection of arginine metabolism: Measurement of nitric oxide synthesis and arginase activity in vascular tissue. Nitric Oxide 16, 1&ndash;9.</p>
  • <p>Chen, X., Xu, J., Feng, Z., Fan, M., Han, J., and Yang, Z. (2010). Simvastatin combined with nifedipine enhances endothelial cell protection by inhibiting ROS generation and activating Akt phosphorylation. Acta Pharmacol. Sin. 31, 813&ndash;820.</p>
  • <p>Chen, C.-A., Wang, T.-Y., Varadharaj, S., Reyes, L.A., Hemann, C., Talukder, M.A.H., Chen, Y.-R., Druhan, L.J., and Zweier, J.L. (2010). S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature 468, 1115&ndash;1118.</p>
  • <p>Crabtree, M.J., Tatham, A.L., Al-Wakeel, Y., Warrick, N., Hale, A.B., Cai, S., Channon, K.M., and Alp, N.J. (2009). Quantitative regulation of intracellular endothelial nitric-oxide synthase (eNOS) coupling by both tetrahydrobiopterin-eNOS stoichiometry and biopterin redox status: insights from cells with tet-regulated GTP cyclohydrolase I expression. J. Biol. Chem. 284, 1136&ndash;1144.</p>
  • <p>Chen CA,&nbsp;Lin CH,&nbsp;Druhan LJ,&nbsp;Wang TY,&nbsp;Chen YR,&nbsp;Zweier JL. &nbsp;Superoxide induces endothelial nitric-oxide synthase protein thiyl radical formation, a novel mechanism regulating&nbsp;eNOS&nbsp;function and coupling. &nbsp;J Biol Chem.&nbsp;2011 286(33):29098-107.</p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p>Du, Y., Navab, M., Shen, M., Hill, J., Pakbin, P., Sioutas, C., Hsiai, T.K., and Li, R. (2013). Ambient ultrafine particles reduce endothelial nitric oxide production via S-glutathionylation of eNOS. Biochem. Biophys. Res. Commun. 436, 462&ndash;466.</p>
  • <p>Dumitrescu, C., Biondi, R., Xia, Y., Cardounel, A.J., Druhan, L.J., Ambrosio, G., and Zweier, J.L. (2007). Myocardial ischemia results in tetrahydrobiopterin (BH4) oxidation with impaired endothelial function ameliorated by BH4. Proc. Natl. Acad. Sci. U. S. A. 104, 15081&ndash;15086.</p>
  • <p>Itoh, Y., Ma, F.H., Hoshi, H., Oka, M., Noda, K., Ukai, Y., Kojima, H., Nagano, T., and Toda, N. (2000). Determination and bioimaging method for nitric oxide in biological specimens by diaminofluorescein fluorometry. Anal. Biochem. 287, 203&ndash;209.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Luo, Z., Fujio, Y., Kureishi, Y., Rudic, R.D., Daumerie, G., Fulton, D., Sessa, W.C., and Walsh, K. (2000). Acute modulation of endothelial Akt/PKB activity alters nitric oxide&ndash;dependent vasomotor activity in vivo. J. Clin. Invest. 106, 493&ndash;499.</p>
  • <p>Nagata, N., Momose, K., and Ishida, Y. (1999). Inhibitory effects of catecholamines and anti-oxidants on the fluorescence reaction of 4,5-diaminofluorescein, DAF-2, a novel indicator of nitric oxide. J. Biochem. (Tokyo) 125, 658&ndash;661.</p>
  • <p>Qiu, W., Kass, D.A., Hu, Q., and Ziegelstein, R.C. (2001). Determinants of shear stress-stimulated endothelial nitric oxide production assessed in real-time by 4,5-diaminofluorescein fluorescence. Biochem. Biophys. Res. Commun. 286, 328&ndash;335.</p>
  • <p>Pastore, A., and Piemonte, F. (2013). Protein glutathionylation in cardiovascular diseases. Int. J. Mol. Sci. 14, 20845&ndash;20876.</p>
  • <p>Xia, Y., Cardounel, A.J., Vanin, A.F., and Zweier, J.L. (2000). Electron paramagnetic resonance spectroscopy with N-methyl-D-glucamine dithiocarbamate iron complexes distinguishes nitric oxide and nitroxyl anion in a redox-dependent manner: applications in identifying nitrogen monoxide products from nitric oxide synthase. Free Radic. Biol. Med. 29, 793&ndash;797.</p>
  • <p>Schuppe I, Mold&eacute;us P, and Cotgreave IA. Protein-specific S-thiolation in human endothelial cells during oxidative stress. (1992) Biochem. Pharmacol. 44: 1757&ndash;1764.</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <div>
  • <div>
  • <h4><a href="/events/937">937: KE7 : Impaired, Vasodilation</a><br></h4>
  • <h5>Short Name: Impaired, Vasodilation</h5>
  • <h4><a href="/events/935">Event: 935: KE2 : Decrease, GTPCH-1</a><br></h4>
  • <h5>Short Name: Decrease, GTPCH-1</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>vasodilation</td>
  • <td>blood vessel</td>
  • <td>abnormal</td>
  • <td>proteasome complex disassembly</td>
  • <td>GTP cyclohydrolase 1</td>
  • <td>decreased</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">149: Peptide Oxidation Leading to Hypertension</a></td>
  • <td><a href="/aops/149">Aop:149 - Peptide Oxidation Leading to Hypertension</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Organization</h4>
  • <h4>Biological Context</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Organ</td>
  • <td>Cellular</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • <h4>Organ term</h4>
  • <h4>Cell term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Organ term</th>
  • <th>Cell term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>circulatory system</td>
  • <td>endothelial cell of vascular tree</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h4>Evidence Supporting Applicability of this Event</h4>
  • <h4>Domain of Applicability</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Oryctolagus cuniculus</td>
  • <td>Oryctolagus cuniculus</td>
  • <td>Weak</td>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9986" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Moderate</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Moderate</td>
  • <td>Low</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <div>
  • <p>Vasodilation has been observed in humans, rabbits, mice and rats.</p>
  • <p>Several studies showed decreased GTPCH-1 activity and/or protein expression in cardiac reperfusion patients, bovine endothelial cells, a mouse model of diabetes and a rat model of hypertension (Cervantes-P&eacute;rez et al., 2012; Abdelghany et al., 2017; Jayaram et al., 2015; Zhao et al., 2013).</p>
  • <p>Furthermore, mice deficient in GTPCH-1&nbsp;demonstrate decreased BH4 bioavailability, increased eNOS uncoupling, pulmonary vascular resistance and pulmonary hypertension (Belik et al. 2011, Nandi et al. 2005, Khoo et al. 2005).</p>
  • <br>
  • </div>
  • <!-- event text -->
  • <h4>How this Key Event Works</h4>
  • <p>Vasodilation refers to the widening or increase in the diameter of blood vessels (e.g. large arteries, large veins, small arterioles) that is caused by the relaxation of vascular smooth muscle cells (VSMCs) within the walls of blood vessels, thus increasing blood flow and decreasing arterial blood pressure and heart rate (Siddiqui, 2011). VSMC relaxation is regulated through a number of mechanisms, including cyclic GMP-dependent hyperpolarization and relaxation via nitric oxide (NO), cAMP-dependent hyperpolarization via prostaglandins, and stimulation of potassium channels via endothelial-derived hyperpolarizing factors (Durand and Gutterman, 2013). Under oxidative stress, decreased NO bioavailability results in impaired vasodilation, which is associated with cardiovascular diseases such as hypertension (Silva et al., 2012).</p>
  • <h4>Key Event Description</h4>
  • <p>Guanosine triphosphate cyclohydrolase-1 (GTPCH-1)&nbsp;is the rate-limiting enzyme in the de novo biosynthesis of tetrahydrobiopterin (BH4), an essential cofactor for endothelial nitric oxide synthase (eNOS) and nitric oxide generation (Wang et al., 2008). GTPCH-1 catalyzes the rearrangement of GTP to 7-dihydroneopterin triphosphate, which is converted to BH4 through sequential actions of pyruvoyl tetrahydrobiopterin synthase and sepiapterin reductase. GTPCH-1 activity is regulated in a negative feedback by levels of BH4 which promotes binding of GTPCH-1 with its inhibitor GTPCH feedback regulatory protein (GFRP), but phosphorylation of GTPCH-1 reduces its binding to GFRP and prevents this negative feedback (Chen et al., 2011). &nbsp;Loss or inactivation of GTPCH-1 results in decreased BH4 levels, which causes eNOS uncoupling.</p>
  • <br>
  • <h4>How it is Measured or Detected</h4>
  • <p>Endothelium-dependent vasodilation can be measured using invasive and non-invasive methods (Raitakari and Celermajer, 2000). For the invasive approach, vasodilation is measured after intra-arterial pharmacologic stimulation with substances that enhance NO release (e.g. acetylcholine, bradykinin). The non-invasive ultrasound-based method evaluates flow-mediated vasodilation (FMD) in the superficial arteries, such as brachial, radial, or femoral vessels.</p>
  • <p>Guidelines for the measurement of FMD have been published (Corretti et al. 2002).</p>
  • <p>The activity of GTPCH-1 can be detected through the quantification of neopterin by high-performance liquid chromatography (HPLC) after the conversion of enzymatically formed dihydroneopterin triphosphate into neopterin by sequential iodine oxidation and dephosphorylation.</p>
  • <br>
  • <h4>References</h4>
  • <p>Guidelines for the ultrasound assessment of endothelial-dependent flow-mediated vasodilation of the brachial artery</p>
  • <p>AbdelGhany, T., Ismail, R., Elmahdy, M., Mansoor F, Zweier J, Lowe, F., and Zweier, JL. (2017). Cigarette Smoke Constituents Cause Endothelial Nitric Oxide Synthase Dysfunction and Uncoupling due to Depletion of Tetrahydrobiopterin with Degradation of GTP Cyclohydrolase.&nbsp;&nbsp;Nitric Oxide&nbsp;(Under review).</p>
  • <p>Corretti&nbsp;MC, Anderson&nbsp;TJ, Benjamin&nbsp;EJ, Celermajer D, Charbonneau F, Creager MA,&nbsp;Deanfield J,&nbsp;Drexler H, Gerhard-Herman M, Herrington D, Vallance P, Vita J, Vogel R. &nbsp;Guidelines for the ultrasound assessment of endothelial-dependent flow-mediated vasodilation of the brachial artery. &nbsp;Journal of the American College of Cardiology,&nbsp;2002,&nbsp;39&nbsp;(2)&nbsp;257-265</p>
  • <p>Belik J,&nbsp;McIntyre BA,&nbsp;Enomoto M,&nbsp;Pan J,&nbsp;Grasemann H,&nbsp;Vasquez-Vivar J. &nbsp;Pulmonary hypertension in the newborn GTP cyclohydrolase I-deficient mouse. &nbsp;Free Radic Biol Med.&nbsp;2011, 51(12):2227-33.</p>
  • <p>Durand, M.J., and Gutterman, D.D. (2013). Diversity in mechanisms of endothelium-dependent vasodilation in health and disease. Microcirc. N. Y. N 1994 20, 239&ndash;247.</p>
  • <p>Cervantes-P&eacute;rez, L.G., Ibarra-Lara, M. de la L., Escalante, B., Del Valle-Mondrag&oacute;n, L., Vargas-Robles, H., P&eacute;rez-Severiano, F., Pastel&iacute;n, G., and S&aacute;nchez-Mendoza, M.A. (2012). Endothelial nitric oxide synthase impairment is restored by clofibrate treatment in an animal model of hypertension. Eur. J. Pharmacol. 685, 108&ndash;115.</p>
  • <p>Raitakari, O.T., and Celermajer, D.S. (2000). Flow-mediated dilatation. Br. J. Clin. Pharmacol. 50, 397&ndash;404.</p>
  • <p>Chen, W., Li, L., Brod, T., Saeed, O., Thabet, S., Jansen, T., Dikalov, S., Weyand, C., Goronzy, J., and Harrison, D.G. (2011). Role of increased guanosine triphosphate cyclohydrolase-1 expression and tetrahydrobiopterin levels upon T cell activation. J. Biol. Chem. 286, 13846&ndash;13851.</p>
  • <p>Siddiqui, A. (2011). Effects of Vasodilation and Arterial Resistance on Cardiac Output. J. Clin. Exp. Cardiol. 02.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Silva, B.R., Pernomian, L., and Bendhack, L.M. (2012). Contribution of oxidative stress to endothelial dysfunction in hypertension. Front. Physiol. 3, 441.</p>
  • <p>Khoo JP, Zhao L, Alp NJ, Bendall JK, Nicoli T, Rockett K, et al. Pivotal role for endothelial tetrahydrobiopterin in pulmonary hypertension.&nbsp;Circulation.&nbsp;2005;111:2126&ndash;33</p>
  • <p>Nandi M, Miller A, Stidwill R, Jacques TS, Lam AA, Haworth S, et al. Pulmonary hypertension in a GTP-cyclohydrolase 1-deficient mouse.&nbsp;Circulation.&nbsp;2005;111:2086&ndash;90</p>
  • <p>Wang, S., Xu, J., Song, P., Wu, Y., Zhang, J., Chul Choi, H., and Zou, M.-H. (2008). Acute inhibition of guanosine triphosphate cyclohydrolase 1 uncouples endothelial nitric oxide synthase and elevates blood pressure. Hypertension 52, 484&ndash;490.</p>
  • <p>Zhao, Y., Wu, J., Zhu, H., Song, P., and Zou, M.-H. (2013). Peroxynitrite-dependent zinc release and inactivation of guanosine 5&rsquo;-triphosphate cyclohydrolase 1 instigate its ubiquitination in diabetes. Diabetes 62, 4247&ndash;4256.</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <div>
  • <div>
  • <h4><a href="/events/951">951: KE8 : Increase, Vascular Resistance</a><br></h4>
  • <h5>Short Name: Increase, Vascular Resistance</h5>
  • <h4><a href="/events/934">Event: 934: KE3 : Decrease, Tetrahydrobiopterin</a><br></h4>
  • <h5>Short Name: Decrease, Tetrahydrobiopterin</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>increased systemic vascular resistance</td>
  • <td></td>
  • <td>increased</td>
  • <td>biosynthetic process</td>
  • <td>5,6,7,8-tetrahydrobiopterin</td>
  • <td>decreased</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">149: Peptide Oxidation Leading to Hypertension</a></td>
  • <td><a href="/aops/149">Aop:149 - Peptide Oxidation Leading to Hypertension</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • <h4>Stressors</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Name</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Reactive oxygen species</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Organization</h4>
  • <h4>Biological Context</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Organ</td>
  • <td>Cellular</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • <h4>Organ term</h4>
  • <h4>Cell term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Organ term</th>
  • <th>Cell term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>circulatory system</td>
  • <td>endothelial cell of vascular tree</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h4>Evidence Supporting Applicability of this Event</h4>
  • <h4>Domain of Applicability</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Weak</td>
  • <td>Low</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <div>
  • <p>Increased SVR was observed in humans, pigs and rats (Siddiqui 2011,&nbsp;Dikalova et al. 2016,&nbsp;</p>
  • <p>&nbsp;</p>
  • <p>Decreased BH4 is observed in humans (Jayaram et al., 2015), cows (Abdelghany et al., 2017; Whitsett et al., 2007, Wang et al., 2008), mice (Adlam et al., 2012; Chuaiphichai et al., 2014; Crabtree et al., 2009; Tatham et al., 2009; Wang et al., 2008) and rats (Cervantes-P&eacute;rez et al., 2012).</p>
  • <br>
  • </div>
  • <!-- event text -->
  • <h4>How this Key Event Works</h4>
  • <p>Vascular resistance is the resistance to blood flow in the circulatory system (Siddiqui, 2011). Systemic vascular resistance (SVR), also known as total peripheral resistance, refers specifically to the resistance to blood flow offered by the peripheral circulation. Vasodilation decreases SVR while vasoconstriction or <a href="/wiki/index.php/Event:937" title="Event:937">impaired vasodilation</a> increases SVR.</p>
  • <h4>Key Event Description</h4>
  • <p>Tetrahydrobiopterin (BH4) is an essential cofactor for a group of enzymes including aromatic acid hydroxylases, nitric oxide synthase (NOS) isoforms, and alkylglycerol monooxygenase (Wang et al., 2014). BH4 is synthesized from guanosine triphosphate through sequential reactions catalyzed by enzymes <a href="/wiki/index.php/Event:935" title="Event:935">GTPCH-1</a>, pyruvoyl tetrahydropterin synthase, and sepiapterin reductase (Tatham et al., 2009). During NOS catalysis, BH4 donates electrons to the ferrous-dioxygen complex in the oxygenase domain, leading to oxidation of L-arginine to N-hydroxy-Larginine and eventually conversion to citrulline and nitric oxide production (Chen et al., 2011; Crabtree et al., 2009). BH4 also stabilizes dimers of NOS isoforms, which is required for their enzymatic activity. When BH4 levels are decreased or limited, for example under oxidative stress conditions, BH4 can be oxidized to dihydrobiopterin (BH2) and then converted to biopterin. This reduction in BH4 availability results in NOS uncoupling where NOS is uncoupled from L-arginine oxidation and superoxide (or other reactive species) is produced rather than nitric oxide (Carnicer et al., 2012). Decreased BH4 have been demonstrated in a variety of vascular diseases such as hypertension, diabetes and atherosclerosis where endothelial dysfunction occurs.</p>
  • <br>
  • <h4>How it is Measured or Detected</h4>
  • <p>Vascular resistance cannot be measured by any direct means, but can be calculated using a formula: (mean arterial pressure minus mean right arterial pressure) divided by cardiac output (Siddiqui 2011).</p>
  • <p>Levels of BH4, BH2 and biopterin levels can be determined by reverse-phase high-performance liquid chromatography (HPLC) followed by electrochemical detection (for BH4) and fluorescence detection (for BH2 and biopterin) (Howells et al., 1986).</p>
  • <p>A LC-MS/MS method has been published by Zhao et al. (2009), which was validated for detection in&nbsp;human, monkey, dog, rabbit, rat and mouse plasma, and used to support a successful drug approval submission.</p>
  • <p>ELISA kits for BH4 are also commercially available.</p>
  • <p>In each case, care must be taken to protect the sample from oxidation, and BH4 is highly redox sensitive. &nbsp;Dithioerythritol is commonly used as a preservation agent.</p>
  • <p>&nbsp;</p>
  • <br>
  • <h4>References</h4>
  • <p>Siddiqui, A. (2011). Effects of Vasodilation and Arterial Resistance on Cardiac Output. J. Clin. Exp. Cardiol. 02.</p>
  • <p>AbdelGhany, T., Ismail, R., Elmahdy, M., Mansoor F, Zweier J, Lowe, F., and Zweier, JL. (2017). Cigarette Smoke Constituents Cause Endothelial Nitric Oxide Synthase Dysfunction and Uncoupling due to Depletion of Tetrahydrobiopterin with Degradation of GTP Cyclohydrolase.&nbsp;&nbsp;Nitric Oxide&nbsp;(Under review).</p>
  • <p>Dikalova A,&nbsp;Aschner JL,&nbsp;Kaplowitz MR,&nbsp;Summar M,&nbsp;Fike CD. &nbsp;Tetrahydrobiopterin oral therapy recouples&nbsp;eNOS&nbsp;and ameliorates chronic hypoxia-induced pulmonary hypertension in newborn pigs. &nbsp;Am J Physiol Lung Cell Mol Physiol.&nbsp;2016, 1;311(4):L743-L753.</p>
  • <p>Adlam, D., Herring, N., Douglas, G., De Bono, J.P., Li, D., Danson, E.J., Tatham, A., Lu, C.-J., Jennings, K.A., Cragg, S.J., et al. (2012). Regulation of &beta;-adrenergic control of heart rate by GTP-cyclohydrolase 1 (GCH1) and tetrahydrobiopterin. Cardiovasc. Res. 93, 694&ndash;701.</p>
  • <p>Carnicer, R., Hale, A.B., Suffredini, S., Liu, X., Reilly, S., Zhang, M.H., Surdo, N.C., Bendall, J.K., Crabtree, M.J., Lim, G.B.S., et al. (2012). Cardiomyocyte GTP cyclohydrolase 1 and tetrahydrobiopterin increase NOS1 activity and accelerate myocardial relaxation. Circ. Res. 111, 718&ndash;727.</p>
  • <p>Cervantes-P&eacute;rez, L.G., Ibarra-Lara, M. de la L., Escalante, B., Del Valle-Mondrag&oacute;n, L., Vargas-Robles, H., P&eacute;rez-Severiano, F., Pastel&iacute;n, G., and S&aacute;nchez-Mendoza, M.A. (2012). Endothelial nitric oxide synthase impairment is restored by clofibrate treatment in an animal model of hypertension. Eur. J. Pharmacol. 685, 108&ndash;115.</p>
  • <p>Chen, W., Li, L., Brod, T., Saeed, O., Thabet, S., Jansen, T., Dikalov, S., Weyand, C., Goronzy, J., and Harrison, D.G. (2011). Role of increased guanosine triphosphate cyclohydrolase-1 expression and tetrahydrobiopterin levels upon T cell activation. J. Biol. Chem. 286, 13846&ndash;13851.</p>
  • <p>Crabtree, M.J., Tatham, A.L., Al-Wakeel, Y., Warrick, N., Hale, A.B., Cai, S., Channon, K.M., and Alp, N.J. (2009). Quantitative regulation of intracellular endothelial nitric-oxide synthase (eNOS) coupling by both tetrahydrobiopterin-eNOS stoichiometry and biopterin redox status: insights from cells with tet-regulated GTP cyclohydrolase I expression. J. Biol. Chem. 284, 1136&ndash;1144.</p>
  • <p>Chuaiphichai, S., McNeill, E., Douglas, G., Crabtree, M.J., Bendall, J.K., Hale, A.B., Alp, N.J., and Channon, K.M. (2014). Cell-autonomous role of endothelial GTP cyclohydrolase 1 and tetrahydrobiopterin in blood pressure regulation. Hypertension 64, 530&ndash;540.</p>
  • <p>Howells, D.W., Smith, I., and Hyland, K. (1986). Estimation of tetrahydrobiopterin and other pterins in cerebrospinal fluid using reversed-phase high-performance liquid chromatography with electrochemical and fluorescence detection. J. Chromatogr. 381, 285&ndash;294.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Tatham, A.L., Crabtree, M.J., Warrick, N., Cai, S., Alp, N.J., and Channon, K.M. (2009). GTP cyclohydrolase I expression, protein, and activity determine intracellular tetrahydrobiopterin levels, independent of GTP cyclohydrolase feedback regulatory protein expression. J. Biol. Chem. 284, 13660&ndash;13668.</p>
  • <p>Wang, Q., Yang, M., Xu, H., and Yu, J. (2014). Tetrahydrobiopterin improves endothelial function in cardiovascular disease: a systematic review. Evid.-Based Complement. Altern. Med. ECAM 2014, 850312.</p>
  • <p>Whitsett, J., Picklo, M.J., and Vasquez-Vivar, J. (2007). 4-Hydroxy-2-nonenal increases superoxide anion radical in endothelial cells via stimulated GTP cyclohydrolase proteasomal degradation. Arterioscler. Thromb. Vasc. Biol. 27, 2340&ndash;2347.</p>
  • <p>Zhao Y,&nbsp;Cao J,&nbsp;Chen YS,&nbsp;Zhu Y,&nbsp;Patrick C,&nbsp;Chien B,&nbsp;Cheng A,&nbsp;Foehr ED. &nbsp;Detection of tetrahydrobiopterin by LC-MS/MS in plasma from multiple species. &nbsp;Bioanalysis.&nbsp;2009;1(5):895-903.</p>
  • <p>&nbsp;</p>
  • <p>&nbsp;</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <div>
  • <div>
  • <h4><a href="/events/973">973: KE5 : Decrease, AKT/eNOS activity</a><br></h4>
  • <h5>Short Name: Decrease, AKT/eNOS activity</h5>
  • <h4><a href="/events/932">Event: 932: KE4 : Uncoupling, eNOS</a><br></h4>
  • <h5>Short Name: Uncoupling, eNOS</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>catalytic activity</td>
  • <td></td>
  • <td>nitric oxide synthase, endothelial</td>
  • <td>decreased</td>
  • </tr>
  • <tr>
  • <td>catalytic activity</td>
  • <td>AKT kinase</td>
  • <td>decreased</td>
  • <td>functional change</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">149: Peptide Oxidation Leading to Hypertension</a></td>
  • <td><a href="/aops/149">Aop:149 - Peptide Oxidation Leading to Hypertension</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • <h4>Stressors</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Name</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Reactive oxygen species</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Organization</h4>
  • <h4>Biological Context</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Cellular</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • <h4>Cell term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Cell term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>endothelial cell of vascular tree</td>
  • <td>endothelial cell</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h4>Evidence Supporting Applicability of this Event</h4>
  • <h4>Domain of Applicability</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Strong</td>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Strong</td>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <div>
  • <p>Decreased Akt and eNOS activity was observed in humans, cows, mice and rats following exposure to stressors.</p>
  • <p>Cigarette smoke exposure was shown to inhibit the phosphorylation of AKT and eNOS in VEGF-stimulated human umbilical vein endothelial cells (HUVECs), resulting in decreased NO levels (Michaud et al. 2006).</p>
  • <p>In rat aortic rings, exposure to methylglyoxal and high concentrations of glucose decreased endothelium-dependent relaxation. &nbsp;Further experiments in rat endothelial cells and HUVECs demonstrated a reduction in eNOS phosphorylation and&nbsp;activity, and reduced NO levels in response to the same stressors&nbsp;(Dhar et al. 2010).</p>
  • <p>In bovine aortic endothelial cells, AKT and eNOS phosphorylation were decreased following exposure to the peroxynitrite source; SIN-1, with an associated reduction in NO bioavailability. &nbsp;These effects were ameliorated by treatment with the ROS scavenger DMPO (Das et al. 2014).</p>
  • <p>eNOS knockout mice are routinely used as models of hypertension. &nbsp;Such mice display reduced bioavailability of NO and impaired vasodilation (Huang et al. 1995).</p>
  • <p>Reduced AKT/eNOS phosphorylation was reported under conditions of hyperglycaemia (in mice) and in HUVECs following treatment with high concentrations of glucose. &nbsp;Aortic rings from hyperglycaemic mice demonstrated impaired vasodilation. &nbsp;Resveratrol treatment was shown to improve vasodilation and eNOS phosphorylation in wild-type mice, but not AKT knockout mice. &nbsp;Transfection of HUVECs with AKT siRNA abolished resveratrol-enhanced eNOS phosphorylation and NO release&nbsp;(Li et al. 2017),</p>
  • <p>eNOS uncoupling has been demonstrated in humans, cows, mice and rats (Chen et al., 2010; Crabtree et al., 2009; De Pascali et al., 2014; Du et al., 2013; Jayaram et al., 2015).</p>
  • <br>
  • </div>
  • <!-- event text -->
  • <h4>How this Key Event Works</h4>
  • <p>Endothelial nitric oxide synthase (eNOS) is responsible for the generation of nitric oxide (NO), which is an important regulator of vascular homeostasis. The activity of eNOS can be regulated through calmodulin-mediated dimerization, tetrahydrobiopterin-mediated conversion of L-arginine to L-citrulline, protein-protein interactions with heat shock protein 90 and caveolin, S-nitrosylation, acetylation and phosphorylation (Atochin et al., 2007; Qian and Fulton, 2013). eNOS has been shown to be phosphorylated at multiple sites including tyrosine (Y), serine (Ser) and threonine (Thr) residues. Serine-threonine protein kinase AKT, a multifunctional regulator of cellular processes like glucose metabolism and proliferation, can directly phosphorylate eNOS at Ser1177/Ser1179, leading to increased eNOS enzymatic activity and subsequent NO production (Dimmeler et al., 1999; Fulton et al., 1999). Inhibition of AKT&nbsp;or a mutation of the AKT phosphorylation site on eNOS attenuates eNOS phosphorylation and its activity, resulting in decreased NO bioavailability and endothelial dysfunction (Dimmler et al. 1999)</p>
  • <h4>Key Event Description</h4>
  • <p>Endothelial nitric oxide synthase (eNOS) is responsible for the generation of vascular nitric oxide (NO), a protective molecule that is involved in the regulation of endothelium-dependent vasodilation, vascular tone, and blood pressure (F&ouml;rstermann and M&uuml;nzel, 2006). To generate NO, eNOS hydroxylates L-arginine to N-hydroxy-L-arginine and then oxidizes N-hydroxy-L-arginine to L-citrulline and NO. This enzymatic process requires NADPH, Ca2+/calmondulin, flavin mononucleotide, flavin adenine dinucleotide and its cofactor <a href="/wiki/index.php/Event:934" title="Event:934">tetrahydrobiopterin (BH4)</a>. Limiting BH4 levels or <a href="/wiki/index.php/Event:927" title="Event:927">S-glutathionylation of eNOS</a> can lead to eNOS uncoupling in which eNOS produces superoxide (or other reactive oxygen species) and less NO. The uncoupling of eNOS has been demonstrated to cause endothelial dysfunction, and is implicated in a number of cardiovascular diseases such as hypertension, atherosclerosis, hypercholesterolemia, and diabetes mellitus (Dumitrescu et al., 2007).</p>
  • <br>
  • <h4>How it is Measured or Detected</h4>
  • <p>Western blot analysis can be performed to determine the expression levels of phosphorylated eNOS, phosphorylated Akt, total Akt and total eNOS proteins using the appropriate anti-phospho-eNOS, anti-phospho-Akt, anti-eNOS, and anti-Akt antibodies. Alternatively, eNOS activity can be measured using the conversion of L-arginine to L-citrulline assay.</p>
  • <p>ELISA kits for AKT/eNOS and phospho AKT/eNOS expression are commercially available.</p>
  • <p>The activity of eNOS can be measured indirectly through superoxide and NO production. Superoxides can be detected using several standard methods including lucigenin-enhanced chemiluminescence (M&uuml;nzel et al., 2002; Tarpey et al., 1999), electron paramagnetic resonance (EPR) spin-trapping (Roubaud et al., 1997), and HPLC/fluorescence detector-based assay using dihydroethidium (Fink et al., 2004; Zhao et al., 2003). NO production can be measured through the conversion of L-arginine to L-citrulline (de Bono et al., 2007) , in situ fluorescent signal detection with fluorescent indicator DAF-2 DA (Itoh et al., 2000; Nagata et al., 1999; Qiu et al., 2001), EPR spin-trapping (Xia et al., 2000), and the determination of total nitrate and nitrite concentration (Crabtree et al., 2009; Du et al., 2013).</p>
  • <br>
  • <h4>References</h4>
  • <p>Atochin, D.N., Wang, A., Liu, V.W.T., Critchlow, J.D., Dantas, A.P.V., Looft-Wilson, R., Murata, T., Salomone, S., Shin, H.K., Ayata, C., et al. (2007). The phosphorylation state of eNOS modulates vascular reactivity and outcome of cerebral ischemia in vivo. J. Clin. Invest. 117, 1961&ndash;1967.</p>
  • <p>de Bono, J.P., Warrick, N., Bendall, J.K., Channon, K.M., and Alp, N.J. (2007). Radiochemical HPLC detection of arginine metabolism: Measurement of nitric oxide synthesis and arginase activity in vascular tissue. Nitric Oxide 16, 1&ndash;9.</p>
  • <p>Das A,&nbsp;Gopalakrishnan B,&nbsp;Druhan LJ,&nbsp;Wang TY,&nbsp;De Pascali F,&nbsp;Rockenbauer A,&nbsp;Racoma I,&nbsp;Varadharaj S,&nbsp;Zweier JL,&nbsp;Cardounel AJ,&nbsp;Villamena FA. &nbsp;Reversal&nbsp;of&nbsp;SIN-1-induced&nbsp;eNOS&nbsp;dysfunction&nbsp;by the&nbsp;spin&nbsp;trap,&nbsp;DMPO, in&nbsp;bovine&nbsp;aortic&nbsp;endothelialcells&nbsp;via&nbsp;eNOS&nbsp;phosphorylation. &nbsp;Br J Pharmacol.&nbsp;2014, 171(9):2321-34. doi: 10.1111/bph.12572.</p>
  • <p>Chen, X., Xu, J., Feng, Z., Fan, M., Han, J., and Yang, Z. (2010). Simvastatin combined with nifedipine enhances endothelial cell protection by inhibiting ROS generation and activating Akt phosphorylation. Acta Pharmacol. Sin. 31, 813&ndash;820.</p>
  • <p>Dhar A,&nbsp;Dhar I,&nbsp;Desai KM,&nbsp;Wu L. &nbsp;Methylglyoxal&nbsp;scavengers attenuate endothelial dysfunction induced by&nbsp;methylglyoxal&nbsp;and high concentrations of glucose. &nbsp;Br J Pharmacol.&nbsp;2010, 161(8):1843-56.</p>
  • <p>Crabtree, M.J., Tatham, A.L., Al-Wakeel, Y., Warrick, N., Hale, A.B., Cai, S., Channon, K.M., and Alp, N.J. (2009). Quantitative regulation of intracellular endothelial nitric-oxide synthase (eNOS) coupling by both tetrahydrobiopterin-eNOS stoichiometry and biopterin redox status: insights from cells with tet-regulated GTP cyclohydrolase I expression. J. Biol. Chem. 284, 1136&ndash;1144.</p>
  • <p>Dimmeler, S., Fleming, I., Fisslthaler, B., Hermann, C., Busse, R., and Zeiher, A.M. (1999). Activation of nitric oxide synthase in endothelial cells by Akt-dependent phosphorylation. Nature 399, 601&ndash;605.</p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p>Fulton, D., Gratton, J.P., McCabe, T.J., Fontana, J., Fujio, Y., Walsh, K., Franke, T.F., Papapetropoulos, A., and Sessa, W.C. (1999). Regulation of endothelium-derived nitric oxide production by the protein kinase AKT. Nature 399, 597&ndash;601.</p>
  • <p>Du, Y., Navab, M., Shen, M., Hill, J., Pakbin, P., Sioutas, C., Hsiai, T.K., and Li, R. (2013). Ambient ultrafine particles reduce endothelial nitric oxide production via S-glutathionylation of eNOS. Biochem. Biophys. Res. Commun. 436, 462&ndash;466.</p>
  • <p>Huang PL,&nbsp;Huang Z,&nbsp;Mashimo H,&nbsp;Bloch KD,&nbsp;Moskowitz MA,&nbsp;Bevan JA,&nbsp;Fishman MC. &nbsp; &nbsp;Hypertension&nbsp;in&nbsp;mice&nbsp;lacking&nbsp;the&nbsp;gene&nbsp;for&nbsp;endothelial nitric oxide synthase. &nbsp;Nature.&nbsp;1995, 377(6546):239-42.</p>
  • <p>Dumitrescu, C., Biondi, R., Xia, Y., Cardounel, A.J., Druhan, L.J., Ambrosio, G., and Zweier, J.L. (2007). Myocardial ischemia results in tetrahydrobiopterin (BH4) oxidation with impaired endothelial function ameliorated by BH4. Proc. Natl. Acad. Sci. U. S. A. 104, 15081&ndash;15086.</p>
  • <p>Li JY,&nbsp;Huang WQ,&nbsp;Tu RH,&nbsp;Zhong GQ,&nbsp;Luo BB,&nbsp;He Y. &nbsp;Resveratrol rescues hyperglycemia-induced endothelial dysfunction via activation of&nbsp;Akt. &nbsp;Acta Pharmacol Sin.&nbsp;2017, 38(2):182-191.</p>
  • <p>Fink, B., Laude, K., McCann, L., Doughan, A., Harrison, D.G., and Dikalov, S. (2004). Detection of intracellular superoxide formation in endothelial cells and intact tissues using dihydroethidium and an HPLC-based assay. Am. J. Physiol. Cell Physiol. 287, C895&ndash;C902.</p>
  • <p>Michaud SE,&nbsp;Dussault S,&nbsp;Groleau J,&nbsp;Haddad P,&nbsp;Rivard A.J. Cigarette smoke exposure impairs VEGF-induced endothelial cell migration: role of NO and reactive oxygen species. &nbsp;Mol Cell Cardiol.&nbsp;2006 Aug;41(2):275-84.</p>
  • <p>F&ouml;rstermann, U., and M&uuml;nzel, T. (2006). Endothelial nitric oxide synthase in vascular disease: from marvel to menace. Circulation 113, 1708&ndash;1714.</p>
  • <p>Qian, J., and Fulton, D. (2013). Post-translational regulation of endothelial nitric oxide synthase in vascular endothelium. Oxid. Physiol. 4, 347.</p>
  • <p>Itoh, Y., Ma, F.H., Hoshi, H., Oka, M., Noda, K., Ukai, Y., Kojima, H., Nagano, T., and Toda, N. (2000). Determination and bioimaging method for nitric oxide in biological specimens by diaminofluorescein fluorometry. Anal. Biochem. 287, 203&ndash;209.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>M&uuml;nzel, T., Afanas&rsquo;ev, I.B., Kleschyov, A.L., and Harrison, D.G. (2002). Detection of Superoxide in Vascular Tissue. Arterioscler. Thromb. Vasc. Biol. 22, 1761&ndash;1768.</p>
  • <p>Nagata, N., Momose, K., and Ishida, Y. (1999). Inhibitory effects of catecholamines and anti-oxidants on the fluorescence reaction of 4,5-diaminofluorescein, DAF-2, a novel indicator of nitric oxide. J. Biochem. (Tokyo) 125, 658&ndash;661.</p>
  • <p>Qiu, W., Kass, D.A., Hu, Q., and Ziegelstein, R.C. (2001). Determinants of shear stress-stimulated endothelial nitric oxide production assessed in real-time by 4,5-diaminofluorescein fluorescence. Biochem. Biophys. Res. Commun. 286, 328&ndash;335.</p>
  • <p>Roubaud, V., Sankarapandi, S., Kuppusamy, P., Tordo, P., and Zweier, J.L. (1997). Quantitative measurement of superoxide generation using the spin trap 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide. Anal. Biochem. 247, 404&ndash;411.</p>
  • <p>Tarpey, M.M., White, C.R., Suarez, E., Richardson, G., Radi, R., and Freeman, B.A. (1999). Chemiluminescent Detection of Oxidants in Vascular Tissue Lucigenin But Not Coelenterazine Enhances Superoxide Formation. Circ. Res. 84, 1203&ndash;1211.</p>
  • <p>Xia, Y., Cardounel, A.J., Vanin, A.F., and Zweier, J.L. (2000). Electron paramagnetic resonance spectroscopy with N-methyl-D-glucamine dithiocarbamate iron complexes distinguishes nitric oxide and nitroxyl anion in a redox-dependent manner: applications in identifying nitrogen monoxide products from nitric oxide synthase. Free Radic. Biol. Med. 29, 793&ndash;797.</p>
  • <p>Zhao, H., Kalivendi, S., Zhang, H., Joseph, J., Nithipatikom, K., V&aacute;squez-Vivar, J., and Kalyanaraman, B. (2003). Superoxide reacts with hydroethidine but forms a fluorescent product that is distinctly different from ethidium: potential implications in intracellular fluorescence detection of superoxide. Free Radic. Biol. Med. 34, 1359&ndash;1368.</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <!-- ao table -->
  • <h3>Adverse Outcomes</h3>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Title</th>
  • <th>Short name</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/events/952">Hypertension</a></td>
  • <td>Hypertension</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <div>
  • <h4><a href="/events/952">952: Hypertension</a><br></h4>
  • <h5>Short Name: Hypertension</h5>
  • <h4><a href="/events/933">Event: 933: KE6 : Depletion, Nitric Oxide</a><br></h4>
  • <h5>Short Name: Depletion, Nitric Oxide</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>hypertension</td>
  • <td></td>
  • <td>increased</td>
  • <td>nitric oxide biosynthetic process</td>
  • <td>nitric oxide</td>
  • <td>decreased</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">149: Peptide Oxidation Leading to Hypertension</a></td>
  • <td>AdverseOutcome</td>
  • <td><a href="/aops/149">Aop:149 - Peptide Oxidation Leading to Hypertension</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • <h4>Stressors</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Name</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Reactive oxygen species</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Organization</h4>
  • <h4>Biological Context</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Individual</td>
  • <td>Cellular</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • <h4>Organ term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Organ term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>blood</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h4>Evidence Supporting Applicability of this Event</h4>
  • <h4>Domain of Applicability</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <div>
  • <p>Animal models including mouse and rat models are routinely used to study hypertension, and have been shown to reflect human physiology relating to hypertension&nbsp;(Leong et al., 2015).</p>
  • <p>NO depletion was observed in humans, cows, mice and rats (Chen et al., 2010; Crabtree et al., 2009; De Pascali et al., 2014; Du et al., 2013; Jayaram et al., 2015).</p>
  • <br>
  • </div>
  • <!-- event text -->
  • <h4>How this Key Event Works</h4>
  • <p>Hypertension is an important cardiovascular risk factor and considered one of the leading causes of cardiovascular morbidity and mortality (Kizhakekuttu and Widlansky, 2010). It is defined as a chronic elevation in blood pressure and is characterized by elevated systemic vascular resistance due to dysregulated vasomotor function and structural remodeling (Lee and Griendling, 2008). Although many genetic and environmental factors contribute to the development to hypertension, oxidative stress appears to be the main pathway involved in its pathogenesis. Excessive reactive oxygen species (ROS) contributes to <a href="/wiki/index.php/Event:932" title="Event:932">endothelial nitric oxide synthase (eNOS) uncoupling</a>, resulting in increased superoxide production but <a href="/wiki/index.php/Event:933" title="Event:933">decreased nitric oxide</a> (NO), a critical regulator of vascular homeostasis (Silva et al., 2012). Depletion of NO leads to <a href="/wiki/index.php/Event:937" title="Event:937">impaired endothelium-dependent vasodilation</a>, thus promoting endothelial dysfunction, which is a hallmark of hypertension.</p>
  • <h4>Key Event Description</h4>
  • <p>Nitric oxide (NO), constitutively produced by endothelial nitric oxide synthase (eNOS), is an important regulator of vascular homeostasis. Endothelial-derived NO promotes vasodilation and protects against atherogenesis through the inhibition of vascular smooth muscle cell proliferation and migration, platelet aggregation and adhesion, and leukocyte adherence. Its effects have an influence on vascular resistance, blood pressure, vascular remodeling and angiogenesis (Luo et al., 2000). Dysfunctional eNOS as a result of <a href="/wiki/index.php/Event:932" title="Event:932">eNOS uncoupling</a> leads to a decrease or loss of NO bioavailability and an elevation of superoxide production (Crabtree et al., 2009). The imbalance of NO and superoxide is associated with many disorders,&nbsp;such as hypertension, atherosclerosis, hypercholesterolemia, and diabetes mellitus.</p>
  • <br>
  • <h4>How it is Measured or Detected</h4>
  • <p>Arterial blood pressure is commonly measured using a sphygmomanometer, which provides systolic and diastolic blood pressure measurements in millimeters of mercury (mmHg).</p>
  • <p>Pathological hypertension is characterised according to current guidelines;&nbsp;https://www.nice.org.uk/guidance/cg127/evidence</p>
  • <p>Stage 1 hypertension : Clinic blood pressure is 140/90&nbsp;mmHg or higher&nbsp;and&nbsp;subsequent ambulatory blood pressure monitoring (ABPM) daytime average or home blood pressure monitoring (HBPM) average blood pressure is 135/85&nbsp;mmHg or higher.</p>
  • <p>Stage 2 hypertension : Clinic blood pressure is 160/100&nbsp;mmHg or higher&nbsp;and&nbsp;subsequent ABPM daytime average or HBPM average blood pressure is 150/95&nbsp;mmHg or higher.</p>
  • <p>Severe hypertension : Clinic systolic blood pressure is 180&nbsp;mmHg or higher&nbsp;or&nbsp;clinic diastolic blood pressure is 110&nbsp;mmHg or higher.</p>
  • <p>NO production can be measured through the conversion of L-arginine to L-citrulline (de Bono et al., 2007) , in situ fluorescent signal detection with fluorescent indicator DAF-2 DA (Itoh et al., 2000; Nagata et al., 1999; Qiu et al., 2001), EPR spin-trapping (Xia et al., 2000), and the determination of total nitrate and nitrite concentration (Crabtree et al., 2009; Du et al., 2013).</p>
  • <br>
  • <h4>References</h4>
  • <p>Durand, M.J., and Gutterman, D.D. (2013). Diversity in mechanisms of endothelium-dependent vasodilation in health and disease. Microcirc. N. Y. N 1994 20, 239&ndash;247.</p>
  • <p>de Bono, J.P., Warrick, N., Bendall, J.K., Channon, K.M., and Alp, N.J. (2007). Radiochemical HPLC detection of arginine metabolism: Measurement of nitric oxide synthesis and arginase activity in vascular tissue. Nitric Oxide 16, 1&ndash;9.</p>
  • <p>Kizhakekuttu, T.J., and Widlansky, M.E. (2010). Natural antioxidants and hypertension: promise and challenges. Cardiovasc. Ther. 28, e20&ndash;e32.</p>
  • <p>Chen, X., Xu, J., Feng, Z., Fan, M., Han, J., and Yang, Z. (2010). Simvastatin combined with nifedipine enhances endothelial cell protection by inhibiting ROS generation and activating Akt phosphorylation. Acta Pharmacol. Sin. 31, 813&ndash;820.</p>
  • <p>Leong, X.-F., Ng, C.-Y., Jaarin, K., Leong, X.-F., Ng, C.-Y., and Jaarin, K. (2015). Animal Models in Cardiovascular Research: Hypertension and Atherosclerosis, Animal Models in Cardiovascular Research: Hypertension and Atherosclerosis. BioMed Res. Int. BioMed Res. Int. 2015, 2015, e528757.</p>
  • <p>Crabtree, M.J., Tatham, A.L., Al-Wakeel, Y., Warrick, N., Hale, A.B., Cai, S., Channon, K.M., and Alp, N.J. (2009). Quantitative regulation of intracellular endothelial nitric-oxide synthase (eNOS) coupling by both tetrahydrobiopterin-eNOS stoichiometry and biopterin redox status: insights from cells with tet-regulated GTP cyclohydrolase I expression. J. Biol. Chem. 284, 1136&ndash;1144.</p>
  • <p>Silva, B.R., Pernomian, L., and Bendhack, L.M. (2012). Contribution of oxidative stress to endothelial dysfunction in hypertension. Front. Physiol. 3, 441.</p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p>Du, Y., Navab, M., Shen, M., Hill, J., Pakbin, P., Sioutas, C., Hsiai, T.K., and Li, R. (2013). Ambient ultrafine particles reduce endothelial nitric oxide production via S-glutathionylation of eNOS. Biochem. Biophys. Res. Commun. 436, 462&ndash;466.</p>
  • <p>Dumitrescu, C., Biondi, R., Xia, Y., Cardounel, A.J., Druhan, L.J., Ambrosio, G., and Zweier, J.L. (2007). Myocardial ischemia results in tetrahydrobiopterin (BH4) oxidation with impaired endothelial function ameliorated by BH4. Proc. Natl. Acad. Sci. U. S. A. 104, 15081&ndash;15086.</p>
  • <p>Itoh, Y., Ma, F.H., Hoshi, H., Oka, M., Noda, K., Ukai, Y., Kojima, H., Nagano, T., and Toda, N. (2000). Determination and bioimaging method for nitric oxide in biological specimens by diaminofluorescein fluorometry. Anal. Biochem. 287, 203&ndash;209.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Luo, Z., Fujio, Y., Kureishi, Y., Rudic, R.D., Daumerie, G., Fulton, D., Sessa, W.C., and Walsh, K. (2000). Acute modulation of endothelial Akt/PKB activity alters nitric oxide&ndash;dependent vasomotor activity in vivo. J. Clin. Invest. 106, 493&ndash;499.</p>
  • <p>Nagata, N., Momose, K., and Ishida, Y. (1999). Inhibitory effects of catecholamines and anti-oxidants on the fluorescence reaction of 4,5-diaminofluorescein, DAF-2, a novel indicator of nitric oxide. J. Biochem. (Tokyo) 125, 658&ndash;661.</p>
  • <p>Qiu, W., Kass, D.A., Hu, Q., and Ziegelstein, R.C. (2001). Determinants of shear stress-stimulated endothelial nitric oxide production assessed in real-time by 4,5-diaminofluorescein fluorescence. Biochem. Biophys. Res. Commun. 286, 328&ndash;335.</p>
  • <p>Xia, Y., Cardounel, A.J., Vanin, A.F., and Zweier, J.L. (2000). Electron paramagnetic resonance spectroscopy with N-methyl-D-glucamine dithiocarbamate iron complexes distinguishes nitric oxide and nitroxyl anion in a redox-dependent manner: applications in identifying nitrogen monoxide products from nitric oxide synthase. Free Radic. Biol. Med. 29, 793&ndash;797.</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <div>
  • <div>
  • <h4><a href="/events/937">Event: 937: KE7 : Impaired, Vasodilation</a><br></h4>
  • <h5>Short Name: Impaired, Vasodilation</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>vasodilation</td>
  • <td>blood vessel</td>
  • <td>abnormal</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <!-- rel table -->
  • <h3>Scientific evidence supporting the linkages in the AOP</h3>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Upstream Event</th>
  • <th>Relationship Type</th>
  • <th>Downstream Event</th>
  • <th>Evidence</th>
  • <th>Quantitative Understanding</th>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Peptide Oxidation</td>
  • <td>directly leads to</td>
  • <td>KE2 : Decrease, GTPCH-1</td>
  • <td>Moderate</td>
  • <td>Weak</td>
  • </tr>
  • <tr>
  • <td>KE1 : S-Glutathionylation, eNOS</td>
  • <td>directly leads to</td>
  • <td>KE4 : Uncoupling, eNOS</td>
  • <td>Strong</td>
  • <td>Moderate</td>
  • </tr>
  • <tr>
  • <td>KE2 : Decrease, GTPCH-1</td>
  • <td>directly leads to</td>
  • <td>KE3 : Decrease, Tetrahydrobiopterin</td>
  • <td>Strong</td>
  • <td>Strong</td>
  • </tr>
  • <tr>
  • <td>KE3 : Decrease, Tetrahydrobiopterin</td>
  • <td>directly leads to</td>
  • <td>KE4 : Uncoupling, eNOS</td>
  • <td>Strong</td>
  • <td>Strong</td>
  • </tr>
  • <tr>
  • <td>KE4 : Uncoupling, eNOS</td>
  • <td>directly leads to</td>
  • <td>KE6 : Depletion, Nitric Oxide</td>
  • <td>Strong</td>
  • <td>Strong</td>
  • </tr>
  • <tr>
  • <td>KE6 : Depletion, Nitric Oxide</td>
  • <td>directly leads to</td>
  • <td>KE7 : Impaired, Vasodilation</td>
  • <td>Strong</td>
  • <td>Moderate</td>
  • </tr>
  • <tr>
  • <td>KE7 : Impaired, Vasodilation</td>
  • <td>directly leads to</td>
  • <td>KE8 : Increase, Vascular Resistance</td>
  • <td>Moderate</td>
  • <td>Weak</td>
  • <td><a href="/aops/149">Aop:149 - Peptide Oxidation Leading to Hypertension</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Context</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <td>KE8 : Increase, Vascular Resistance</td>
  • <td>directly leads to</td>
  • <td>Hypertension</td>
  • <td>Moderate</td>
  • <td>Weak</td>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>KE5 : Decrease, AKT/eNOS activity</td>
  • <td>directly leads to</td>
  • <td>KE6 : Depletion, Nitric Oxide</td>
  • <td>Strong</td>
  • <td>Strong</td>
  • <td>Organ</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • <h4>Organ term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <td>Peptide Oxidation</td>
  • <td>directly leads to</td>
  • <td>KE5 : Decrease, AKT/eNOS activity</td>
  • <td>Strong</td>
  • <td>Moderate</td>
  • <th>Organ term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Peptide Oxidation</td>
  • <td>directly leads to</td>
  • <td>KE1 : S-Glutathionylation, eNOS</td>
  • <td>Moderate</td>
  • <td>Weak</td>
  • <td>circulatory system</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <!-- Evidence for relationship links section, this lists the relationships and then supports them -->
  • <div id="evidence_supporting_links">
  • <hr>
  • <div>
  • <h4><a href="/relationships/980">Peptide Oxidation leads to Decrease, GTPCH-1</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Directness</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>directly leads to</th>
  • <th>Moderate </th>
  • <th>Weak</th>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <h4>Domain of Applicability</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Moderate</td>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Moderate</td>
  • <td>Oryctolagus cuniculus</td>
  • <td>Oryctolagus cuniculus</td>
  • <td>Low</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9986" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>Several studies showed decreased GTPCH-1 activity and/or protein expression under oxidative stress in cardiac reperfusion patients, bovine endothelial cells, a mouse model of diabetes and a rat model of hypertension (Cervantes-P&eacute;rez et al., 2012; AbdelGhany et al., (under review); Jayaram et al., 2015; Zhao et al., 2013).</p>
  • <h4>How Does This Key Event Relationship Work</h4>
  • <p>Exposure to known inducers of oxidative stress such as cigarette smoke extract (AbdelGhany et al., under review) or peroxynitrite (Zhao et al., 2013) causes the loss of GTPCH-1 activity, resulting in decreased levels of tetrahydrobiopterin (BH4) and subsequent uncoupling of endothelial nitric oxide synthase (eNOS).</p>
  • <!-- if nothing shows up in any of these fields, then weight of evidence will not be displayed -->
  • <h4>Weight of Evidence</h4>
  • <strong>Biological Plausibility</strong>
  • <p>Several studies demonstrated that GTPCH-1 is also affected by oxidative stress, which provides evidence for moderate biological plausibility. In vitro exposure to chemically synthesized peroxynitrite inhibited GTPCH-1 activity in a dose-dependent manner (Zhao et al., 2013). This inhibition as well as increased ubiquitination of GTPCH-1 were observed in cultured bovine aortic endothelial cells (BAECs) and streptozotocin-induced diabetes in mice following peroxynitrite treatment. Ubiquitination of GTPCH-1 leads to its degradation, which is equivalent to a decrease in GTPCH-1. In another study, GTPCH-1 levels were reduced by cigarette smoke extract (CSE) exposure in BAECs (AbdelGhany et al., under review). There is also evidence that CSE promoted GTPCH-1 degradation by increasing proteasomal activity. Furthermore,&nbsp;cardiac reperfusion patients experienced oxidative stress which was associated with reduced GTPCH-1 activity (Jayaram et al., 2015).</p>
  • <div>
  • <p>Vasodilation has been observed in humans, rabbits, mice and rats.</p>
  • <br>
  • </div>
  • <strong>Empirical Support for Linkage</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>In a rat model of aortic coarctation-associated hypertension, increased ROS (1.75 nmol DCF/mg protein/min) and decreased GTPCH-1 protein expression (western blot band intensity of 0.0087&plusmn;0.005) were observed (Cervantes-P&eacute;rez et al., 2012). However, following clofibrate treatment (100 mg/kg/day, 7 days), ROS was reduced by approximately 30% and GTPCH-1 band intensity increased to 0.0087&plusmn;0.005. This study provides evidence that there is a dependent change between oxidative stress and GTPCH-1, but we found no dose-response relationship, so empirical support for this KER is weak.</p>
  • <strong>Uncertainties or Inconsistencies</strong>
  • <p>No uncertainties or inconsistencies were found for this KER.</p>
  • <!-- event text -->
  • <h4>Key Event Description</h4>
  • <p>Vasodilation refers to the widening or increase in the diameter of blood vessels (e.g. large arteries, large veins, small arterioles) that is caused by the relaxation of vascular smooth muscle cells (VSMCs) within the walls of blood vessels, thus increasing blood flow and decreasing arterial blood pressure and heart rate (Siddiqui, 2011). VSMC relaxation is regulated through a number of mechanisms, including cyclic GMP-dependent hyperpolarization and relaxation via nitric oxide (NO), cAMP-dependent hyperpolarization via prostaglandins, and stimulation of potassium channels via endothelial-derived hyperpolarizing factors (Durand and Gutterman, 2013). Under oxidative stress, decreased NO bioavailability results in impaired vasodilation, which is associated with cardiovascular diseases such as hypertension (Silva et al., 2012).</p>
  • <br>
  • <strong>Quantitative Understanding of the Linkage</strong>
  • <p><em>Is it known how much change in the first event is needed to impact the second? Are there known modulators of the response-response relationships? Are there models or extrapolation approaches that help describe those relationships? </em></p>
  • <h4>How it is Measured or Detected</h4>
  • <p>Endothelium-dependent vasodilation can be measured using invasive and non-invasive methods (Raitakari and Celermajer, 2000). For the invasive approach, vasodilation is measured after intra-arterial pharmacologic stimulation with substances that enhance NO release (e.g. acetylcholine, bradykinin). The non-invasive ultrasound-based method evaluates flow-mediated vasodilation (FMD) in the superficial arteries, such as brachial, radial, or femoral vessels.</p>
  • <p>As the relationship between oxidative stress and GTPCH-1 is not well-studied, there is limited quantitative understanding of this linkage.</p>
  • <p>Guidelines for the measurement of FMD have been published (Corretti et al. 2002).</p>
  • <br>
  • <!--<!% unless aop_rel.relationship.relationship_taxons.blank? %>-->
  • <!--<!%= render 'snapshot_taxons', taxons: aop_rel.relationship.relationship_taxons %>-->
  • <!--<!% unless aop_rel.relationship.taxon_evidence.blank? %>-->
  • <!--<h3>Evidence Supporting Taxonomic Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
  • <!--<!% end %>-->
  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>AbdelGhany, T., Ismail, R., Elmahdy, M., Mansoor F, Zweier J, Lowe, F., and Zweier, JL. (2017). Cigarette Smoke Constituents Cause Endothelial Nitric Oxide Synthase Dysfunction and Uncoupling due to Depletion of Tetrahydrobiopterin with Degradation of GTP Cyclohydrolase.&nbsp;&nbsp;Nitric Oxide&nbsp;(Under review)</p>
  • <p>Guidelines for the ultrasound assessment of endothelial-dependent flow-mediated vasodilation of the brachial artery</p>
  • <p>Cervantes-P&eacute;rez, L.G., Ibarra-Lara, M. de la L., Escalante, B., Del Valle-Mondrag&oacute;n, L., Vargas-Robles, H., P&eacute;rez-Severiano, F., Pastel&iacute;n, G., and S&aacute;nchez-Mendoza, M.A. (2012). Endothelial nitric oxide synthase impairment is restored by clofibrate treatment in an animal model of hypertension. Eur. J. Pharmacol. 685, 108&ndash;115.</p>
  • <p>Corretti&nbsp;MC, Anderson&nbsp;TJ, Benjamin&nbsp;EJ, Celermajer D, Charbonneau F, Creager MA,&nbsp;Deanfield J,&nbsp;Drexler H, Gerhard-Herman M, Herrington D, Vallance P, Vita J, Vogel R. &nbsp;Guidelines for the ultrasound assessment of endothelial-dependent flow-mediated vasodilation of the brachial artery. &nbsp;Journal of the American College of Cardiology,&nbsp;2002,&nbsp;39&nbsp;(2)&nbsp;257-265</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49</p>
  • <p>Durand, M.J., and Gutterman, D.D. (2013). Diversity in mechanisms of endothelium-dependent vasodilation in health and disease. Microcirc. N. Y. N 1994 20, 239&ndash;247.</p>
  • <p>Zhao, Y., Wu, J., Zhu, H., Song, P., and Zou, M.-H. (2013). Peroxynitrite-dependent zinc release and inactivation of guanosine 5&rsquo;-triphosphate cyclohydrolase 1 instigate its ubiquitination in diabetes. Diabetes 62, 4247&ndash;4256.</p>
  • <p>Raitakari, O.T., and Celermajer, D.S. (2000). Flow-mediated dilatation. Br. J. Clin. Pharmacol. 50, 397&ndash;404.</p>
  • <p>Siddiqui, A. (2011). Effects of Vasodilation and Arterial Resistance on Cardiac Output. J. Clin. Exp. Cardiol. 02.</p>
  • <p>Silva, B.R., Pernomian, L., and Bendhack, L.M. (2012). Contribution of oxidative stress to endothelial dysfunction in hypertension. Front. Physiol. 3, 441.</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <br>
  • <div>
  • <h4><a href="/relationships/955">S-Glutathionylation, eNOS leads to Uncoupling, eNOS</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <div>
  • <div>
  • <h4><a href="/events/951">Event: 951: KE8 : Increase, Vascular Resistance</a><br></h4>
  • <h5>Short Name: Increase, Vascular Resistance</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Directness</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>directly leads to</th>
  • <th>Strong </th>
  • <th>Moderate</th>
  • <td>increased systemic vascular resistance</td>
  • <td></td>
  • <td>increased</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">Aop:149 - Peptide Oxidation Leading to Hypertension</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Context</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Organ</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • <h4>Organ term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Organ term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>circulatory system</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h4>Domain of Applicability</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Strong</td>
  • <td>Low</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>The evidence supporting this key event relationship are from human subjects, HAECs, BAECs, and SHR rats (Chen et al., 2010; De Pascali et al., 2014; Du et al., 2013; Jayaram et al., 2015).</p>
  • <h4>How Does This Key Event Relationship Work</h4>
  • <p>Oxidative stress can trigger S-glutathionylation of eNOS at cysteine residues Cys689 and Cys908, which are known to be critical for normal eNOS function (Zweier et al., 2011). S-glutathionylation directly causes eNOS uncoupling, a state in which eNOS switches from producing NO to generating superoxide, thus impairing endothelium-dependent vasodilation and contributing to endothelial dysfunction. Uncoupling of eNOS via S-glutathionylation is different from BH4-mediated eNOS uncoupling in that superoxide is produced in the reductase domain rather than the oxygenase domain and superoxide generation cannot be inhibited by L-NG-nitroarginine methyl ester (L-NAME), suggesting that S-glutathionylation occurs independent of calcium/calmodulin and heme.</p>
  • <!-- if nothing shows up in any of these fields, then weight of evidence will not be displayed -->
  • <h4>Weight of Evidence</h4>
  • <strong>Biological Plausibility</strong>
  • <p><em>In vitro</em> experiments showed that S-glutathionylation of eNOS significantly decreased NO activity in dose-dependent manner and greatly increased superoxide generation (Chen et al., 2010). NOS inhibitor L-NAME partially blocked superoxide generation. These results were observed in bovine aortic endothelial cells (BAECs) and in aortae of spontaneously hypertensive (SHR) rats. Treatment of BAECs with 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), an inhibitor of glutathione reductase, induced eNOS S-glutathionylation, resulting in decreased NO and increased superoxide generation. Gene silencing of eNOS in BAECs also induced superoxide production. In aortae of SHR rats, S-glutathionylation as determined by immunohistology was associated with a decrease in endothelium-dependent vasodilation as a result of decreased NO. Exposure of human aortic endothelial cells (HAECs) to ultrafine particles (UFP) caused S-glutathionylation of eNOS and a dose-dependent decrease in NO production (Du et al., 2013). Decreased NO production was found to be mediated by S-glutathionylation since overexpression of glutaredoxin, an inhibitor of S-glutathionylation, significantly reduced UPF-mediated decrease in NO production. Cardiac reperfusion patients exhibited decreased eNOS activity which was identified to be a result of eNOS S-glutathionylation (Jayaram et al., 2015). Additional evidence was observed in BAECs undergoing hypoxia and reoxygenation in which eNOS S-glutathionylation increased by three-fold compared to control cells and NO production was decreased (De Pascali et al., 2014). These effects were reversed with treatment of N-acetyl-l-cysteine, which increased cellular concentration of GSH. These results demonstrate a clear interaction between&nbsp;S-glutathionylation and eNOS uncoupling, therefore this link has strong biological plausibility.</p>
  • <strong>Empirical Support for Linkage</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>Treatment with BCNU (25 &mu;M, 80 &mu;M) resulted in increased eNOS S-glutathionylation, increased superoxide generation and decreased NO production in a dose-dependent manner in BAECs (Chen et al., 2010).</p>
  • <p>Exposure to hypoxia/reoxygenation and treatment with angiotensin II (Ang II) demonstrated a response-response relationship between eNOS S-glutathionylation and superoxide generation in human and bovine endothelial cells (De Pascali et al., 2014; Galougahi et al., 2014). Following hypoxia and reoxygenation in BAECs, increased S-glutathionylation of eNOS (0.3 to 1 arbitray units; measured by western blot), decreased NO production (100% to 34.2 &plusmn; 1.7%) and increased superoxide generation (relative intensity of 3.5 to 49.2; by fluorescence) were observed (De Pascali et al., 2014). Treatment with 100 nmol/L Ang II for 1 hour caused an increase in both eNOS S-glutathionylation (from 1 to 1.42 fold) and superoxide generation (from 81.1 to 113 fluorescence signal unit) (Galougahi et al., 2014).</p>
  • <div>
  • <p>Increased SVR was observed in humans, pigs and rats (Siddiqui 2011,&nbsp;Dikalova et al. 2016,&nbsp;</p>
  • <p>Overall, there is moderate empirical support for eNOS S-glutathionylation leading to eNOS uncoupling.</p>
  • <p>&nbsp;</p>
  • <br>
  • </div>
  • <strong>Uncertainties or Inconsistencies</strong>
  • <p>No uncertainties or inconsistencies were found for this KER.</p>
  • <!-- event text -->
  • <h4>Key Event Description</h4>
  • <p>Vascular resistance is the resistance to blood flow in the circulatory system (Siddiqui, 2011). Systemic vascular resistance (SVR), also known as total peripheral resistance, refers specifically to the resistance to blood flow offered by the peripheral circulation. Vasodilation decreases SVR while vasoconstriction or <a href="/wiki/index.php/Event:937" title="Event:937">impaired vasodilation</a> increases SVR.</p>
  • <br>
  • <strong>Quantitative Understanding of the Linkage</strong>
  • <p><em>Is it known how much change in the first event is needed to impact the second? Are there known modulators of the response-response relationships? Are there models or extrapolation approaches that help describe those relationships? </em></p>
  • <p>No study explored how much change in S-glutathionylation of eNOS is required to lead to eNOS uncoupling. One study in HUVECs showed treatment with 100nM/L Ang II increased S-glutathionylated eNOS protein expression&nbsp;by 1.42 fold, and was associated with an increase&nbsp;in superoxide generation of nearly 50% at the same dose.&nbsp; DTT (which removes PR-SG residues)&nbsp;was shown to partially reverse this effect&nbsp;(Galougahi et al., 2014). Ang II and hypoxia/reoxygenation were demonstrated to be modulators of the response-response relationship between S-glutathionylation of eNOS and eNOS uncoupling as these two stressors caused a change in both key events (De Pascali et al., 2014; Galougahi et al., 2014). More experiments using other stressors or oxidants are needed to further understand of this relationship.</p>
  • <h4>How it is Measured or Detected</h4>
  • <p>Vascular resistance cannot be measured by any direct means, but can be calculated using a formula: (mean arterial pressure minus mean right arterial pressure) divided by cardiac output (Siddiqui 2011).</p>
  • <br>
  • <!--<!% unless aop_rel.relationship.relationship_taxons.blank? %>-->
  • <!--<!%= render 'snapshot_taxons', taxons: aop_rel.relationship.relationship_taxons %>-->
  • <!--<!% unless aop_rel.relationship.taxon_evidence.blank? %>-->
  • <!--<h3>Evidence Supporting Taxonomic Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
  • <!--<!% end %>-->
  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>Chen, C.-A., Wang, T.-Y., Varadharaj, S., Reyes, L.A., Hemann, C., Talukder, M.A.H., Chen, Y.-R., Druhan, L.J., and Zweier, J.L. (2010). S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature 468, 1115&ndash;1118.</p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p>Du, Y., Navab, M., Shen, M., Hill, J., Pakbin, P., Sioutas, C., Hsiai, T.K., and Li, R. (2013). Ambient ultrafine particles reduce endothelial nitric oxide production via S-glutathionylation of eNOS. Biochem. Biophys. Res. Commun. 436, 462&ndash;466.</p>
  • <p>Siddiqui, A. (2011). Effects of Vasodilation and Arterial Resistance on Cardiac Output. J. Clin. Exp. Cardiol. 02.</p>
  • <p>Galougahi, K.K., Liu, C.-C., Gentile, C., Kok, C., Nunez, A., Garcia, A., Fry, N.A.S., Davies, M.J., Hawkins, C.L., Rasmussen, H.H., et al. (2014). Glutathionylation Mediates Angiotensin II&ndash;Induced eNOS Uncoupling, Amplifying NADPH Oxidase-Dependent Endothelial Dysfunction. J. Am. Heart Assoc. 3, e000731.</p>
  • <p>Dikalova A,&nbsp;Aschner JL,&nbsp;Kaplowitz MR,&nbsp;Summar M,&nbsp;Fike CD. &nbsp;Tetrahydrobiopterin oral therapy recouples&nbsp;eNOS&nbsp;and ameliorates chronic hypoxia-induced pulmonary hypertension in newborn pigs. &nbsp;Am J Physiol Lung Cell Mol Physiol.&nbsp;2016, 1;311(4):L743-L753.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>&nbsp;</p>
  • <p>Zweier, J.L., Chen, C.-A., and Druhan, L.J. (2011). S-glutathionylation reshapes our understanding of endothelial nitric oxide synthase uncoupling and nitric oxide/reactive oxygen species-mediated signaling. Antioxid. Redox Signal. 14, 1769&ndash;1775.</p>
  • <p>&nbsp;</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <br>
  • <div>
  • <h4><a href="/relationships/952">Decrease, GTPCH-1 leads to Decrease, Tetrahydrobiopterin</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <div>
  • <div>
  • <h4><a href="/events/973">Event: 973: KE5 : Decrease, AKT/eNOS activity</a><br></h4>
  • <h5>Short Name: Decrease, AKT/eNOS activity</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Directness</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>directly leads to</th>
  • <th>Strong </th>
  • <th>Strong</th>
  • <td>catalytic activity</td>
  • <td>nitric oxide synthase, endothelial</td>
  • <td>decreased</td>
  • </tr>
  • <tr>
  • <td>catalytic activity</td>
  • <td>AKT kinase</td>
  • <td>decreased</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">Aop:149 - Peptide Oxidation Leading to Hypertension</a></td>
  • <td>KeyEvent</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • <h4>Stressors</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Name</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Reactive oxygen species</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Context</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Cellular</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • <h4>Cell term</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Cell term</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>endothelial cell of vascular tree</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h4>Domain of Applicability</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Strong</td>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Strong</td>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>The relationship between GTPCH-1 and BH4 is supported in humans (Jayaram et al., 2015), cows (Abdelghany&nbsp;et al., 2017; Whitsett et al., 2007, Wang et al., 2008), mice (Adlam et al., 2012; Chuaiphichai et al., 2014; Crabtree et al., 2009; Tatham et al., 2009; Wang et al., 2008) and rats (Cervantes-P&eacute;rez et al., 2012).</p>
  • <h4>How Does This Key Event Relationship Work</h4>
  • <p>Guanosine triphosphate cyclohydrolase-1 (GTPCH-1) is the rate-limiting enzyme in the de novo biosynthesis of BH4, which is an essential cofactor for eNOS and NO generation (Wang et al., 2008). Oxidative stress can disrupt and decrease GTPCH-1 activity, leading to decreased BH4 levels and subsequent uncoupling of eNOS.</p>
  • <!-- if nothing shows up in any of these fields, then weight of evidence will not be displayed -->
  • <h4>Weight of Evidence</h4>
  • <strong>Biological Plausibility</strong>
  • <p>As GTPCH-1 is required for BH4 biosynthesis, there is strong biological plausibility for this relationship.</p>
  • <p>Many studies demonstrated that deletion of GTPCH-1 led to the deficiency of BH4 in endothelial cells. In the hph-1 mouse model, cardiac GTPCH-1 enzymatic activity was reduced by 90% compared to wild-type mice, which led to 60% reduction in BH4 levels (Adlam et al., 2012). In another mouse model of endothelial-targeted deletion of GTPCH-1, BH4 levels in lung, heart and aorta were significantly decreased compared to wild-type mice (Chuaiphichai et al., 2014). GTPCH-1 siRNA significantly reduced GTPCH-1 enzyme activity and BH4 levels in murine sEnd.1 and aortic endothelial cells (Crabtree et al., 2009; Tatham et al., 2009; Wang et al., 2008). The selective GTPCH-1 inhibitor diaminohydroxypyrimidine (DAHP) reduced levels of BH4 in bovine aortic endothelial cells (BAECs) (Wang et al., 2008). Also, transgenic overexpression of GTPCH-1 increased BH4 protein levels in murine hearts and aortas, leading to enhanced eNOS activity (Alp et al., 2003; Carnicer et al., 2012).</p>
  • <strong>Empirical Support for Linkage</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>Exposure to a wide range of stimuli led to a decrease in both GTPCH-1 expression and activity and a decrease in BH4 levels, indicating strong empirical support between these two key events. An assumption that a decrease in GTPCH-1 expression and activity also results in a decrease in BH4 levels was made for these studies.&nbsp;</p>
  • <div>
  • <p>Decreased Akt and eNOS activity was observed in humans, cows, mice and rats following exposure to stressors.</p>
  • <p>After myocardial reperfusion, GTPCH-1 activity decreased from 100% to 50% and arterial BH4 levels was reduced by 32% (Jayaram et al., 2015).</p>
  • <p>Cigarette smoke exposure was shown to inhibit the phosphorylation of AKT and eNOS in VEGF-stimulated human umbilical vein endothelial cells (HUVECs), resulting in decreased NO levels (Michaud et al. 2006).</p>
  • <p>IL6/TNF/LPS (4 ng/mL, 10 nmol/L, 80 ng/mL) stimulation for 24 hours induced a 3-fold upregulation of GTPCH-1 gene expression and 3- to 4-fold increase in vascular BH4 (Antoniades et al., 2011). For this evidence, an assumption that a change in GTPCH-1 gene expression would affect GTPCH-1 enzyme activity was made.</p>
  • <p>In rat aortic rings, exposure to methylglyoxal and high concentrations of glucose decreased endothelium-dependent relaxation. &nbsp;Further experiments in rat endothelial cells and HUVECs demonstrated a reduction in eNOS phosphorylation and&nbsp;activity, and reduced NO levels in response to the same stressors&nbsp;(Dhar et al. 2010).</p>
  • <p>Cigarette smoke extract (CSE, 5%) exposure for 4 hours significantly decreased BH4 levels from 100% to 50% and GTPCH-1 expression from 100% to 51%, while a lower concentration of 2.5% CSE did not cause a significant change in bovine aortic endothelial cells (Abdelghany&nbsp;et al., 2017).</p>
  • <p>In bovine aortic endothelial cells, AKT and eNOS phosphorylation were decreased following exposure to the peroxynitrite source; SIN-1, with an associated reduction in NO bioavailability. &nbsp;These effects were ameliorated by treatment with the ROS scavenger DMPO (Das et al. 2014).</p>
  • <p>In BAECs, stimulation with 25 &mu;M 4-hydroxy-2-nonenal decreased BH4 levels from 10 pmol/mg protein to 6.1 pmol/mg protein and GTPCH-1 activity from 27 pmol/hr/mg protein to 23 pmol/hr/mg protein (Whitsett et al., 2007).</p>
  • <p>eNOS knockout mice are routinely used as models of hypertension. &nbsp;Such mice display reduced bioavailability of NO and impaired vasodilation (Huang et al. 1995).</p>
  • <p>In a rat hypertensive model, the decrease in GTPCH-1 protein expression (via densitometry of western blot band intensity, 0.089 control to 0.0087) and BH4 (33.71&plusmn;3.318 pmol control to 17.144&plusmn;2.251 pmol) induced by aortic coarctation was attenuated by clofibrate treatment (GTPCH-1: 0.088&plusmn;0.022 band intensity, BH4: 32.534&plusmn;5.809 pmol) (Cervantes-P&eacute;rez et al., 2012).</p>
  • <p>Reduced AKT/eNOS phosphorylation was reported under conditions of hyperglycaemia (in mice) and in HUVECs following treatment with high concentrations of glucose. &nbsp;Aortic rings from hyperglycaemic mice demonstrated impaired vasodilation. &nbsp;Resveratrol treatment was shown to improve vasodilation and eNOS phosphorylation in wild-type mice, but not AKT knockout mice. &nbsp;Transfection of HUVECs with AKT siRNA abolished resveratrol-enhanced eNOS phosphorylation and NO release&nbsp;(Li et al. 2017),</p>
  • <br>
  • </div>
  • <strong>Uncertainties or Inconsistencies</strong>
  • <p>There are no uncertainties or inconsistencies.</p>
  • <strong>Quantitative Understanding of the Linkage</strong>
  • <p><em>Is it known how much change in the first event is needed to impact the second? Are there known modulators of the response-response relationships? Are there models or extrapolation approaches that help describe those relationships? </em></p>
  • <!-- event text -->
  • <h4>Key Event Description</h4>
  • <p>Endothelial nitric oxide synthase (eNOS) is responsible for the generation of nitric oxide (NO), which is an important regulator of vascular homeostasis. The activity of eNOS can be regulated through calmodulin-mediated dimerization, tetrahydrobiopterin-mediated conversion of L-arginine to L-citrulline, protein-protein interactions with heat shock protein 90 and caveolin, S-nitrosylation, acetylation and phosphorylation (Atochin et al., 2007; Qian and Fulton, 2013). eNOS has been shown to be phosphorylated at multiple sites including tyrosine (Y), serine (Ser) and threonine (Thr) residues. Serine-threonine protein kinase AKT, a multifunctional regulator of cellular processes like glucose metabolism and proliferation, can directly phosphorylate eNOS at Ser1177/Ser1179, leading to increased eNOS enzymatic activity and subsequent NO production (Dimmeler et al., 1999; Fulton et al., 1999). Inhibition of AKT&nbsp;or a mutation of the AKT phosphorylation site on eNOS attenuates eNOS phosphorylation and its activity, resulting in decreased NO bioavailability and endothelial dysfunction (Dimmler et al. 1999)</p>
  • <br>
  • <h4>How it is Measured or Detected</h4>
  • <p>Western blot analysis can be performed to determine the expression levels of phosphorylated eNOS, phosphorylated Akt, total Akt and total eNOS proteins using the appropriate anti-phospho-eNOS, anti-phospho-Akt, anti-eNOS, and anti-Akt antibodies. Alternatively, eNOS activity can be measured using the conversion of L-arginine to L-citrulline assay.</p>
  • <p>Based on the relationship between GTPCH-1 and BH4, it would be possible that any change in GTPCH-1 activity would affect BH4 biosynthesis. The studies above showed that there are many modulators of the response-response relationships including cardiac reperfusion (Jayaram et al., 2015), cytokines (Antoniades et al., 2011), CSE (Abdelghany&nbsp;et al., 2017), and 4-hydroxy-2-nonenal (Whitsett et al., 2007).</p>
  • <p>ELISA kits for AKT/eNOS and phospho AKT/eNOS expression are commercially available.</p>
  • <br>
  • <!--<!% unless aop_rel.relationship.relationship_taxons.blank? %>-->
  • <!--<!%= render 'snapshot_taxons', taxons: aop_rel.relationship.relationship_taxons %>-->
  • <!--<!% unless aop_rel.relationship.taxon_evidence.blank? %>-->
  • <!--<h3>Evidence Supporting Taxonomic Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
  • <!--<!% end %>-->
  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>AbdelGhany, T., Ismail, R., Elmahdy, M., Mansoor F, Zweier J, Lowe, F., and Zweier, JL. (2017). Cigarette Smoke Constituents Cause Endothelial Nitric Oxide Synthase Dysfunction and Uncoupling due to Depletion of Tetrahydrobiopterin with Degradation of GTP Cyclohydrolase.&nbsp;&nbsp;Nitric Oxide&nbsp;(Under review).</p>
  • <p>Adlam, D., Herring, N., Douglas, G., De Bono, J.P., Li, D., Danson, E.J., Tatham, A., Lu, C.-J., Jennings, K.A., Cragg, S.J., et al. (2012). Regulation of &beta;-adrenergic control of heart rate by GTP-cyclohydrolase 1 (GCH1) and tetrahydrobiopterin. Cardiovasc. Res. 93, 694&ndash;701.</p>
  • <p>Alp, N.J., Mussa, S., Khoo, J., Cai, S., Guzik, T., Jefferson, A., Goh, N., Rockett, K.A., and Channon, K.M. (2003). Tetrahydrobiopterin-dependent preservation of nitric oxide-mediated endothelial function in diabetes by targeted transgenic GTP-cyclohydrolase I overexpression. J. Clin. Invest. 112, 725&ndash;735.</p>
  • <p>Antoniades, C., Cunnington, C., Antonopoulos, A., Neville, M., Margaritis, M., Demosthenous, M., Bendall, J., Hale, A., Cerrato, R., Tousoulis, D., et al. (2011). Induction of vascular GTP-cyclohydrolase I and endogenous tetrahydrobiopterin synthesis protect against inflammation-induced endothelial dysfunction in human atherosclerosis. Circulation 124, 1860&ndash;1870.</p>
  • <p>Carnicer, R., Hale, A.B., Suffredini, S., Liu, X., Reilly, S., Zhang, M.H., Surdo, N.C., Bendall, J.K., Crabtree, M.J., Lim, G.B.S., et al. (2012). Cardiomyocyte GTP cyclohydrolase 1 and tetrahydrobiopterin increase NOS1 activity and accelerate myocardial relaxation. Circ. Res. 111, 718&ndash;727.</p>
  • <p>Atochin, D.N., Wang, A., Liu, V.W.T., Critchlow, J.D., Dantas, A.P.V., Looft-Wilson, R., Murata, T., Salomone, S., Shin, H.K., Ayata, C., et al. (2007). The phosphorylation state of eNOS modulates vascular reactivity and outcome of cerebral ischemia in vivo. J. Clin. Invest. 117, 1961&ndash;1967.</p>
  • <p>Cervantes-P&eacute;rez, L.G., Ibarra-Lara, M. de la L., Escalante, B., Del Valle-Mondrag&oacute;n, L., Vargas-Robles, H., P&eacute;rez-Severiano, F., Pastel&iacute;n, G., and S&aacute;nchez-Mendoza, M.A. (2012). Endothelial nitric oxide synthase impairment is restored by clofibrate treatment in an animal model of hypertension. Eur. J. Pharmacol. 685, 108&ndash;115.</p>
  • <p>Das A,&nbsp;Gopalakrishnan B,&nbsp;Druhan LJ,&nbsp;Wang TY,&nbsp;De Pascali F,&nbsp;Rockenbauer A,&nbsp;Racoma I,&nbsp;Varadharaj S,&nbsp;Zweier JL,&nbsp;Cardounel AJ,&nbsp;Villamena FA. &nbsp;Reversal&nbsp;of&nbsp;SIN-1-induced&nbsp;eNOS&nbsp;dysfunction&nbsp;by the&nbsp;spin&nbsp;trap,&nbsp;DMPO, in&nbsp;bovine&nbsp;aortic&nbsp;endothelialcells&nbsp;via&nbsp;eNOS&nbsp;phosphorylation. &nbsp;Br J Pharmacol.&nbsp;2014, 171(9):2321-34. doi: 10.1111/bph.12572.</p>
  • <p>Chen, C.-A., Lin, C.-H., Druhan, L.J., Wang, T.-Y., Chen, Y.-R., and Zweier, J.L. (2011). Superoxide induces endothelial nitric-oxide synthase protein thiyl radical formation, a novel mechanism regulating eNOS function and coupling. J. Biol. Chem. 286, 29098&ndash;29107.</p>
  • <p>Dhar A,&nbsp;Dhar I,&nbsp;Desai KM,&nbsp;Wu L. &nbsp;Methylglyoxal&nbsp;scavengers attenuate endothelial dysfunction induced by&nbsp;methylglyoxal&nbsp;and high concentrations of glucose. &nbsp;Br J Pharmacol.&nbsp;2010, 161(8):1843-56.</p>
  • <p>Chuaiphichai, S., McNeill, E., Douglas, G., Crabtree, M.J., Bendall, J.K., Hale, A.B., Alp, N.J., and Channon, K.M. (2014). Cell-autonomous role of endothelial GTP cyclohydrolase 1 and tetrahydrobiopterin in blood pressure regulation. Hypertension 64, 530&ndash;540.</p>
  • <p>Dimmeler, S., Fleming, I., Fisslthaler, B., Hermann, C., Busse, R., and Zeiher, A.M. (1999). Activation of nitric oxide synthase in endothelial cells by Akt-dependent phosphorylation. Nature 399, 601&ndash;605.</p>
  • <p>Crabtree, M.J., Tatham, A.L., Al-Wakeel, Y., Warrick, N., Hale, A.B., Cai, S., Channon, K.M., and Alp, N.J. (2009). Quantitative regulation of intracellular endothelial nitric-oxide synthase (eNOS) coupling by both tetrahydrobiopterin-eNOS stoichiometry and biopterin redox status: insights from cells with tet-regulated GTP cyclohydrolase I expression. J. Biol. Chem. 284, 1136&ndash;1144.</p>
  • <p>Fulton, D., Gratton, J.P., McCabe, T.J., Fontana, J., Fujio, Y., Walsh, K., Franke, T.F., Papapetropoulos, A., and Sessa, W.C. (1999). Regulation of endothelium-derived nitric oxide production by the protein kinase AKT. Nature 399, 597&ndash;601.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Huang PL,&nbsp;Huang Z,&nbsp;Mashimo H,&nbsp;Bloch KD,&nbsp;Moskowitz MA,&nbsp;Bevan JA,&nbsp;Fishman MC. &nbsp; &nbsp;Hypertension&nbsp;in&nbsp;mice&nbsp;lacking&nbsp;the&nbsp;gene&nbsp;for&nbsp;endothelial nitric oxide synthase. &nbsp;Nature.&nbsp;1995, 377(6546):239-42.</p>
  • <p>Tatham, A.L., Crabtree, M.J., Warrick, N., Cai, S., Alp, N.J., and Channon, K.M. (2009). GTP cyclohydrolase I expression, protein, and activity determine intracellular tetrahydrobiopterin levels, independent of GTP cyclohydrolase feedback regulatory protein expression. J. Biol. Chem. 284, 13660&ndash;13668.</p>
  • <p>Li JY,&nbsp;Huang WQ,&nbsp;Tu RH,&nbsp;Zhong GQ,&nbsp;Luo BB,&nbsp;He Y. &nbsp;Resveratrol rescues hyperglycemia-induced endothelial dysfunction via activation of&nbsp;Akt. &nbsp;Acta Pharmacol Sin.&nbsp;2017, 38(2):182-191.</p>
  • <p>Wang, S., Xu, J., Song, P., Wu, Y., Zhang, J., Chul Choi, H., and Zou, M.-H. (2008). Acute inhibition of guanosine triphosphate cyclohydrolase 1 uncouples endothelial nitric oxide synthase and elevates blood pressure. Hypertension 52, 484&ndash;490.</p>
  • <p>Michaud SE,&nbsp;Dussault S,&nbsp;Groleau J,&nbsp;Haddad P,&nbsp;Rivard A.J. Cigarette smoke exposure impairs VEGF-induced endothelial cell migration: role of NO and reactive oxygen species. &nbsp;Mol Cell Cardiol.&nbsp;2006 Aug;41(2):275-84.</p>
  • <p>Whitsett, J., Picklo, M.J., and Vasquez-Vivar, J. (2007). 4-Hydroxy-2-nonenal increases superoxide anion radical in endothelial cells via stimulated GTP cyclohydrolase proteasomal degradation. Arterioscler. Thromb. Vasc. Biol. 27, 2340&ndash;2347.</p>
  • <p>Qian, J., and Fulton, D. (2013). Post-translational regulation of endothelial nitric oxide synthase in vascular endothelium. Oxid. Physiol. 4, 347.</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <br>
  • <div>
  • <h4><a href="/relationships/953">Decrease, Tetrahydrobiopterin leads to Uncoupling, eNOS</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <h3>List of Adverse Outcomes in this AOP</h3>
  • <div>
  • <div>
  • <h4><a href="/events/952">Event: 952: Hypertension</a><br></h4>
  • <h5>Short Name: Hypertension</h5>
  • </div>
  • <h4>Key Event Component</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Directness</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • <tr>
  • <th>Process</th>
  • <th>Object</th>
  • <th>Action</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>directly leads to</th>
  • <th>Strong </th>
  • <th>Strong</th>
  • <td>hypertension</td>
  • <td></td>
  • <td>increased</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <div>
  • <!-- loop to find all aops that use this event -->
  • <h4>AOPs Including This Key Event</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP ID and Name</th>
  • <th>Event Type</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td><a href="/aops/149">Aop:149 - Peptide Oxidation Leading to Hypertension</a></td>
  • <td>AdverseOutcome</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- loop to find stressors under event -->
  • <div>
  • <h4>Stressors</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Name</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Reactive oxygen species</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <br>
  • <!-- biological organization -->
  • <div>
  • <h4>Biological Context</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Level of Biological Organization</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Individual</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end of bio org -->
  • <!-- cell term -->
  • <div>
  • </div>
  • <!-- end of cell term -->
  • <!-- organ term -->
  • <div>
  • </div>
  • <!-- end of organ term -->
  • <!-- Evidence for Perturbation of This Event by Stressors -->
  • <!-- end Evidence for Perturbation of This Event by Stressors -->
  • <h4>Domain of Applicability</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <!-- loop to find taxonomic applicability under event -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <!-- life stages -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- end life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>The relationship between BH4 depletion and eNOS uncoupling was observed in humans (Jayaram et al., 2015), cows (Wang et al., 2008; Whitsett et al., 2007), mice (Chuaiphichai et al., 2014; Crabtree et al., 2009) and rats (Cervantes-P&eacute;rez et al., 2012; Dumitrescu et al., 2007).</p>
  • <div>
  • <p>Animal models including mouse and rat models are routinely used to study hypertension, and have been shown to reflect human physiology relating to hypertension&nbsp;(Leong et al., 2015).</p>
  • <h4>How Does This Key Event Relationship Work</h4>
  • <p>Oxidative stress leads to the excessive oxidation and depletion of BH4, resulting in eNOS uncoupling where eNOS produces superoxide rather than nitric oxide (F&ouml;rstermann and M&uuml;nzel, 2006).</p>
  • <br>
  • </div>
  • <!-- if nothing shows up in any of these fields, then weight of evidence will not be displayed -->
  • <h4>Weight of Evidence</h4>
  • <strong>Biological Plausibility</strong>
  • <p>BH4 is an essential cofactor for eNOS and is required for its enzymatic activity to produce NO. The depletion of BH4 leading to eNOS uncoupling is well-studied, thus there is strong biological plausibility for this link.</p>
  • <p>Two mouse studies showed limited BH4 availability induced eNOS uncoupling by reducing eNOS activity, leading to decreased nitric oxide and increased superoxide. In the mouse endothelial cell line sEnd.1, BH4 deficiency induced eNOS uncoupling as determined by superoxide production and impaired vasodilation (Crabtree et al., 2009). In primary aortic endothelial cells of GTPCH1-knockout mice, BH4 depletion significantly reduced eNOS activity, increased basal superoxide production and decreased NO bioactivity (Chuaiphichai et al., 2014). In rat hearts, BH4 content and eNOS activity were decreased in a time-dependent manner following myocardial ischemia with a marked decline after thirty minutes, while superoxide generation increased (Dumitrescu et al., 2007).</p>
  • <p>BAECs undergoing hypoxia and reoxygenation had decreased BH4 and decreased NO production, which was partially restored by treatment with the xanthine oxidase inhibitor oxypurinol, N-acetyl-l-cysteine (NAC) and NAC+BH4 (De Pascali et al., 2014). Inhibition of BH4 due to treatment with 4-HNE decreased eNOS activity and NO production in BAECs (Whitsett et al., 2007).</p>
  • <p>Many studies demonstrated that BH4 treatment reduced eNOS-mediated superoxide generation and increased NO formation in bovine, mouse, and rat endothelium (Chen et al., 2011; De Pascali et al., 2014; Landmesser et al., 2003; Ozaki et al., 2002; Shinozaki et al., 2000). Clinical studies reported improvement endothelial function in cardiovascular disease after treatment with BH4 (Wang et al., 2014). Oral treatment with BH4 in hypertensive patients significantly decreased blood pressure.</p>
  • <!-- event text -->
  • <h4>Key Event Description</h4>
  • <p>Hypertension is an important cardiovascular risk factor and considered one of the leading causes of cardiovascular morbidity and mortality (Kizhakekuttu and Widlansky, 2010). It is defined as a chronic elevation in blood pressure and is characterized by elevated systemic vascular resistance due to dysregulated vasomotor function and structural remodeling (Lee and Griendling, 2008). Although many genetic and environmental factors contribute to the development to hypertension, oxidative stress appears to be the main pathway involved in its pathogenesis. Excessive reactive oxygen species (ROS) contributes to <a href="/wiki/index.php/Event:932" title="Event:932">endothelial nitric oxide synthase (eNOS) uncoupling</a>, resulting in increased superoxide production but <a href="/wiki/index.php/Event:933" title="Event:933">decreased nitric oxide</a> (NO), a critical regulator of vascular homeostasis (Silva et al., 2012). Depletion of NO leads to <a href="/wiki/index.php/Event:937" title="Event:937">impaired endothelium-dependent vasodilation</a>, thus promoting endothelial dysfunction, which is a hallmark of hypertension.</p>
  • <br>
  • <strong>Empirical Support for Linkage</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>Multiple studies demonstrated a strong dependency between BH4 and eNOS uncoupling; decreased BH4 along with decreased eNOS activity, decreased NO production or increased superoxide generation were observed following various perturbations.</p>
  • <p>Prolonged myocardial ischemia (&gt;30 min) in isolated rat hearts caused extensive BH4 depletion (95% depletion), which was paralleled by decreased eNOS activity (58% reduction) and increased superoxide generation from &lt;0.01 relative fluorescence unit/mg protein to 0.3 measured using a fluorescence detector-based assay (Dumitrescu et al., 2007). Similarly, cardiac reperfusion patients exhibited a reduction in BH4 levels by 32%, decreased eNOS activity by 40% and increased superoxide production from 37.83 to 65.02 light unit/s/mg as detected using lucigenin-enhanced chemiluminescence (Jayaram et al., 2015).</p>
  • <h4>How it is Measured or Detected</h4>
  • <p>Arterial blood pressure is commonly measured using a sphygmomanometer, which provides systolic and diastolic blood pressure measurements in millimeters of mercury (mmHg).</p>
  • <p>In BAECs treated with 10 mmol/L DAHP or 25 &mu;M 4-HNE for 4 hours, BH4 levels decreased (control: 10 pmol/mg, DAHP: 4.8 pmol/mg, 4-HNE: 6.1 pmol/mg). At 24 hours, a reduction in NO (control: 1678 pmol/mg, DAHP: 1274 pmol/mg, 4-HNE: 1106 pmol/mg) and increased superoxide formation (control: 59 pmol/mg, DAHP: 97 pmol/mg, 4-HNE: 122 pmol/mg) were observed (Whitsett et al., 2007). Another study in BAECs using DAHP showed similar results for BH4 (control: 20.5 pmol/mg, DAHP: 11.8 pmol/mg), superoxide (control: 100%, DAHP: 257%), and NO (control: 96%, DAHP: 60%) (Wang et al., 2008).</p>
  • <p>Pathological hypertension is characterised according to current guidelines;&nbsp;https://www.nice.org.uk/guidance/cg127/evidence</p>
  • <p>In BAECs undergoing hypoxia and reoxygenation (H/R), treatment with oxypurinol increased BH4 levels from 6.1 &plusmn; 0.9 pmol/mg protein (after H/R) to 11.9 &plusmn; 0.8 pmol/mg protein and increased NO from 34.2 &plusmn; 1.7% to 63.7 &plusmn; 3.0%, demonstrating that these key events are modulated together (De Pascali et al., 2014).</p>
  • <p>Stage 1 hypertension : Clinic blood pressure is 140/90&nbsp;mmHg or higher&nbsp;and&nbsp;subsequent ambulatory blood pressure monitoring (ABPM) daytime average or home blood pressure monitoring (HBPM) average blood pressure is 135/85&nbsp;mmHg or higher.</p>
  • <p>In a rat model of hypertension, BH4 depletion and eNOS uncoupling induced by aortic coarctation (AoCo) was&nbsp;reversed by treatment with clofibrate; eNOS activity increased from 6.927&plusmn;3.475 ng L-citrulline/mg protein/30 min to 23.2 &plusmn; 9.034 ng L-citrulline/mg protein/30 min, BH4 levels increased from 17.144&plusmn;2.251 pmol to 32.534&plusmn;5.809 pmol, and superoxide decreased from relative fluorescence intensity of 32.22&plusmn;2.903 to 24.59&plusmn;1.124 (Cervantes-P&eacute;rez et al., 2012). Note that normal eNOS activity converts L-arginine to L-citrulline and produces NO; thus L-citrulline is an indirect measure of eNOS activity.</p>
  • <strong>Uncertainties or Inconsistencies</strong>
  • <p>The uncoupling of eNOS may also occur through other mechanisms such as S-glutathionylation of eNOS and depletion of L-arginine (Zweier et al., 2011).</p>
  • <strong>Quantitative Understanding of the Linkage</strong>
  • <p><em>Is it known how much change in the first event is needed to impact the second? Are there known modulators of the response-response relationships? Are there models or extrapolation approaches that help describe those relationships? </em></p>
  • <p>Stage 2 hypertension : Clinic blood pressure is 160/100&nbsp;mmHg or higher&nbsp;and&nbsp;subsequent ABPM daytime average or HBPM average blood pressure is 150/95&nbsp;mmHg or higher.</p>
  • <p>As BH4 is required for normal eNOS function, it could be possible that any change in BH4 may affect eNOS function. The studies above demonstrated that there are many modulators of the response-response relationships including cardiac reperfusion (Jayaram et al., 2015), DAHP (Wang et al., 2008; Whitsett et al., 2007), and 4-HNE (Whitsett et al., 2007).</p>
  • <p>Severe hypertension : Clinic systolic blood pressure is 180&nbsp;mmHg or higher&nbsp;or&nbsp;clinic diastolic blood pressure is 110&nbsp;mmHg or higher.</p>
  • <br>
  • <!--<!% unless aop_rel.relationship.relationship_taxons.blank? %>-->
  • <!--<!%= render 'snapshot_taxons', taxons: aop_rel.relationship.relationship_taxons %>-->
  • <!--<!% unless aop_rel.relationship.taxon_evidence.blank? %>-->
  • <!--<h3>Evidence Supporting Taxonomic Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
  • <!--<!% end %>-->
  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>Cervantes-P&eacute;rez, L.G., Ibarra-Lara, M. de la L., Escalante, B., Del Valle-Mondrag&oacute;n, L., Vargas-Robles, H., P&eacute;rez-Severiano, F., Pastel&iacute;n, G., and S&aacute;nchez-Mendoza, M.A. (2012). Endothelial nitric oxide synthase impairment is restored by clofibrate treatment in an animal model of hypertension. Eur. J. Pharmacol. 685, 108&ndash;115.</p>
  • <p>Chen, C.-A., Lin, C.-H., Druhan, L.J., Wang, T.-Y., Chen, Y.-R., and Zweier, J.L. (2011). Superoxide induces endothelial nitric-oxide synthase protein thiyl radical formation, a novel mechanism regulating eNOS function and coupling. J. Biol. Chem. 286, 29098&ndash;29107.</p>
  • <p>Chuaiphichai, S., McNeill, E., Douglas, G., Crabtree, M.J., Bendall, J.K., Hale, A.B., Alp, N.J., and Channon, K.M. (2014). Cell-autonomous role of endothelial GTP cyclohydrolase 1 and tetrahydrobiopterin in blood pressure regulation. Hypertension 64, 530&ndash;540.</p>
  • <p>Crabtree, M.J., Tatham, A.L., Al-Wakeel, Y., Warrick, N., Hale, A.B., Cai, S., Channon, K.M., and Alp, N.J. (2009). Quantitative regulation of intracellular endothelial nitric-oxide synthase (eNOS) coupling by both tetrahydrobiopterin-eNOS stoichiometry and biopterin redox status: insights from cells with tet-regulated GTP cyclohydrolase I expression. J. Biol. Chem. 284, 1136&ndash;1144.</p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p>Dumitrescu, C., Biondi, R., Xia, Y., Cardounel, A.J., Druhan, L.J., Ambrosio, G., and Zweier, J.L. (2007). Myocardial ischemia results in tetrahydrobiopterin (BH4) oxidation with impaired endothelial function ameliorated by BH4. Proc. Natl. Acad. Sci. U. S. A. 104, 15081&ndash;15086.</p>
  • <p>F&ouml;rstermann, U., and M&uuml;nzel, T. (2006). Endothelial nitric oxide synthase in vascular disease: from marvel to menace. Circulation 113, 1708&ndash;1714.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Landmesser, U., Dikalov, S., Price, S.R., McCann, L., Fukai, T., Holland, S.M., Mitch, W.E., and Harrison, D.G. (2003). Oxidation of tetrahydrobiopterin leads to uncoupling of endothelial cell nitric oxide synthase in hypertension. J. Clin. Invest. 111, 1201&ndash;1209.</p>
  • <p>Ozaki, M., Kawashima, S., Yamashita, T., Hirase, T., Namiki, M., Inoue, N., Hirata, K., Yasui, H., Sakurai, H., Yoshida, Y., et al. (2002). Overexpression of endothelial nitric oxide synthase accelerates atherosclerotic lesion formation in apoE-deficient mice. J. Clin. Invest. 110, 331&ndash;340.</p>
  • <p>Shinozaki, K., Nishio, Y., Okamura, T., Yoshida, Y., Maegawa, H., Kojima, H., Masada, M., Toda, N., Kikkawa, R., and Kashiwagi, A. (2000). Oral administration of tetrahydrobiopterin prevents endothelial dysfunction and vascular oxidative stress in the aortas of insulin-resistant rats. Circ. Res. 87, 566&ndash;573.</p>
  • <p>Durand, M.J., and Gutterman, D.D. (2013). Diversity in mechanisms of endothelium-dependent vasodilation in health and disease. Microcirc. N. Y. N 1994 20, 239&ndash;247.</p>
  • <p>Wang, Q., Yang, M., Xu, H., and Yu, J. (2014). Tetrahydrobiopterin improves endothelial function in cardiovascular disease: a systematic review. Evid.-Based Complement. Altern. Med. ECAM 2014, 850312.</p>
  • <p>Kizhakekuttu, T.J., and Widlansky, M.E. (2010). Natural antioxidants and hypertension: promise and challenges. Cardiovasc. Ther. 28, e20&ndash;e32.</p>
  • <p>Wang, S., Xu, J., Song, P., Wu, Y., Zhang, J., Chul Choi, H., and Zou, M.-H. (2008). Acute inhibition of guanosine triphosphate cyclohydrolase 1 uncouples endothelial nitric oxide synthase and elevates blood pressure. Hypertension 52, 484&ndash;490.</p>
  • <p>Leong, X.-F., Ng, C.-Y., Jaarin, K., Leong, X.-F., Ng, C.-Y., and Jaarin, K. (2015). Animal Models in Cardiovascular Research: Hypertension and Atherosclerosis, Animal Models in Cardiovascular Research: Hypertension and Atherosclerosis. BioMed Res. Int. BioMed Res. Int. 2015, 2015, e528757.</p>
  • <p>Whitsett, J., Picklo, M.J., and Vasquez-Vivar, J. (2007). 4-Hydroxy-2-nonenal increases superoxide anion radical in endothelial cells via stimulated GTP cyclohydrolase proteasomal degradation. Arterioscler. Thromb. Vasc. Biol. 27, 2340&ndash;2347.</p>
  • <p>Silva, B.R., Pernomian, L., and Bendhack, L.M. (2012). Contribution of oxidative stress to endothelial dysfunction in hypertension. Front. Physiol. 3, 441.</p>
  • <br>
  • <!-- end event text -->
  • </div>
  • <br>
  • <div>
  • <h2>Appendix 2</h2>
  • <h2>List of Key Event Relationships in the AOP</h2>
  • <!-- Evidence for relationship links section, this lists the relationships and then supports them -->
  • <div id="evidence_supporting_links">
  • <hr>
  • <h3>List of Adjacent Key Event Relationships</h3>
  • <div>
  • <h4><a href="/relationships/956">Uncoupling, eNOS leads to Depletion, Nitric Oxide</a></h4>
  • <h4><a href="/relationships/980">Relationship: 980: Peptide Oxidation leads to Decrease, GTPCH-1</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Directness</th>
  • <th>Adjacency</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>directly leads to</th>
  • <th>Strong </th>
  • <th>Strong</th>
  • <th>adjacent</th>
  • <th>Moderate </th>
  • <th>Low</th>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Strong</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>The relationship between NO depletion and eNOS uncoupling was observed in humans, cows, and rats as demonstrated by the above studies.</p>
  • <p>Several studies showed decreased GTPCH-1 activity and/or protein expression under oxidative stress in cardiac reperfusion patients, bovine endothelial cells, a mouse model of diabetes and a rat model of hypertension (Cervantes-P&eacute;rez et al., 2012; AbdelGhany et al., (under review); Jayaram et al., 2015; Zhao et al., 2013).</p>
  • <h4>How Does This Key Event Relationship Work</h4>
  • <p>The uncoupling of eNOS occurs through a number of mechanisms like BH4 depletion or S-glutathionylation of eNOS and leads to a reduction in NO bioavailability and an elevation in superoxide production, contributing to endothelial dysfunction (Zweier et al., 2011).</p>
  • <h4>Key Event Relationship Description</h4>
  • <p>Exposure to known inducers of oxidative stress such as cigarette smoke extract (AbdelGhany et al., under review) or peroxynitrite (Zhao et al., 2013) causes the loss of GTPCH-1 activity, resulting in decreased levels of tetrahydrobiopterin (BH4) and subsequent uncoupling of endothelial nitric oxide synthase (eNOS).</p>
  • <!-- if nothing shows up in any of these fields, then weight of evidence will not be displayed -->
  • <h4>Weight of Evidence</h4>
  • <!-- if nothing shows up in any of these fields, then evidence supporting this KER will not be displayed -->
  • <h4>Evidence Supporting this KER</h4>
  • <strong>Biological Plausibility</strong>
  • <p>It is well-established that uncoupling of eNOS causes eNOS to switch from producing NO to generating superoxides (F&ouml;rstermann and M&uuml;nzel, 2006). Many studies that reported <a href="/wiki/index.php/Relationship:953" title="Relationship:953">BH4 depletion leading to eNOS uncoupling</a> or <a href="/wiki/index.php/Relationship:955" title="Relationship:955">S-glutathionylation leading to eNOS uncoupling</a> measured levels of NO and superoxide which are indicative of eNOS uncoupling.</p>
  • <p>Several studies demonstrated that GTPCH-1 is also affected by oxidative stress, which provides evidence for moderate biological plausibility. In vitro exposure to chemically synthesized peroxynitrite inhibited GTPCH-1 activity in a dose-dependent manner (Zhao et al., 2013). This inhibition as well as increased ubiquitination of GTPCH-1 were observed in cultured bovine aortic endothelial cells (BAECs) and streptozotocin-induced diabetes in mice following peroxynitrite treatment. Ubiquitination of GTPCH-1 leads to its degradation, which is equivalent to a decrease in GTPCH-1. In another study, GTPCH-1 levels were reduced by cigarette smoke extract (CSE) exposure in BAECs (AbdelGhany et al., under review). There is also evidence that CSE promoted GTPCH-1 degradation by increasing proteasomal activity. Furthermore,&nbsp;cardiac reperfusion patients experienced oxidative stress which was associated with reduced GTPCH-1 activity (Jayaram et al., 2015).</p>
  • <strong>Empirical Support for Linkage</strong>
  • <strong>Empirical Evidence</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>Multiple experiments demonstrated that eNOS uncoupling results in increased superoxide formation and decreased NO production, which provide strong empirical support for this KER.</p>
  • <p>Prolonged myocardial ischemia (&gt;30 min) in isolated rat hearts decreased eNOS activity (58% reduction) and increased superoxide generation from &lt;0.01 relative fluorescence unit/mg to 0.3 as measured using a fluorescence detector-based assay (Dumitrescu et al., 2007). Similarly, cardiac reperfusion patients had decreased eNOS activity (40% reduction) and increased superoxide production from 37.83 to 65.02 light unit/s/mg as detected using lucigenin-enhanced chemiluminescence (Jayaram et al., 2015).</p>
  • <p>In BAECs, treatment with 10 mmol/L DAHP or 25 &mu;M 4-HNE for 24 hours led to a reduction in NO (control: 1678 pmol/mg, DAHP: 1274 pmol/mg, 4-HNE: 1106 pmol/mg) and increased superoxide formation (control: 59 pmol/mg, DAHP: 97 pmol/mg, 4-HNE: 122 pmol/mg) (Whitsett et al., 2007). Similar results were observed in another study using DAHP-treated BAECs; superoxide was increased (control: 100%, DAHP: 257%) and NO was decreased (control: 96%, DAHP: 60%) (Wang et al., 2008).</p>
  • <p>In BAECs undergoing hypoxia and reoxygenation (H/R), NO decreased from 34.2&plusmn;1.7% to 63.7&plusmn;3.0% and superoxide increased from relative intensity of 3.5 to 49.2, demonstrating that these key events are modulated together (De Pascali et al., 2014).</p>
  • <p>BCNU (25 &mu;M, 80 &mu;M) resulted in increased superoxide (25 &mu;M: 432 fluorescence intensity, 80 &mu;M:759 fluorescence intensity) and decreased NO (25 &mu;M: 61%, 80 &mu;M:36%) in a dose-dependent manner in BAECs (Chen et al., 2010).</p>
  • <p>Exposure to 50 &mu;mol/L peroxynitrite for 3 hours led to an increase in superoxide from 2 nmol/min/well to 8.6 nmol/min/well and a decrease in NO from 90% to 6.5% in BAECs (Zou et al., 2002).</p>
  • <p>In a rat model of aortic coarctation-associated hypertension, increased ROS (1.75 nmol DCF/mg protein/min) and decreased GTPCH-1 protein expression (western blot band intensity of 0.0087&plusmn;0.005) were observed (Cervantes-P&eacute;rez et al., 2012). However, following clofibrate treatment (100 mg/kg/day, 7 days), ROS was reduced by approximately 30% and GTPCH-1 band intensity increased to 0.0087&plusmn;0.005. This study provides evidence that there is a dependent change between oxidative stress and GTPCH-1, but we found no dose-response relationship, so empirical support for this KER is weak.</p>
  • <strong>Uncertainties and Inconsistencies</strong>
  • <p>No uncertainties or inconsistencies were found for this KER.</p>
  • <strong>Uncertainties or Inconsistencies</strong>
  • <p>There are no uncertainties or inconsistencies.</p>
  • <strong>Quantitative Understanding of the Linkage</strong>
  • <p><em>Is it known how much change in the first event is needed to impact the second? Are there known modulators of the response-response relationships? Are there models or extrapolation approaches that help describe those relationships? </em></p>
  • <p>The uncoupling of eNOS automatically results in NO depletion and increased superoxide production. The experimental studies above included a number of modulators of the response-response relationship, such as peroxynitrite, BCNU, and DAHP.</p>
  • <!--<!% unless aop_rel.relationship.relationship_taxons.blank? %>-->
  • <!--<!%= render 'snapshot_taxons', taxons: aop_rel.relationship.relationship_taxons %>-->
  • <!--<!% unless aop_rel.relationship.taxon_evidence.blank? %>-->
  • <!--<h3>Evidence Supporting Taxonomic Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
  • <!--<!% end %>-->
  • <!--<!%= render 'snapshot_taxons', taxons: aop_rel.relationship.relationship_taxons %>-->
  • <!--<!% unless aop_rel.relationship.taxon_evidence.blank? %>-->
  • <!--<h3>Domain of Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
  • <!--<!% end %>-->
  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>Chen, X., Xu, J., Feng, Z., Fan, M., Han, J., and Yang, Z. (2010). Simvastatin combined with nifedipine enhances endothelial cell protection by inhibiting ROS generation and activating Akt phosphorylation. Acta Pharmacol. Sin. 31, 813&ndash;820.</p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p>Dumitrescu, C., Biondi, R., Xia, Y., Cardounel, A.J., Druhan, L.J., Ambrosio, G., and Zweier, J.L. (2007). Myocardial ischemia results in tetrahydrobiopterin (BH4) oxidation with impaired endothelial function ameliorated by BH4. Proc. Natl. Acad. Sci. U. S. A. 104, 15081&ndash;15086.</p>
  • <p>F&ouml;rstermann, U., and M&uuml;nzel, T. (2006). Endothelial nitric oxide synthase in vascular disease: from marvel to menace. Circulation 113, 1708&ndash;1714.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Wang, S., Xu, J., Song, P., Wu, Y., Zhang, J., Chul Choi, H., and Zou, M.-H. (2008). Acute inhibition of guanosine triphosphate cyclohydrolase 1 uncouples endothelial nitric oxide synthase and elevates blood pressure. Hypertension 52, 484&ndash;490.</p>
  • <p>AbdelGhany, T., Ismail, R., Elmahdy, M., Mansoor F, Zweier J, Lowe, F., and Zweier, JL. (2017). Cigarette Smoke Constituents Cause Endothelial Nitric Oxide Synthase Dysfunction and Uncoupling due to Depletion of Tetrahydrobiopterin with Degradation of GTP Cyclohydrolase.&nbsp;&nbsp;Nitric Oxide&nbsp;(Under review)</p>
  • <p>Whitsett, J., Picklo, M.J., and Vasquez-Vivar, J. (2007). 4-Hydroxy-2-nonenal increases superoxide anion radical in endothelial cells via stimulated GTP cyclohydrolase proteasomal degradation. Arterioscler. Thromb. Vasc. Biol. 27, 2340&ndash;2347.</p>
  • <p>Cervantes-P&eacute;rez, L.G., Ibarra-Lara, M. de la L., Escalante, B., Del Valle-Mondrag&oacute;n, L., Vargas-Robles, H., P&eacute;rez-Severiano, F., Pastel&iacute;n, G., and S&aacute;nchez-Mendoza, M.A. (2012). Endothelial nitric oxide synthase impairment is restored by clofibrate treatment in an animal model of hypertension. Eur. J. Pharmacol. 685, 108&ndash;115.</p>
  • <p>Zou, M.-H., Hou, X.-Y., Shi, C.-M., Nagata, D., Walsh, K., and Cohen, R.A. (2002). Modulation by peroxynitrite of Akt- and AMP-activated kinase-dependent Ser1179 phosphorylation of endothelial nitric oxide synthase. J. Biol. Chem. 277, 32552&ndash;32557.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49</p>
  • <p>Zweier, J.L., Chen, C.-A., and Druhan, L.J. (2011). S-glutathionylation reshapes our understanding of endothelial nitric oxide synthase uncoupling and nitric oxide/reactive oxygen species-mediated signaling. Antioxid. Redox Signal. 14, 1769&ndash;1775.</p>
  • <p>Zhao, Y., Wu, J., Zhu, H., Song, P., and Zou, M.-H. (2013). Peroxynitrite-dependent zinc release and inactivation of guanosine 5&rsquo;-triphosphate cyclohydrolase 1 instigate its ubiquitination in diabetes. Diabetes 62, 4247&ndash;4256.</p>
  • </div>
  • <br>
  • <div>
  • <div>
  • <h4><a href="/relationships/958">Depletion, Nitric Oxide leads to Impaired, Vasodilation</a></h4>
  • <h4><a href="/relationships/955">Relationship: 955: S-Glutathionylation, eNOS leads to Uncoupling, eNOS</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Directness</th>
  • <th>Adjacency</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>directly leads to</th>
  • <th>Strong </th>
  • <th>adjacent</th>
  • <th>High </th>
  • <th>Moderate</th>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Strong</td>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Oryctolagus cuniculus</td>
  • <td>Oryctolagus cuniculus</td>
  • <td>Moderate</td>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9986" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>This relationship between NO depletion and impaired vasodilation was shown in humans (Li et al. 1993, Heitzer et al. 2000), rabbits (Luo et al. 2000), mice (Luo et al. 2000, Wang et al. 2008)&nbsp;and rats (Paulis et al. 2008, Li et al. 1993, S&eacute;lley et al. 2014, Chen et al. 2010).</p>
  • <p>The evidence supporting this key event relationship are from human subjects, HAECs, BAECs, and SHR rats (Chen et al., 2010; De Pascali et al., 2014; Du et al., 2013; Jayaram et al., 2015).</p>
  • <h4>How Does This Key Event Relationship Work</h4>
  • <p>Nitric oxide (NO)&nbsp;is a critical endothelium-derived hyperpolarising factor (EDHF), responsible for relaxation of vascular smooth muscle and vasodilation.&nbsp; The primary regulator of endothelial vasodilator function via NO is vascular shear; the frictional force exerted on the vascular wall during the flow of blood through the vessel. Vascular shear opens calcium channels on endothelial cells, and leads to the calcium-dependent activation of eNOS and thus NO production.&nbsp; NO then diffuses to the underlying vascular smooth muscle, where it activates soluble guanylate cyclase, causing an increase in cyclic guanosine monophosphate (cGMP), potassium ion efflux, hyperpolarization and smooth muscle relaxation (Giles et al. 2012).</p>
  • <p>Depletion of vascular NO bioavailability causes an imbalance in the maintenance of vascular tone, which shifts in favour of vasoconstriction, and hence elevates blood pressure (Kojda et al. 1999).&nbsp; Under oxidative stress, decreased NO bioavailability results in impaired endothelium-dependent vasodilation (Silva et al., 2012).</p>
  • <h4>Key Event Relationship Description</h4>
  • <p>Oxidative stress can trigger S-glutathionylation of eNOS at cysteine residues Cys689 and Cys908, which are known to be critical for normal eNOS function (Zweier et al., 2011). S-glutathionylation directly causes eNOS uncoupling, a state in which eNOS switches from producing NO to generating superoxide, thus impairing endothelium-dependent vasodilation and contributing to endothelial dysfunction. Uncoupling of eNOS via S-glutathionylation is different from BH4-mediated eNOS uncoupling in that superoxide is produced in the reductase domain rather than the oxygenase domain and superoxide generation cannot be inhibited by L-NG-nitroarginine methyl ester (L-NAME), suggesting that S-glutathionylation occurs independent of calcium/calmodulin and heme.</p>
  • <!-- if nothing shows up in any of these fields, then weight of evidence will not be displayed -->
  • <h4>Weight of Evidence</h4>
  • <!-- if nothing shows up in any of these fields, then evidence supporting this KER will not be displayed -->
  • <h4>Evidence Supporting this KER</h4>
  • <strong>Biological Plausibility</strong>
  • <p>Vasodilation is caused by the relaxation of vascular smooth muscle cells within the walls of blood vessels, which is regulated through a number of mechanisms, including cyclic GMP-dependent hyperpolarization and relaxation via NO. Thus, alterations to NO levels have an influence on vasodilation (Silva et al., 2012). Many animal studies demonstrated that inhibition of NO via eNOS inhibitors impaired acetylcholine-induced endothelium-dependent vasodilation, providing strong biological plausibility for this key event relationship.</p>
  • <p>Constitutively active AKT, which has been shown to phosphorylate and activate eNOS, increased resting vessel diameter in a rabbit femoral artery model, but infusion with eNOS inhibitor L-NAME reversed this effect (Luo et al., 2000). Dominant negative AKT, which prevents eNOS activation, also blocked vasodilation in response to the endothelium-dependent agonist acetylcholine (ACh) in rabbit arteries and mouse aortas. Since acetylcholine induces endothelium-dependent vasodilation, the assumption is that when eNOS inhibitors blocks Ach-induced vasodilation, NO production is decreased and vasodilation is impaired. L-NAME abolished endothelium-dependent relaxations induced by ACh in aortas of wild-type and hph-1 mice (Li et al., 2007). ACh induced a dose-dependent endothelium-dependent relaxation in rat small mesenteric arteries, but the NO-dependent component of ACh-induced relaxation was decreased in L-NAME-treated rats (Paulis et al., 2008). After L-NAME cessation, the NO-dependent component was restored to above control levels. L-NAME diminished eNOS activity by 66% and its cessation restore NOS activity to control levels in rat aortas. Another study showed that L-NAME caused a mild but significant decrease in vasodilation in rat thoracic aorta (S&eacute;lley et al., 2014).</p>
  • <p><em>In vitro</em> experiments showed that S-glutathionylation of eNOS significantly decreased NO activity in dose-dependent manner and greatly increased superoxide generation (Chen et al., 2010). NOS inhibitor L-NAME partially blocked superoxide generation. These results were observed in bovine aortic endothelial cells (BAECs) and in aortae of spontaneously hypertensive (SHR) rats. Treatment of BAECs with 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), an inhibitor of glutathione reductase, induced eNOS S-glutathionylation, resulting in decreased NO and increased superoxide generation. Gene silencing of eNOS in BAECs also induced superoxide production. In aortae of SHR rats, S-glutathionylation as determined by immunohistology was associated with a decrease in endothelium-dependent vasodilation as a result of decreased NO. Exposure of human aortic endothelial cells (HAECs) to ultrafine particles (UFP) caused S-glutathionylation of eNOS and a dose-dependent decrease in NO production (Du et al., 2013). Decreased NO production was found to be mediated by S-glutathionylation since overexpression of glutaredoxin, an inhibitor of S-glutathionylation, significantly reduced UPF-mediated decrease in NO production. Cardiac reperfusion patients exhibited decreased eNOS activity which was identified to be a result of eNOS S-glutathionylation (Jayaram et al., 2015). Additional evidence was observed in BAECs undergoing hypoxia and reoxygenation in which eNOS S-glutathionylation increased by three-fold compared to control cells and NO production was decreased (De Pascali et al., 2014). These effects were reversed with treatment of N-acetyl-l-cysteine, which increased cellular concentration of GSH. These results demonstrate a clear interaction between&nbsp;S-glutathionylation and eNOS uncoupling, therefore this link has strong biological plausibility.</p>
  • <strong>Empirical Support for Linkage</strong>
  • <strong>Empirical Evidence</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>There is moderate empirical support for depletion of NO leading to impaired vasodilation based on several studies measuring decreased NO and vasodilation following a number of perturbations. In rat small mesenteric arteries, treatment with L-NAME for five weeks decreased NO-dependent relaxation from 28% to 7.5%, while diminishing eNOS activity by 66% (Paulis et al, 2008). In rat aortic rings, BCNU treatment for 4 hours caused a dose-dependent decrease in NO production (control: 100%, 25 &mu;M: 61%, 80 &mu;M: 36%), and also decreased vasodilation from 69% control to 37% (at 80 &mu;M) (Chen et al., 2010). Similarly, treatment with 10 mM DAHP for 24 hours caused a decrease in both NO (from 100% to 61%) and vasodilation (from 87% to 42%) (Wang et al., 2008).</p>
  • <p>Treatment with BCNU (25 &mu;M, 80 &mu;M) resulted in increased eNOS S-glutathionylation, increased superoxide generation and decreased NO production in a dose-dependent manner in BAECs (Chen et al., 2010).</p>
  • <p>In&nbsp;humans, application of L-NAME and/or N<sup>G</sup>-monomethyl-L-arginine (L-NMMA) resulted in elevated mean arterial pressure via impairment of NO-mediated vasodilation.&nbsp; Application of both drugs compounded the effect (Sander et al. 1999).&nbsp; BH4 administration reversed NO depletion by L-NMMA and improved forearm venous blood flow in chronic smokers (Heizter et al. 2000).&nbsp; In human skin, acetylcholine-mediated vasodilation was shown to be modulated by NO and impaired by the eNOS inhibitor; L-NAME (Kellogg et al. 2005).&nbsp; Intravenous infusion&nbsp;of the eNOS substrate L-arginine was shown to decrease blood pressure and total peripheral resistance.&nbsp;&nbsp;The onset and the duration of the vasodilator effect of L-arginine and its effects on endogenous NO production closely corresponded to the plasma concentration half-life of l-arginine, and was associatedwith a concomminant elevation of urinary nitrate and cyclic GMP excretion (Bode-B&ouml;ger et al. 1998).&nbsp; Finally, in chronic smokers with impaired flow-mediated dilation (FMD;&nbsp;5.6&plusmn;3.0% vs. 8.1&plusmn;3.7% for non-smokers&nbsp;P&lt;0.01), administration of the essential eNOS cofactor; BH4, improved FMD in both cohorts (6.6&plusmn;3.3% vs. 9.8&plusmn;3.2%; P&lt;0.01) and smoking cessation for 1 week also improved FMD (from 5.0&plusmn;2.9 to 7.8&plusmn;3.2%;P&lt;0.01).&nbsp; This data indicates that improvement in arterial vasodilation was partially improved by BH4 administration (Taylor et al. 2016).&nbsp; Another study by Carnevale et al. (2016) in which current smokers switched to an e-cigarette product for 1 week showed significant reductions in FMD impairment compared to baseline, which was associated with improvement in levels of 8-iso-prostagladin F2&alpha; (a biomarker of oxidative stress).</p>
  • <p>Exposure to hypoxia/reoxygenation and treatment with angiotensin II (Ang II) demonstrated a response-response relationship between eNOS S-glutathionylation and superoxide generation in human and bovine endothelial cells (De Pascali et al., 2014; Galougahi et al., 2014). Following hypoxia and reoxygenation in BAECs, increased S-glutathionylation of eNOS (0.3 to 1 arbitray units; measured by western blot), decreased NO production (100% to 34.2 &plusmn; 1.7%) and increased superoxide generation (relative intensity of 3.5 to 49.2; by fluorescence) were observed (De Pascali et al., 2014). Treatment with 100 nmol/L Ang II for 1 hour caused an increase in both eNOS S-glutathionylation (from 1 to 1.42 fold) and superoxide generation (from 81.1 to 113 fluorescence signal unit) (Galougahi et al., 2014).</p>
  • <p>Overall, there is moderate empirical support for eNOS S-glutathionylation leading to eNOS uncoupling.</p>
  • <strong>Uncertainties and Inconsistencies</strong>
  • <p>No uncertainties or inconsistencies were found for this KER.</p>
  • <strong>Uncertainties or Inconsistencies</strong>
  • <p>While the effect of NO depletion on impaired vasodilation is clear, the NO pathway does not appear to be solely responsible for this phenomenon.&nbsp; Vascular tone is a balance between relaxation and constriction factors.&nbsp; Studies have shown that when NO bioavailability is decreased, COX-mediated pro-inflammatory factors such as prostaglandins (Kellogg et al. 2005, L&uuml;scher and Vanhoutte 1986) and Endothelin-1 (Taddei et al. 2003)&nbsp; contribute to a shift in vascular tone towards vasoconstiction.&nbsp; The&nbsp;review by Silva et al. (2012) discusses the roles of various pathways and their effects on vascular tone.&nbsp; The relative contribution of these mechanisms towards vascular tone is currently unknown.</p>
  • <strong>Quantitative Understanding of the Linkage</strong>
  • <p><em>Is it known how much change in the first event is needed to impact the second? Are there known modulators of the response-response relationships? Are there models or extrapolation approaches that help describe those relationships? </em></p>
  • <p>In rat aortic rings, BCNU treatment for 4 hours caused a dose-dependent decrease in NO production (control: 100%, 25 &mu;M: 61%, 80 &mu;M: 36%), and also decreased vasodilation from 69% control to 37% (at 80 &mu;M) (Chen et al., 2010). Similarly, treatment with 10 mM DAHP for 24 hours caused a decrease in both NO (from 100% to 61%) and vasodilation (from 87% to 42%) (Wang et al., 2008).</p>
  • <p>Hence, it appears that a decrease&nbsp;of 40%&nbsp;NO relative to baseline&nbsp;conditions appears sufficient to impact vasodilation since the experiments show that NO decreases to near 60% after applying the stressors. The studies used several perturbations that were able to modulate NO production and vasodilation simultaneously including L-NAME, DAHP and BCNU.&nbsp;&nbsp;The evidence is qualitative however.</p>
  • <p>Furthermore, as NO is a highly volatile substance, researchers are not able to treat models/subjects with specific doses to observe the corresponding biological effects.&nbsp; NO donors such as sodium nitroprusside and glyceryl trinitrate are often used to generate&nbsp;NO in humans and animal studies, however this is indepedent of the vascular endothelium.&nbsp; More stable metabolites of NO&nbsp;are often measured in vivo to estimate NO turnover e.g. Nitrate/nitrite, however the conversion kinetics&nbsp;of NO to these metabolites is unclear and hence limited to qualitative comparisons.&nbsp; An example is the study by Bode-B&ouml;ger et al. (1998) which investigated the pharmacokinetics of the eNOS substrate L-arginine and its subsequent effects upon vasodilation (measured using total peripheral resistance and blood pressure as surrogate endpoints).&nbsp;&nbsp;Plasma l-arginine levels increased to (mean&plusmn;s.e.mean) 6223&plusmn;407 (range, 5100&ndash;7680) and 822&plusmn;59 (527&ndash;955) &mu;mol&nbsp;l&minus;1&nbsp;after intravenous infusion of 30&nbsp;g and 6&nbsp;g l-arginine, respectively, and to 310&plusmn;152 (118&ndash;1219) &mu;mol&nbsp;l&minus;1&nbsp;after oral ingestion of 6&nbsp;g l-arginine. Oral bioavailability of l-arginine was 68&plusmn;9 (51&ndash;87)%. Clearance was 544&plusmn;24 (440&ndash;620), 894&plusmn;164 (470&ndash;1190), and 1018&plusmn;230 (710&ndash;2130)&nbsp;ml&nbsp;min&minus;1, and elimination half-life was calculated as 41.6&plusmn;2.3 (34&ndash;55), 59.6&plusmn;9.1 (24&ndash;98), and 79.5&plusmn;9.3 (50&ndash;121) min, respectively, for 30&nbsp;g i.v., 6&nbsp;g i.v., and 6&nbsp;g p.o. of l-arginine. Blood pressure and total peripheral resistance were significantly decreased after intravenous infusion of 30&nbsp;g l-arginine by 4.4&plusmn;1.4% and 10.4&plusmn;3.6%, respectively, but were not significantly changed after oral or intravenous administration of 6&nbsp;g l-arginine. l-arginine (30&nbsp;g) also significantly increased urinary nitrate and cyclic GMP excretion rates by 97&plusmn;28 and 66&plusmn;20%, respectively. After infusion of 6&nbsp;g l-arginine, urinary nitrate excretion also significantly increased, (nitrate by 47&plusmn;12% [P&lt;0.05], cyclic GMP by 67&plusmn;47% [P=ns]), although to a lesser and more variable extent than after 30&nbsp;g of l-arginine. The onset and the duration of the vasodilator effect of l-arginine and its effects on endogenous NO production closely corresponded to the plasma concentration half-life of l-arginine, as indicated by an equilibration half-life of 6&plusmn;2 (3.7&ndash;8.4) min between plasma concentration and effect in pharmacokinetic-pharmacodynamic analysis, and the lack of hysteresis in the plasma concentration-versus-effect plot.</p>
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  • <!--<h3>Evidence Supporting Taxonomic Applicability</h3>-->
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  • <!--<h3>Domain of Applicability</h3>-->
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  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>Bode-B&ouml;ger SM,&nbsp;B&ouml;ger RH,&nbsp;Galland A,&nbsp;Tsikas D,&nbsp;Fr&ouml;lich JC.&nbsp; L-arginine-induced&nbsp;vasodilation&nbsp;in&nbsp;healthy&nbsp;humans:&nbsp;pharmacokinetic-pharmacodynamicrelationship.&nbsp; Br J Clin Pharmacol.&nbsp;1998, 46(5):489-97.</p>
  • <p>Carnevale R, Sciarretta S, Violi F et al.&nbsp; Acute Impact of Tobacco vs Electronic Cigarette Smoking on Oxidative Stress and Vascular Function.&nbsp; Chest. (2016) 150(3):606-12.</p>
  • <p>Chen, C.-A., Wang, T.-Y., Varadharaj, S., Reyes, L.A., Hemann, C., Talukder, M.A.H., Chen, Y.-R., Druhan, L.J., and Zweier, J.L. (2010). S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature 468, 1115&ndash;1118.</p>
  • <p>Giles TD, Sander GE, Nossaman BD et al. Impaired vasodilation in the pathogenesis of hypertension: focus on nitric oxide, endothelial-derived hyperpolarizing factors, and prostaglandins.&nbsp; J Clin Hypertens (2012) 14(4):198-205.</p>
  • <p>Heitzer T,&nbsp;Brockhoff C,&nbsp;Mayer B,&nbsp;Warnholtz A,&nbsp;Mollnau H,&nbsp;Henne S,&nbsp;Meinertz T,&nbsp;M&uuml;nzel T.&nbsp; Tetrahydrobiopterin improves endothelium-dependent&nbsp;vasodilation&nbsp;in chronic smokers : evidence for a dysfunctional&nbsp;nitric oxide&nbsp;synthase.&nbsp; Circ Res.&nbsp;2000, 86(2):E36-41.</p>
  • <p>Kellogg DL Jr,&nbsp;Zhao JL,&nbsp;Coey U,&nbsp;Green JV.&nbsp; Acetylcholine-induced&nbsp;vasodilation&nbsp;is mediated by&nbsp;nitric oxide&nbsp;and prostaglandins in&nbsp;human&nbsp;skin.&nbsp; J Appl Physiol (1985).&nbsp;2005, 98(2):629-32.</p>
  • <p>Kojda G, Laursen JB, Ramasamy S et al. Protein expression, vascular reactivity and soluble guanylate cyclase activity in mice lacking the endothelial cell nitric oxide synthase: contributions of NOS isoforms to blood pressure and heart rate control.&nbsp; Cardiovasc Res. (1999) 42(1):206-13.</p>
  • <p>Li, J., Zhou, Z., Jiang, D.-J., Li, D., Tan, B., Liu, H., and Li, Y.-J. (2007). Reduction of NO- and EDHF-mediated vasodilatation in hypertension: role of asymmetric dimethylarginine. Clin. Exp. Hypertens. N. Y. N 1993 29, 489&ndash;501.</p>
  • <p>L&uuml;scher T. F., Vanhoutte P. M. (1986).&nbsp;Endothelium-dependent contractions to acetylcholine in the aorta of the spontaneously hypertensive rat.&nbsp;Hypertension&nbsp;8, 344&ndash;348</p>
  • <p>Luo, Z., Fujio, Y., Kureishi, Y., Rudic, R.D., Daumerie, G., Fulton, D., Sessa, W.C., and Walsh, K. (2000). Acute modulation of endothelial AKT/PKB activity alters nitric oxide&ndash;dependent vasomotor activity in vivo. J. Clin. Invest. 106, 493&ndash;499.</p>
  • <p>Paulis, L., Zicha, J., Kunes, J., Hojna, S., Behuliak, M., Celec, P., Kojsova, S., Pechanova, O., and Simko, F. (2008). Regression of L-NAME-induced hypertension: the role of nitric oxide and endothelium-derived constricting factor. Hypertens. Res. Off. J. Jpn. Soc. Hypertens. 31, 793&ndash;803.</p>
  • <p>Sander M, Chavoshan B, Victor RG.&nbsp; A large blood pressure-raising effect of nitric oxide synthase inhibition in humans.&nbsp; Hypertension. (1999) 33(4):937-42.</p>
  • <p>Chen, C.-A., Wang, T.-Y., Varadharaj, S., Reyes, L.A., Hemann, C., Talukder, M.A.H., Chen, Y.-R., Druhan, L.J., and Zweier, J.L. (2010). S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature 468, 1115&ndash;1118.</p>
  • <p>S&eacute;lley, E., Kun, S., Szij&aacute;rt&oacute;, I.A., Laczy, B., Kov&aacute;cs, T., F&uuml;l&ouml;p, F., Wittmann, I., and Moln&aacute;r, G.A. (2014). Exenatide induces aortic vasodilation increasing hydrogen sulphide, carbon monoxide and nitric oxide production. Cardiovasc. Diabetol. 13, 69.</p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p>Silva, B.R., Pernomian, L., and Bendhack, L.M. (2012). Contribution of oxidative stress to endothelial dysfunction in hypertension. Front. Physiol. 3, 441.</p>
  • <p>Du, Y., Navab, M., Shen, M., Hill, J., Pakbin, P., Sioutas, C., Hsiai, T.K., and Li, R. (2013). Ambient ultrafine particles reduce endothelial nitric oxide production via S-glutathionylation of eNOS. Biochem. Biophys. Res. Commun. 436, 462&ndash;466.</p>
  • <p>Taddei S., Ghiadoni L., Virdis A., Versari D., Salvetti A. (2003).&nbsp;Mechanisms of endothelial dysfunction: clinical significance and preventive non-pharmacological therapeutic strategies.&nbsp;Curr. Pharm. Des.&nbsp;9, 2385&ndash;2402</p>
  • <p>Galougahi, K.K., Liu, C.-C., Gentile, C., Kok, C., Nunez, A., Garcia, A., Fry, N.A.S., Davies, M.J., Hawkins, C.L., Rasmussen, H.H., et al. (2014). Glutathionylation Mediates Angiotensin II&ndash;Induced eNOS Uncoupling, Amplifying NADPH Oxidase-Dependent Endothelial Dysfunction. J. Am. Heart Assoc. 3, e000731.</p>
  • <p>Taylor BA, Zaleski AL, Dornelas EA et al.&nbsp; The impact of tetrahydrobiopterin administration on endothelial function before and after smoking cessation in chronic smokers.&nbsp; Hypertens Res. (2016) 39(3):144-50.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Wang, S., Xu, J., Song, P., Wu, Y., Zhang, J., Chul Choi, H., and Zou, M.-H. (2008). Acute inhibition of guanosine triphosphate cyclohydrolase 1 uncouples endothelial nitric oxide synthase and elevates blood pressure. Hypertension 52, 484&ndash;490.</p>
  • <p>Zweier, J.L., Chen, C.-A., and Druhan, L.J. (2011). S-glutathionylation reshapes our understanding of endothelial nitric oxide synthase uncoupling and nitric oxide/reactive oxygen species-mediated signaling. Antioxid. Redox Signal. 14, 1769&ndash;1775.</p>
  • </div>
  • <br>
  • <div>
  • <div>
  • <h4><a href="/relationships/982">Impaired, Vasodilation leads to Increase, Vascular Resistance</a></h4>
  • <h4><a href="/relationships/952">Relationship: 952: Decrease, GTPCH-1 leads to Decrease, Tetrahydrobiopterin</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Directness</th>
  • <th>Adjacency</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>directly leads to</th>
  • <th>Moderate </th>
  • <th>Weak</th>
  • <th>adjacent</th>
  • <th>High </th>
  • <th>High</th>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Moderate</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Weak</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Weak</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Not Specified</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>This relationship between impaired vasodilation and SVR was shown in human and rat studies.</p>
  • <p>The relationship between GTPCH-1 and BH4 is supported in humans (Jayaram et al., 2015), cows (Abdelghany&nbsp;et al., 2017; Whitsett et al., 2007, Wang et al., 2008), mice (Adlam et al., 2012; Chuaiphichai et al., 2014; Crabtree et al., 2009; Tatham et al., 2009; Wang et al., 2008) and rats (Cervantes-P&eacute;rez et al., 2012).</p>
  • <h4>How Does This Key Event Relationship Work</h4>
  • <p>Vasodilation decreases systemic vascular resistance (SVR; also known previously as Total Peripheral Resistance; TPR), the resistance to blood flow offered by the peripheral circulation, and blood pressure through relaxation of vascular smooth muscle cells (VSMCs) (Siddiqui, 2011). When vasodilation is impaired due to decreased NO availability, SVR and blood pressure become elevated.</p>
  • <h4>Key Event Relationship Description</h4>
  • <p>Guanosine triphosphate cyclohydrolase-1 (GTPCH-1) is the rate-limiting enzyme in the de novo biosynthesis of BH4, which is an essential cofactor for eNOS and NO generation (Wang et al., 2008). Oxidative stress can disrupt and decrease GTPCH-1 activity, leading to decreased BH4 levels and subsequent uncoupling of eNOS.</p>
  • <!-- if nothing shows up in any of these fields, then weight of evidence will not be displayed -->
  • <h4>Weight of Evidence</h4>
  • <p>The overall weight of evidence for the KER was rated &quot;moderate&quot; due the fact that acute pharmacological manipulation of the NO pathway resulted in corresponding changes in SVR.&nbsp; However, in the context of the development of hypertension, the chronic effects of impaired vasodilation are much less clear.</p>
  • <!-- if nothing shows up in any of these fields, then evidence supporting this KER will not be displayed -->
  • <h4>Evidence Supporting this KER</h4>
  • <strong>Biological Plausibility</strong>
  • <p>It is well-accepted that vasodilation and SVR are negatively correlated; blood flow is increased when blood vessels dilate due to a decrease in vascular resistance (Siddiqui, 2011). When vasodilation is impaired, SVR increases, in turn increasing blood pressure. Agents that cause hyperpolarization are potent vasodilators and activate&nbsp;potassium channels, while factors causing depolarization increase vascular tone (Nelson, 1990). Vascular tone is governed by the&nbsp;contractile activity of VSMCs in the walls of small arteries and arterioles, and is the major determinant of the resistance to blood flow through the circulation (Jackson, 2000). VSMCs from hypertensive animals have decreased functional voltage-gated potassium channels, which may contribute to depolarization. Two studies demonstrated that blockade of potassium channels completely inhibited NO-dependent vasodilation and increased SVR (Dessy et al., 2004; Berg et al., 2011). Inhibitors of eNOS activity (L-NAME, L-NMMA), which have been shown to decrease acetylcholine-induced vasorelaxation in animal studies (Li et al., 2007; Paulis et al., 2008), also caused an increase in SVR in human studies (Wilkinson et al., 2002; McVeigh et al., 2001; Brett et al., 1998). Overall, these results provide strong biological plausibility for this link.</p>
  • <p>As GTPCH-1 is required for BH4 biosynthesis, there is strong biological plausibility for this relationship.</p>
  • <p>Many studies demonstrated that deletion of GTPCH-1 led to the deficiency of BH4 in endothelial cells. In the hph-1 mouse model, cardiac GTPCH-1 enzymatic activity was reduced by 90% compared to wild-type mice, which led to 60% reduction in BH4 levels (Adlam et al., 2012). In another mouse model of endothelial-targeted deletion of GTPCH-1, BH4 levels in lung, heart and aorta were significantly decreased compared to wild-type mice (Chuaiphichai et al., 2014). GTPCH-1 siRNA significantly reduced GTPCH-1 enzyme activity and BH4 levels in murine sEnd.1 and aortic endothelial cells (Crabtree et al., 2009; Tatham et al., 2009; Wang et al., 2008). The selective GTPCH-1 inhibitor diaminohydroxypyrimidine (DAHP) reduced levels of BH4 in bovine aortic endothelial cells (BAECs) (Wang et al., 2008). Also, transgenic overexpression of GTPCH-1 increased BH4 protein levels in murine hearts and aortas, leading to enhanced eNOS activity (Alp et al., 2003; Carnicer et al., 2012).</p>
  • <strong>Empirical Support for Linkage</strong>
  • <strong>Empirical Evidence</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>No direct evidence was found for this linkage; thus the empirical support is weak. One study indirectly showed a relationship between NO depletion and SVR, where infusion of L-NMMA (1.0 mg/kg/min) caused an increase in SVR by 63% and a 65% reduction in NO in 11 healthy volunteers after three minutes (Stamler et al., 1994). In rat small mesenteric arteries, treatment with L-NAME for five weeks decreased NO-dependent relaxation from 30% to less than 10%, but increased systolic and diastolic blood pressure to 26% and 40%, respectively (Paulis et al, 2008). Meta-analysis of clinical trials showing that infusion of the NO donor sodium nitroprusside led to a dose-dependent reduction in SVR (Eugene et al., 2016).&nbsp; In a study of 400 post-menopausal women with mild-moderate hypertension and impaired FMD, anti-hypertensive therapy for 6 months was shown to improve FMD by &gt;10% in 250 women, whereas in 150 women, FMD did not significantly change (&lt;10% change, Modena et al. 2002).&nbsp; Whilst the study demonstrates a linkage between vasodilation and hypertension in a large proportion of the study population, it raises a&nbsp;question regarding causality i.e. did NO-independent medication lower blood pressure, which in turn improved FMD?&nbsp; Furthermore, over a third of the population showed no change.&nbsp; It is possible that the medications used directly influenced key components of the NO (or other) pathways.&nbsp; Since the study did not elaborate on the medicatons used by the study cohort, this is impossible to determine.</p>
  • <p>Exposure to a wide range of stimuli led to a decrease in both GTPCH-1 expression and activity and a decrease in BH4 levels, indicating strong empirical support between these two key events. An assumption that a decrease in GTPCH-1 expression and activity also results in a decrease in BH4 levels was made for these studies.&nbsp;</p>
  • <p>After myocardial reperfusion, GTPCH-1 activity decreased from 100% to 50% and arterial BH4 levels was reduced by 32% (Jayaram et al., 2015).</p>
  • <p>IL6/TNF/LPS (4 ng/mL, 10 nmol/L, 80 ng/mL) stimulation for 24 hours induced a 3-fold upregulation of GTPCH-1 gene expression and 3- to 4-fold increase in vascular BH4 (Antoniades et al., 2011). For this evidence, an assumption that a change in GTPCH-1 gene expression would affect GTPCH-1 enzyme activity was made.</p>
  • <p>Cigarette smoke extract (CSE, 5%) exposure for 4 hours significantly decreased BH4 levels from 100% to 50% and GTPCH-1 expression from 100% to 51%, while a lower concentration of 2.5% CSE did not cause a significant change in bovine aortic endothelial cells (Abdelghany&nbsp;et al., 2017).</p>
  • <p>In BAECs, stimulation with 25 &mu;M 4-hydroxy-2-nonenal decreased BH4 levels from 10 pmol/mg protein to 6.1 pmol/mg protein and GTPCH-1 activity from 27 pmol/hr/mg protein to 23 pmol/hr/mg protein (Whitsett et al., 2007).</p>
  • <p>In a rat hypertensive model, the decrease in GTPCH-1 protein expression (via densitometry of western blot band intensity, 0.089 control to 0.0087) and BH4 (33.71&plusmn;3.318 pmol control to 17.144&plusmn;2.251 pmol) induced by aortic coarctation was attenuated by clofibrate treatment (GTPCH-1: 0.088&plusmn;0.022 band intensity, BH4: 32.534&plusmn;5.809 pmol) (Cervantes-P&eacute;rez et al., 2012).</p>
  • <strong>Uncertainties and Inconsistencies</strong>
  • <p>There are no uncertainties or inconsistencies.</p>
  • <strong>Uncertainties or Inconsistencies</strong>
  • <p>As mentioned above, acute pharmacological manipulation of the NO pathway results in expected changes in SVR.&nbsp; However,&nbsp;the link between chronically impaired vasodilation&nbsp;and SVR (the context of this AOP) is much less clear due to gaps in the literature.&nbsp; Epidemiological studies tend to investigate linkages between impaired vasodilation and cardiovascular events, as opposed to SVR and/or hypertension - making assessment of this KER difficult.</p>
  • <p>Furthermore, the complexity in the mechanisms influencing vascular re-modelling over time has hampered&nbsp;understanding of the phenomenon to date.&nbsp; The study by Modena et al. 2002 highlights that members of the general population respond differently to hypertensive therapy in the context of FMD improvement.</p>
  • <strong>Quantitative Understanding of the Linkage</strong>
  • <p><em>Is it known how much change in the first event is needed to impact the second? Are there known modulators of the response-response relationships? Are there models or extrapolation approaches that help describe those relationships? </em></p>
  • <p>There is very limited&nbsp;quantitative understanding of this linkage, however the evidence is directed towards &quot;cardiovascular events&quot;, as opposed to specifically SVR and/or hypertension.&nbsp; A meta analysis by Ras et al. (2013) stated that &quot;a 1% increase in FMD corresponds to a 14% decrease in future CVD events.&quot;.&nbsp;&nbsp;A total of 23 studies including 14,753 subjects were eligible for inclusion in the meta-analysis. For studies reporting continuous risk estimates, the pooled overall CVD risk was 0.92 (95%CI: 0.88; 0.95) per 1% higher FMD. The observed association seemed stronger (P-value&lt;0.01) in diseased populations than in asymptomatic populations (0.87 (95%CI: 0.83; 0.92) and 0.96 (95%CI: 0.92; 1.00) per 1% higher FMD, respectively). For studies reporting categorical risk estimates, the pooled overall CVD risk for high vs. low FMD was similar in both types of populations, on average 0.49 (95%CI: 0.39; 0.62).</p>
  • <p>Similarly, Yeboah et al. (2007) concluded that FMD was a predictor of future cardiovascular (CVD) events and that systolic blood&nbsp;pressure per unit S.D.&nbsp;was a signficant (p=0.02) risk factor.&nbsp; However, they reported that FMD added little to current risk prediction scores for future CVD events.</p>
  • <!--<!% unless aop_rel.relationship.relationship_taxons.blank? %>-->
  • <!--<!%= render 'snapshot_taxons', taxons: aop_rel.relationship.relationship_taxons %>-->
  • <!--<!% unless aop_rel.relationship.taxon_evidence.blank? %>-->
  • <!--<h3>Evidence Supporting Taxonomic Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
  • <!--<!% end %>-->
  • <!--<!%= render 'snapshot_taxons', taxons: aop_rel.relationship.relationship_taxons %>-->
  • <!--<!% unless aop_rel.relationship.taxon_evidence.blank? %>-->
  • <!--<h3>Domain of Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
  • <!--<!% end %>-->
  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>Berg, T., and Jensen, J. (2011). Simultaneous parasympathetic and sympathetic activation reveals altered autonomic control of heart rate, vascular tension, and epinephrine release in anesthetized hypertensive rats. Front. Neurol. 2, 71.</p>
  • <p>Brett, S.E., Cockcroft, J.R., Mant, T.G., Ritter, J.M., and Chowienczyk, P.J. (1998). Haemodynamic effects of inhibition of nitric oxide synthase and of L-arginine at rest and during exercise. J. Hypertens. 16, 429&ndash;435.</p>
  • <p>Dessy, C., Moniotte, S., Ghisdal, P., Havaux, X., Noirhomme, P., and Balligand, J.L. (2004). Endothelial beta3-adrenoceptors mediate vasorelaxation of human coronary microarteries through nitric oxide and endothelium-dependent hyperpolarization. Circulation 110, 948&ndash;954.</p>
  • <p>AbdelGhany, T., Ismail, R., Elmahdy, M., Mansoor F, Zweier J, Lowe, F., and Zweier, JL. (2017). Cigarette Smoke Constituents Cause Endothelial Nitric Oxide Synthase Dysfunction and Uncoupling due to Depletion of Tetrahydrobiopterin with Degradation of GTP Cyclohydrolase.&nbsp;&nbsp;Nitric Oxide&nbsp;(Under review).</p>
  • <p>Eugene, A.R. (2016). The influences of nitric oxide, epinephrine, and dopamine on vascular tone: dose-response modeling and simulations. Hosp. Chron. Nosokomeiaka Chron. 11, 1&ndash;8.</p>
  • <p>Adlam, D., Herring, N., Douglas, G., De Bono, J.P., Li, D., Danson, E.J., Tatham, A., Lu, C.-J., Jennings, K.A., Cragg, S.J., et al. (2012). Regulation of &beta;-adrenergic control of heart rate by GTP-cyclohydrolase 1 (GCH1) and tetrahydrobiopterin. Cardiovasc. Res. 93, 694&ndash;701.</p>
  • <p>Jackson, W.F. (2000). Ion channels and vascular tone. Hypertension 35, 173&ndash;178.</p>
  • <p>Alp, N.J., Mussa, S., Khoo, J., Cai, S., Guzik, T., Jefferson, A., Goh, N., Rockett, K.A., and Channon, K.M. (2003). Tetrahydrobiopterin-dependent preservation of nitric oxide-mediated endothelial function in diabetes by targeted transgenic GTP-cyclohydrolase I overexpression. J. Clin. Invest. 112, 725&ndash;735.</p>
  • <p>Li, J., Zhou, Z., Jiang, D.-J., Li, D., Tan, B., Liu, H., and Li, Y.-J. (2007). Reduction of NO- and EDHF-mediated vasodilatation in hypertension: role of asymmetric dimethylarginine. Clin. Exp. Hypertens. N. Y. N 1993 29, 489&ndash;501.</p>
  • <p>Antoniades, C., Cunnington, C., Antonopoulos, A., Neville, M., Margaritis, M., Demosthenous, M., Bendall, J., Hale, A., Cerrato, R., Tousoulis, D., et al. (2011). Induction of vascular GTP-cyclohydrolase I and endogenous tetrahydrobiopterin synthesis protect against inflammation-induced endothelial dysfunction in human atherosclerosis. Circulation 124, 1860&ndash;1870.</p>
  • <p>McVeigh, G.E., Allen, P.B., Morgan, D.R., Hanratty, C.G., and Silke, B. (2001). Nitric oxide modulation of blood vessel tone identified by arterial waveform analysis. Clin. Sci. Lond. Engl. 1979 100, 387&ndash;393.</p>
  • <p>Carnicer, R., Hale, A.B., Suffredini, S., Liu, X., Reilly, S., Zhang, M.H., Surdo, N.C., Bendall, J.K., Crabtree, M.J., Lim, G.B.S., et al. (2012). Cardiomyocyte GTP cyclohydrolase 1 and tetrahydrobiopterin increase NOS1 activity and accelerate myocardial relaxation. Circ. Res. 111, 718&ndash;727.</p>
  • <p>Modena MG,&nbsp;Bonetti L,&nbsp;Coppi F,&nbsp;Bursi F,&nbsp;Rossi R Prognostic&nbsp;role&nbsp;of&nbsp;reversible&nbsp;endothelial&nbsp;dysfunction&nbsp;in&nbsp;hypertensive&nbsp;postmenopausal&nbsp;women.&nbsp; J Am Coll Cardiol.&nbsp;2002, 40(3):505-10.</p>
  • <p>Cervantes-P&eacute;rez, L.G., Ibarra-Lara, M. de la L., Escalante, B., Del Valle-Mondrag&oacute;n, L., Vargas-Robles, H., P&eacute;rez-Severiano, F., Pastel&iacute;n, G., and S&aacute;nchez-Mendoza, M.A. (2012). Endothelial nitric oxide synthase impairment is restored by clofibrate treatment in an animal model of hypertension. Eur. J. Pharmacol. 685, 108&ndash;115.</p>
  • <p>Nelson, M.T., Patlak, J.B., Worley, J.F., and Standen, N.B. (1990). Calcium channels, potassium channels, and voltage dependence of arterial smooth muscle tone. Am. J. Physiol. 259, C3&ndash;C18.</p>
  • <p>Chen, C.-A., Lin, C.-H., Druhan, L.J., Wang, T.-Y., Chen, Y.-R., and Zweier, J.L. (2011). Superoxide induces endothelial nitric-oxide synthase protein thiyl radical formation, a novel mechanism regulating eNOS function and coupling. J. Biol. Chem. 286, 29098&ndash;29107.</p>
  • <p>Paulis, L., Zicha, J., Kunes, J., Hojna, S., Behuliak, M., Celec, P., Kojsova, S., Pechanova, O., and Simko, F. (2008). Regression of L-NAME-induced hypertension: the role of nitric oxide and endothelium-derived constricting factor. Hypertens. Res. Off. J. Jpn. Soc. Hypertens. 31, 793&ndash;803.</p>
  • <p>Chuaiphichai, S., McNeill, E., Douglas, G., Crabtree, M.J., Bendall, J.K., Hale, A.B., Alp, N.J., and Channon, K.M. (2014). Cell-autonomous role of endothelial GTP cyclohydrolase 1 and tetrahydrobiopterin in blood pressure regulation. Hypertension 64, 530&ndash;540.</p>
  • <p>Ras&nbsp;RT,&nbsp;Streppel MT,&nbsp;Draijer R,&nbsp;Zock PL.&nbsp; Flow-mediated dilation and cardiovascular risk prediction: a systematic review with meta-analysis.&nbsp; Int J Cardiol.&nbsp;2013, 168(1):344-51.</p>
  • <p>Crabtree, M.J., Tatham, A.L., Al-Wakeel, Y., Warrick, N., Hale, A.B., Cai, S., Channon, K.M., and Alp, N.J. (2009). Quantitative regulation of intracellular endothelial nitric-oxide synthase (eNOS) coupling by both tetrahydrobiopterin-eNOS stoichiometry and biopterin redox status: insights from cells with tet-regulated GTP cyclohydrolase I expression. J. Biol. Chem. 284, 1136&ndash;1144.</p>
  • <p>Siddiqui, A. (2011). Effects of Vasodilation and Arterial Resistance on Cardiac Output. J. Clin. Exp. Cardiol. 02.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Stamler, J.S., Loh, E., Roddy, M.A., Currie, K.E., and Creager, M.A. (1994). Nitric oxide regulates basal systemic and pulmonary vascular resistance in healthy humans. Circulation 89, 2035&ndash;2040.</p>
  • <p>Tatham, A.L., Crabtree, M.J., Warrick, N., Cai, S., Alp, N.J., and Channon, K.M. (2009). GTP cyclohydrolase I expression, protein, and activity determine intracellular tetrahydrobiopterin levels, independent of GTP cyclohydrolase feedback regulatory protein expression. J. Biol. Chem. 284, 13660&ndash;13668.</p>
  • <p>Wilkinson, I.B., MacCallum, H., Cockcroft, J.R., and Webb, D.J. (2002). Inhibition of basal nitric oxide synthesis increases aortic augmentation index and pulse wave velocity in vivo. Br. J. Clin. Pharmacol. 53, 189&ndash;192.</p>
  • <p>Wang, S., Xu, J., Song, P., Wu, Y., Zhang, J., Chul Choi, H., and Zou, M.-H. (2008). Acute inhibition of guanosine triphosphate cyclohydrolase 1 uncouples endothelial nitric oxide synthase and elevates blood pressure. Hypertension 52, 484&ndash;490.</p>
  • <p>Yeboah J, Crouse JR, Hsu FC, Burke GL, Herrington DM.&nbsp; Brachial flow-mediated dilation predicts incident cardiovascular events<br />
  • &nbsp;in older adults: the Cardiovascular Health Study. Circulation. 2007;115:2390&ndash;7.</p>
  • <p>Whitsett, J., Picklo, M.J., and Vasquez-Vivar, J. (2007). 4-Hydroxy-2-nonenal increases superoxide anion radical in endothelial cells via stimulated GTP cyclohydrolase proteasomal degradation. Arterioscler. Thromb. Vasc. Biol. 27, 2340&ndash;2347.</p>
  • </div>
  • <br>
  • <div>
  • <div>
  • <h4><a href="/relationships/983">Increase, Vascular Resistance leads to Hypertension</a></h4>
  • <h4><a href="/relationships/953">Relationship: 953: Decrease, Tetrahydrobiopterin leads to Uncoupling, eNOS</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Directness</th>
  • <th>Adjacency</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>directly leads to</th>
  • <th>Moderate </th>
  • <th>Weak</th>
  • <th>adjacent</th>
  • <th>High </th>
  • <th>High</th>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Weak</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Adults</td>
  • <td>Strong</td>
  • <td>All life stages</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>Studies supporting this key event relationship were performed in humans and rats.</p>
  • <p>The relationship between BH4 depletion and eNOS uncoupling was observed in humans (Jayaram et al., 2015), cows (Wang et al., 2008; Whitsett et al., 2007), mice (Chuaiphichai et al., 2014; Crabtree et al., 2009) and rats (Cervantes-P&eacute;rez et al., 2012; Dumitrescu et al., 2007).</p>
  • <h4>How Does This Key Event Relationship Work</h4>
  • <p>Hypertension is characterized partly by elevated systemic vascular resistance which is caused by alterations to vascular tone (towards vasoconstriction) over time&nbsp;(Lee and Griendling, 2008).&nbsp; As blood vessels constrict, the available volume in the vessel lumen for blood flow&nbsp;is restricted, resulting in elevated blood pressure.</p>
  • <p>Note : The role of the heart in the maintenace (and change) of blood pressure over time is not part of this AOP, however it is of critical importance for the development of hypertension.</p>
  • <h4>Key Event Relationship Description</h4>
  • <p>Oxidative stress leads to the excessive oxidation and depletion of BH4, resulting in eNOS uncoupling where eNOS produces superoxide rather than nitric oxide (F&ouml;rstermann and M&uuml;nzel, 2006).</p>
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  • <h4>Weight of Evidence</h4>
  • <!-- if nothing shows up in any of these fields, then evidence supporting this KER will not be displayed -->
  • <h4>Evidence Supporting this KER</h4>
  • <strong>Biological Plausibility</strong>
  • <p>It is well-established that increased systemic vascular resistance (SVR), increased vascular stiffness and increased vascular reactivity contribute to the pathophysiology of hypertension (Fo&euml;x and Sear, 2004; Mayet and Hughes, 2003; Brandes et al., 2014); thus biological plausibility is strong for increased SVR leading to hypertension. This is observed in patients with hypertension (Chan et al., 2016).</p>
  • <p>BH4 is an essential cofactor for eNOS and is required for its enzymatic activity to produce NO. The depletion of BH4 leading to eNOS uncoupling is well-studied, thus there is strong biological plausibility for this link.</p>
  • <p>Two mouse studies showed limited BH4 availability induced eNOS uncoupling by reducing eNOS activity, leading to decreased nitric oxide and increased superoxide. In the mouse endothelial cell line sEnd.1, BH4 deficiency induced eNOS uncoupling as determined by superoxide production and impaired vasodilation (Crabtree et al., 2009). In primary aortic endothelial cells of GTPCH1-knockout mice, BH4 depletion significantly reduced eNOS activity, increased basal superoxide production and decreased NO bioactivity (Chuaiphichai et al., 2014). In rat hearts, BH4 content and eNOS activity were decreased in a time-dependent manner following myocardial ischemia with a marked decline after thirty minutes, while superoxide generation increased (Dumitrescu et al., 2007).</p>
  • <p>BAECs undergoing hypoxia and reoxygenation had decreased BH4 and decreased NO production, which was partially restored by treatment with the xanthine oxidase inhibitor oxypurinol, N-acetyl-l-cysteine (NAC) and NAC+BH4 (De Pascali et al., 2014). Inhibition of BH4 due to treatment with 4-HNE decreased eNOS activity and NO production in BAECs (Whitsett et al., 2007).</p>
  • <p>Many studies demonstrated that BH4 treatment reduced eNOS-mediated superoxide generation and increased NO formation in bovine, mouse, and rat endothelium (Chen et al., 2011; De Pascali et al., 2014; Landmesser et al., 2003; Ozaki et al., 2002; Shinozaki et al., 2000). Clinical studies reported improvement endothelial function in cardiovascular disease after treatment with BH4 (Wang et al., 2014). Oral treatment with BH4 in hypertensive patients significantly decreased blood pressure.</p>
  • <strong>Empirical Support for Linkage</strong>
  • <strong>Empirical Evidence</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>Empirical support for SVR leading to hypertension is moderate based on several human studies showing a dose-dependent change in SVR and hypertension following treatment with eNOS inhibitors L-NMMA and L-NAME.&nbsp; Whilst the acute changes in SVR on blood pressure in the context of endothelial NO production is well characterised, the linkage between chronically elevated SVR and hypertension is more complicated, due to the roles of the heart, nervous system and kidneys.&nbsp; Given that this AOP is focused on endothelial NO production, other AOPs are required to capture the roles of these other important biological processes on chronic SVR changes and hypertension, hence the rating of &quot;moderate&quot;.</p>
  • <p>Multiple studies demonstrated a strong dependency between BH4 and eNOS uncoupling; decreased BH4 along with decreased eNOS activity, decreased NO production or increased superoxide generation were observed following various perturbations.</p>
  • <p>Intravenous infusion of L-NMMA (0.1, 1.0, 3.0 mg/kg/min) for 15 minutes led to a dose-dependent increase in SVR and mean arterial pressure compared to saline infusion in eight healthy men (Wilkinson et al., 2002). SVR increased from 31&plusmn;2 (arbitrary units) at baseline to 57&plusmn;4 at the highest dose, while mean arterial pressure increased from 86&plusmn;2 mmHg baseline to 91&plusmn;2 mmHg at the highest concentration. Infusion of L-NMMA (1.0 mg/kg/min) caused an increase in systolic blood pressure by 15% and SVR by 63% in 11 healthy volunteers after three minutes (Stamler et al., 1994). The study also demonstrated a dose-dependent increase in both events with five different doses of L-NMMA (0.01, 0.03, 0.1, 0.3 and 1 mg/kg/min). In another study,&nbsp;fifteen healthy men were&nbsp;intravenously infused with L-NAME &nbsp;at lower doses (0.25-0.75 mg/kg) for eight minutes, resulting in increased arterial pressure and SVR (McVeigh et al., 2001). Infusion with L-arginine, an eNOS substrate, restored function, specifically SVR and small artery compliance to baseline. L-NMMA (3 mg/kg) for five minutes increased mean arterial pressure by 10% and increased SVR by 46% in eight healthy subjects (Haynes et al., 1993). One experiment in rats showed that L-NAME increased SVR, aortic stiffness, and blood pressure (Nakmareong et al., 2012). Overall, these studies demonstrated that SVR and hypertension are modulated together by eNOS inhibitors.</p>
  • <p>Prolonged myocardial ischemia (&gt;30 min) in isolated rat hearts caused extensive BH4 depletion (95% depletion), which was paralleled by decreased eNOS activity (58% reduction) and increased superoxide generation from &lt;0.01 relative fluorescence unit/mg protein to 0.3 measured using a fluorescence detector-based assay (Dumitrescu et al., 2007). Similarly, cardiac reperfusion patients exhibited a reduction in BH4 levels by 32%, decreased eNOS activity by 40% and increased superoxide production from 37.83 to 65.02 light unit/s/mg as detected using lucigenin-enhanced chemiluminescence (Jayaram et al., 2015).</p>
  • <p>In BAECs treated with 10 mmol/L DAHP or 25 &mu;M 4-HNE for 4 hours, BH4 levels decreased (control: 10 pmol/mg, DAHP: 4.8 pmol/mg, 4-HNE: 6.1 pmol/mg). At 24 hours, a reduction in NO (control: 1678 pmol/mg, DAHP: 1274 pmol/mg, 4-HNE: 1106 pmol/mg) and increased superoxide formation (control: 59 pmol/mg, DAHP: 97 pmol/mg, 4-HNE: 122 pmol/mg) were observed (Whitsett et al., 2007). Another study in BAECs using DAHP showed similar results for BH4 (control: 20.5 pmol/mg, DAHP: 11.8 pmol/mg), superoxide (control: 100%, DAHP: 257%), and NO (control: 96%, DAHP: 60%) (Wang et al., 2008).</p>
  • <p>In BAECs undergoing hypoxia and reoxygenation (H/R), treatment with oxypurinol increased BH4 levels from 6.1 &plusmn; 0.9 pmol/mg protein (after H/R) to 11.9 &plusmn; 0.8 pmol/mg protein and increased NO from 34.2 &plusmn; 1.7% to 63.7 &plusmn; 3.0%, demonstrating that these key events are modulated together (De Pascali et al., 2014).</p>
  • <p>In a rat model of hypertension, BH4 depletion and eNOS uncoupling induced by aortic coarctation (AoCo) was&nbsp;reversed by treatment with clofibrate; eNOS activity increased from 6.927&plusmn;3.475 ng L-citrulline/mg protein/30 min to 23.2 &plusmn; 9.034 ng L-citrulline/mg protein/30 min, BH4 levels increased from 17.144&plusmn;2.251 pmol to 32.534&plusmn;5.809 pmol, and superoxide decreased from relative fluorescence intensity of 32.22&plusmn;2.903 to 24.59&plusmn;1.124 (Cervantes-P&eacute;rez et al., 2012). Note that normal eNOS activity converts L-arginine to L-citrulline and produces NO; thus L-citrulline is an indirect measure of eNOS activity.</p>
  • <strong>Uncertainties and Inconsistencies</strong>
  • <p>The uncoupling of eNOS may also occur through other mechanisms such as S-glutathionylation of eNOS and depletion of L-arginine (Zweier et al., 2011).</p>
  • <strong>Uncertainties or Inconsistencies</strong>
  • <p>One study showed that infusion of L-NMMA (6 mg/kg) resulted in increased SVR and only a modest increase in blood pressure.&nbsp; Changes in diastolic blood pressure were observed to be&nbsp;more pronounced in healthy men, than systolic blood pressure (Brett et al., 1998), and infusion of L-arginine (an eNOS substrate) had no significant effect.</p>
  • <p>As mentioned above, other AOPs are necessary to capture understanding and assess the evidence surrounding the roles of the heart, kidney and nervous system in order to get the full picture of the linkage between chronic changes in SVR and hypertension.</p>
  • <strong>Quantitative Understanding of the Linkage</strong>
  • <p><em>Is it known how much change in the first event is needed to impact the second? Are there known modulators of the response-response relationships? Are there models or extrapolation approaches that help describe those relationships? </em></p>
  • <p>Mean arterial pressure is calculated from cardiac output and SVR. Therefore, theoretically, any change in SVR will impact the mean arterial pressure. The studies mentioned above showed that a small change for SVR such as from 31&plusmn;2 (arbitrary units) at baseline to 35 &plusmn; 2 was able to change the arterial pressure from 86&plusmn;2 mmHg to 91&plusmn;2 mmHg (Wilkinson et al., 2002). This trend was also observed by Stamler et al. (1994) and McVeigh et al. (2001).</p>
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  • <!--<h3>Domain of Applicability</h3>-->
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  • <h4>References</h4>
  • <p>Brandes, R.P. (2014). Endothelial dysfunction and hypertension. Hypertension 64, 924&ndash;928.</p>
  • <p>Cervantes-P&eacute;rez, L.G., Ibarra-Lara, M. de la L., Escalante, B., Del Valle-Mondrag&oacute;n, L., Vargas-Robles, H., P&eacute;rez-Severiano, F., Pastel&iacute;n, G., and S&aacute;nchez-Mendoza, M.A. (2012). Endothelial nitric oxide synthase impairment is restored by clofibrate treatment in an animal model of hypertension. Eur. J. Pharmacol. 685, 108&ndash;115.</p>
  • <p>Brett, S.E., Cockcroft, J.R., Mant, T.G., Ritter, J.M., and Chowienczyk, P.J. (1998). Haemodynamic effects of inhibition of nitric oxide synthase and of L-arginine at rest and during exercise. J. Hypertens. 16, 429&ndash;435.</p>
  • <p>Chen, C.-A., Lin, C.-H., Druhan, L.J., Wang, T.-Y., Chen, Y.-R., and Zweier, J.L. (2011). Superoxide induces endothelial nitric-oxide synthase protein thiyl radical formation, a novel mechanism regulating eNOS function and coupling. J. Biol. Chem. 286, 29098&ndash;29107.</p>
  • <p>Chan, S.S., Tse, M.M., Chan, C.P., Tai, M.C., Graham, C.A., and Rainer, T.H. (2016). Haemodynamic changes in emergency department patients with poorly controlled hypertension. Hong Kong Med. J. Xianggang Yi Xue Za Zhi Hong Kong Acad. Med. 22, 116&ndash;123.</p>
  • <p>Chuaiphichai, S., McNeill, E., Douglas, G., Crabtree, M.J., Bendall, J.K., Hale, A.B., Alp, N.J., and Channon, K.M. (2014). Cell-autonomous role of endothelial GTP cyclohydrolase 1 and tetrahydrobiopterin in blood pressure regulation. Hypertension 64, 530&ndash;540.</p>
  • <p>Fo&euml;x, P., and Sear, J.W. (2004). Hypertension: pathophysiology and treatment. Contin. Educ. Anaesth. Crit. Care Pain 4, 71&ndash;75.</p>
  • <p>Crabtree, M.J., Tatham, A.L., Al-Wakeel, Y., Warrick, N., Hale, A.B., Cai, S., Channon, K.M., and Alp, N.J. (2009). Quantitative regulation of intracellular endothelial nitric-oxide synthase (eNOS) coupling by both tetrahydrobiopterin-eNOS stoichiometry and biopterin redox status: insights from cells with tet-regulated GTP cyclohydrolase I expression. J. Biol. Chem. 284, 1136&ndash;1144.</p>
  • <p>Haynes, W.G., Noon, J.P., Walker, B.R., and Webb, D.J. (1993). Inhibition of nitric oxide synthesis increases blood pressure in healthy humans. J. Hypertens. 11, 1375&ndash;1380.</p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p>Lee, M.Y., and Griendling, K.K. (2008). Redox signaling, vascular function, and hypertension. Antioxid. Redox Signal. 10, 1045&ndash;1059.</p>
  • <p>Dumitrescu, C., Biondi, R., Xia, Y., Cardounel, A.J., Druhan, L.J., Ambrosio, G., and Zweier, J.L. (2007). Myocardial ischemia results in tetrahydrobiopterin (BH4) oxidation with impaired endothelial function ameliorated by BH4. Proc. Natl. Acad. Sci. U. S. A. 104, 15081&ndash;15086.</p>
  • <p>Mayet, J., and Hughes, A. (2003). Cardiac and vascular pathophysiology in hypertension. Heart Br. Card. Soc. 89, 1104&ndash;1109.</p>
  • <p>F&ouml;rstermann, U., and M&uuml;nzel, T. (2006). Endothelial nitric oxide synthase in vascular disease: from marvel to menace. Circulation 113, 1708&ndash;1714.</p>
  • <p>McVeigh, G.E., Allen, P.B., Morgan, D.R., Hanratty, C.G., and Silke, B. (2001). Nitric oxide modulation of blood vessel tone identified by arterial waveform analysis. Clin. Sci. Lond. Engl. 1979 100, 387&ndash;393.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Nakmareong, S., Kukongviriyapan, U., Pakdeechote, P., Kukongviriyapan, V., Kongyingyoes, B., Donpunha, W., Prachaney, P., and Phisalaphong, C. (2012). Tetrahydrocurcumin alleviates hypertension, aortic stiffening and oxidative stress in rats with nitric oxide deficiency. Hypertens. Res. Off. J. Jpn. Soc. Hypertens. 35, 418&ndash;425.</p>
  • <p>Landmesser, U., Dikalov, S., Price, S.R., McCann, L., Fukai, T., Holland, S.M., Mitch, W.E., and Harrison, D.G. (2003). Oxidation of tetrahydrobiopterin leads to uncoupling of endothelial cell nitric oxide synthase in hypertension. J. Clin. Invest. 111, 1201&ndash;1209.</p>
  • <p>Silva, B.R., Pernomian, L., and Bendhack, L.M. (2012). Contribution of oxidative stress to endothelial dysfunction in hypertension. Front. Physiol. 3, 441.</p>
  • <p>Ozaki, M., Kawashima, S., Yamashita, T., Hirase, T., Namiki, M., Inoue, N., Hirata, K., Yasui, H., Sakurai, H., Yoshida, Y., et al. (2002). Overexpression of endothelial nitric oxide synthase accelerates atherosclerotic lesion formation in apoE-deficient mice. J. Clin. Invest. 110, 331&ndash;340.</p>
  • <p>Stamler, J.S., Loh, E., Roddy, M.A., Currie, K.E., and Creager, M.A. (1994). Nitric oxide regulates basal systemic and pulmonary vascular resistance in healthy humans. Circulation 89, 2035&ndash;2040.</p>
  • <p>Shinozaki, K., Nishio, Y., Okamura, T., Yoshida, Y., Maegawa, H., Kojima, H., Masada, M., Toda, N., Kikkawa, R., and Kashiwagi, A. (2000). Oral administration of tetrahydrobiopterin prevents endothelial dysfunction and vascular oxidative stress in the aortas of insulin-resistant rats. Circ. Res. 87, 566&ndash;573.</p>
  • <p>Wilkinson, I.B., MacCallum, H., Cockcroft, J.R., and Webb, D.J. (2002). Inhibition of basal nitric oxide synthesis increases aortic augmentation index and pulse wave velocity in vivo. Br. J. Clin. Pharmacol. 53, 189&ndash;192.</p>
  • <p>Wang, Q., Yang, M., Xu, H., and Yu, J. (2014). Tetrahydrobiopterin improves endothelial function in cardiovascular disease: a systematic review. Evid.-Based Complement. Altern. Med. ECAM 2014, 850312.</p>
  • <p>Wang, S., Xu, J., Song, P., Wu, Y., Zhang, J., Chul Choi, H., and Zou, M.-H. (2008). Acute inhibition of guanosine triphosphate cyclohydrolase 1 uncouples endothelial nitric oxide synthase and elevates blood pressure. Hypertension 52, 484&ndash;490.</p>
  • <p>Whitsett, J., Picklo, M.J., and Vasquez-Vivar, J. (2007). 4-Hydroxy-2-nonenal increases superoxide anion radical in endothelial cells via stimulated GTP cyclohydrolase proteasomal degradation. Arterioscler. Thromb. Vasc. Biol. 27, 2340&ndash;2347.</p>
  • </div>
  • <br>
  • <div>
  • <div>
  • <h4><a href="/relationships/988">Decrease, AKT/eNOS activity leads to Depletion, Nitric Oxide</a></h4>
  • <h4><a href="/relationships/956">Relationship: 956: Uncoupling, eNOS leads to Depletion, Nitric Oxide</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Directness</th>
  • <th>Adjacency</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>directly leads to</th>
  • <th>Strong </th>
  • <th>Strong</th>
  • <th>adjacent</th>
  • <th>High </th>
  • <th>High</th>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Strong</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>The relationship between decreased AKT/eNOS activity and NO depletion is supported by studies performed in humans, cows, and rats.</p>
  • <p>The relationship between NO depletion and eNOS uncoupling was observed in humans, cows, and rats as demonstrated by the above studies.</p>
  • <h4>How Does This Key Event Relationship Work</h4>
  • <p>AKT can phosphorylate eNOS which leads to increased eNOS enzymatic activity and subsequent NO production (Dimmeler et al., 1999; Fulton et al., 1999). Inhibition of AKT&nbsp;attenuates eNOS phosphorylation and its activity, resulting in decreased NO bioavailability and endothelial dysfunction.</p>
  • <h4>Key Event Relationship Description</h4>
  • <p>The uncoupling of eNOS occurs through a number of mechanisms like BH4 depletion or S-glutathionylation of eNOS and leads to a reduction in NO bioavailability and an elevation in superoxide production, contributing to endothelial dysfunction (Zweier et al., 2011).</p>
  • <!-- if nothing shows up in any of these fields, then weight of evidence will not be displayed -->
  • <h4>Weight of Evidence</h4>
  • <!-- if nothing shows up in any of these fields, then evidence supporting this KER will not be displayed -->
  • <h4>Evidence Supporting this KER</h4>
  • <strong>Biological Plausibility</strong>
  • <p>Two studies demonstrated that AKT&nbsp;can directly phosphorylate eNOS at Ser1177/Ser1179, leading to increased eNOS enzymatic activity and subsequent NO production in COS cells (Dimmeler et al., 1999; Fulton et al., 1999). Expressing active AKT&nbsp;in bovine microvascular endothelial cells also induced NO release (Fulton et al., 1999). Inhibition of Akt or a mutation of eNOS at AKT-sensitive sites attenuated eNOS phosphorylation and its activity, resulting in decreased NO bioavailability and endothelial dysfunction (Dimmeler et al., 1999; Fulton et al., 1999; Uruno et al., 2005). Treatment of BAECs with SIN-1 resulted in a reduction in both eNOS activity and NO production (Das et al., 2014). Cigarette smoke extract (CSE) treatment inhibited AKT&nbsp;and eNOS and NO release in VEGF-stimulated HUVECs (Michaud et al., 2006), and methylglyoxal and high glucose reduced eNOS bradykinin-stimulated eNOS activity and NO production in HUVECs (Dhar et al., 2010). Overall, the biological plausibility for decreased AKT/eNOS activity leading to NO depletion is strong.</p>
  • <p>It is well-established that uncoupling of eNOS causes eNOS to switch from producing NO to generating superoxides (F&ouml;rstermann and M&uuml;nzel, 2006). Many studies that reported <a href="/wiki/index.php/Relationship:953" title="Relationship:953">BH4 depletion leading to eNOS uncoupling</a> or <a href="/wiki/index.php/Relationship:955" title="Relationship:955">S-glutathionylation leading to eNOS uncoupling</a> measured levels of NO and superoxide which are indicative of eNOS uncoupling.</p>
  • <strong>Empirical Support for Linkage</strong>
  • <strong>Empirical Evidence</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>Empirical support for this linkage is strong since various stressors (ischemia, peroxynitrite, SIN-1, insulin+orotic acidura, etc.) can cause a decrease in AKT&nbsp;or eNOS activity which then leads to increased eNOS uncoupling and decreased NO levels.</p>
  • <p>Treatment of HUVECs with 30 &mu;M methylglyoxal (MG) and 25 mM high glucose (HG) concentrations for 24 hours caused a decrease in eNOS activity (control: 100%, MG: 55%, HG: 66%), while a shorter treatment for 3 hours caused a decrease in NO production (control: 20 &mu;M, MG: 12 &mu;M, HG: 16 &mu;M) (Dhar et al., 2010).</p>
  • <p>Multiple experiments demonstrated that eNOS uncoupling results in increased superoxide formation and decreased NO production, which provide strong empirical support for this KER.</p>
  • <p>Treatment of BAECs with SIN-1 for 2 hours decreased eNOS activity from 100% to 70% and NO production from 100% to 53% (Das et al., 2014).</p>
  • <p>Prolonged myocardial ischemia (&gt;30 min) in isolated rat hearts decreased eNOS activity (58% reduction) and increased superoxide generation from &lt;0.01 relative fluorescence unit/mg to 0.3 as measured using a fluorescence detector-based assay (Dumitrescu et al., 2007). Similarly, cardiac reperfusion patients had decreased eNOS activity (40% reduction) and increased superoxide production from 37.83 to 65.02 light unit/s/mg as detected using lucigenin-enhanced chemiluminescence (Jayaram et al., 2015).</p>
  • <p>In HUVECs, exposure to 5 &mu;mol/L peroxynitrite for 5 minutes reduced AKT&nbsp;phosphorylation by 30% (Song et al., 2007) while exposure to 50 &mu;mol/L peroxynitrite for 3 hours reduced NO production from 90% to 6.5% (Zou et al., 2002), demonstrating that AKT&nbsp;phosphorylation occurs before NO production at a lower dose.</p>
  • <p>In BAECs, treatment with 10 mmol/L DAHP or 25 &mu;M 4-HNE for 24 hours led to a reduction in NO (control: 1678 pmol/mg, DAHP: 1274 pmol/mg, 4-HNE: 1106 pmol/mg) and increased superoxide formation (control: 59 pmol/mg, DAHP: 97 pmol/mg, 4-HNE: 122 pmol/mg) (Whitsett et al., 2007). Similar results were observed in another study using DAHP-treated BAECs; superoxide was increased (control: 100%, DAHP: 257%) and NO was decreased (control: 96%, DAHP: 60%) (Wang et al., 2008).</p>
  • <p>Prolonged myocardial ischemia (&gt;30 min) in isolated rat hearts caused decreased eNOS activity (58% reduction) and increased superoxide generation (from relative fluorescence unit of &lt;0.01 to 0.3), suggesting a depletion in NO production (Dumitrescu et al., 2007).</p>
  • <p>In BAECs undergoing hypoxia and reoxygenation (H/R), NO decreased from 34.2&plusmn;1.7% to 63.7&plusmn;3.0% and superoxide increased from relative intensity of 3.5 to 49.2, demonstrating that these key events are modulated together (De Pascali et al., 2014).</p>
  • <p>Treatment of HUVECs with 50 &mu;M insulin for 30 minutes increased phosphorylation of AKT&nbsp;(control: 1, insulin: 5.7) and eNOS (control: 1, insulin: 1.9) as well as NO production (control: 100%, insulin: 157%), but these effects were reversed by additional treatment with 100 &mu;M orotic aciduria (OA) for one hour (Akt: 1.8; eNOS: 0.28; NO: 106%) (Choi et al., 2015). Similar results were observed with addition of uric acid to insulin-treated HUVECs (Choi et al., 2014).</p>
  • <p>BCNU (25 &mu;M, 80 &mu;M) resulted in increased superoxide (25 &mu;M: 432 fluorescence intensity, 80 &mu;M:759 fluorescence intensity) and decreased NO (25 &mu;M: 61%, 80 &mu;M:36%) in a dose-dependent manner in BAECs (Chen et al., 2010).</p>
  • <p>Treatment with all-trans retinoic acid (ATRA) and retinoic acid receptor agonist Am580 at 1 &mu;mol/L for 48 hours also increased AKT&nbsp;(control: 100%, ATRA: 304%, Am580: 212%) and eNOS (control: 100%, ATRA: 276%, Am580: 209%) phosphorylation and NO production (control: 100%, ATRA: 170%, Am580: 169%) in human endothelial cells (Uruno et al., 2005).</p>
  • <p>Exposure to 50 &mu;mol/L peroxynitrite for 3 hours led to an increase in superoxide from 2 nmol/min/well to 8.6 nmol/min/well and a decrease in NO from 90% to 6.5% in BAECs (Zou et al., 2002).</p>
  • <strong>Uncertainties and Inconsistencies</strong>
  • <p>There are no uncertainties or inconsistencies.</p>
  • <strong>Uncertainties or Inconsistencies</strong>
  • <p>In the context of this AOP, decreased activity of AKT is likely due to proteasomal degradation following exposure to a (oxidative) stressor (Abdehghany et al. 2017).&nbsp; However, decreased eNOS activity could be due to multiple causes, highlighted in this AOP.&nbsp; Firstly, if AKT expression levels are reduced, it follows that eNOS phosphorylation will be reduced.&nbsp; Secondly, similar to AKT, eNOS itself has been shown to be susceptible to proteasomal degradation (Abdehghany et al. 2017).&nbsp; Thirdly, depletion of BH4 and/or S-glutathionylation has been shown to uncouple eNOS, leading to reduced NO levels and increased superoxide levels.&nbsp; The relative contribution of&nbsp;each of these eNOS perturbation routes to NO depletion is currently unknown.</p>
  • <strong>Quantitative Understanding of the Linkage</strong>
  • <p><em>Is it known how much change in the first event is needed to impact the second? Are there known modulators of the response-response relationships? Are there models or extrapolation approaches that help describe those relationships? </em></p>
  • <p>In two studies (Song et al, 2007; Das et al., 2014), it appears that a minimum of 30% reduction in eNOS activity or AKT&nbsp;phosphorylation caused a change in NO production in as little as five minutes. Other studies showed that 50-60% reduction in AKT&nbsp;phosphorylation/eNOS activity will lead to decreased NO bioavailability (Dhar et al., 2010; Dumitrescu et al., 2007). The studies above demonstrated that there are several known modulators for these two key events including peroxynitrite, high glucose, methylglyoxal, insulin and ATRA.</p>
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  • <!--<h3>Evidence Supporting Taxonomic Applicability</h3>-->
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  • <!--<h3>Domain of Applicability</h3>-->
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  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>AbdelGhany, T., Ismail, R., Elmahdy, M., Mansoor F, Zweier J, Lowe, F., and Zweier, JL. (2017). Cigarette Smoke Constituents Cause Endothelial Nitric Oxide Synthase Dysfunction and Uncoupling due to Depletion of Tetrahydrobiopterin with Degradation of GTP Cyclohydrolase.&nbsp;&nbsp;Nitric Oxide&nbsp;(Under review).</p>
  • <p>Choi, Y.-J., Yoon, Y., Lee, K.-Y., Hien, T.T., Kang, K.W., Kim, K.-C., Lee, J., Lee, M.-Y., Lee, S.M., Kang, D.-H., et al. (2014). Uric acid induces endothelial dysfunction by vascular insulin resistance associated with the impairment of nitric oxide synthesis. FASEB J. 28, 3197&ndash;3204.</p>
  • <p>Choi, Y.-J., Yoon, Y., Lee, K.-Y., Kang, Y.-P., Lim, D.K., Kwon, S.W., Kang, K.-W., Lee, S.-M., and Lee, B.-H. (2015). Orotic Acid Induces Hypertension Associated with Impaired Endothelial Nitric Oxide Synthesis. Toxicol. Sci. 144, 307&ndash;317.</p>
  • <p>Das, A., Gopalakrishnan, B., Druhan, L.J., Wang, T.-Y., De Pascali, F., Rockenbauer, A., Racoma, I., Varadharaj, S., Zweier, J.L., Cardounel, A.J., et al. (2014). Reversal of SIN-1-induced eNOS dysfunction by the spin trap, DMPO, in bovine aortic endothelial cells via eNOS phosphorylation. Br. J. Pharmacol. 171, 2321&ndash;2334.</p>
  • <p>Dhar, A., Dhar, I., Desai, K.M., and Wu, L. (2010). Methylglyoxal scavengers attenuate endothelial dysfunction induced by methylglyoxal and high concentrations of glucose. Br. J. Pharmacol. 161, 1843&ndash;1856.</p>
  • <p>Chen, X., Xu, J., Feng, Z., Fan, M., Han, J., and Yang, Z. (2010). Simvastatin combined with nifedipine enhances endothelial cell protection by inhibiting ROS generation and activating Akt phosphorylation. Acta Pharmacol. Sin. 31, 813&ndash;820.</p>
  • <p>Dimmeler, S., Fleming, I., Fisslthaler, B., Hermann, C., Busse, R., and Zeiher, A.M. (1999). Activation of nitric oxide synthase in endothelial cells by Akt-dependent phosphorylation. Nature 399, 601&ndash;605.</p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p>Dumitrescu, C., Biondi, R., Xia, Y., Cardounel, A.J., Druhan, L.J., Ambrosio, G., and Zweier, J.L. (2007). Myocardial ischemia results in tetrahydrobiopterin (BH4) oxidation with impaired endothelial function ameliorated by BH4. Proc. Natl. Acad. Sci. U. S. A. 104, 15081&ndash;15086.</p>
  • <p>Fulton, D., Gratton, J.P., McCabe, T.J., Fontana, J., Fujio, Y., Walsh, K., Franke, T.F., Papapetropoulos, A., and Sessa, W.C. (1999). Regulation of endothelium-derived nitric oxide production by the protein kinase Akt. Nature 399, 597&ndash;601.</p>
  • <p>Michaud, S.E., Dussault, S., Groleau, J., Haddad, P., and Rivard, A. (2006). Cigarette smoke exposure impairs VEGF-induced endothelial cell migration: role of NO and reactive oxygen species. J. Mol. Cell. Cardiol. 41, 275&ndash;284.</p>
  • <p>F&ouml;rstermann, U., and M&uuml;nzel, T. (2006). Endothelial nitric oxide synthase in vascular disease: from marvel to menace. Circulation 113, 1708&ndash;1714.</p>
  • <p>Song, P., Wu, Y., Xu, J., Xie, Z., Dong, Y., Zhang, M., and Zou, M.-H. (2007). Reactive nitrogen species induced by hyperglycemia suppresses Akt signaling and triggers apoptosis by upregulating phosphatase PTEN (phosphatase and tensin homologue deleted on chromosome 10) in an LKB1-dependent manner. Circulation 116, 1585&ndash;1595.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Song, P., Xie, Z., Wu, Y., Xu, J., Dong, Y., and Zou, M.-H. (2008). Protein kinase Czeta-dependent LKB1 serine 428 phosphorylation increases LKB1 nucleus export and apoptosis in endothelial cells. J. Biol. Chem. 283, 12446&ndash;12455.</p>
  • <p>Wang, S., Xu, J., Song, P., Wu, Y., Zhang, J., Chul Choi, H., and Zou, M.-H. (2008). Acute inhibition of guanosine triphosphate cyclohydrolase 1 uncouples endothelial nitric oxide synthase and elevates blood pressure. Hypertension 52, 484&ndash;490.</p>
  • <p>Uruno, A., Sugawara, A., Kanatsuka, H., Kagechika, H., Saito, A., Sato, K., Kudo, M., Takeuchi, K., and Ito, S. (2005). Upregulation of nitric oxide production in vascular endothelial cells by all-trans retinoic acid through the phosphoinositide 3-kinase/Akt pathway. Circulation 112, 727&ndash;736.</p>
  • <p>Whitsett, J., Picklo, M.J., and Vasquez-Vivar, J. (2007). 4-Hydroxy-2-nonenal increases superoxide anion radical in endothelial cells via stimulated GTP cyclohydrolase proteasomal degradation. Arterioscler. Thromb. Vasc. Biol. 27, 2340&ndash;2347.</p>
  • <p>Zou, M.-H., Hou, X.-Y., Shi, C.-M., Nagata, D., Walsh, K., and Cohen, R.A. (2002). Modulation by peroxynitrite of Akt- and AMP-activated kinase-dependent Ser1179 phosphorylation of endothelial nitric oxide synthase. J. Biol. Chem. 277, 32552&ndash;32557.</p>
  • <p>Zweier, J.L., Chen, C.-A., and Druhan, L.J. (2011). S-glutathionylation reshapes our understanding of endothelial nitric oxide synthase uncoupling and nitric oxide/reactive oxygen species-mediated signaling. Antioxid. Redox Signal. 14, 1769&ndash;1775.</p>
  • </div>
  • <br>
  • <div>
  • <div>
  • <h4><a href="/relationships/992">Peptide Oxidation leads to Decrease, AKT/eNOS activity</a></h4>
  • <h4><a href="/relationships/958">Relationship: 958: Depletion, Nitric Oxide leads to Impaired, Vasodilation</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Directness</th>
  • <th>Adjacency</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>directly leads to</th>
  • <th>Strong </th>
  • <th>adjacent</th>
  • <th>High </th>
  • <th>Moderate</th>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>Oryctolagus cuniculus</td>
  • <td>Oryctolagus cuniculus</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9986" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Weak</td>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Weak</td>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>The relationship between oxidative stress and decreased AKT/eNOS activity is supported by studies performed in humans, cows, mice and rats.</p>
  • <p>This relationship between NO depletion and impaired vasodilation was shown in humans (Li et al. 1993, Heitzer et al. 2000), rabbits (Luo et al. 2000), mice (Luo et al. 2000, Wang et al. 2008)&nbsp;and rats (Paulis et al. 2008, Li et al. 1993, S&eacute;lley et al. 2014, Chen et al. 2010).</p>
  • <h4>How Does This Key Event Relationship Work</h4>
  • <p>Exposure to known inducers of oxidative stress causes the phosphorylation of AKT&nbsp;and eNOS, leading to a decrease in their activities.</p>
  • <h4>Key Event Relationship Description</h4>
  • <p>Nitric oxide (NO)&nbsp;is a critical endothelium-derived hyperpolarising factor (EDHF), responsible for relaxation of vascular smooth muscle and vasodilation.&nbsp; The primary regulator of endothelial vasodilator function via NO is vascular shear; the frictional force exerted on the vascular wall during the flow of blood through the vessel. Vascular shear opens calcium channels on endothelial cells, and leads to the calcium-dependent activation of eNOS and thus NO production.&nbsp; NO then diffuses to the underlying vascular smooth muscle, where it activates soluble guanylate cyclase, causing an increase in cyclic guanosine monophosphate (cGMP), potassium ion efflux, hyperpolarization and smooth muscle relaxation (Giles et al. 2012).</p>
  • <p>Depletion of vascular NO bioavailability causes an imbalance in the maintenance of vascular tone, which shifts in favour of vasoconstriction, and hence elevates blood pressure (Kojda et al. 1999).&nbsp; Under oxidative stress, decreased NO bioavailability results in impaired endothelium-dependent vasodilation (Silva et al., 2012).</p>
  • <!-- if nothing shows up in any of these fields, then weight of evidence will not be displayed -->
  • <h4>Weight of Evidence</h4>
  • <!-- if nothing shows up in any of these fields, then evidence supporting this KER will not be displayed -->
  • <h4>Evidence Supporting this KER</h4>
  • <strong>Biological Plausibility</strong>
  • <p>Multiple experimental studies reported modulations in Akt and eNOS phosphorylation/activity following oxidative stress, thus providing strong biological plausibility for this key event relationship.</p>
  • <p>In HUVECs, peroxynitrite significantly inhibited AKT&nbsp;phosphorylation at Ser473 and AKT&nbsp;activity (Song et al., 2007, 2008; Zou et al., 2002). However, Zou et al. (2002) found that peroxynitrite increased eNOS phosphorylation at Ser1199, but decreased NO availability, suggesting eNOS phosphorylation may not depend on AKT. Treatment of BAECs with SIN-1, a source of peroxynitrite, inhibited eNOS activity by 30% and AKT/eNOS phosphorylation (Das et al., 2014).</p>
  • <p>High glucose concentrations can also inhibit AKT&nbsp;phosphorylation (Song et al., 2008). HUVECs treated with methylglyoxal and high concentrations of glucose exhibited reduced bradykinin-stimulated eNOS activity via Ser1177 phosphorylation (Dhar et al., 2010), while hyperglycemia inhibited eNOS activity in BAECs (Du et al., 2001). In EA.hy926 endothelial cells, methylglyoxal treatment results in reduced eNOS phosphorylation at Ser1177 (Su et al., 2013). High-fat diet-induced obesity in mice caused an increase in ROS and a reduction in AKT&nbsp;and eNOS phosphorylation compared to non-obese mice (Du et al., 2013).</p>
  • <p>Vasodilation is caused by the relaxation of vascular smooth muscle cells within the walls of blood vessels, which is regulated through a number of mechanisms, including cyclic GMP-dependent hyperpolarization and relaxation via NO. Thus, alterations to NO levels have an influence on vasodilation (Silva et al., 2012). Many animal studies demonstrated that inhibition of NO via eNOS inhibitors impaired acetylcholine-induced endothelium-dependent vasodilation, providing strong biological plausibility for this key event relationship.</p>
  • <p>Treatment with cigarette smoke extract (CSE) also inhibited Akt and eNOS in VEGF-stimulated HUVECs (Michaud et al., 2006). Myocardial ischemia decreased phosphorylated AKT and eNOS in spontaneously hypertension (SHR) rats compared to sham animals (Zhang et al., 2014).</p>
  • <p>Constitutively active AKT, which has been shown to phosphorylate and activate eNOS, increased resting vessel diameter in a rabbit femoral artery model, but infusion with eNOS inhibitor L-NAME reversed this effect (Luo et al., 2000). Dominant negative AKT, which prevents eNOS activation, also blocked vasodilation in response to the endothelium-dependent agonist acetylcholine (ACh) in rabbit arteries and mouse aortas. Since acetylcholine induces endothelium-dependent vasodilation, the assumption is that when eNOS inhibitors blocks Ach-induced vasodilation, NO production is decreased and vasodilation is impaired. L-NAME abolished endothelium-dependent relaxations induced by ACh in aortas of wild-type and hph-1 mice (Li et al., 2007). ACh induced a dose-dependent endothelium-dependent relaxation in rat small mesenteric arteries, but the NO-dependent component of ACh-induced relaxation was decreased in L-NAME-treated rats (Paulis et al., 2008). After L-NAME cessation, the NO-dependent component was restored to above control levels. L-NAME diminished eNOS activity by 66% and its cessation restore NOS activity to control levels in rat aortas. Another study showed that L-NAME caused a mild but significant decrease in vasodilation in rat thoracic aorta (S&eacute;lley et al., 2014).</p>
  • <strong>Empirical Support for Linkage</strong>
  • <strong>Empirical Evidence</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>Treatment with 30 &mu;M methylglyoxal (MG) and 25 mM glucose (HG) for 24 hours caused an increase in ROS (measured by DCF fluorescence in arbitrary units; control: 0.77, MG: 1.2, HG: 1) and a decrease in eNOS activity (control: 100%, MG: 55%, HG: 66%) in HUVECs (Dhar et al., 2010). SIN-1 treatment for two hours increased ROS (100% to 290%) and decreased eNOS activity (100% to 70%) (Das et al., 2014). An increase in ROS and decrease in eNOS activity was observed following treatment with 100 &mu;mol/L H<sub>2</sub>O<sub>2</sub> (Chen et al., 2010). Another study showed that 5 &mu;M H<sub>2</sub>O<sub>2</sub> treatment initially increased eNOS Ser1179 phosphorylation and activity, but after the peak increase at 30 minutes, eNOS Ser1179 phosphorylation dramatically decreased (Hu et al., 2008). A similar trend was observed for AKT&nbsp;phosphorylation. Treatment with 100 &mu;M/L H<sub>2</sub>O<sub>2</sub> for 30 minutes inhibited eNOS expression (Chen et al., 2010), assuming that eNOS expression translates to eNOS activity. Treatment with H<sub>2</sub>O<sub>2</sub> (400 &micro;M, 30 min) increased eNOS phosphorylation at the inhibitory site Thr495, suggesting eNOS activity decreased (Guterbaum, 2013).</p>
  • <p>There is moderate empirical support for depletion of NO leading to impaired vasodilation based on several studies measuring decreased NO and vasodilation following a number of perturbations. In rat small mesenteric arteries, treatment with L-NAME for five weeks decreased NO-dependent relaxation from 28% to 7.5%, while diminishing eNOS activity by 66% (Paulis et al, 2008). In rat aortic rings, BCNU treatment for 4 hours caused a dose-dependent decrease in NO production (control: 100%, 25 &mu;M: 61%, 80 &mu;M: 36%), and also decreased vasodilation from 69% control to 37% (at 80 &mu;M) (Chen et al., 2010). Similarly, treatment with 10 mM DAHP for 24 hours caused a decrease in both NO (from 100% to 61%) and vasodilation (from 87% to 42%) (Wang et al., 2008).</p>
  • <p>In&nbsp;humans, application of L-NAME and/or N<sup>G</sup>-monomethyl-L-arginine (L-NMMA) resulted in elevated mean arterial pressure via impairment of NO-mediated vasodilation.&nbsp; Application of both drugs compounded the effect (Sander et al. 1999).&nbsp; BH4 administration reversed NO depletion by L-NMMA and improved forearm venous blood flow in chronic smokers (Heizter et al. 2000).&nbsp; In human skin, acetylcholine-mediated vasodilation was shown to be modulated by NO and impaired by the eNOS inhibitor; L-NAME (Kellogg et al. 2005).&nbsp; Intravenous infusion&nbsp;of the eNOS substrate L-arginine was shown to decrease blood pressure and total peripheral resistance.&nbsp;&nbsp;The onset and the duration of the vasodilator effect of L-arginine and its effects on endogenous NO production closely corresponded to the plasma concentration half-life of l-arginine, and was associatedwith a concomminant elevation of urinary nitrate and cyclic GMP excretion (Bode-B&ouml;ger et al. 1998).&nbsp; Finally, in chronic smokers with impaired flow-mediated dilation (FMD;&nbsp;5.6&plusmn;3.0% vs. 8.1&plusmn;3.7% for non-smokers&nbsp;P&lt;0.01), administration of the essential eNOS cofactor; BH4, improved FMD in both cohorts (6.6&plusmn;3.3% vs. 9.8&plusmn;3.2%; P&lt;0.01) and smoking cessation for 1 week also improved FMD (from 5.0&plusmn;2.9 to 7.8&plusmn;3.2%;P&lt;0.01).&nbsp; This data indicates that improvement in arterial vasodilation was partially improved by BH4 administration (Taylor et al. 2016).&nbsp; Another study by Carnevale et al. (2016) in which current smokers switched to an e-cigarette product for 1 week showed significant reductions in FMD impairment compared to baseline, which was associated with improvement in levels of 8-iso-prostagladin F2&alpha; (a biomarker of oxidative stress).</p>
  • <strong>Uncertainties and Inconsistencies</strong>
  • <p>While the effect of NO depletion on impaired vasodilation is clear, the NO pathway does not appear to be solely responsible for this phenomenon.&nbsp; Vascular tone is a balance between relaxation and constriction factors.&nbsp; Studies have shown that when NO bioavailability is decreased, COX-mediated pro-inflammatory factors such as prostaglandins (Kellogg et al. 2005, L&uuml;scher and Vanhoutte 1986) and Endothelin-1 (Taddei et al. 2003)&nbsp; contribute to a shift in vascular tone towards vasoconstiction.&nbsp; The&nbsp;review by Silva et al. (2012) discusses the roles of various pathways and their effects on vascular tone.&nbsp; The relative contribution of these mechanisms towards vascular tone is currently unknown.</p>
  • <strong>Uncertainties or Inconsistencies</strong>
  • <p>There are many studies examining the effect of H<sub>2</sub>O<sub>2</sub> on AKT/eNOS phosphorylation, but there are conflicting results. Exposure to H<sub>2</sub>O<sub>2</sub> for 30 minutes resulted in an increase in AKT/eNOS phosphorylation, but its concentration was much higher at 200 &mu;M (Barbosa et al., 2013) compared to 5 &mu;M in Hu et al. (2008). Another study found that treatment with 50 &mu;M H<sub>2</sub>O<sub>2</sub> increased eNOS phosphorylation at Ser1177 (Kumar et al., 2010). Results from studies with H<sub>2</sub>O<sub>2</sub> as a source of ROS may not be universally applicable to this key event relationship.</p>
  • <strong>Quantitative Understanding of the Linkage</strong>
  • <p><em>Is it known how much change in the first event is needed to impact the second? Are there known modulators of the response-response relationships? Are there models or extrapolation approaches that help describe those relationships? </em></p>
  • <p>Dhar et al. (2010) showed that an increase in ROS of &gt;30% due to high glucose and methylglyoxal treatment was able to decrease eNOS activity, while Das et al. (2014) showed a three-fold increase in ROS led to a 30% reduction in eNOS activity. SIN-1, high glucose, methylglyoxal and H<sub>2</sub>O<sub>2</sub> were demonstrated to modulate both key events at the same time.</p>
  • <!--<!% unless aop_rel.relationship.relationship_taxons.blank? %>-->
  • <!--<!%= render 'snapshot_taxons', taxons: aop_rel.relationship.relationship_taxons %>-->
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  • <!--<h3>Evidence Supporting Taxonomic Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
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  • <!--<!%= render 'snapshot_taxons', taxons: aop_rel.relationship.relationship_taxons %>-->
  • <!--<!% unless aop_rel.relationship.taxon_evidence.blank? %>-->
  • <!--<h3>Domain of Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
  • <!--<!% end %>-->
  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>Barbosa, V.A., Luciano, T.F., Marques, S.O., Vitto, M.F., Souza, D.R., Silva, L.A., Santos, J.P.A., Moreira, J.C., Dal-Pizzol, F., Lira, F.S., et al. (2013). Acute exercise induce endothelial nitric oxide synthase phosphorylation via Akt and AMP-activated protein kinase in aorta of rats: Role of reactive oxygen species. Int. J. Cardiol. 167, 2983&ndash;2988.</p>
  • <p>Bode-B&ouml;ger SM,&nbsp;B&ouml;ger RH,&nbsp;Galland A,&nbsp;Tsikas D,&nbsp;Fr&ouml;lich JC.&nbsp; L-arginine-induced&nbsp;vasodilation&nbsp;in&nbsp;healthy&nbsp;humans:&nbsp;pharmacokinetic-pharmacodynamicrelationship.&nbsp; Br J Clin Pharmacol.&nbsp;1998, 46(5):489-97.</p>
  • <p>Chen, X., Xu, J., Feng, Z., Fan, M., Han, J., and Yang, Z. (2010). Simvastatin combined with nifedipine enhances endothelial cell protection by inhibiting ROS generation and activating Akt phosphorylation. Acta Pharmacol. Sin. 31, 813&ndash;820.</p>
  • <p>Carnevale R, Sciarretta S, Violi F et al.&nbsp; Acute Impact of Tobacco vs Electronic Cigarette Smoking on Oxidative Stress and Vascular Function.&nbsp; Chest. (2016) 150(3):606-12.</p>
  • <p>Dhar, A., Dhar, I., Desai, K.M., and Wu, L. (2010). Methylglyoxal scavengers attenuate endothelial dysfunction induced by methylglyoxal and high concentrations of glucose. Br. J. Pharmacol. 161, 1843&ndash;1856.</p>
  • <p>Chen, C.-A., Wang, T.-Y., Varadharaj, S., Reyes, L.A., Hemann, C., Talukder, M.A.H., Chen, Y.-R., Druhan, L.J., and Zweier, J.L. (2010). S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature 468, 1115&ndash;1118.</p>
  • <p>Das, A., Gopalakrishnan, B., Druhan, L.J., Wang, T.-Y., De Pascali, F., Rockenbauer, A., Racoma, I., Varadharaj, S., Zweier, J.L., Cardounel, A.J., et al. (2014). Reversal of SIN-1-induced eNOS dysfunction by the spin trap, DMPO, in bovine aortic endothelial cells via eNOS phosphorylation. Br. J. Pharmacol. 171, 2321&ndash;2334.</p>
  • <p>Giles TD, Sander GE, Nossaman BD et al. Impaired vasodilation in the pathogenesis of hypertension: focus on nitric oxide, endothelial-derived hyperpolarizing factors, and prostaglandins.&nbsp; J Clin Hypertens (2012) 14(4):198-205.</p>
  • <p>Du, J., Fan, L.M., Mai, A., and Li, J.-M. (2013a). Crucial roles of Nox2-derived oxidative stress in deteriorating the function of insulin receptors and endothelium in dietary obesity of middle-aged mice. Br. J. Pharmacol. 170, 1064&ndash;1077.</p>
  • <p>Heitzer T,&nbsp;Brockhoff C,&nbsp;Mayer B,&nbsp;Warnholtz A,&nbsp;Mollnau H,&nbsp;Henne S,&nbsp;Meinertz T,&nbsp;M&uuml;nzel T.&nbsp; Tetrahydrobiopterin improves endothelium-dependent&nbsp;vasodilation&nbsp;in chronic smokers : evidence for a dysfunctional&nbsp;nitric oxide&nbsp;synthase.&nbsp; Circ Res.&nbsp;2000, 86(2):E36-41.</p>
  • <p>Du, X.L., Edelstein, D., Dimmeler, S., Ju, Q., Sui, C., and Brownlee, M. (2001). Hyperglycemia inhibits endothelial nitric oxide synthase activity by posttranslational modification at the Akt site. J. Clin. Invest. 108, 1341&ndash;1348.</p>
  • <p>Kellogg DL Jr,&nbsp;Zhao JL,&nbsp;Coey U,&nbsp;Green JV.&nbsp; Acetylcholine-induced&nbsp;vasodilation&nbsp;is mediated by&nbsp;nitric oxide&nbsp;and prostaglandins in&nbsp;human&nbsp;skin.&nbsp; J Appl Physiol (1985).&nbsp;2005, 98(2):629-32.</p>
  • <p>Guterbaum, T.J., Braunstein, T.H., Fossum, A., Holstein-Rathlou, N.-H., Torp-Pedersen, C.T., and Dom&iacute;nguez, H. (2013). Endothelial nitric oxide synthase phosphorylation at Threonine 495 and mitochondrial reactive oxygen species formation in response to a high H₂O₂ concentration. J. Vasc. Res. 50, 410&ndash;420.</p>
  • <p>Kojda G, Laursen JB, Ramasamy S et al. Protein expression, vascular reactivity and soluble guanylate cyclase activity in mice lacking the endothelial cell nitric oxide synthase: contributions of NOS isoforms to blood pressure and heart rate control.&nbsp; Cardiovasc Res. (1999) 42(1):206-13.</p>
  • <p>Hu, Z., Chen, J., Wei, Q., and Xia, Y. (2008). Bidirectional actions of hydrogen peroxide on endothelial nitric-oxide synthase phosphorylation and function: co-commitment and interplay of Akt and AMPK. J. Biol. Chem. 283, 25256&ndash;25263.</p>
  • <p>Li, J., Zhou, Z., Jiang, D.-J., Li, D., Tan, B., Liu, H., and Li, Y.-J. (2007). Reduction of NO- and EDHF-mediated vasodilatation in hypertension: role of asymmetric dimethylarginine. Clin. Exp. Hypertens. N. Y. N 1993 29, 489&ndash;501.</p>
  • <p>Kumar, S., Sud, N., Fonseca, F.V., Hou, Y., and Black, S.M. (2010). Shear stress stimulates nitric oxide signaling in pulmonary arterial endothelial cells via a reduction in catalase activity: role of protein kinase C delta. Am. J. Physiol. Lung Cell. Mol. Physiol. 298, L105&ndash;L116.</p>
  • <p>L&uuml;scher T. F., Vanhoutte P. M. (1986).&nbsp;Endothelium-dependent contractions to acetylcholine in the aorta of the spontaneously hypertensive rat.&nbsp;Hypertension&nbsp;8, 344&ndash;348</p>
  • <p>Michaud, S.E., Dussault, S., Groleau, J., Haddad, P., and Rivard, A. (2006). Cigarette smoke exposure impairs VEGF-induced endothelial cell migration: role of NO and reactive oxygen species. J. Mol. Cell. Cardiol. 41, 275&ndash;284.</p>
  • <p>Luo, Z., Fujio, Y., Kureishi, Y., Rudic, R.D., Daumerie, G., Fulton, D., Sessa, W.C., and Walsh, K. (2000). Acute modulation of endothelial AKT/PKB activity alters nitric oxide&ndash;dependent vasomotor activity in vivo. J. Clin. Invest. 106, 493&ndash;499.</p>
  • <p>Song, P., Wu, Y., Xu, J., Xie, Z., Dong, Y., Zhang, M., and Zou, M.-H. (2007). Reactive nitrogen species induced by hyperglycemia suppresses Akt signaling and triggers apoptosis by upregulating phosphatase PTEN (phosphatase and tensin homologue deleted on chromosome 10) in an LKB1-dependent manner. Circulation 116, 1585&ndash;1595.</p>
  • <p>Paulis, L., Zicha, J., Kunes, J., Hojna, S., Behuliak, M., Celec, P., Kojsova, S., Pechanova, O., and Simko, F. (2008). Regression of L-NAME-induced hypertension: the role of nitric oxide and endothelium-derived constricting factor. Hypertens. Res. Off. J. Jpn. Soc. Hypertens. 31, 793&ndash;803.</p>
  • <p>Song, P., Xie, Z., Wu, Y., Xu, J., Dong, Y., and Zou, M.-H. (2008). Protein kinase Czeta-dependent LKB1 serine 428 phosphorylation increases LKB1 nucleus export and apoptosis in endothelial cells. J. Biol. Chem. 283, 12446&ndash;12455.</p>
  • <p>Sander M, Chavoshan B, Victor RG.&nbsp; A large blood pressure-raising effect of nitric oxide synthase inhibition in humans.&nbsp; Hypertension. (1999) 33(4):937-42.</p>
  • <p>Su, Y., Qadri, S.M., Wu, L., and Liu, L. (2013). Methylglyoxal modulates endothelial nitric oxide synthase-associated functions in EA.hy926 endothelial cells. Cardiovasc. Diabetol. 12, 134.</p>
  • <p>S&eacute;lley, E., Kun, S., Szij&aacute;rt&oacute;, I.A., Laczy, B., Kov&aacute;cs, T., F&uuml;l&ouml;p, F., Wittmann, I., and Moln&aacute;r, G.A. (2014). Exenatide induces aortic vasodilation increasing hydrogen sulphide, carbon monoxide and nitric oxide production. Cardiovasc. Diabetol. 13, 69.</p>
  • <p>Zhang, W., Han, Y., Meng, G., Bai, W., Xie, L., Lu, H., Shao, Y., Wei, L., Pan, S., Zhou, S., et al. (2014). Direct renin inhibition with aliskiren protects against myocardial ischemia/reperfusion injury by activating nitric oxide synthase signaling in spontaneously hypertensive rats. J. Am. Heart Assoc. 3, e000606.</p>
  • <p>Silva, B.R., Pernomian, L., and Bendhack, L.M. (2012). Contribution of oxidative stress to endothelial dysfunction in hypertension. Front. Physiol. 3, 441.</p>
  • <p>Zou, M.-H., Hou, X.-Y., Shi, C.-M., Nagata, D., Walsh, K., and Cohen, R.A. (2002). Modulation by peroxynitrite of Akt- and AMP-activated kinase-dependent Ser1179 phosphorylation of endothelial nitric oxide synthase. J. Biol. Chem. 277, 32552&ndash;32557.</p>
  • <p>Taddei S., Ghiadoni L., Virdis A., Versari D., Salvetti A. (2003).&nbsp;Mechanisms of endothelial dysfunction: clinical significance and preventive non-pharmacological therapeutic strategies.&nbsp;Curr. Pharm. Des.&nbsp;9, 2385&ndash;2402</p>
  • <p>Taylor BA, Zaleski AL, Dornelas EA et al.&nbsp; The impact of tetrahydrobiopterin administration on endothelial function before and after smoking cessation in chronic smokers.&nbsp; Hypertens Res. (2016) 39(3):144-50.</p>
  • <p>Wang, S., Xu, J., Song, P., Wu, Y., Zhang, J., Chul Choi, H., and Zou, M.-H. (2008). Acute inhibition of guanosine triphosphate cyclohydrolase 1 uncouples endothelial nitric oxide synthase and elevates blood pressure. Hypertension 52, 484&ndash;490.</p>
  • </div>
  • <br>
  • <div>
  • <div>
  • <h4><a href="/relationships/1698">Peptide Oxidation leads to S-Glutathionylation, eNOS</a></h4>
  • <h4><a href="/relationships/982">Relationship: 982: Impaired, Vasodilation leads to Increase, Vascular Resistance</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Directness</th>
  • <th>Adjacency</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>directly leads to</th>
  • <th>adjacent</th>
  • <th>Moderate </th>
  • <th>Weak</th>
  • <th>Low</th>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>rat</td>
  • <td>Rattus norvegicus</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>human</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>Homo sapiens</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>mouse</td>
  • <td>Mus musculus</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>cow</td>
  • <td>Bos taurus</td>
  • <td>Strong</td>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Low</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>Strong</td>
  • <td>Low</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>Strong</td>
  • <td>Not Specified</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>The above evidence demonstrates similar responses to stressors in cows, mice and humans.&nbsp; A functional response using&nbsp;aortic rings was demonstrated in rats.</p>
  • <p>This relationship between impaired vasodilation and SVR was shown in human and rat studies.</p>
  • <h4>How Does This Key Event Relationship Work</h4>
  • <p>Oxidation of GSH results in the formation of a disulphide-bridged glutathione dimer (GSSG).&nbsp; GSSG is either rapidly re-reduced back to GSH by nicotinamide adenine dinucleotide phosphate (NADPH)-dependent GSSG-reductases or extruded from the cell by adenosine triphosphate (ATP)-dependent translocases. However, when these mechanisms become overwhelmed by high local oxidant concentrations, GSSG can interact with protein thiol groups to form protein-GSSG adducts, a process termed S-glutathionylation. Interestingly, glutathione disulfide-protein formation has been suggested to occur with a certain degree of specificity to cellular proteins, since protein thiol groups exhibit a considerable heterogeneity in terms of their individual pK<sub>a</sub> values and their location in protein structures <span style="font-size:10.8333px">(</span>Schuppe et al. 1992).&nbsp; The oxidation of GSH to GSSG elevates levels of GSSG, which then covalently bind to critical serine residues on endothelial nitric oxide synthase (eNOS; Chen et al 2010, Du et al. 2013, De Pascali et al. 2014).</p>
  • <h4>Key Event Relationship Description</h4>
  • <p>Vasodilation decreases systemic vascular resistance (SVR; also known previously as Total Peripheral Resistance; TPR), the resistance to blood flow offered by the peripheral circulation, and blood pressure through relaxation of vascular smooth muscle cells (VSMCs) (Siddiqui, 2011). When vasodilation is impaired due to decreased NO availability, SVR and blood pressure become elevated.</p>
  • <!-- if nothing shows up in any of these fields, then weight of evidence will not be displayed -->
  • <h4>Weight of Evidence</h4>
  • <p>&nbsp;</p>
  • <p>&nbsp;</p>
  • <!-- if nothing shows up in any of these fields, then evidence supporting this KER will not be displayed -->
  • <h4>Evidence Supporting this KER</h4>
  • <p>The overall weight of evidence for the KER was rated &quot;moderate&quot; due the fact that acute pharmacological manipulation of the NO pathway resulted in corresponding changes in SVR.&nbsp; However, in the context of the development of hypertension, the chronic effects of impaired vasodilation are much less clear.</p>
  • <strong>Biological Plausibility</strong>
  • <p>When antioxidant defence&nbsp;mechanisms become overwhelmed by high local oxidant concentrations, GSSG can interact with protein thiol groups to form protein-GSSG adducts, a process termed S-glutathionylation<span style="font-size:10.8333px">&nbsp;(</span>Schuppe et al. 1992).</p>
  • <p>It is well-accepted that vasodilation and SVR are negatively correlated; blood flow is increased when blood vessels dilate due to a decrease in vascular resistance (Siddiqui, 2011). When vasodilation is impaired, SVR increases, in turn increasing blood pressure. Agents that cause hyperpolarization are potent vasodilators and activate&nbsp;potassium channels, while factors causing depolarization increase vascular tone (Nelson, 1990). Vascular tone is governed by the&nbsp;contractile activity of VSMCs in the walls of small arteries and arterioles, and is the major determinant of the resistance to blood flow through the circulation (Jackson, 2000). VSMCs from hypertensive animals have decreased functional voltage-gated potassium channels, which may contribute to depolarization. Two studies demonstrated that blockade of potassium channels completely inhibited NO-dependent vasodilation and increased SVR (Dessy et al., 2004; Berg et al., 2011). Inhibitors of eNOS activity (L-NAME, L-NMMA), which have been shown to decrease acetylcholine-induced vasorelaxation in animal studies (Li et al., 2007; Paulis et al., 2008), also caused an increase in SVR in human studies (Wilkinson et al., 2002; McVeigh et al., 2001; Brett et al., 1998). Overall, these results provide strong biological plausibility for this link.</p>
  • <strong>Empirical Evidence</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>Hypoxia/reoxygenation-induced oxidative stress (associated with ischaemia-reperfusion injury) was shown to deplete GSH in bovine aortic endothelial cells, which led to S-glutathionylation of eNOS and eNOS uncoupling.&nbsp; This phenomenon was partially reversible, in bovine aortic endothelial cells and rat aortic rings, by raising intracellular GSH levels upon administration of N-acetylcysteine (Chen et al. 2010, DePascali et al. 2014)<span style="font-size:10.8333px">.</span></p>
  • <p>No direct evidence was found for this linkage; thus the empirical support is weak. One study indirectly showed a relationship between NO depletion and SVR, where infusion of L-NMMA (1.0 mg/kg/min) caused an increase in SVR by 63% and a 65% reduction in NO in 11 healthy volunteers after three minutes (Stamler et al., 1994). In rat small mesenteric arteries, treatment with L-NAME for five weeks decreased NO-dependent relaxation from 30% to less than 10%, but increased systolic and diastolic blood pressure to 26% and 40%, respectively (Paulis et al, 2008). Meta-analysis of clinical trials showing that infusion of the NO donor sodium nitroprusside led to a dose-dependent reduction in SVR (Eugene et al., 2016).&nbsp; In a study of 400 post-menopausal women with mild-moderate hypertension and impaired FMD, anti-hypertensive therapy for 6 months was shown to improve FMD by &gt;10% in 250 women, whereas in 150 women, FMD did not significantly change (&lt;10% change, Modena et al. 2002).&nbsp; Whilst the study demonstrates a linkage between vasodilation and hypertension in a large proportion of the study population, it raises a&nbsp;question regarding causality i.e. did NO-independent medication lower blood pressure, which in turn improved FMD?&nbsp; Furthermore, over a third of the population showed no change.&nbsp; It is possible that the medications used directly influenced key components of the NO (or other) pathways.&nbsp; Since the study did not elaborate on the medicatons used by the study cohort, this is impossible to determine.</p>
  • <strong>Empirical Support for Linkage</strong>
  • <p>Chen and colleagues (2010) have shown that eNOS is particularly sensitive to S-glutathionylation at cysteine residues 689 and 908 of the reductase domain, a phenomenon that is dose-dependent with application of exogenous GSSG.&nbsp; This finding was corroborated by Peng <em>et al.</em>&nbsp;(2015) using mutated eNOS constructs in <em>E. coli</em>, demonstrating that superoxide was produced by the eNOS phosphorylation site in the reductase domain.</p>
  • <strong>Uncertainties and Inconsistencies</strong>
  • <p>As mentioned above, acute pharmacological manipulation of the NO pathway results in expected changes in SVR.&nbsp; However,&nbsp;the link between chronically impaired vasodilation&nbsp;and SVR (the context of this AOP) is much less clear due to gaps in the literature.&nbsp; Epidemiological studies tend to investigate linkages between impaired vasodilation and cardiovascular events, as opposed to SVR and/or hypertension - making assessment of this KER difficult.</p>
  • <p>Wu <em>et al.</em>&nbsp;(2014) studied responses in human lung microvascular endothelial cells to lipopolysaccharide (LPS) <em>in vitro</em>.&nbsp; Upon LPS administration, NADPH oxidase 2 (NOX2) expression levels were increased with a subsequent rise in superoxide production, which led to S-glutathionylation of eNOS.&nbsp;&nbsp;Furthermore, in mice, co-immunoprecipitation studies revealed that NOX2 associated with eNOS, and that S-glutathionylation in response to LPS was much more apparent in elderly animals compared to younger animals.&nbsp; Similar observations were made by De Pascali <em>et al.</em>&nbsp;(2014) following hypoxia-induced oxidative stress in bovine aortic endothelial cells.</p>
  • <p>Furthermore, the complexity in the mechanisms influencing vascular re-modelling over time has hampered&nbsp;understanding of the phenomenon to date.&nbsp; The study by Modena et al. 2002 highlights that members of the general population respond differently to hypertensive therapy in the context of FMD improvement.</p>
  • <strong>Uncertainties or Inconsistencies</strong>
  • <p>Quantitative data for humans is very limited.</p>
  • <strong>Quantitative Understanding of the Linkage</strong>
  • <p>Chen <em>et al.</em>&nbsp;(2013) demonstrated that co-administration of glutaredoxin-1 and GSH reversed GSSG-mediated eNOS S-glutathionylation and restored eNOS-mediated NO production, also in bovine aortic cells.&nbsp; Interestingly, inhibition of eNOS function occurred when the GSH/GSSG ratio was &gt;0.2 and function was restored at a ratio of &lt;0.1.</p>
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  • <!--<h3>Domain of Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
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  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>Chen CA, Wang TY, Varadharaj, S <em>et al</em>. S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature (2010) 468: 1115&ndash;1118.</p>
  • <p>Berg, T., and Jensen, J. (2011). Simultaneous parasympathetic and sympathetic activation reveals altered autonomic control of heart rate, vascular tension, and epinephrine release in anesthetized hypertensive rats. Front. Neurol. 2, 71.</p>
  • <p>Chen CA, De Pascali F, Basye A <em>et al.</em> Redox modulation of endothelial nitric oxide synthase by glutaredoxin-1 through reversible oxidative post-translational modification.&nbsp; (2013)&nbsp; Biochemistry. 52(38):6712-23</p>
  • <p>Brett, S.E., Cockcroft, J.R., Mant, T.G., Ritter, J.M., and Chowienczyk, P.J. (1998). Haemodynamic effects of inhibition of nitric oxide synthase and of L-arginine at rest and during exercise. J. Hypertens. 16, 429&ndash;435.</p>
  • <p>Dhar A, Dhar I, Desai KM <em>et al.</em>&nbsp; Methylglyoxal scavengers attenuate endothelial dysfunction induced by methylglyoxal and high concentrations of glucose. (2010) Br. J. Pharmacol. 161: 1843&ndash;1856.</p>
  • <p>Dessy, C., Moniotte, S., Ghisdal, P., Havaux, X., Noirhomme, P., and Balligand, J.L. (2004). Endothelial beta3-adrenoceptors mediate vasorelaxation of human coronary microarteries through nitric oxide and endothelium-dependent hyperpolarization. Circulation 110, 948&ndash;954.</p>
  • <p>De Pascali F, Hemann C, Samons, K <em>et al.</em> Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (2014). 53 : 3679&ndash;3688.</p>
  • <p>Eugene, A.R. (2016). The influences of nitric oxide, epinephrine, and dopamine on vascular tone: dose-response modeling and simulations. Hosp. Chron. Nosokomeiaka Chron. 11, 1&ndash;8.</p>
  • <p>Du Y, Navab M, Shen M, <em>et al.</em>&nbsp; Ambient ultrafine particles reduce endothelial nitric oxide production via S-glutathionylation of eNOS. Biochem. Biophys. Res. Commun. (2013) 436 : 462&ndash;466.</p>
  • <p>Jackson, W.F. (2000). Ion channels and vascular tone. Hypertension 35, 173&ndash;178.</p>
  • <p>Peng H, Zhuang Y, Chen Y <em>et al.</em>&nbsp; The Characteristics and Regulatory Mechanisms of Superoxide Generation from eNOS Reductase Domain.&nbsp; (2015) PLoS One. 10(10):e0140365</p>
  • <p>Li, J., Zhou, Z., Jiang, D.-J., Li, D., Tan, B., Liu, H., and Li, Y.-J. (2007). Reduction of NO- and EDHF-mediated vasodilatation in hypertension: role of asymmetric dimethylarginine. Clin. Exp. Hypertens. N. Y. N 1993 29, 489&ndash;501.</p>
  • <p>Schuppe I, Mold&eacute;us P, and Cotgreave IA. Protein-specific S-thiolation in human endothelial cells during oxidative stress. (1992) Biochem. Pharmacol. 44: 1757&ndash;1764.</p>
  • <p>McVeigh, G.E., Allen, P.B., Morgan, D.R., Hanratty, C.G., and Silke, B. (2001). Nitric oxide modulation of blood vessel tone identified by arterial waveform analysis. Clin. Sci. Lond. Engl. 1979 100, 387&ndash;393.</p>
  • <p>Wu F, Szczepaniak WS, Shiva S <em>et al.</em>&nbsp; Nox2-dependent glutathionylation of endothelial NOS leads to uncoupled superoxide production and endothelial barrier dysfunction in acute lung injury.&nbsp; (2014) Am J Physiol Lung Cell Mol Physiol. 307(12):L987-97.</p>
  • <p>Modena MG,&nbsp;Bonetti L,&nbsp;Coppi F,&nbsp;Bursi F,&nbsp;Rossi R Prognostic&nbsp;role&nbsp;of&nbsp;reversible&nbsp;endothelial&nbsp;dysfunction&nbsp;in&nbsp;hypertensive&nbsp;postmenopausal&nbsp;women.&nbsp; J Am Coll Cardiol.&nbsp;2002, 40(3):505-10.</p>
  • <p>Nelson, M.T., Patlak, J.B., Worley, J.F., and Standen, N.B. (1990). Calcium channels, potassium channels, and voltage dependence of arterial smooth muscle tone. Am. J. Physiol. 259, C3&ndash;C18.</p>
  • <p>Paulis, L., Zicha, J., Kunes, J., Hojna, S., Behuliak, M., Celec, P., Kojsova, S., Pechanova, O., and Simko, F. (2008). Regression of L-NAME-induced hypertension: the role of nitric oxide and endothelium-derived constricting factor. Hypertens. Res. Off. J. Jpn. Soc. Hypertens. 31, 793&ndash;803.</p>
  • <p>Ras&nbsp;RT,&nbsp;Streppel MT,&nbsp;Draijer R,&nbsp;Zock PL.&nbsp; Flow-mediated dilation and cardiovascular risk prediction: a systematic review with meta-analysis.&nbsp; Int J Cardiol.&nbsp;2013, 168(1):344-51.</p>
  • <p>Siddiqui, A. (2011). Effects of Vasodilation and Arterial Resistance on Cardiac Output. J. Clin. Exp. Cardiol. 02.</p>
  • <p>Stamler, J.S., Loh, E., Roddy, M.A., Currie, K.E., and Creager, M.A. (1994). Nitric oxide regulates basal systemic and pulmonary vascular resistance in healthy humans. Circulation 89, 2035&ndash;2040.</p>
  • <p>Wilkinson, I.B., MacCallum, H., Cockcroft, J.R., and Webb, D.J. (2002). Inhibition of basal nitric oxide synthesis increases aortic augmentation index and pulse wave velocity in vivo. Br. J. Clin. Pharmacol. 53, 189&ndash;192.</p>
  • <p>Yeboah J, Crouse JR, Hsu FC, Burke GL, Herrington DM.&nbsp; Brachial flow-mediated dilation predicts incident cardiovascular events<br />
  • &nbsp;in older adults: the Cardiovascular Health Study. Circulation. 2007;115:2390&ndash;7.</p>
  • </div>
  • <br>
  • </div>
  • <!-- end relationship loop -->
  • <div>
  • <!-- graphical representation -->
  • <h2>Graphical Representation</h2>
  • <img src="https://aopwiki.org/system/dragonfly/production/2017/08/17/62rd12uxde_AOP_149_JPG.jpg" , height="500" , width="700"> </img>
  • <!-- end graphical representation -->
  • </div>
  • <!-- end summary -->
  • <h4><a href="/relationships/983">Relationship: 983: Increase, Vascular Resistance leads to Hypertension</a></h4>
  • <!-- Overall assessment section, *** what is included here? *** -->
  • <div id="overall_assessment">
  • <h2>Overall Assessment of the AOP</h2>
  • <hr>
  • <h4>AOPs Referencing Relationship</h4>
  • <h3>Domain of Applicability</h3>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td></td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Adjacency</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>adjacent</th>
  • <th>Moderate </th>
  • <th>Low</th>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <strong>Taxonomic Applicability</strong>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>human</td>
  • <td>Homo sapiens</td>
  • <td>Strong</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>mouse</td>
  • <td>Mus musculus</td>
  • <td>Moderate</td>
  • <td>Homo sapiens</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>rat</td>
  • <td>Rattus norvegicus</td>
  • <td>Moderate</td>
  • <td>Rattus norvegicus</td>
  • <td>Low</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>cow</td>
  • <td>Bos taurus</td>
  • <td>Moderate</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • <td>Adults</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <strong>Sex Applicability</strong>
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td></td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <p><a href="#Life_Stage_Applicability">Life Stage Applicability</a>, <a href="#Taxonomic_Applicability"> Taxonomic Applicability</a>, <a href="#Sex_Applicability"> Sex Applicability</a><br />
  • <em>Elaborate on the domains of applicability listed in the summary section above. Specifically, provide the literature supporting, or excluding, certain domains. </em></p>
  • <p>This proposed AOP is applicable to males and females, however a single study did suggest that KE responses may vary with sex (Scotland et al. 2005).&nbsp;</p>
  • <p>The AOP is relevant for all life stages, as vascular oxidative stress (a common cause of the MIE)&nbsp;can occur at all life stages. &nbsp;However, the AO via this mechanism is likely limited to adults, due to a loss of vaso-protective mechanisms with age (a major&nbsp;risk factor) and cardiovascular re-modelling over time. &nbsp;Hypertension is known to occur in adolescents and children, however the condition can be due to a variety of causes, such as renal disease and cardiovascular complications in early life.&nbsp; For example, persistant pulmonary hypertension is&nbsp;a consequence of failed pulmonary vascular transition at birth and leads to pulmonary&nbsp;hypertension&nbsp;with shunting of deoxygenated blood across the ductus arteriosus&nbsp;and foramen ovale,&nbsp;resulting in severe hypoxemia. &nbsp;The condition is improved by administration of nitric oxide by inhalation (Lai et al. 2017). &nbsp;While the initial cause of the hypertension is not vascular oxidative stress, ROS-mediated perturbations of the vascular endothelium, which arise as a result of the&nbsp;hypoxic conditions, contribute to the development of hypertension, and may have health implications for the neonate later in&nbsp;life (de Wijs-Meijler et al. 2017). &nbsp;Therefore, the physiology described in the AOP is relevant to the condition, but not strictly causative.</p>
  • <p>The KEs in this AOP are well-documented and well-studied in humans, cows and rodents. &nbsp;Similar outcomes are observed across these species following chemical exposure and other phenomena which cause vascular oxidative stress (e.g. hypoxia, ischaemia/reperfusion).</p>
  • <p>The specific amino-acid residues involved in post-translational modification of eNOS do differ across species, however the functional effect is similar. &nbsp;For example, human eNOS is phosphorylated by AKT at serine residue 1177&nbsp;(Reviewed by Heiss et al. 2014), whereas bovine eNOS is phosphoryated by AKT at serine 1179 (Chen et al. 2017).</p>
  • <h3>Essentiality of the Key Events</h3>
  • <p><a href="#Molecular_Initiating_Event">Molecular Initiating Event Summary</a>, <a href="#Key_Events"> Key Event Summary</a><br />
  • <em>Provide an overall assessment of the essentiality for the key events in the AOP. Support calls for individual key events can be included in the molecular initiating event, key event, and adverse outcome tables above. </em></p>
  • <p>The essentiality for this AOP is strong since there is direct evidence from multiple experimental studies showing that downstream key events can be prevented or inhibited if an upstream key event is blocked. An increase in intracellular glutathione, rather than the oxidized form, prevented S-glutathionylation (De Pascali et al., 2014), and inhibiting S-glutathionylation attenuated ultrafine particle-induced reduction in NO production (Du et al., 2013). In addition, increasing GTPCH-1, BH4 and Akt/eNOS activity increased eNOS activity and NO production but decreased superoxide generation (Fulton et al., 1999; Alp et al., 2003; Carnicer et al., 2012; Antoniades et al., 2011; Chen et al., 2011; De Pascali et al., 2014; Landmessar et al., 2003; Shinozaki et al., 2000; Ozaki et al., 2002). Infusion of NO donor sodium nitroprusside reduced vascular resistance, while sodium nitrite increased vasodilation (Eugene, 2016; Sindlier et al., 2014).</p>
  • <p>The essentiality of increased vascular resistance to hypertension is moderate due to the involvement of the heart in regulating blood pressure. &nbsp;While increased vascular resistance is a hallmark of essential hypertension, Mayet and Hughes (2003) discussed the roles of the heart and vascular resistance and concluded that both are of importance, and should not be viewed in isolation. &nbsp;Their conclusion is drawn from observations in younger adults with elevated blood pressure, who have a normal vascular resistance, but higher cardiac index. &nbsp;Over time, such adults often go on to change to a phenotype with a lower cardiac index and high vascular resistance and a diagnosis of chronic hypertension. &nbsp;Such changes are likely due to vascular remodelling processes with advancing age, however the specifics of this process are not well understood.</p>
  • <p>&nbsp;</p>
  • <p>&nbsp;</p>
  • <p>&nbsp;</p>
  • <p>&nbsp;</p>
  • <table border="0" cellpadding="0" cellspacing="0" style="width:1322px">
  • <tbody>
  • </tbody>
  • </table>
  • <table border="0" cellpadding="0" cellspacing="0" style="width:1132px">
  • <tbody>
  • <tr>
  • <td rowspan="2" style="height:140px; text-align:center; width:128px"><strong>Support for Essentiality of KEs</strong></td>
  • <td rowspan="2" style="text-align:center; width:203px"><strong>KE Description</strong></td>
  • <td style="text-align:center; width:148px"><strong>Defining Question</strong></td>
  • <td style="text-align:center; width:205px"><strong>High</strong></td>
  • <td style="text-align:center; width:203px"><strong>Moderate</strong></td>
  • <td style="text-align:center; width:245px"><strong>Low</strong></td>
  • </tr>
  • <tr>
  • <td style="height:108px; text-align:center; width:148px"><strong>Are downstream KEs and/or the AO prevented if an upstream KE is blocked?</strong></td>
  • <td style="text-align:center; width:205px"><strong>Direct evidence from experimental studies illustrating essentiality for at least one of the important KEs.</strong></td>
  • <td style="text-align:center; width:203px"><strong>Indirect evidence that sufficient modification of an expected modulating factor attenuates or augments a KE.</strong></td>
  • <td style="text-align:center; width:245px"><strong>No or contradictory experimental evidence of the essentiality of any of the KEs.</strong></td>
  • </tr>
  • <tr>
  • <td style="height:104px; width:128px">Peptide Oxidation (MIE)</td>
  • <td style="width:203px">Generation of localised ROS and other oxidative species in the vascular endothelium leads to oxidation of amino acid residues of peptides.&nbsp;&nbsp;</td>
  • <td style="width:148px">&nbsp;</td>
  • <td colspan="3" style="width:653px">&nbsp;</td>
  • </tr>
  • <tr>
  • <td style="height:125px; width:128px">S-glutathionylation of eNOS (KE1)</td>
  • <td style="width:203px">The oxidation of GSH into GSSG, elevates levels of GSSG, which then covalently bind to critical serine residues on the eNOS enzyme&nbsp;</td>
  • <td style="text-align:center; width:148px">High</td>
  • <td colspan="3" style="width:653px"><span style="font-size:14px">Protein S-Glutathionylation can only occur following covalent binding of GSSG.&nbsp; GSSG commonly binds to protein thiol residues, which can affect protein function&nbsp;(Bendall et al 2014).&nbsp;S-glutathionylation of eNOS is observed under conditions of oxidative stress, at cysteine residues 689 and 908 leading to eNOS uncoupling&nbsp;(Chen et al. 2010).&nbsp; Application of Glutaredoxin-1 or dithiothreitol removes S-glutathionylation and restores eNOS function&nbsp;(Chen et al. 2013, Jayaram et al. 2015) &nbsp;Essentiality rating is high due to direct experimental evidence showing removal of S-glutathionylation of eNOS restores NO production.</span></td>
  • </tr>
  • <tr>
  • <td style="height:105px; width:128px">Loss of GTPCH-1 (KE2)</td>
  • <td style="width:203px">Oxidative damage to GTPCH-1 has been shown to lead to reduced expression/activity of the enzyme</td>
  • <td style="text-align:center; width:148px">High</td>
  • <td colspan="3" style="width:653px">
  • <p>&nbsp;</p>
  • <p><span style="font-size:14px">Numerous studies have demonstrated that deletion of GTPCH-1 led to the deficiency of BH4 in bovine and murine endothelial cells (Tatham et al. 2009, Crabtree et al. 2009, Adlam et al. 2012)&nbsp;and in knockout mice&nbsp;(Chen et al. 2011).&nbsp; Gene silencing and GTPCH-1 inhibition lead to elevated blood pressure in experimental animals (Wang et al. 2008, Mitchell et al. 2003).&nbsp; Essentiality rating is high due to sunstantial experimental evidence of GTPCH-1 blocking on downstream KEs.</span></p>
  • </td>
  • </tr>
  • <tr>
  • <td style="height:100px; width:128px">Decreased BH4 (KE3)</td>
  • <td style="width:203px">Oxidation of BH4 itself to BH2, and/or reduced bioavailability of BH4 by a reduction in de novo synthesis by GTPCH-1.</td>
  • <td style="text-align:center; width:148px">High</td>
  • <td colspan="3" style="width:653px">
  • <p>&nbsp;</p>
  • <p><span style="font-size:14px">Depletion of BH4 has been extensively reported to uncouple eNOS, decrease NO bioavailability and increase superoxide production.&nbsp; Application of exogenous BH4 has been reported to reverse this phenomenon (De Pascali et al. 2014, Zweier et al. 2011, Dumitrescu et al. 2007, Tiefenbacher et al. 1996,&nbsp;V&aacute;squez-Vivar et al. 2002)&nbsp;&nbsp; Essentiality rating is high due to extensive direct experimental evidence.</span></p>
  • </td>
  • </tr>
  • <tr>
  • <td style="height:112px; width:128px">eNOS Uncoupling (KE4)</td>
  • <td style="width:203px">eNOS uncoupling is characterised by a loss of NO production and superoxide production as eNOS dimerisation is lost</td>
  • <td style="text-align:center; width:148px">High</td>
  • <td colspan="3" style="width:653px"><span style="font-size:14px">eNOS uncoupling leads to reduction in NO bioavailability.&nbsp; Reversal of eNOS uncoupling is achieved by supplementation with BH4 or removal of eNOS S-glutathionylation (as described earlier), which restores NO production (Du et al. 2013b,&nbsp;Nozik-Grayck et al. 2014,&nbsp;Talukder&nbsp;et al. 2011, Su et al. 2013).&nbsp; Essentiality rating is high due to direct experimental evidence.</span></td>
  • </tr>
  • <tr>
  • <td style="height:142px; width:128px">Loss of AKT/eNOS (KE5)</td>
  • <td style="width:203px">In humans, AKT phosphorylates eNOS at serine residue 1177, leading to eNOS activation and NO production.&nbsp; Loss of AKT/eNOS function depletes NO bioavailability.</td>
  • <td style="text-align:center">High</td>
  • <td colspan="3" style="width:653px">
  • <p><span style="font-size:14px">eNOS knockout mice are routinely used as models of hypertension&nbsp;(Huang&nbsp;2000).&nbsp; Aortic rings from eNOS knockout mice do not relax following application of acetylcholine, but do relax upon application of NO donor sodium nitroprusside (Huang et al. 1995).&nbsp; Many animal studies demonstrated that inhibition of NO via eNOS inhibitors impaired endothelium-dependent vasodilation (Li et al. 2007, Paulis et al. 2008, Luo et al. 2000,&nbsp;S&eacute;lley&nbsp;et al. 2014).&nbsp; Inhibition of Akt or mutant eNOS attenuated eNOS phosphorylation in human and bovine cells, resulting in decreased NO bioavailability (Michaud et al. 2006, Dhar et al. 2010, Dimmeler et al. 1999, Fulton et al. 1999, Das et al. 2014, Uruno et al. 2005).&nbsp; Essentiality rating is high due to extensive direct experimental evidence.</span></p>
  • </td>
  • </tr>
  • <tr>
  • <td style="height:187px; width:128px">NO Depletion (KE6)</td>
  • <td style="width:203px">Nitric oxide is a potent vasodilator radical released by endothelial (cell) eNOS.&nbsp; NO signalling leads to potassium ion efflux and hyperpolarization of vascular smooth muscle cells and blood vessel relaxation.&nbsp; Reduction in NO bioavailability blunts this response</td>
  • <td style="text-align:center">Moderate</td>
  • <td colspan="3" style="width:653px">
  • <p>&nbsp;</p>
  • <p><span style="font-size:14px">Depletion of NO bioavailability by pharmacological blockade of AKT and eNOS is widely reported to shift vascular tone towards a more vasoconstrictive phenotype, leading to hypertension (Li et al. 2007, Paulis et al. 2008, Scotland et al. 2005, Luo et al. 2000,&nbsp;S&eacute;lley&nbsp;et al. 2014, Haynes et al. 1993).&nbsp; However gender differences in experimental animals were reported to influence vasodilation by different mechanisms&nbsp;(Scotland et al. 2005).&nbsp; Furthermore, compensatory mechanisms affecting vascular tone are evident when bioavailability of NO is decreased (Brandes 2014, Durand et al. 2013).&nbsp;&nbsp; Essentiality rating is moderate due to extensive evidence from pharmacological blocking studies in humans and animals which show impairment of vasodilation and elevated BP.&nbsp; However, compensatory effects of other biological mechanisms on downstream KEs are evident as are gender differences in experimental animals.</span></p>
  • </td>
  • </tr>
  • <tr>
  • <td style="height:167px; width:128px">Impaired vasodilation (KE7)</td>
  • <td style="width:203px">
  • <p>&nbsp;</p>
  • <p>Vasodilation is mediated by vascular smooth muscle cells and characterized by potassium ion efflux and hyperpolarization of the tissue in response to signalling from the nervous system and chemical mediators released by the vascular endothelium</p>
  • </td>
  • <td style="text-align:center; width:148px">Moderate</td>
  • <td colspan="3" style="width:653px"><span style="font-size:14px">It is well accepted that vasodilation and systemic vascular resistance (SVR) are negatively correlated.&nbsp; Blood flow volume is increased when blood vessels dilate due to decreased vascular resistance (Siddiqui 2011). When vasodilation is impaired as a result of NO depletion or blockade of potassium channels, vascular stiffness and SVR increase (Berg et al. 2011, Brett et al. 1998, Dessey et al. 2004, Li et al. 2007, McVeigh et al. 2001, Paulis et al. 2008, Wilkinson et al. 2002).&nbsp; Essentiality rating is high due to extensive direct experimental evidence.</span></td>
  • </tr>
  • <tr>
  • <td style="height:116px; width:128px">Increased vascular resistance (KE8)</td>
  • <td style="width:203px">Increases in vascular tone as a result of impaired vasodilation.&nbsp; Commonly referred to as systemic vascular resistance (SVR) or total peripheral resistance (TPR).</td>
  • <td style="text-align:center; width:148px">Moderate</td>
  • <td colspan="3" style="width:653px"><span style="font-size:14px">It is well established that increased SVR (TPR), increased vascular stiffness and increased vascular reactivity contribute to hypertension (Brandes 2014,&nbsp;Fo&euml;x&nbsp;and Sear 2004, Mayet and Hughes 2003). In patients with hypertension, SVR was elevated in approximately 66% of enrolled patients (Chan et al. 2016).&nbsp; However, due to the critical role of cardiac output in blood pressure regulation (Brandes 2014, Mayet and Hughes 2003), and other compensatory mechanisms described earlier, the essentiality for the AO is moderate.</span></td>
  • </tr>
  • <tr>
  • <td style="height:89px; width:128px">Hypertension (AO)</td>
  • <td style="width:203px">Chronic elevation of systolic and/or diastolic blood pressure in systemic circulation or localised organs</td>
  • <td style="width:148px">&nbsp;</td>
  • <td colspan="3" style="width:653px">&nbsp;</td>
  • </tr>
  • </tbody>
  • </table>
  • <p>&nbsp;</p>
  • <h3>Weight of Evidence Summary</h3>
  • <p><a href="#Relationships_Among_Key_Events_and_the_Adverse_Outcome">Summary Table</a><br />
  • <em>Provide an overall summary of the weight of evidence based on the evaluations of the individual linkages from the Key Event Relationship pages. </em></p>
  • <p>Quantitative measurements with dose and time response data from published studies cited below can be found here: <a href="/wiki/index.php/File:Hypertension_Empirical_Support_Concordance_Table.pdf" title="File:Hypertension Empirical Support Concordance Table.pdf">File:Hypertension Empirical Support Concordance Table.pdf</a>.</p>
  • <table border="1" class="table">
  • <tbody>
  • <tr>
  • <th rowspan="2">Support for Biological Plausibility of KERs</th>
  • <th>Defining Question</th>
  • <th>High (Strong)</th>
  • <th>Moderate</th>
  • <th>Low (Weak)</th>
  • </tr>
  • <tr>
  • <td>Is there a mechanistic relationship between KEup and KEdown consistent with established biological knowledge?</td>
  • <td>Extensive understanding of the KER based on previous documentation and broad acceptance.</td>
  • <td>KER is plausible based on analogy to,accepted biological relationships, but scientific understanding is incomplete.</td>
  • <td>Empirical support for association between KEs, but the structural or functional relationship between them is not understood.</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to Glutathione, Oxidation:</td>
  • <td>Strong</td>
  • <td colspan="3">Multiple studies demonstrated that oxidative stress leads to the oxidation of glutathione (GSH) in the vascular endothelium. Exposure to a number of oxidants, including tert-butyl hydroperoxide, hydrogen peroxide, diamide, methylglyoxal, glucose, ischemia, and ultrafine particles caused a decrease in levels of GSH, which is indicative of its oxidation in human, bovine, and rat endothelial cells (De Pascali et al., 2014; Dhar et al., 2010; Du et al., 2013b; Montecinos et al., 2007; Park, 2013; Schuppe et al., 1992; van Gorp et al., 1999, 2002).</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to AKT/eNOS activity, Decrease</td>
  • <td>Strong</td>
  • <td colspan="3">Multiple experimental studies reported decreased Akt and eNOS phosphorylation/activity following oxidative stress as a consequence of exposure to peroxynitrite, high glucose, methylglyoxal, high fat, cigarette smoke extract (CSE) and ischemia in humans, bovine, mouse and rat endothelial cells (Das et al., 2014; Dhar et al., 2010; Du et al., 2001; Du et al., 2013; Michaud et al., 2006; Song et al., 2007, 2008; Su et al., 2013; Zhang et al., 2014; Zou et al., 2002).</td>
  • </tr>
  • <tr>
  • <td>AKT/eNOS activity, Decrease Directly Leads to Nitric Oxide, Depletion</td>
  • <td>Strong</td>
  • <td colspan="3">Several studies demonstrated that Akt can directly phosphorylate eNOS, leading to increased eNOS enzymatic activity and subsequent NO production (Dimmeler et al., 1999; Fulton et al., 1999). Inhibition of Akt or mutant eNOS attenuated eNOS phosphorylation in human and bovine cells, resulting in decreased NO bioavailability (Das et al., 2014; Dhar et al. 2010; Dimmeler et al., 1999; Fulton et al., 1999; Michaud et al., 2006; Uruno et al., 2005).</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to GTPCH-1, Decrease</td>
  • <td>Moderate</td>
  • <td colspan="3">Several studies demonstrated that GTPCH-1, the rate-limiting enzyme for BH4 synthesis, is affected by oxidative stress. GTPCH-1 expression or activity was inhibited by peroxynitrite and CSE (Abdelghany et al., 2017; Zhao et al., 2003). Cardiac reperfusion patients who experienced oxidative stress had reduced GTPCH-1 activity (Jayaram et al., 2015).</td>
  • </tr>
  • <tr>
  • <td>GTPCH-1, Decrease Directly Leads to Tetrahydrobiopterin, Decrease</td>
  • <td>Strong</td>
  • <td colspan="3">Many studies demonstrated that GTPCH-1 deletion led to reduced bioavailability of BH4 in bovine and murine endothelial cells (Adlam et al., 2012; Chen et al., 2011; Chuaiphichai et al., 2014; Crabtree et al., 2009; Tatham et al., 2009; Wang et al., 2008). Several studies also showed that overexpression of GTPCH-1 in human and mouse endothelium increased BH4 levels and eNOS activity (Alp et al., 2003; Antoniades et al., 2011; Carnicer et al., 2012).</td>
  • </tr>
  • <tr>
  • <td>Tetrahydrobiopterin, Decrease Directly Leads to eNOS, Uncoupling</td>
  • <td>Strong</td>
  • <td colspan="3">The depletion of BH4 leading to eNOS uncoupling is well-studied. Several studies showed reduced levels of BH4 induced eNOS uncoupling by reducing eNOS activity, decreasing NO and increasing superoxide levels in bovine, murine, and rat endothelium&nbsp;(Chuaiphichai et al., 2014; Crabtree et al., 2009; De Pascali et al., 2014; Dumitrescu et al., 2007; Whitsett et al., 2007). Many studies demonstrated that BH4 treatment improved endothelial function by reducing eNOS-mediated superoxide generation and increasing NO formation in human, bovine, mouse, and rat endothelium (Chen et al., 2011; De Pascali et al., 2014; Landmesser et al., 2003; Ozaki et al., 2002; Shinozaki et al., 2000; Wang et al., 2014).</td>
  • </tr>
  • <tr>
  • <td>Glutathione, Oxidation Directly Leads to eNOS, S-Glutathionylation</td>
  • <td>Strong</td>
  • <td colspan="3">Glutathione oxidation as determined by increased oxidized GSSG or decreased GSH levels caused S-glutathionylation of eNOS in bovine and human aortic endothelial cells, and in hypertensive rats and mice (Chen et al., 2010; De Pascali et al., 2014; Du et al., 2013).</td>
  • </tr>
  • <tr>
  • <td>eNOS, S-Glutathionylation Directly Leads to eNOS, Uncoupling</td>
  • <td>Strong</td>
  • <td colspan="3"><em>In vitro</em> experiments showed that S-glutathionylation of eNOS significantly decreased NO activity and greatly increased superoxide generation (Chen et al., 2010). These results were observed in bovine and human aortic endothelial cells as well as in vessels of spontaneously hypertensive rats and cardiac reperfusion patients (De Pascali et al., 2014; Du et al., 2013; Jayaram et al., 2015).</td>
  • </tr>
  • <tr>
  • <td>eNOS, Uncoupling Directly Leads to Nitric Oxide, Depletion</td>
  • <td>Strong</td>
  • <td colspan="3">It is well-established that uncoupling of eNOS causes eNOS to switch from producing NO to generating superoxides (F&ouml;rstermann and M&uuml;nzel, 2006). Studies reporting eNOS uncoupling as a result of BH4 depletion or S-glutathionylation measured levels of NO and superoxide which are indicative of eNOS uncoupling (Chen et al. 2010; De Pascali et al., 2014, Du et al., 2013; Whitsett et al., 2007).</td>
  • </tr>
  • <tr>
  • <td>Nitric Oxide, Depletion Directly Leads to Vasodilation, impaired</td>
  • <td>Strong</td>
  • <td colspan="3">Vasodilation is caused by the relaxation of vascular smooth muscle cells within the walls of blood vessels, and is regulated through a number of mechanisms, including cyclic GMP-dependent hyperpolarization and relaxation via NO. Thus, alterations to NO levels have an influence on vasodilation (Silva et al., 2012). Many animal studies demonstrated that inhibition of NO via eNOS inhibitors impaired endothelium-dependent vasodilation (Li et al., 2007; Luo et al., 2000; Paulis et al., 2008; S&eacute;lley et al., 2014).</td>
  • </tr>
  • <tr>
  • <td>Vasodilation, impaired Directly Leads to Vascular resistance, Increase</td>
  • <td>Strong</td>
  • <td colspan="3">It is well-accepted that vasodilation and systemic vascular resistance (SVR) are negatively correlated; blood flow is increased when blood vessels dilate due to decreased vascular resistance (Siddiqui, 2011). When vasodilation is impaired as a result of NO depletion or changes in potassium channels, vascular tone and SVR increase (Berg and Jensen, 2011; Brett et al., 1998; Dessy et al., 2004; Li et al., 2007; McVeigh et al., 2001; Paulis et al., 2008; Wilkinson et al., 2002).</td>
  • </tr>
  • <tr>
  • <td>Vascular resistance, Increase Directly Leads to Hypertension, N/A</td>
  • <td>Strong</td>
  • <td colspan="3">It is well-established that increased systemic vascular resistance (SVR), increased vascular stiffness and increased vascular reactivity contribute to hypertension (Brandes, 2014; Fo&euml;x and Sear, 2004; Mayet and Hughes, 2003). In patients with hypertension, SVR was elevated in about 66% of enrolled patients (Chan et al., 2016).</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <!-- end sex terms -->
  • <p>Studies supporting this key event relationship were performed in humans and rats.</p>
  • <h4>Key Event Relationship Description</h4>
  • <p>Hypertension is characterized partly by elevated systemic vascular resistance which is caused by alterations to vascular tone (towards vasoconstriction) over time&nbsp;(Lee and Griendling, 2008).&nbsp; As blood vessels constrict, the available volume in the vessel lumen for blood flow&nbsp;is restricted, resulting in elevated blood pressure.</p>
  • <p>&nbsp;</p>
  • <p>Note : The role of the heart in the maintenace (and change) of blood pressure over time is not part of this AOP, however it is of critical importance for the development of hypertension.</p>
  • <!-- if nothing shows up in any of these fields, then evidence supporting this KER will not be displayed -->
  • <h4>Evidence Supporting this KER</h4>
  • <strong>Biological Plausibility</strong>
  • <p>It is well-established that increased systemic vascular resistance (SVR), increased vascular stiffness and increased vascular reactivity contribute to the pathophysiology of hypertension (Fo&euml;x and Sear, 2004; Mayet and Hughes, 2003; Brandes et al., 2014); thus biological plausibility is strong for increased SVR leading to hypertension. This is observed in patients with hypertension (Chan et al., 2016).</p>
  • <strong>Empirical Evidence</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <table border="1" class="table">
  • <tbody>
  • <tr>
  • <th rowspan="2">Empirical Support for KERs</th>
  • <th>Defining Question</th>
  • <th>High (Strong)</th>
  • <th>Moderate</th>
  • <th>Low (Weak)</th>
  • </tr>
  • <tr>
  • <td>Does empirical evidence support that a change in KEup leads to an appropriate change in KEdown? Does KEup occur at lower doses, earlier time points, and higher in incidence than KEdown&nbsp;? Inconsistencies?</td>
  • <td>Multiple studies showing dependent change in both events following exposure to a wide range of specific stressors. No or few critical data gaps or conflicting data.</td>
  • <td>Demonstrated dependent change in both events following exposure to a small number of stressors. Some inconsistencies with expected pattern that can be explained by various factors.</td>
  • <td>Limited or no studies reporting dependent change in both events following exposure to a specific stressor; and/or significant inconsistencies in empirical support across taxa and species</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to Glutathione, Oxidation:</td>
  • <td>Moderate</td>
  • <td colspan="3">Many studies demonstrated a dose-dependent relationship between known inducers of oxidative stress (tert-butyl hydroperoxide, hydrogen peroxide, methylglyoxal, high glucose, and ultrafine particles) and reduced GSH levels in human and rat studies (Dhar et al., 2010; Du et al., 2013; Montecinos et al., 2007; Park et al., 2013; van Gorp et al., 1999).</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to AKT/eNOS activity, Decrease</td>
  • <td>Moderate</td>
  • <td colspan="3">eNOS activity and reactive oxygen species (ROS) were modulated in the opposite manner by several stressors (methylglyoxal, high glucose, SIN-1, hydrogen peroxide) in human and bovine endothelial cells, resulting in increased ROS and decreased eNOS activity (Das et al., 2014; Dhar et al., 2010; Chen et al., 2010).</td>
  • </tr>
  • <tr>
  • <td>AKT/eNOS activity, Decrease Directly Leads to Nitric Oxide, Depletion</td>
  • <td>Strong</td>
  • <td colspan="3">Various stress inducers (ischemia, peroxynitrite, SIN-1, insulin plus orotic acidura, etc.) showed that a decrease in AKT and/or eNOS activity led to increased eNOS uncoupling and decreased NO (Choi et al., 2014, 2015; Das et al., 2014; Dhar et al., 2010; Dumitrescu et al., 2007 Uruno et al., 2005).</td>
  • </tr>
  • <tr>
  • <td>Oxidative Stress, Increase Directly Leads to GTPCH-1, Decrease</td>
  • <td>Weak</td>
  • <td colspan="3">One study in a rat model of aortic coarctation-associated hypertension provides evidence that there is a interdependence between oxidative stress and GTPCH-1 with increased ROS and decreased GTPCH-1 expression following a perturbation, but there is no dose-response or temporal data (Cervantes-P&eacute;rez et al., 2012).</td>
  • </tr>
  • <tr>
  • <td>GTPCH-1, Decrease Directly Leads to Tetrahydrobiopterin, Decrease</td>
  • <td>Strong</td>
  • <td colspan="3">Exposure to a wide range of perturbations (e.g. CSE, 4-hydroxy-2-nonenal, cytokines) led to a decrease in both GTPCH-1 activity/expression and BH4 levels in human, cows and rats (Antoniades et al., 2011; Cervantes-P&eacute;rez et al., 2012; Chen et al., 2011; Ismail et al., 2015; Jayaram et al., 2015; Whitsett et al., 2007).</td>
  • </tr>
  • <tr>
  • <td>Tetrahydrobiopterin, Decrease Directly Leads to eNOS, Uncoupling</td>
  • <td>Strong</td>
  • <td colspan="3">Multiple studies demonstrated strong dependency between BH4 and eNOS uncoupling; decreased BH4 along with decreased eNOS activity, decreased NO production or increased superoxide generation were observed after various perturbations (Cervantes-P&eacute;rez et al., 2012; De Pascali et al., 2014; Dumitrescu et al., 2007; Jayaram et al., 2015; Whitsett et al., 2007; Wang et al., 2008).</td>
  • </tr>
  • <tr>
  • <td>Glutathione, Oxidation Directly Leads to eNOS, S-Glutathionylation</td>
  • <td>Moderate</td>
  • <td colspan="3">Treatment with GSSG induced a dose-dependent increase in human eNOS S-glutathionylation <em>in vitro</em> whereas 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) treatment also resulted in increased eNOS S-glutathionylation in a dose-dependent manner in bovine aortic endothelial cells (Chen et al., 2010). Exposure to ultrafine particles or hypoxia/reoxygenation in human or bovine aortic endothelial cells respectively, demonstrated a response-response relationship between reduced glutathione (GSH) and eNOS S-glutathionylation &nbsp;(Du et al., 2013; De Pascali et al., 2014).</td>
  • </tr>
  • <tr>
  • <td>eNOS, S-Glutathionylation Directly Leads to eNOS, Uncoupling</td>
  • <td>Moderate</td>
  • <td colspan="3">Treatment with BCNU resulted in increased eNOS S-glutathionylation, increased superoxide generation and decreased NO production in a dose-dependent manner in bovine aortic endothelial cells (Chen et al., 2010). Exposure to hypoxia/reoxygenation and treatment with angiotensin II demonstrated a response-response relationship between eNOS S-glutathionylation and superoxide generation in human and bovine endothelial cells (De Pascali et al., 2014; Galougahi et al., 2014).</td>
  • </tr>
  • <tr>
  • <td>eNOS, Uncoupling Directly Leads to Nitric Oxide, Depletion</td>
  • <td>Strong</td>
  • <td colspan="3">Multiple experiments demonstrated that eNOS uncoupling results in increased superoxide formation and decreased NO production (Chen et al., 2010; De Pascali et al., 2014; Dumitrescu et al., 2007; Wang et al., 2008; Whitsett et al., 2007; Zou et al., 2002).</td>
  • </tr>
  • <tr>
  • <td>Nitric Oxide, Depletion Directly Leads to Vasodilation, impaired</td>
  • <td>Moderate</td>
  • <td colspan="3">Treatment with eNOS inhibitors and two other stressors, BCNU and diaminohydroxypyrimidine (DAHP), caused a decrease in both NO production and vasodilation (Chen et al., 2010; Paulis et al, 2008; Wang et al., 2008).</td>
  • </tr>
  • <tr>
  • <td>Vasodilation, impaired Directly Leads to Vascular resistance, Increase</td>
  • <td>Weak</td>
  • <td colspan="3">No direct evidence was found for this KER, but there is indirect support. Treatment with eNOS inhibitor L-NG-monomethyl arginine citrate (L-NMMA) caused an increase in SVR and a reduction in NO (Stamler et al., 1994), while L-NG-nitroarginine methyl ester (L-NAME) decreased NO-dependent relaxation and increased blood pressure (Paulis et al, 2008). Infusion of NO donor sodium nitroprusside led to dose-dependent reductions in SVR (Eugene, 2016).</td>
  • </tr>
  • <tr>
  • <td>Vascular resistance, Increase Directly Leads to Hypertension, N/A</td>
  • <td>Moderate</td>
  • <td colspan="3">Several human studies showed a dose-dependent change in SVR and hypertension following treatment with eNOS inhibitors (Brett et al., 1998; Haynes et al., 1993; McVeigh et al., 2001; Stamler et al., 1994; Wilkinson et al., 2002).</td>
  • </tr>
  • </tbody>
  • </table>
  • <p>Empirical support for SVR leading to hypertension is moderate based on several human studies showing a dose-dependent change in SVR and hypertension following treatment with eNOS inhibitors L-NMMA and L-NAME.&nbsp; Whilst the acute changes in SVR on blood pressure in the context of endothelial NO production is well characterised, the linkage between chronically elevated SVR and hypertension is more complicated, due to the roles of the heart, nervous system and kidneys.&nbsp; Given that this AOP is focused on endothelial NO production, other AOPs are required to capture the roles of these other important biological processes on chronic SVR changes and hypertension, hence the rating of &quot;moderate&quot;.</p>
  • <p>Intravenous infusion of L-NMMA (0.1, 1.0, 3.0 mg/kg/min) for 15 minutes led to a dose-dependent increase in SVR and mean arterial pressure compared to saline infusion in eight healthy men (Wilkinson et al., 2002). SVR increased from 31&plusmn;2 (arbitrary units) at baseline to 57&plusmn;4 at the highest dose, while mean arterial pressure increased from 86&plusmn;2 mmHg baseline to 91&plusmn;2 mmHg at the highest concentration. Infusion of L-NMMA (1.0 mg/kg/min) caused an increase in systolic blood pressure by 15% and SVR by 63% in 11 healthy volunteers after three minutes (Stamler et al., 1994). The study also demonstrated a dose-dependent increase in both events with five different doses of L-NMMA (0.01, 0.03, 0.1, 0.3 and 1 mg/kg/min). In another study,&nbsp;fifteen healthy men were&nbsp;intravenously infused with L-NAME &nbsp;at lower doses (0.25-0.75 mg/kg) for eight minutes, resulting in increased arterial pressure and SVR (McVeigh et al., 2001). Infusion with L-arginine, an eNOS substrate, restored function, specifically SVR and small artery compliance to baseline. L-NMMA (3 mg/kg) for five minutes increased mean arterial pressure by 10% and increased SVR by 46% in eight healthy subjects (Haynes et al., 1993). One experiment in rats showed that L-NAME increased SVR, aortic stiffness, and blood pressure (Nakmareong et al., 2012). Overall, these studies demonstrated that SVR and hypertension are modulated together by eNOS inhibitors.</p>
  • <h3>Quantitative Consideration</h3>
  • <p><a href="#Relationships_Among_Key_Events_and_the_Adverse_Outcome">Summary Table</a><br />
  • <em>Provide an overall discussion of the quantitative information available for this AOP. Support calls for the individual relationships can be included in the Key Event Relationship table above. </em></p>
  • <strong>Uncertainties and Inconsistencies</strong>
  • <p>One study showed that infusion of L-NMMA (6 mg/kg) resulted in increased SVR and only a modest increase in blood pressure.&nbsp; Changes in diastolic blood pressure were observed to be&nbsp;more pronounced in healthy men, than systolic blood pressure (Brett et al., 1998), and infusion of L-arginine (an eNOS substrate) had no significant effect.</p>
  • <p>Overall, there is a good amount of quantitative data available for this AOP as demonstrated by the weight of evidence tables, which shows that empirical support ranges from weak to strong. Four key event relationships (KERs) have strong quantitative support: decreased AKT/eNOS activity =&gt; NO depletion, decreased GTPCH-1 =&gt; decreased BH4, decreased BH4 =&gt; eNOS uncoupling and eNOS uncoupling =&gt; NO depletion. The relationships between these key events are well described in the literature as GTPCH-1 is the rate-limiting enzyme for BH4 synthesis (Wang et al., 2008), BH4 is an essential cofactor for eNOS and function (Wang et al., 2014), and the uncoupling of eNOS causes it to generate superoxide instead of NO (Carnicer et al., 2012). As they are functionally interconnected, many studies measure these key events together; thus providing strong support for their dependency.</p>
  • <p>As mentioned above, other AOPs are necessary to capture understanding and assess the evidence surrounding the roles of the heart, kidney and nervous system in order to get the full picture of the linkage between chronic changes in SVR and hypertension.</p>
  • <!--<!% unless aop_rel.relationship.relationship_taxons.blank? %>-->
  • <!--<!%= render 'snapshot_taxons', taxons: aop_rel.relationship.relationship_taxons %>-->
  • <!--<!% unless aop_rel.relationship.taxon_evidence.blank? %>-->
  • <!--<h3>Domain of Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
  • <!--<!% end %>-->
  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>Brandes, R.P. (2014). Endothelial dysfunction and hypertension. Hypertension 64, 924&ndash;928.</p>
  • <p>Two key events have limited quantitative support (oxidative stress =&gt; decreased GTPCH-1, impaired vasodilation =&gt; increased vascular resistance). Generally, the oxidation of BH4 rather than decreased GTPCH-1 is measured when cells are under oxidative stress, and ROS are assumed to be increased so no quantitative measures are taken. For vasodilation and vascular resistance, there appears to be a correlative relationship, where increased vasodilation would mean decreased vascular resistance and vice versa, so studies do not measure both key events. Several studies showed that treatment with eNOS inhibitors led to increased vascular resistance, suggesting impaired vasodilation (Li et al., 2007; McVeigh et al., 2001; Paulis et al., 2008; Wilkinson et al., 2002).</p>
  • <p>Brett, S.E., Cockcroft, J.R., Mant, T.G., Ritter, J.M., and Chowienczyk, P.J. (1998). Haemodynamic effects of inhibition of nitric oxide synthase and of L-arginine at rest and during exercise. J. Hypertens. 16, 429&ndash;435.</p>
  • <p>The other KERs have moderate quantitative support, meaning there were studies showing a dependent change in both key events following treatment with stressors. Several studies measured decreased NO and vasodilation following perturbations to eNOS inhibitors, BCNU and DAHP (Chen et al., 2010; Paulis et al, 2008; Wang et al., 2008).</p>
  • <p>Chan, S.S., Tse, M.M., Chan, C.P., Tai, M.C., Graham, C.A., and Rainer, T.H. (2016). Haemodynamic changes in emergency department patients with poorly controlled hypertension. Hong Kong Med. J. Xianggang Yi Xue Za Zhi Hong Kong Acad. Med. 22, 116&ndash;123.</p>
  • <p>In general, experiments with both dose-dependent and temporal response data following a stressor are not readily available for all KERs as most measurements are generally taken at one time point after a perturbation, or the measurements are for one key event, not both. However, an exhaustive literature search was not performed. An ideal experiment would be to treat cells with three to four stressors at increasing concentrations and measure the key events at different time intervals; thus providing a greater understanding of the temporal and dose-dependent responses between the key events.</p>
  • <p>Fo&euml;x, P., and Sear, J.W. (2004). Hypertension: pathophysiology and treatment. Contin. Educ. Anaesth. Crit. Care Pain 4, 71&ndash;75.</p>
  • <p>Haynes, W.G., Noon, J.P., Walker, B.R., and Webb, D.J. (1993). Inhibition of nitric oxide synthesis increases blood pressure in healthy humans. J. Hypertens. 11, 1375&ndash;1380.</p>
  • <p>Lee, M.Y., and Griendling, K.K. (2008). Redox signaling, vascular function, and hypertension. Antioxid. Redox Signal. 10, 1045&ndash;1059.</p>
  • <p>Mayet, J., and Hughes, A. (2003). Cardiac and vascular pathophysiology in hypertension. Heart Br. Card. Soc. 89, 1104&ndash;1109.</p>
  • <p>McVeigh, G.E., Allen, P.B., Morgan, D.R., Hanratty, C.G., and Silke, B. (2001). Nitric oxide modulation of blood vessel tone identified by arterial waveform analysis. Clin. Sci. Lond. Engl. 1979 100, 387&ndash;393.</p>
  • <p>Nakmareong, S., Kukongviriyapan, U., Pakdeechote, P., Kukongviriyapan, V., Kongyingyoes, B., Donpunha, W., Prachaney, P., and Phisalaphong, C. (2012). Tetrahydrocurcumin alleviates hypertension, aortic stiffening and oxidative stress in rats with nitric oxide deficiency. Hypertens. Res. Off. J. Jpn. Soc. Hypertens. 35, 418&ndash;425.</p>
  • <p>Silva, B.R., Pernomian, L., and Bendhack, L.M. (2012). Contribution of oxidative stress to endothelial dysfunction in hypertension. Front. Physiol. 3, 441.</p>
  • <p>Stamler, J.S., Loh, E., Roddy, M.A., Currie, K.E., and Creager, M.A. (1994). Nitric oxide regulates basal systemic and pulmonary vascular resistance in healthy humans. Circulation 89, 2035&ndash;2040.</p>
  • <p>Wilkinson, I.B., MacCallum, H., Cockcroft, J.R., and Webb, D.J. (2002). Inhibition of basal nitric oxide synthesis increases aortic augmentation index and pulse wave velocity in vivo. Br. J. Clin. Pharmacol. 53, 189&ndash;192.</p>
  • </div>
  • <br>
  • <div>
  • <h4><a href="/relationships/988">Relationship: 988: Decrease, AKT/eNOS activity leads to Depletion, Nitric Oxide</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Adjacency</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>adjacent</th>
  • <th>High </th>
  • <th>High</th>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>The relationship between decreased AKT/eNOS activity and NO depletion is supported by studies performed in humans, cows, and rats.</p>
  • <h4>Key Event Relationship Description</h4>
  • <p>AKT can phosphorylate eNOS which leads to increased eNOS enzymatic activity and subsequent NO production (Dimmeler et al., 1999; Fulton et al., 1999). Inhibition of AKT&nbsp;attenuates eNOS phosphorylation and its activity, resulting in decreased NO bioavailability and endothelial dysfunction.</p>
  • </div>
  • <!-- potential consierations, text as entered by author -->
  • <div id="considerations_for_potential_applicaitons">
  • <h2>Considerations for Potential Applications of the AOP (optional)</h2>
  • <hr>
  • <p>One of the most widely reported hypertension risk factors is tobacco smoking.&nbsp; While smoking cessation remains the best way to reduce the harmful effects of tobacco smoking, tobacco harm reduction is being considered by some regulators (e.g. US Food and Drug Administration; FDA) as a complementary strategy to reduce smoking-related disease burden.&nbsp; The US FDA has published guidance on assessing a &ldquo;modified risk tobacco product&rdquo; (MRTP), either through demonstration of reduced toxicant exposure or reduction in health risks<span style="font-size:10.8333px">&nbsp;<span style="font-family:arial,helvetica,sans-serif">(FDA 2012)</span></span>. By gaining an understanding of how, and to what extent tobacco smoke initiates biological mechanisms of hypertension, such knowledge could be utilised as a baseline for comparison purposes in toxicological assessments of the risk reduction potential of e-cigarettes.&nbsp;</p>
  • <p>Given the wide variety of data requirements to support such risk assessments e.g. human exposure studies, behavioural studies, efficacy studies, <em>in vitro</em> studies, clinical biomarker studies, pharmacokinetic studies, quality of life studies etc., a data integration framework is required to organise these data types into a comprehensive story that (i) characterises the toxicological problem, (ii) demonstrates the likely outcome of an intervention, and (iii) can be utilised to monitor the performance of any intervention over time.&nbsp; Adverse outcome pathways offer scientists and regulators a way to inform future Integrated Approaches to Testing and Assessment (IATA) for the risk assessment/harm reduction potential of electronic cigarettes and heated tobacco products.</p>
  • <p>&nbsp;</p>
  • <p>Other applications could include informing risk assessments for long term exposure to airborne pollution, in the context of helping to set acceptable exposure limits in urban environments for specific pollutants e.g. diesel exhuast particles.&nbsp; Furthermore, health supplements which purport to benefit the cardiovascular system in the context of&nbsp;hypertension risk, could also be assessed for efficacy.</p>
  • <!-- if nothing shows up in any of these fields, then evidence supporting this KER will not be displayed -->
  • <h4>Evidence Supporting this KER</h4>
  • </div>
  • <strong>Biological Plausibility</strong>
  • <p>Two studies demonstrated that AKT&nbsp;can directly phosphorylate eNOS at Ser1177/Ser1179, leading to increased eNOS enzymatic activity and subsequent NO production in COS cells (Dimmeler et al., 1999; Fulton et al., 1999). Expressing active AKT&nbsp;in bovine microvascular endothelial cells also induced NO release (Fulton et al., 1999). Inhibition of Akt or a mutation of eNOS at AKT-sensitive sites attenuated eNOS phosphorylation and its activity, resulting in decreased NO bioavailability and endothelial dysfunction (Dimmeler et al., 1999; Fulton et al., 1999; Uruno et al., 2005). Treatment of BAECs with SIN-1 resulted in a reduction in both eNOS activity and NO production (Das et al., 2014). Cigarette smoke extract (CSE) treatment inhibited AKT&nbsp;and eNOS and NO release in VEGF-stimulated HUVECs (Michaud et al., 2006), and methylglyoxal and high glucose reduced eNOS bradykinin-stimulated eNOS activity and NO production in HUVECs (Dhar et al., 2010). Overall, the biological plausibility for decreased AKT/eNOS activity leading to NO depletion is strong.</p>
  • <!-- reference section, text as of right now but should be changed to be handled as table -->
  • <div id="references">
  • <h2>References</h2>
  • <hr>
  • <p>AbdelGhany, T., Ismail, R., Elmahdy, M., Mansoor F, Zweier J, Lowe, F., and Zweier, JL. (2017). Cigarette Smoke Constituents Cause Endothelial Nitric Oxide Synthase Dysfunction and Uncoupling due to Depletion of Tetrahydrobiopterin with Degradation of GTP Cyclohydrolase.&nbsp;&nbsp;Nitric Oxide&nbsp;(Under review).</p>
  • <p>Adlam, D., Herring, N., Douglas, G., De Bono, J.P., Li, D., Danson, E.J., Tatham, A., Lu, C.-J., Jennings, K.A., Cragg, S.J., et al. (2012). Regulation of &beta;-adrenergic control of heart rate by GTP-cyclohydrolase 1 (GCH1) and tetrahydrobiopterin. Cardiovasc. Res. 93, 694&ndash;701.</p>
  • <p>Alp, N.J., Mussa, S., Khoo, J., Cai, S., Guzik, T., Jefferson, A., Goh, N., Rockett, K.A., and Channon, K.M. (2003). Tetrahydrobiopterin-dependent preservation of nitric oxide-mediated endothelial function in diabetes by targeted transgenic GTP-cyclohydrolase I overexpression. J. Clin. Invest. 112, 725&ndash;735.</p>
  • <p>Antoniades, C., Cunnington, C., Antonopoulos, A., Neville, M., Margaritis, M., Demosthenous, M., Bendall, J., Hale, A., Cerrato, R., Tousoulis, D., et al. (2011). Induction of vascular GTP-cyclohydrolase I and endogenous tetrahydrobiopterin synthesis protect against inflammation-induced endothelial dysfunction in human atherosclerosis. Circulation 124, 1860&ndash;1870.</p>
  • <p>Bendall JK, Douglas G, McNeill E <em>et al.</em>&nbsp; Tetrahydrobiopterin in cardiovascular health and disease.&nbsp; Antioxid Redox Signal. (2014) 20(18):3040-77.</p>
  • <p>Berg, T., and Jensen, J. (2011). Simultaneous parasympathetic and sympathetic activation reveals altered autonomic control of heart rate, vascular tension, and epinephrine release in anesthetized hypertensive rats. Front. Neurol. 2, 71.</p>
  • <p>Berlett BS,&nbsp;Stadtman ER. Protein&nbsp;oxidation&nbsp;in&nbsp;aging,&nbsp;disease, and&nbsp;oxidative stress.&nbsp;&nbsp;J Biol Chem.&nbsp;1997, 272(33):20313-6.</p>
  • <p>Bhatnagar, A., Whitsel, L.P., Ribisl, K.M., Bullen, C., Chaloupka, F., Piano, M.R., Robertson, R.M., McAuley, T., Goff, D., Benowitz, N., et al. (2014). Electronic cigarettes: a policy statement from the American Heart Association. Circulation 130, 1418&ndash;1436.</p>
  • <p>Brandes, R.P. (2014). Endothelial dysfunction and hypertension. Hypertension 64, 924&ndash;928.</p>
  • <p>Brett, S.E., Cockcroft, J.R., Mant, T.G., Ritter, J.M., and Chowienczyk, P.J. (1998). Haemodynamic effects of inhibition of nitric oxide synthase and of L-arginine at rest and during exercise. J. Hypertens. 16, 429&ndash;435.</p>
  • <strong>Empirical Evidence</strong>
  • <p><em>Include consideration of temporal concordance here </em></p>
  • <p>Carnicer, R., Hale, A.B., Suffredini, S., Liu, X., Reilly, S., Zhang, M.H., Surdo, N.C., Bendall, J.K., Crabtree, M.J., Lim, G.B.S., et al. (2012). Cardiomyocyte GTP cyclohydrolase 1 and tetrahydrobiopterin increase NOS1 activity and accelerate myocardial relaxation. Circ. Res. 111, 718&ndash;727.</p>
  • <p>Empirical support for this linkage is strong since various stressors (ischemia, peroxynitrite, SIN-1, insulin+orotic acidura, etc.) can cause a decrease in AKT&nbsp;or eNOS activity which then leads to increased eNOS uncoupling and decreased NO levels.</p>
  • <p>Cervantes-P&eacute;rez, L.G., Ibarra-Lara, M. de la L., Escalante, B., Del Valle-Mondrag&oacute;n, L., Vargas-Robles, H., P&eacute;rez-Severiano, F., Pastel&iacute;n, G., and S&aacute;nchez-Mendoza, M.A. (2012). Endothelial nitric oxide synthase impairment is restored by clofibrate treatment in an animal model of hypertension. Eur. J. Pharmacol. 685, 108&ndash;115.</p>
  • <p>Treatment of HUVECs with 30 &mu;M methylglyoxal (MG) and 25 mM high glucose (HG) concentrations for 24 hours caused a decrease in eNOS activity (control: 100%, MG: 55%, HG: 66%), while a shorter treatment for 3 hours caused a decrease in NO production (control: 20 &mu;M, MG: 12 &mu;M, HG: 16 &mu;M) (Dhar et al., 2010).</p>
  • <p>Chan, S.S., Tse, M.M., Chan, C.P., Tai, M.C., Graham, C.A., and Rainer, T.H. (2016). Haemodynamic changes in emergency department patients with poorly controlled hypertension. Hong Kong Med. J. Xianggang Yi Xue Za Zhi Hong Kong Acad. Med. 22, 116&ndash;123.</p>
  • <p>Treatment of BAECs with SIN-1 for 2 hours decreased eNOS activity from 100% to 70% and NO production from 100% to 53% (Das et al., 2014).</p>
  • <p>Chen, C.-A., Wang, T.-Y., Varadharaj, S., Reyes, L.A., Hemann, C., Talukder, M.A.H., Chen, Y.-R., Druhan, L.J., and Zweier, J.L. (2010a). S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature 468, 1115&ndash;1118.</p>
  • <p>In HUVECs, exposure to 5 &mu;mol/L peroxynitrite for 5 minutes reduced AKT&nbsp;phosphorylation by 30% (Song et al., 2007) while exposure to 50 &mu;mol/L peroxynitrite for 3 hours reduced NO production from 90% to 6.5% (Zou et al., 2002), demonstrating that AKT&nbsp;phosphorylation occurs before NO production at a lower dose.</p>
  • <p>Chen, C.-A., Lin, C.-H., Druhan, L.J., Wang, T.-Y., Chen, Y.-R., and Zweier, J.L. (2011). Superoxide induces endothelial nitric-oxide synthase protein thiyl radical formation, a novel mechanism regulating eNOS function and coupling. J. Biol. Chem. 286, 29098&ndash;29107.</p>
  • <p>Prolonged myocardial ischemia (&gt;30 min) in isolated rat hearts caused decreased eNOS activity (58% reduction) and increased superoxide generation (from relative fluorescence unit of &lt;0.01 to 0.3), suggesting a depletion in NO production (Dumitrescu et al., 2007).</p>
  • <p>Chen CA, De Pascali F, Basye A <em>et al.</em> Redox modulation of endothelial nitric oxide synthase by glutaredoxin-1 through reversible oxidative post-translational modification.&nbsp; (2013)&nbsp; Biochemistry. 52(38):6712-23</p>
  • <p>Treatment of HUVECs with 50 &mu;M insulin for 30 minutes increased phosphorylation of AKT&nbsp;(control: 1, insulin: 5.7) and eNOS (control: 1, insulin: 1.9) as well as NO production (control: 100%, insulin: 157%), but these effects were reversed by additional treatment with 100 &mu;M orotic aciduria (OA) for one hour (Akt: 1.8; eNOS: 0.28; NO: 106%) (Choi et al., 2015). Similar results were observed with addition of uric acid to insulin-treated HUVECs (Choi et al., 2014).</p>
  • <p>Chen Y,&nbsp;Jiang B,&nbsp;Zhuang Y,&nbsp;Peng H,&nbsp;Chen W. &nbsp;Differential effects of heat shock protein 90 and serine 1179&nbsp;phosphorylation&nbsp;on endothelial nitric oxide synthase activity and on its cofactors. &nbsp;PLoS One.&nbsp;2017 12(6):e0179978.</p>
  • <p>Treatment with all-trans retinoic acid (ATRA) and retinoic acid receptor agonist Am580 at 1 &mu;mol/L for 48 hours also increased AKT&nbsp;(control: 100%, ATRA: 304%, Am580: 212%) and eNOS (control: 100%, ATRA: 276%, Am580: 209%) phosphorylation and NO production (control: 100%, ATRA: 170%, Am580: 169%) in human endothelial cells (Uruno et al., 2005).</p>
  • <strong>Uncertainties and Inconsistencies</strong>
  • <p>In the context of this AOP, decreased activity of AKT is likely due to proteasomal degradation following exposure to a (oxidative) stressor (Abdehghany et al. 2017).&nbsp; However, decreased eNOS activity could be due to multiple causes, highlighted in this AOP.&nbsp; Firstly, if AKT expression levels are reduced, it follows that eNOS phosphorylation will be reduced.&nbsp; Secondly, similar to AKT, eNOS itself has been shown to be susceptible to proteasomal degradation (Abdehghany et al. 2017).&nbsp; Thirdly, depletion of BH4 and/or S-glutathionylation has been shown to uncouple eNOS, leading to reduced NO levels and increased superoxide levels.&nbsp; The relative contribution of&nbsp;each of these eNOS perturbation routes to NO depletion is currently unknown.</p>
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  • <!--<!% unless aop_rel.relationship.taxon_evidence.blank? %>-->
  • <!--<h3>Domain of Applicability</h3>-->
  • <!--<!%== aop_rel.relationship.taxon_evidence %>-->
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  • <!--<!% end %>-->
  • <h4>References</h4>
  • <p>AbdelGhany, T., Ismail, R., Elmahdy, M., Mansoor F, Zweier J, Lowe, F., and Zweier, JL. (2017). Cigarette Smoke Constituents Cause Endothelial Nitric Oxide Synthase Dysfunction and Uncoupling due to Depletion of Tetrahydrobiopterin with Degradation of GTP Cyclohydrolase.&nbsp;&nbsp;Nitric Oxide&nbsp;(Under review).</p>
  • <p>Choi, Y.-J., Yoon, Y., Lee, K.-Y., Hien, T.T., Kang, K.W., Kim, K.-C., Lee, J., Lee, M.-Y., Lee, S.M., Kang, D.-H., et al. (2014). Uric acid induces endothelial dysfunction by vascular insulin resistance associated with the impairment of nitric oxide synthesis. FASEB J. 28, 3197&ndash;3204.</p>
  • <p>Choi, Y.-J., Yoon, Y., Lee, K.-Y., Kang, Y.-P., Lim, D.K., Kwon, S.W., Kang, K.-W., Lee, S.-M., and Lee, B.-H. (2015). Orotic Acid Induces Hypertension Associated with Impaired Endothelial Nitric Oxide Synthesis. Toxicol. Sci. 144, 307&ndash;317.</p>
  • <p>Chuaiphichai, S., McNeill, E., Douglas, G., Crabtree, M.J., Bendall, J.K., Hale, A.B., Alp, N.J., and Channon, K.M. (2014). Cell-autonomous role of endothelial GTP cyclohydrolase 1 and tetrahydrobiopterin in blood pressure regulation. Hypertension 64, 530&ndash;540.</p>
  • <p>Crabtree, M.J., Tatham, A.L., Al-Wakeel, Y., Warrick, N., Hale, A.B., Cai, S., Channon, K.M., and Alp, N.J. (2009). Quantitative regulation of intracellular endothelial nitric-oxide synthase (eNOS) coupling by both tetrahydrobiopterin-eNOS stoichiometry and biopterin redox status: insights from cells with tet-regulated GTP cyclohydrolase I expression. J. Biol. Chem. 284, 1136&ndash;1144.</p>
  • <p>Das, A., Gopalakrishnan, B., Druhan, L.J., Wang, T.-Y., De Pascali, F., Rockenbauer, A., Racoma, I., Varadharaj, S., Zweier, J.L., Cardounel, A.J., et al. (2014). Reversal of SIN-1-induced eNOS dysfunction by the spin trap, DMPO, in bovine aortic endothelial cells via eNOS phosphorylation. Br. J. Pharmacol. 171, 2321&ndash;2334.</p>
  • <p>De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679&ndash;3688.</p>
  • <p>Dessy, C., Moniotte, S., Ghisdal, P., Havaux, X., Noirhomme, P., and Balligand, J.L. (2004). Endothelial beta3-adrenoceptors mediate vasorelaxation of human coronary microarteries through nitric oxide and endothelium-dependent hyperpolarization. Circulation 110, 948&ndash;954.</p>
  • <p>Dhar, A., Dhar, I., Desai, K.M., and Wu, L. (2010). Methylglyoxal scavengers attenuate endothelial dysfunction induced by methylglyoxal and high concentrations of glucose. Br. J. Pharmacol. 161, 1843&ndash;1856.</p>
  • <p>Dharmashankar, K., and Widlansky, M.E. (2010). Vascular endothelial function and hypertension: insights and directions. Curr. Hypertens. Rep. 12, 448&ndash;455.</p>
  • <p>Dimmeler, S., Fleming, I., Fisslthaler, B., Hermann, C., Busse, R., and Zeiher, A.M. (1999). Activation of nitric oxide synthase in endothelial cells by Akt-dependent phosphorylation. Nature 399, 601&ndash;605.</p>
  • <p>Du, J., Fan, L.M., Mai, A., and Li, J.-M. (2013). Crucial roles of Nox2-derived oxidative stress in deteriorating the function of insulin receptors and endothelium in dietary obesity of middle-aged mice. Br. J. Pharmacol. 170, 1064&ndash;1077.</p>
  • <p>Du, X.L., Edelstein, D., Dimmeler, S., Ju, Q., Sui, C., and Brownlee, M. (2001). Hyperglycemia inhibits endothelial nitric oxide synthase activity by posttranslational modification at the Akt site. J. Clin. Invest. 108, 1341&ndash;1348.</p>
  • <p>Du, Y., Navab, M., Shen, M., Hill, J., Pakbin, P., Sioutas, C., Hsiai, T.K., and Li, R. (2013b). Ambient ultrafine particles reduce endothelial nitric oxide production via S-glutathionylation of eNOS. Biochem. Biophys. Res. Commun. 436, 462&ndash;466.</p>
  • <p>Dumitrescu, C., Biondi, R., Xia, Y., Cardounel, A.J., Druhan, L.J., Ambrosio, G., and Zweier, J.L. (2007). Myocardial ischemia results in tetrahydrobiopterin (BH4) oxidation with impaired endothelial function ameliorated by BH4. Proc. Natl. Acad. Sci. U. S. A. 104, 15081&ndash;15086.</p>
  • <p>Durand MJ, Gutterman DD.&nbsp; Diversity in mechanisms of endothelium-dependent vasodilation in health and disease.&nbsp; Microcirculation. (2013) 20(3):239-47</p>
  • <p>Eugene, A.R. (2016). The influences of nitric oxide, epinephrine, and dopamine on vascular tone: dose-response modeling and simulations. Hosp. Chron. Nosokomeiaka Chron. 11, 1&ndash;8.</p>
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  • <p>Fo&euml;x, P., and Sear, J.W. (2004). Hypertension: pathophysiology and treatment. Contin. Educ. Anaesth. Crit. Care Pain 4, 71&ndash;75.</p>
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  • <p>Galougahi, K.K., Liu, C.-C., Gentile, C., Kok, C., Nunez, A., Garcia, A., Fry, N.A.S., Davies, M.J., Hawkins, C.L., Rasmussen, H.H., et al. (2014). Glutathionylation Mediates Angiotensin II&ndash;Induced eNOS Uncoupling, Amplifying NADPH Oxidase-Dependent Endothelial Dysfunction. J. Am. Heart Assoc. 3, e000731.</p>
  • <p>van Gorp, R.M.A., Heeneman, S., Broers, J.L.V., Bronnenberg, N.M.H.J., van Dam-Mieras, M.C.E., and Heemskerk, J.W.M. (2002). Glutathione oxidation in calcium- and p38 MAPK-dependent membrane blebbing of endothelial cells. Biochim. Biophys. Acta 1591, 129&ndash;138.</p>
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  • <p>Higashi, Y., Maruhashi, T., Noma, K., and Kihara, Y. (2014). Oxidative stress and endothelial dysfunction: clinical evidence and therapeutic implications. Trends Cardiovasc. Med. 24, 165&ndash;169.</p>
  • <p>Huang PL.&nbsp; Mouse models of nitric oxide synthase deficiency. (2000) J Am Soc Nephrol. 11 Suppl 16:S120-3.</p>
  • <p>Huang PL, Huang Z, Mashimo H <em>et al.</em> Hypertension in mice lacking the gene for endothelial nitric oxide synthase.&nbsp; (1995) Nature. 377(6546):239-42.</p>
  • <p>Jayaram, R., Goodfellow, N., Zhang, M.H., Reilly, S., Crabtree, M., De Silva, R., Sayeed, R., and Casadei, B. (2015). Molecular mechanisms of myocardial nitroso-redox imbalance during on-pump cardiac surgery. Lancet Lond. Engl. 385 Suppl 1, S49.</p>
  • <p>Lai M,&nbsp;Chu S,&nbsp;Lakshminrusimha S,&nbsp;Lin H. &nbsp;Beyond the inhaled nitric oxide in persistent pulmonary&nbsp;hypertension&nbsp;of the newborn. &nbsp;Pediatr Neonatol.&nbsp;2017&nbsp;pii: S1875-9572(17)30480-1. doi: 10.1016/j.pedneo.2016.09.011. [Epub ahead of print]</p>
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  • <p>Lerman, L.O., Nath, K.A., Rodriguez-Porcel, M., Krier, J.D., Schwartz, R.S., Napoli, C., and Romero, J.C. (2001). Increased oxidative stress in experimental renovascular hypertension. Hypertension 37, 541&ndash;546.</p>
  • <p>Li, J., Zhou, Z., Jiang, D.-J., Li, D., Tan, B., Liu, H., and Li, Y.-J. (2007). Reduction of NO- and EDHF-mediated vasodilatation in hypertension: role of asymmetric dimethylarginine. Clin. Exp. Hypertens. N. Y. N 1993 29, 489&ndash;501.</p>
  • <h2><span style="font-family:arial,helvetica,sans-serif"><span style="font-size:14px">Lowe FJ., Luettich K, Talikka M, Hoang V, Haswell LE., Hoeng J, and Gaca MD. Development of an Adverse Outcome Pathway for the Onset of Hypertension by Oxidative Stress-Mediated Perturbation of Endothelial Nitric Oxide Bioavailability. &nbsp;Applied In Vitro Toxicology. (2017), 3(1): 131-148.&nbsp;&nbsp;</span></span></h2>
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  • <p>Michaud, S.E., Dussault, S., Groleau, J., Haddad, P., and Rivard, A. (2006). Cigarette smoke exposure impairs VEGF-induced endothelial cell migration: role of NO and reactive oxygen species. J. Mol. Cell. Cardiol. 41, 275&ndash;284.</p>
  • <p>Mitchell BM , Dorrance AM, Webb RC. GTP cyclohydrolase 1 inhibition attenuates vasodilation and increases blood pressure in rats.&nbsp; (2003) Am J Physiol Heart Circ Physiol. 285(5):H2165-70.</p>
  • <p>Song, P., Wu, Y., Xu, J., Xie, Z., Dong, Y., Zhang, M., and Zou, M.-H. (2007). Reactive nitrogen species induced by hyperglycemia suppresses Akt signaling and triggers apoptosis by upregulating phosphatase PTEN (phosphatase and tensin homologue deleted on chromosome 10) in an LKB1-dependent manner. Circulation 116, 1585&ndash;1595.</p>
  • <p>Montecinos, V., Guzm&aacute;n, P., Barra, V., Villagr&aacute;n, M., Mu&ntilde;oz-Montesino, C., Sotomayor, K., Escobar, E., Godoy, A., Mardones, L., Sotomayor, P., et al. (2007). Vitamin C is an essential antioxidant that enhances survival of oxidatively stressed human vascular endothelial cells in the presence of a vast molar excess of glutathione. J. Biol. Chem. 282, 15506&ndash;15515.</p>
  • <p>Song, P., Xie, Z., Wu, Y., Xu, J., Dong, Y., and Zou, M.-H. (2008). Protein kinase Czeta-dependent LKB1 serine 428 phosphorylation increases LKB1 nucleus export and apoptosis in endothelial cells. J. Biol. Chem. 283, 12446&ndash;12455.</p>
  • <p>Nozik-Grayck E, Woods C, Taylor JM <em>et al.</em> Selective depletion of vascular EC-SOD augments chronic hypoxic pulmonary hypertension.&nbsp; Am J Physiol Lung Cell Mol Physiol. (2014) 307(11):L868-76.&nbsp;</p>
  • <p>Uruno, A., Sugawara, A., Kanatsuka, H., Kagechika, H., Saito, A., Sato, K., Kudo, M., Takeuchi, K., and Ito, S. (2005). Upregulation of nitric oxide production in vascular endothelial cells by all-trans retinoic acid through the phosphoinositide 3-kinase/Akt pathway. Circulation 112, 727&ndash;736.</p>
  • <p>Zou, M.-H., Hou, X.-Y., Shi, C.-M., Nagata, D., Walsh, K., and Cohen, R.A. (2002). Modulation by peroxynitrite of Akt- and AMP-activated kinase-dependent Ser1179 phosphorylation of endothelial nitric oxide synthase. J. Biol. Chem. 277, 32552&ndash;32557.</p>
  • </div>
  • <br>
  • <div>
  • <h4><a href="/relationships/992">Relationship: 992: Peptide Oxidation leads to Decrease, AKT/eNOS activity</a></h4>
  • <h4>AOPs Referencing Relationship</h4>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>AOP Name</th>
  • <th>Adjacency</th>
  • <th>Weight of Evidence</th>
  • <th>Quantitative Understanding</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <th><a href="/aops/149">Peptide Oxidation Leading to Hypertension</a></th>
  • <th>adjacent</th>
  • <th>High </th>
  • <th>Moderate</th>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • <h4>Evidence Supporting Applicability of this Relationship</h4>
  • <br>
  • <!-- loop to find taxonomic applicability under relationship -->
  • <div>
  • <strong>Taxonomic Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Term</th>
  • <th>Scientific Term</th>
  • <th>Evidence</th>
  • <th>Links</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Homo sapiens</td>
  • <td>Homo sapiens</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9606" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Bos taurus</td>
  • <td>Bos taurus</td>
  • <td>High</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=9913" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Mus musculus</td>
  • <td>Mus musculus</td>
  • <td>Low</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10090" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • <tr>
  • <td>Rattus norvegicus</td>
  • <td>Rattus norvegicus</td>
  • <td>Low</td>
  • <td>
  • <a href="http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=10116" , target="_blank">NCBI</a>
  • </td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for taxons -->
  • <!-- loop to find life stages under relationship -->
  • <div>
  • <strong>Life Stage Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Life Stage</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>All life stages</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end loop for life stages -->
  • <!-- sex terms -->
  • <div>
  • <strong>Sex Applicability</strong>
  • <div class="panel panel-default">
  • <table class="table table-bordered table-striped">
  • <thead>
  • <tr>
  • <th>Sex</th>
  • <th>Evidence</th>
  • </tr>
  • </thead>
  • <tbody>
  • <tr>
  • <td>Unspecific</td>
  • <td>High</td>
  • </tr>
  • </tbody>
  • </table>
  • </div>
  • </div>
  • <!-- end sex terms -->
  • <p>The relationship between oxidative stress and decreased AKT/eNOS activity is supported by studies performed in humans, cows, mice and rats.</p>
  • <h4>Key Event Relationship Description</h4>
  • <p>Exposure to known inducers of oxidative stress causes the phosphorylation of AKT&nbsp;and eNOS, leading to a decrease in their activities.</p>
  • <!-- if nothing shows up in any of these fields, then evidence supporting this KER will not be displayed -->
  • <h4>Evidence Supporting this KER</h4>
  • <strong>Biological Plausibility</strong>
  • <p>Multiple experimental studies reported modulations in Akt and eNOS phosphorylation/activity following oxidative stress, thus providing strong biological plausibility for this key event relationship.</p>
  • <p>Ohara, Y., Peterson, T.E., and Harrison, D.G. (1993). Hypercholesterolemia increases endothelial superoxide anion production. J. Clin. Invest. 91, 2546&ndash;2551.</p>
  • <p>In HUVECs, peroxynitrite significantly inhibited AKT&nbsp;phosphorylation at Ser473 and AKT&nbsp;activity (Song et al., 2007, 2008; Zou et al., 2002). However, Zou et al. (2002) found that peroxynitrite increased eNOS phosphorylation at Ser1199, but decreased NO availability, suggesting eNOS phosphorylation may not depend on AKT. Treatment of BAECs with SIN-1, a source of peroxynitrite, inhibited eNOS activity by 30% and AKT/eNOS phosphorylation (Das et al., 2014).</p>
  • <p>Ozaki, M., Kawashima, S., Yamashita, T., Hirase, T., Namiki, M., Inoue, N., Hirata, K., Yasui, H., Sakurai, H., Yoshida, Y., et al. (2002). Overexpression of endothelial nitric oxide synthase accelerates atherosclerotic lesion formation in apoE-deficient mice. J. Clin. Invest. 110, 331&ndash;340.</p>
  • <p>High glucose concentrations can also inhibit AKT&nbsp;phosphorylation (Song et al., 2008). HUVECs treated with methylglyoxal and high concentrations of glucose exhibited reduced bradykinin-stimulated eNOS activity via Ser1177 phosphorylation (Dhar et al., 2010), while hyperglycemia inhibited eNOS activity in BAECs (Du et al., 2001). In EA.hy926 endothelial cells, methylglyoxal treatment results in reduced eNOS phosphorylation at Ser1177 (Su et al., 2013). High-fat diet-induced obesity in mice caused an increase in ROS and a reduction in AKT&nbsp;and eNOS phosphorylation compared to non-obese mice (Du et al., 2013).</p>
  • <p>Park, W.H. (2013). The effects of exogenous H2O2 on cell death, reactive oxygen species and glutathione levels in calf pulmonary artery and human umbilical vein endothelial cells. Int. J. Mol. Med. 31, 471&ndash;476.</p>
  • <p>Treatment with cigarette smoke extract (CSE) also inhibited Akt and eNOS in VEGF-stimulated HUVECs (Michaud et al., 2006). Myocardial ischemia decreased phosphoryl