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Relationship: 3318

Title

A descriptive phrase which clearly defines the two KEs being considered and the sequential relationship between them (i.e., which is upstream, and which is downstream). More help

Altered Stress Response Signaling leads to Increase, Endothelial Dysfunction

Upstream event

The causing Key Event (KE) in a Key Event Relationship (KER). More help

Altered Stress Response Signaling

Downstream event

The responding Key Event (KE) in a Key Event Relationship (KER). More help

Increase, Endothelial Dysfunction

Key Event Relationship Overview

The utility of AOPs for regulatory application is defined, to a large extent, by the confidence and precision with which they facilitate extrapolation of data measured at low levels of biological organisation to predicted outcomes at higher levels of organisation and the extent to which they can link biological effect measurements to their specific causes.Within the AOP framework, the predictive relationships that facilitate extrapolation are represented by the KERs. Consequently, the overall WoE for an AOP is a reflection in part, of the level of confidence in the underlying series of KERs it encompasses. Therefore, describing the KERs in an AOP involves assembling and organising the types of information and evidence that defines the scientific basis for inferring the probable change in, or state of, a downstream KE from the known or measured state of an upstream KE. More help

AOPs Referencing Relationship

AOP Name	Adjacency	Weight of Evidence	Quantitative Understanding	Point of Contact	Author Status	OECD Status
Deposition of energy leads to abnormal vascular remodeling	adjacent	Moderate	Low	Vinita Chauhan (send email)	Open for citation & comment	WPHA/WNT Endorsed

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) that help to define the biological applicability domain of the KER.In general, this will be dictated by the more restrictive of the two KEs being linked together by the KER. More help

Term	Scientific Term	Evidence	Link
human	Homo sapiens	Low	NCBI
rat	Rattus norvegicus	Moderate	NCBI

Sex Applicability

An indication of the the relevant sex for this KER. More help

Sex	Evidence
Male	Moderate
Female	Low
Hermaphrodite	Low

Life Stage Applicability

An indication of the the relevant life stage(s) for this KER. More help

Term	Evidence
Adult	Moderate
Juvenile	Low

Key Event Relationship Description

Provides a concise overview of the information given below as well as addressing details that aren’t inherent in the description of the KEs themselves. More help

Altered stress response signaling can disrupt cellular homeostasis and induce endothelial dysfunction, characterized by a prolonged state of endothelial activation (Deanfield et al., 2007). Signaling pathways involved in triggering endothelial dysfunction include the p53-p21 pathway, the Akt/phosphtidylinositol-3-kinase (PI3K)/mechanistic target of rapamycin (mTOR) pathway, the RhoA-Rho-kinase pathway, and the acid sphingomyelinase (ASM)/ceramide (Cer) pathway (Venkatsulu et al., 2018; Soloviev et al., 2019; Wang et al., 2016). Activation of the signaling molecule p53 by phosphorylation enhances its stability, leading to cell cycle arrest and premature senescence in endothelial cells and can alternatively lead to a caspase cascade resulting in cellular apoptosis. Activation of the sphingomyelinase ceramide pathway can also contribute to endothelial apoptosis through production of ceramide that activates mitogen-activated protein kinase (MAPK) and extracellular-signal-regulated kinase (ERK). Signaling molecules MAPK and ERK can also be activated as a direct response to a stressor and prompt a cascade of events resulting in endothelial cell apoptosis. Impairment of the Akt/PI3K/mTOR pathway can lead to apoptosis by preventing cell survival signaling and can also lead to downregulation of Rho cytoskeletal proteins for senescence of endothelial cells (Venkatesulu et al., 2018; Soloviev et al., 2019; Nagane et al., 2021; Ramadan et al., 2021; Hughson et al., 2018).

Evidence Collection Strategy

Include a description of the approach for identification and assembly of the evidence base for the KER. For evidence identification, include, for example, a description of the sources and dates of information consulted including expert knowledge, databases searched and associated search terms/strings. Include also a description of study screening criteria and methodology, study quality assessment considerations, the data extraction strategy and links to any repositories/databases of relevant references.Tabular summaries and links to relevant supporting documentation are encouraged, wherever possible. More help

Evidence Supporting this KER

Addresses the scientific evidence supporting KERs in an AOP setting the stage for overall assessment of the AOP. More help

Overall weight of evidence: Moderate

Biological Plausibility

Addresses the biological rationale for a connection between KEupstream and KEdownstream. This field can also incorporate additional mechanistic details that help inform the relationship between KEs, this is useful when it is not practical/pragmatic to represent these details as separate KEs due to the difficulty or relative infrequency with which it is likely to be measured. More help

The biological plausibility of the connection between altered stress response signaling leading to an increase in endothelial dysfunction is well-supported by literature and the mechanisms are generally understood. Multiple signaling pathways influence endothelial function.

Modulation of the Akt/PI3K/mTOR pathway downregulates downstream Rho cytoskeletal proteins, which leads to partial-to-full senescence of endothelial cells, resulting in increased vascular permeability and endothelial dysfunction (Venkatsulu et al., 2018). Modulation of the Akt/PI3K/mTOR pathway also acts upstream of the p53-p21 pathway to mediate endothelial cell senescence (Wang et al., 2016). Unlike in other cell types, in endothelial cells the p53-p21 pathway is more important than the p16-Rb pathway for induction of cell senescence (Wang et al., 2016). Senescent endothelial cells show changes in cell morphology, cell-cycle arrest, and increased senescence-associated β-galactosidase (SA-β-gal) staining. These changes lead to endothelial dysfunction, which results in dysregulation of vasodilation (Wang et al., 2016; Hughson et al., 2018; Ramadan et al., 2021). Phosphorylation of p53 is another important moderator of apoptosis in endothelial cells, as well as the ASM/Cer pathway, where the production of ceramide mediates endothelial apoptosis through sphingomyelinase, activating MAPK and ERK, which prompt a cascade of events culminating in endothelial cell apoptosis, another cellular marker for endothelial dysfunction (Venkatsulu et al., 2018; Soloviev et al., 2019).

Empirical Evidence

Provides specific (citable) evidence that supports the idea of a change in the upstream KE (KEupstream) leading to, or being associated with, a subsequent change in the downstream KE (KEdownstream), assuming the perturbation of KEupstream is sufficient. More help

The empirical evidence supporting this KER was gathered from research utilizing both in vivo and in vitro models. Many in vitro studies have examined the relationship using endothelial cell cultures. Endothelial dysfunction due to altered signaling can be measured by premature endothelial cell senescence, apoptosis and impaired contractile response (Korpela & Liu, 2014). Levels of signaling molecules in pathways such as the Akt/PI3K/mTOR, RhoA-Rho-kinase, and ASM/Cer pathways, and the effect they have on endothelial dysfunction as characterized by endothelial cellular senescence, apoptosis and contractile response, was examined in these studies (Chang et al., 2017; Cheng et al., 2017; Su et al., 2020; Summers et al., 2008; Yentrapalli et al., 2013a; Yentrapalli et al., 2013b).

Dose Concordance

Studies have used in vitro models with acute and chronic doses of radiation administered from 0.23 to 10 Gy to show that the dose at which significant alterations in stress response signaling occurs is concordant with doses at which endothelial dysfunction occurs. Alterations in various signaling pathways, including decreases in the Akt/PI3K/mTOR pathway and increases in the p53-p21 pathway, both occurred at 2.4 and 4 Gy; endothelial cell senescence was found only at 4 Gy (Yentrapalli et al., 2013a). Another study using 4.1 mGy/h chronic gamma irradiation found significant changes in the p53-p21 and ERK signaling pathways after both 2.1 and 4.1 Gy (Yentrapalli et al., 2013b). These changes were associated with an increase in endothelial dysfunction at both doses as well, with only a slight increase in cell senescence after 2.1 Gy (Yentrapalli et al., 2013b). Endothelial cells exposed to a single dose of 10 Gy of X-rays showed an increase in Akt and p-Akt that was correlated with an increase in endothelial dysfunction indicated by a 5-fold increase in apoptosis (Chang et al., 2017).

Various studies also showed that altered signaling and endothelial dysfunction can occur at the same stressor severity during hindlimb unloading (HU). Rats exposed to microgravity for 20 days showed both decreased RhoA signaling molecule and impaired vasodilation (Summers et al., 2008). Two studies observing the effects of microgravity for 4 weeks in rats found that a decrease in the ASM/Cer pathway resulted in better endothelial function, while an increase in the pathway resulted in endothelial dysfunction (Cheng et al., 2017; Su et al., 2020).

Time Concordance

There is limited evidence to suggest a time concordance between altered stress response signaling and endothelial dysfunction. In a study using gamma irradiation in vitro, the p53-p21 and ERK signaling pathways were altered after 3 weeks of chronic gamma irradiation, while cell senescence was increased only slightly after 3 weeks, and increased more after 6 weeks (Yentrapalli et al., 2013b). In a similar study, alterations in various signaling pathways including decreases in the Akt/PI3K/mTOR pathway were found as early as 1 week during chronic gamma irradiation (Yentrapalli et al., 2013a). The earliest significant increase in cellular senescence occurred after 10 weeks (Yentrapalli et al., 2013a).

Incidence Concordance

There is limited support in current literature for an incidence concordance relationship between altered signaling and endothelial dysfunction. One out of the 6 primary research studies used to support this KER demonstrated an average change to endpoints of altered signaling that was greater or equal to that of endothelial dysfunction (Cheng et al., 2017; Yentrapalli et al., 2013b). After 6 weeks of gamma irradiation of HUVECs, altered signaling marker, p21, was increased by 3.5-fold, compared to the 3-fold increase in SA-β-gal staining, a marker for endothelial cell senescence (Yentrapalli et al., 2013b).

Essentiality

Changes in endothelial signaling pathways can trigger endothelial dysfunction. Therefore, in the absence of altered signaling endothelial dysfunction is not expected. Through the use of signaling molecule inhibitors such as Y27632 and dpm, altered signaling can be suppressed which results in reduced endothelial dysfunction (Venkatsulu et al., 2018; Soloviev et al., 2019; Wang et al., 2016). A study observing the effects of Y27632, a Rho kinase inhibitor, found that when rat abdominal aortas were incubated with Y27632 the contractile response, which was hindered by HU, was returned to control levels (Summers et al., 2008). The NFkB inhibitor, PS1145, reduced senescence-like cells almost by 2-fold after 8 Gy of irradiation (Dong et al., 2015). Similar results have been shown by other groups that have looked at endothelial cells incubated in mesenchymal stem cell conditioned media (MSC-CM), which is thought to exhibit therapeutic potential for microvascular injury through angiogenic cytokines. MSC-CM prevented an increase in cleaved capsase-3 and increased both Akt and p-Akt, which was associated with a substantial decrease in apoptosis (Chang et al., 2017). It has also been found that enhancing the ASM/Cer pathway with C6-ceramide increases downstream caspase-3 and endothelial dysfunction, measured by apoptosis, while ASM inhibitors dpm and DOX partially decreased both caspase-3 and apoptosis (Su et al., 2020; Cheng et al., 2017).

Uncertainties and Inconsistencies

Addresses inconsistencies or uncertainties in the relationship including the identification of experimental details that may explain apparent deviations from the expected patterns of concordance. More help

Much of the evidence for this relationship comes from in vitro studies; further work is needed to determine the certainty of the relationship at the tissue level.

Known modulating factors

This table captures specific information on the MF, its properties, how it affects the KER and respective references.1.) What is the modulating factor? Name the factor for which solid evidence exists that it influences this KER. Examples: age, sex, genotype, diet 2.) Details of this modulating factor. Specify which features of this MF are relevant for this KER. Examples: a specific age range or a specific biological age (defined by...); a specific gene mutation or variant, a specific nutrient (deficit or surplus); a sex-specific homone; a certain threshold value (e.g. serum levels of a chemical above...) 3.) Description of how this modulating factor affects this KER. Describe the provable modification of the KER (also quantitatively, if known). Examples: increase or decrease of the magnitude of effect (by a factor of...); change of the time-course of the effect (onset delay by...); alteration of the probability of the effect; increase or decrease of the sensitivity of the downstream effect (by a factor of...) 4.) Provision of supporting scientific evidence for an effect of this MF on this KER. Give a list of references. More help

Modulating factor	Details	Effects on the KER	References
Drug	Ceramide-6 (Cer-6)	Increases downstream caspase-3 and apoptosis	(Cheng et al., 2017; Su et al., 2020)
Media	Mesenchymal stem cell conditioned media	Prevented an increase in cleaved capsase-3, increased both Akt and p-Akt, and decreased apoptosis	(Chang et al., 2017)
Drug	Y27632 (Rho kinase inhibitor)	Recovered contractile response that was attenuated by HU	(Summers et al., 2008)
Drug	Desipramine (dpm) (ASM inhibitor)	Partially decreased caspase-3 and apoptosis	(Cheng et al., 2017; Su et al., 2020)
Drug	Dopexin hydrochloride (DOX) (ASM inhibitor)	Partially decreased caspase-3 and apoptosis	(Cheng et al., 2017; Su et al., 2020)

Quantitative Understanding of the Linkage

Captures information that helps to define how much change in the upstream KE, and/or for how long, is needed to elicit a detectable and defined change in the downstream KE. More help

The following are a few examples of quantitative understanding of the relationship. All data that is represented is statistically significant unless otherwise indicated.

Response-response Relationship

Provides sources of data that define the response-response relationships between the KEs. More help

Dose/incidence concordance

Reference	Experiment Description	Result
Yentrapalli et al., 2013a	In vitro. Human umbilical vein endothelial cells (HUVEC) were irradiated with 137Cs gamma radiation at 1.4 mGy/h or 2.4 mGy/h dose rates over a period of 12 weeks. Signaling molecules were used as measures of altered signaling. Cell senescence was used as a measure of endothelial dysfunction.	1.4 mGy/h dose rate: This dose rate led to alterations in the Akt/PI3K/mTOR pathway including a 3-fold increase in p21, a 0.7-fold decrease in p-Akt and PI3K, and a 0.8-fold decrease in mTOR. However, this dose rate did not induce endothelial dysfunction (no significant changes in SA-β-gal staining). 2.4 mGy/h dose rate: This dose rate led to alterations in the Akt/PI3K/mTOR pathway including a 3-fold increase in p21, a 1.5-fold increase in Akt, a 0.7-fold decrease in p-Akt and PI3K, a 0.8-fold decrease in mTOR and a 0.5-fold decrease in p-ERK. This dose rate induced endothelial dysfunction as indicated by a 2-fold increase in SA-β-gal staining, a marker for endothelial cell senescence.
Yentrapalli et al., 2013b	In vitro. Human umbilical vein endothelial cells (HUVEC) were irradiated with 137Cs gamma radiation at a 4.1 mGy/h dose rate over the course of 6 weeks. Signaling molecules were used as measures of altered signaling. Cell senescence was used as a measure of endothelial dysfunction.	Chronic irradiation led to a 2.5-fold increase in p-p53, a key signaling molecule in initiating endothelial cell senescence, after week 6 (4.13 Gy). There was no significant change in total p53 or p-ERK2 at any time point, while total ERK2 showed a 0.5-fold decrease after 3 weeks (2.07 Gy). p21, which acts downstream of p53, increased 3.5-fold by week 6 (4.13 Gy). Alterations in the above signaling molecules correlated with an increase in endothelial dysfunction as indicated by a 1.5-fold increase in SA-β-gal staining, a marker for endothelial cell senescence, after 3 weeks and 3-fold after 6 weeks (4.13 Gy).
Chang et al., 2017	In vitro. Human umbilical vein endothelial cells (HUVEC) were irradiated with 10 Gy of X-rays. Signaling molecules were used as measures of altered signaling. Apoptosis was used as a measure of endothelial dysfunction.	A 2-fold increase in cleaved caspase-3, part of the apoptosis diated endothelial cells was corr signaling cascade, in the irraelated with an increase in endothelial dysfunction indicated by a 4-fold increase in the percentage of apoptotic cells.
Summers et al., 2008	In vivo. Adult male Wistar rats underwent 20-day HU after which abdominal aortas were harvested. Signaling molecules were used as measures of altered signaling. Contractile response was used as a measure of endothelial dysfunction.	Following HU, signaling molecule RhoA decreased by 0.5-fold and endothelial dysfunction was induced in the form of reduced contractile response to phenylephrine by 0.5-fold.
Su et al., 2020	In vivo. Sprague-Dawley male rats underwent 4 weeks of hindlimb unloading (HU), following which cerebral and mesenteric arteries were harvested. Signaling molecules were used as measures of altered signaling. Apoptosis was used as a measure of endothelial dysfunction.	Simulated microgravity revealed a decrease in signaling molecules ASM and Cer in cerebral arteries but an increase in mesenteric arteries of rats. Caspase-3, a signaling molecule in the apoptosis cascade, decreased 0.5-fold in cerebral arteries and increased 2-fold in mesenteric arteries. This was associated with an increase in endothelial function in cerebral arteries by decreasing apoptosis 0.3-fold and an increase in endothelial dysfunction in mesenteric arteries by increasing apoptosis 2-fold.
Cheng et al., 2017	In vivo. Sprague-Dawley male rats underwent 4 weeks of hindlimb unloading following which carotid arteries were harvested. Signaling molecules were used as measures of altered signaling. Apoptosis was used as a measure of endothelial dysfunction.	2 weeks of simulated microgravity resulted in a 0.6-fold decrease in signaling molecules ASM and caspase-3, which led to a decrease of endothelial dysfunction indicated by 0.5-fold reduced apoptosis in the carotid arteries of rats.

Time-scale

Information regarding the approximate time-scale of the changes in KEdownstream relative to changes in KEupstream (i.e., do effects on KEdownstream lag those on KEupstream by seconds, minutes, hours, or days?). More help

Time concordance

Reference

Experiment Description

Result

Yentrapalli et al., 2013a

In vitro. Human umbilical vein endothelial cells (HUVEC) were irradiated with 137Cs gamma radiation at 1.4 mGy/h or 2.4 mGy/h dose rates over a period of 10 weeks. Signaling molecules were used as measures of altered signaling. Cell senescence was used as a measure of endothelial dysfunction.

1.4 mGy/h dose rate: This dose rate led to alterations in the Akt/PI3K/mTOR pathway including a 3-fold increase in p21 after 10 weeks, a 0.7-fold decrease in p-Akt and PI3K after 6 and 10 weeks, and a 0.8-fold decrease in mTOR after 10 weeks. However, this dose rate did not induce endothelial dysfunction (no significant changes in SA-β-gal staining at any timepoint).

2.4 mGy/h dose rate: This dose rate caused alterations in the Akt/PI3K/mTOR pathway including a 3-fold increase in p21 after 10 weeks, a 1.5-fold increase in Akt after 1 week, a 0.7-fold decrease in p-Akt and PI3K after 6 and 10 weeks, a 0.8-fold decrease in mTOR and a 0.5-fold decrease in p-ERK after 10 weeks. This dose rate induced endothelial dysfunction indicated by a 2-fold increase in SA-β-gal staining, after 12 weeks.

Yentrapalli et al., 2013b

In vitro. Human umbilical vein endothelial cells (HUVEC) were irradiated with 137Cs gamma radiation at a 4.1 mGy/h dose rate over the course of 6 weeks. Signaling molecules were used as measures of altered signaling. Cell senescence was used as a measure of endothelial dysfunction.

Chronic irradiation led to a 2.5-fold increase in p-p53, a key signaling molecule in initiating endothelial cell senescence, after week 6 (4.13 Gy). There was no significant change in total p53 or p-ERK2 at any time point, while total ERK2 showed a 0.5-fold decrease after 3 weeks (2.07 Gy). p21, which acts downstream of p53, increased 3.5-fold by week 6 (4.13 Gy). Alterations in the above signaling molecules correlated with an increase in endothelial dysfunction as indicated by a 1.5-fold increase in SA-β-gal staining after 3 weeks and 3-fold after 6 weeks (4.13 Gy).

Known Feedforward/Feedback loops influencing this KER

Define whether there are known positive or negative feedback mechanisms involved and what is understood about their time-course and homeostatic limits. More help

Not identified.

Domain of Applicability

A free-text section of the KER description that the developers can use to explain their rationale for the taxonomic, life stage, or sex applicability structured terms. More help

Evidence for this KER is supported through in vivo rat and in vitro human studies. The in vivo studies were conducted in male animals, although the relationship is still plausible in females. The in vivo studies were undertaken in adolescent and adult rats.

References

List of the literature that was cited for this KER description. More help

Chang, P. Y. et al. (2017), “MSC-derived cytokines repair radiation-induced intra-villi microvascular injury”, Oncotarget, Vol. 8/50, Impact Journals, Orchard Park, https://doi.org/10.18632/oncotarget.21236.

Cheng, Y. P. et al. (2017), “Acid sphingomyelinase/ceramide regulates carotid intima-media thickness in simulated weightless rats”, Pflugers Archiv European Journal of Physiology, Vol. 469, Springer, New York, https://doi.org/10.1007/s00424-017-1969-z.

Deanfield, J.E., J. P. Halcox and T. J. Rabelink (2007), “Endothelial function and dysfunction: Testing and clinical relevance”, Circulation, Vol. 115/10, Lippincott Williams & Wilkins, Philadelphia, https://doi.org/10.1161/CIRCULATIONAHA.106.652859

Dong, X. et al. (2015), “NEMO modulates radiation-induced endothelial senescence of human umbilical veins through NF-κB signal pathway”, Radiation research, Vol. 183/1, BioOne, Washington, https://doi.org/10.1667/RR13682.1

Hughson, R.L., A. Helm and M. Durante (2018), “Heart in space: Effect of the extraterrestrial environment on the cardiovascular system”, Nature Reviews Cardiology, Vol. 15/3, Nature Portfolio, London, https://doi.org/10.1038/nrcardio.2017.157.

Korpela, E., and S. K. Liu (2014), “Endothelial perturbations and therapeutic strategies in normal tissue radiation damage”, Radiation oncology, Vol. 9, BioMed Central, London, https://doi.org/10.1186/s13014-014-0266-7.

Nagane, M. et al. (2021), “DNA damage response in vascular endothelial senescence: Implication for radiation-induced cardiovascular diseases”, Journal of Radiation Research, Vol. 62/4, Oxford University Press, Oxford, https://doi.org/10.1093/JRR/RRAB032.

Ramadan, R. et al. (2021), “The role of connexin proteins and their channels in radiation-induced atherosclerosis”, Cellular and molecular life sciences: CMLS, Vol. 78/7, Springer, New York, https://doi.org/10.1007/s00018-020-03716-3.

Soloviev, A. I. and I.V. Kizub (2019), “Mechanisms of vascular dysfunction evoked by ionizing radiation and possible targets for its pharmacological correction”, Biochemical pharmacology, Vol. 159, Elsevier, Amsterdam, https://doi.org/10.1016/j.bcp.2018.11.019.

Su, Y. T. et al. (2020), “Acid sphingomyelinase/ceramide mediates structural remodeling of cerebral artery and small mesenteric artery in simulated weightless rats”, Life sciences, Vol. 243, Elsevier, Amsterdam, https://doi.org/10.1016/j.lfs.2019.117253.

Summers, S. M., S. V. Nguyen, and R. E. Purdy (2008), “Hindlimb unweighting induces changes in the RhoA-Rho-kinase pathway of the rat abdominal aorta”, Vascular pharmacology, Vol. 48/4-6, Elsevier, Amsterdam, https://doi.org/10.1016/j.vph.2008.03.006.

Venkatesulu, B. P. et al. (2018), “Radiation-Induced Endothelial Vascular Injury: A Review of Possible Mechanisms”, JACC: Basic to translational science, Vol. 3/4, Elsevier, Amsterdam, https://doi.org/10.1016/j.jacbts.2018.01.014.

Wang, Y., M. Boerma and D. Zhou (2016), “Ionizing Radiation-Induced Endothelial Cell Senescence and Cardiovascular Diseases”, Radiation research, Vol. 186/2, Radiation Research Society, Bozeman, https://doi.org/10.1667/RR14445.1.  

Yentrapalli, R. et al. (2013a), “The PI3K/Akt/mTOR pathway is implicated in the premature senescence of primary human endothelial cells exposed to chronic radiation”, PloS one, Vol. 8/8, PLOS, San Francisco, https://doi.org/10.1371/journal.pone.0070024.

Yentrapalli, R. et al. (2013b), “Quantitative proteomic analysis reveals induction of premature senescence in human umbilical vein endothelial cells exposed to chronic low-dose rate gamma radiation”, Proteomics, Vol. 13/7, John Wiley & Sons, Ltd., Hoboken, https://doi.org/10.1002/pmic.201200463.