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Event: 1496

Key Event Title

The KE title should describe a discrete biological change that can be measured. It should generally define the biological object or process being measured and whether it is increased, decreased, or otherwise definably altered relative to a control state. For example “enzyme activity, decreased”, “hormone concentration, increased”, or “growth rate, decreased”, where the specific enzyme or hormone being measured is defined. More help

Increased, secretion of proinflammatory mediators

Short name
The KE short name should be a reasonable abbreviation of the KE title and is used in labelling this object throughout the AOP-Wiki. The short name should be less than 80 characters in length. More help
Increased proinflammatory mediators

Biological Context

Structured terms, selected from a drop-down menu, are used to identify the level of biological organization for each KE. Note, KEs should be defined within a particular level of biological organization. Only KERs should be used to transition from one level of organization to another. Selection of the level of biological organization defines which structured terms will be available to select when defining the Event Components (below). More help
Level of Biological Organization
Cellular

Cell term

Further information on Event Components and Biological Context may be viewed on the attached pdf.The biological context describes the location/biological environment in which the event takes place.  For molecular/cellular events this would include the cellular context (if known), organ context, and species/life stage/sex for which the event is relevant. For tissue/organ events cellular context is not applicable.  For individual/population events, the organ context is not applicable. More help
Cell term
eukaryotic cell

Organ term

Further information on Event Components and Biological Context may be viewed on the attached pdf.The biological context describes the location/biological environment in which the event takes place.  For molecular/cellular events this would include the cellular context (if known), organ context, and species/life stage/sex for which the event is relevant. For tissue/organ events cellular context is not applicable.  For individual/population events, the organ context is not applicable. More help

Key Event Components

Further information on Event Components and Biological Context may be viewed on the attached pdf.Because one of the aims of the AOP-KB is to facilitate de facto construction of AOP networks through the use of shared KE and KER elements, authors are also asked to define their KEs using a set of structured ontology terms (Event Components). In the absence of structured terms, the same KE can readily be defined using a number of synonymous titles (read by a computer as character strings). In order to make these synonymous KEs more machine-readable, KEs should also be defined by one or more “event components” consisting of a biological process, object, and action with each term originating from one of 22 biological ontologies (Ives, et al., 2017; See List). Biological process describes dynamics of the underlying biological system (e.g., receptor signalling). The biological object is the subject of the perturbation (e.g., a specific biological receptor that is activated or inhibited). Action represents the direction of perturbation of this system (generally increased or decreased; e.g., ‘decreased’ in the case of a receptor that is inhibited to indicate a decrease in the signalling by that receptor).Note that when editing Event Components, clicking an existing Event Component from the Suggestions menu will autopopulate these fields, along with their source ID and description. To clear any fields before submitting the event component, use the 'Clear process,' 'Clear object,' or 'Clear action' buttons. If a desired term does not exist, a new term request may be made via Term Requests. Event components may not be edited; to edit an event component, remove the existing event component and create a new one using the terms that you wish to add. More help
Process Object Action
cytokine production involved in inflammatory response Cytokine increased
chemokine secretion Chemokine increased
complement activation increased
Interleukin increased

Key Event Overview

AOPs Including This Key Event

All of the AOPs that are linked to this KE will automatically be listed in this subsection. This table can be particularly useful for derivation of AOP networks including the KE. Clicking on the name of the AOP will bring you to the individual page for that AOP. More help
AOP Name Role of event in AOP Point of Contact Author Status OECD Status
Substance interaction with the lung cell membrane leading to lung fibrosis KeyEvent Sabina Halappanavar (send email) Under development: Not open for comment. Do not cite EAGMST Under Review
SARS-CoV-2 leads to acute respiratory distress KeyEvent Young Jun Kim (send email) Open for comment. Do not cite Under Development
AT1R, lung fibrosis KeyEvent Young Jun Kim (send email) Under development: Not open for comment. Do not cite Under Development
Dysregulated fibrinolysis/bradykinin leading to hyperinflammation KeyEvent Penny Nymark (send email) Under development: Not open for comment. Do not cite Under Development
Frustrated phagocytosis leads to malignant mesothelioma KeyEvent Penny Nymark (send email) Under development: Not open for comment. Do not cite
TLR9 activation leading to Multi Organ Failure and ARDS KeyEvent Gillina Bezemer (send email) Under development: Not open for comment. Do not cite
Covalent binding to proteins leads to Respiratory Sensitisation/Sensitization/Allergy KeyEvent Kristie Sullivan (send email) Under Development: Contributions and Comments Welcome Under Development
Binding to ACE2 leads to lung fibrosis KeyEvent Young Jun Kim (send email) Open for comment. Do not cite Under Development
Interaction with lung cells leads to lung cancer KeyEvent Penny Nymark (send email) Under development: Not open for comment. Do not cite

Stressors

This is a structured field used to identify specific agents (generally chemicals) that can trigger the KE. Stressors identified in this field will be linked to the KE in a machine-readable manner, such that, for example, a stressor search would identify this as an event the stressor can trigger. NOTE: intermediate or downstream KEs in one AOP may function as MIEs in other AOPs, meaning that stressor information may be added to the KE description, even if it is a downstream KE in the pathway currently under development.Information concerning the stressors that may trigger an MIE can be defined using a combination of structured and unstructured (free-text) fields. For example, structured fields may be used to indicate specific chemicals for which there is evidence of an interaction relevant to this MIE. By linking the KE description to a structured chemical name, it will be increasingly possible to link the MIE to other sources of chemical data and information, enhancing searchability and inter-operability among different data-sources and knowledgebases. The free-text section “Evidence for perturbation of this MIE by stressor” can be used both to identify the supporting evidence for specific stressors triggering the MIE as well as to define broad chemical categories or other properties that classify the stressors able to trigger the MIE for which specific structured terms may not exist. More help

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) can be selected from an ontology. In many cases, individual species identified in these structured fields will be those for which the strongest evidence used in constructing the AOP was available in relation to this KE. More help
Term Scientific Term Evidence Link
mouse Mus musculus High NCBI
rats Rattus norvegicus High NCBI
human Homo sapiens High NCBI

Life Stages

The structured ontology terms for life-stage are more comprehensive than those for taxa, but may still require further description/development and explanation in the free text section. More help
Life stage Evidence
Adults High

Sex Applicability

The authors must select from one of the following: Male, female, mixed, asexual, third gender, hermaphrodite, or unspecific. More help
Term Evidence
Male High
Female High

Key Event Description

A description of the biological state being observed or measured, the biological compartment in which it is measured, and its general role in the biology should be provided. For example, the biological state being measured could be the activity of an enzyme, the expression of a gene or abundance of an mRNA transcript, the concentration of a hormone or protein, neuronal activity, heart rate, etc. The biological compartment may be a particular cell type, tissue, organ, fluid (e.g., plasma, cerebrospinal fluid), etc. The role in the biology could describe the reaction that an enzyme catalyses and the role of that reaction within a given metabolic pathway; the protein that a gene or mRNA transcript codes for and the function of that protein; the function of a hormone in a given target tissue, physiological function of an organ, etc. Careful attention should be taken to avoid reference to other KEs, KERs or AOPs. Only describe this KE as a single isolated measurable event/state. This will ensure that the KE is modular and can be used by other AOPs, thereby facilitating construction of AOP networks. More help

Pro-inflammatory mediators are the chemical and biological molecules that initiate and regulate inflammatory reactions. Pro-inflammatory mediators are secreted following exposure to an inflammogen in a gender/sex or developmental stage independent manner. They are secreted during inflammation in all species. Different types of pro-inflammatory mediators are secreted during innate or adaptive immune responses across various species (Mestas and Hughes, 2004). Cell-derived pro-inflammatory mediators include cytokines, chemokines, and growth factors. Blood derived pro-inflammatory mediators include vasoactive amines, complement activation products and others. These modulators can be grouped based on the cell type that secrete them, their cellular localisation and also based on the type of immune response they trigger. For example, members of the interleukin (IL) family including IL-2, IL-4, IL-7, IL-9, IL-15, IL-21, IL-3, IL-5 and GM-CSF are involved in the adaptive immune responses. The pro-inflammatory cytokines include IL-1 family (IL-1a, IL-1b, IL-1ra, IL-18, IL-36a, IL-36b, IL-36g, IL-36Ra, IL-37), IL-6 family, TNF family, IL-17, and IFNg (Turner et al., 2014). While IL-4 and IL-5 are considered T helper (Th) cell type 2 response, IFNg is suggested to be Th1 type response.

Different types of pro-inflammatory mediators are secreted during innate or adaptive immune responses across various species (Mestas and Hughes, 2004). However, IL-1 family cytokines, IL-4, IL-5, IL-6, TNFa, IFNg are the commonly measured mediators in experimental animals and in humans. Similar gene expression patterns involving inflammation and matrix remodelling are observed in human patients of pulmonary fibrosis and mouse lungs exposed to bleomycin (Kaminski, 2002).

Literature evidence for its perturbation:

Several studies show increased proinflammatory mediators in rodent lungs and bronchoalveolar lavage fluid, and in cell culture supernatants following exposure to a variety of CNT types and other materials. Poland et al., 2008 showed that long and thin CNTs (>5 µm) can elicit asbestos-like pathogenicity through the continual release of pro-inflammatory cytokines and ROS (reactive oxygen species). Exposure to crystalline silica induces release of inflammatory cytokines (TNFa, IL-1, IL-6), transcription factors (NF-kb, AP-1) and kinase signalling pathways in mice that contain NFkB luciferase reporter (Hubbard et al., 2002). Boyles et al., 2015 found that lung responses to long MWCNTs included high expression levels of pro-inflammatory mediators MCP-1, TGF-β1, and TNF-α (Boyles et al., 2015). Bleomycin administration in rodents induces lung inflammation and increased expression of pro-inflammatory mediators (Park et al., 2019). Inflammation induced by bleomycin, paraquat and CNTs is characterised by the altered expression of pro-inflammatory mediators. A large number of NMs induce expression of cytokines and chemokines in lungs of rodents exposed via inhalation (Halappanavar et al., 2011; Husain et al., 2015a). Similarities are observed in gene programs involving pro-inflammatory event is observed in both humans and experimental mice (Zuo et al., 2002).

How It Is Measured or Detected

One of the primary considerations in evaluating AOPs is the relevance and reliability of the methods with which the KEs can be measured. The aim of this section of the KE description is not to provide detailed protocols, but rather to capture, in a sentence or two, per method, the type(s) of measurements that can be employed to evaluate the KE and the relative level of scientific confidence in those measurements. Methods that can be used to detect or measure the biological state represented in the KE should be briefly described and/or cited. These can range from citation of specific validated test guidelines, citation of specific methods published in the peer reviewed literature, or outlines of a general protocol or approach (e.g., a protein may be measured by ELISA).Key considerations regarding scientific confidence in the measurement approach include whether the assay is fit for purpose, whether it provides a direct or indirect measure of the biological state in question, whether it is repeatable and reproducible, and the extent to which it is accepted in the scientific and/or regulatory community. Information can be obtained from the OECD Test Guidelines website and the EURL ECVAM Database Service on Alternative Methods to Animal Experimentation (DB-ALM). ?

The selection of pro-inflammatory mediators for investigation varies based on the expertise of the lab, cell types studied and the availability of the specific antibodies.

qRT-PCR – will measure the abundance of cytokine mRNA in a given sample. The method involves three steps: conversion of RNA into cDNA by reverse transcription method, amplification of cDNA using the PCR, and the real-time detection and quantification of amplified products (amplicons) (Nolan T et al., 2006). Amplicons are detected using fluorescence, increase in which is directly proportional to the amplified PCR product. The number of cycles required per sample to reach a certain threshold of fluorescence (set by the user – usually set in the linear phase of the amplification, and the observed difference in samples to cross the set threshold reflects the initial amount available for amplification) is used to quantify the relative amount in the samples. The amplified products are detected by the DNA intercalating minor groove-binding fluorophore SYBR green, which produces a signal when incorporated into double-stranded amplicons. Since the cDNA is single stranded, the dye does not bind enhancing the specificity of the results. There are other methods such as nested fluorescent probes for detection but SYBR green is widely used. RT-PCR primers specific to several pro-inflammatory mediators in several species including mouse, rat and humans, are readily available commercially.

ELISA assays – permit quantitative measurement of antigens in biological samples. The method is the same as described for the MIE. Both ELISA and qRT-PCR assays are used in vivo and are readily applicable to in vitro cell culture models, where cell culture supernatants or whole cell homogenates are used for ELISA or mRNA assays. Both assays are straight forward, quantitative and require relatively a small amount of input sample.

Apart from assaying single protein or gene at a time, cytokine bead arrays or cytokine PCR arrays can also be used to detect a whole panel of inflammatory mediators in a multiplex method (Husain et al., 2015b). This method is quantitative and especially advantageous when the sample amount available for testing is scarce. Lastly, immunohistochemistry can also be used to detect specific immune cell types producing the pro-inflammatory mediators and its downstream effectors in any given tissue (Costa et al., 2017). Immunohistochemistry results can be used as weight of evidence; however, the technique is not quantitative and depending on the specific antibodies used, the assay sensitivity may also become an issue (Amsen and De Visser, 2009).

Cell models - of varying complexity have been used to assess the expression of pro-inflammatory mediators. Two dimensional submerged monocultures of the main fibrotic effector cells – lung epithelial cells, macrophages, and fibroblasts – have routinely been used in vitro due to the large literature base, and ease of use, but do not adequately mimic the in vivo condition (Sundarakrishnan et al., 2018, Sharma et al., 2016). Recently, the EpiAlveolar in vitro lung model (containing epithelial cells, endothelial cells, and fibroblasts) was used to predict the fibrotic potential of MWCNT, and researchers noted increases in the pro-inflammatory molecules TNF-a, IL-1b, and the pro-fibrotic TGF-b using ELISA assays (Barasova et al., 2020). A similar, but less complicated co-culture model of immortalized human alveolar epithelial cells and IPF patient derived fibroblasts was used to assess pro-fibrotic signalling, and noted enhanced secretion of PDGF and bFGF (basic fibroblast growth factor), as well as evidence for epithelial to mesenchymal transition of epithelial cells in this system (Prasad et al., 2014). Models such as these better capitulate the in vivo pulmonary alveolar capillary, but have lower reproducibility as compared to traditional submerged mono-culture experiments.

Domain of Applicability

This free text section should be used to elaborate on the scientific basis for the indicated domains of applicability and the WoE calls (if provided). While structured terms may be selected to define the taxonomic, life stage and sex applicability (see structured applicability terms, above) of the KE, the structured terms may not adequately reflect or capture the overall biological applicability domain (particularly with regard to taxa). Likewise, the structured terms do not provide an explanation or rationale for the selection. The free-text section on evidence for taxonomic, life stage, and sex applicability can be used to elaborate on why the specific structured terms were selected, and provide supporting references and background information.  More help

Human, mouse, rat

Cytokines are the common pro-inflammatory mediators secreted following inflammogenic stimuli. Cytokines can be defined as diverse group of signaling protein molecules. They are secreted by different cell types in different tissues and in all mammalian species, irrespective of gender, age or sex. A lot of literature is available to support cross species, gender and developmental stage application for this KE. The challenge is the specificity; most cytokines exhibit redundant functions and many are pleotropic.

References

List of the literature that was cited for this KE description. Ideally, the list of references, should conform, to the extent possible, with the OECD Style Guide (https://www.oecd.org/about/publishing/OECD-Style-Guide-Third-Edition.pdf) (OECD, 2015). More help

1. Amsen, D. and De Visser, K. (2009). Approaches to Determine Expression of Inflammatory Cytokines. Methods in molecular biology.. 511th ed. (Clifton, NJ), pp.107-142.

2. Barosova, H., Maione, A. G., Septiadi, D., Sharma, M., Haeni, L., Balog, S., O'Connell, O., Jackson, G. R., Brown, D., Clippinger, A. J., Hayden, P., Petri-Fink, A., Stone, V., & Rothen-Rutishauser, B. (2020). Use of EpiAlveolar Lung Model to Predict Fibrotic Potential of Multiwalled Carbon Nanotubes. ACS nano, 14(4), 3941–3956.

3. Boyles, M. S., Young, L., Brown, D. M., MacCalman, L., Cowie, H., Moisala, A., Smail, F., Smith, P. J., Proudfoot, L., Windle, A. H., & Stone, V. (2015). Multi-walled carbon nanotube induced frustrated phagocytosis, cytotoxicity and pro-inflammatory conditions in macrophages are length dependent and greater than that of asbestos. Toxicology in vitro : an international journal published in association with BIBRA, 29(7), 1513–1528.

4. Costa, P. M., Gosens, I., Williams, A., Farcal, L., Pantano, D., Brown, D. M., Stone, V., Cassee, F. R., Halappanavar, S., & Fadeel, B. (2018). Transcriptional profiling reveals gene expression changes associated with inflammation and cell proliferation following short-term inhalation exposure to copper oxide nanoparticles. Journal of applied toxicology : JAT, 38(3), 385–397.

5. Halappanavar, S., Jackson, P., Williams, A., Jensen, K. A., Hougaard, K. S., Vogel, U., Yauk, C. L., & Wallin, H. (2011). Pulmonary response to surface-coated nanotitanium dioxide particles includes induction of acute phase response genes, inflammatory cascades, and changes in microRNAs: a toxicogenomic study. Environmental and molecular mutagenesis, 52(6), 425–439.

6. Hubbard, A., Timblin, C., Shukla, A., Rincón, M. and Mossman, B. (2002). Activation of NF-κB-dependent gene expression by silica in lungs of luciferase reporter mice. American Journal of Physiology-Lung Cellular and Molecular Physiology, 282(5), pp.L968-L975.

7. Husain, M., Kyjovska, Z. O., Bourdon-Lacombe, J., Saber, A. T., Jensen, K. A., Jacobsen, N. R., Williams, A., Wallin, H., Halappanavar, S., Vogel, U., & Yauk, C. L. (2015a). Carbon black nanoparticles induce biphasic gene expression changes associated with inflammatory responses in the lungs of C57BL/6 mice following a single intratracheal instillation. Toxicology and applied pharmacology, 289(3), 573–588.

8. Husain, M., Wu, D., Saber, A. T., Decan, N., Jacobsen, N. R., Williams, A., Yauk, C. L., Wallin, H., Vogel, U., & Halappanavar, S. (2015b). Intratracheally instilled titanium dioxide nanoparticles translocate to heart and liver and activate complement cascade in the heart of C57BL/6 mice. Nanotoxicology, 9(8), 1013–1022.

9. Kaminski N. (2003). Microarray analysis of idiopathic pulmonary fibrosis. American journal of respiratory cell and molecular biology, 29(3 Suppl), S32–S36.

10. Mestas, J., & Hughes, C. C. (2004). Of mice and not men: differences between mouse and human immunology. Journal of immunology (Baltimore, Md. : 1950), 172(5), 2731–2738.

11. Nolan, T., Hands, R. E., & Bustin, S. A. (2006). Quantification of mRNA using real-time RT-PCR. Nature protocols, 1(3), 1559–1582.

12. Park, S. J., & Im, D. S. (2019). Deficiency of Sphingosine-1-Phosphate Receptor 2 (S1P2) Attenuates Bleomycin-Induced Pulmonary Fibrosis. Biomolecules & therapeutics, 27(3), 318–326. https://doi.org/10.4062/biomolther.2018.131

13. Poland, C. A., Duffin, R., Kinloch, I., Maynard, A., Wallace, W. A., Seaton, A., Stone, V., Brown, S., Macnee, W., & Donaldson, K. (2008). Carbon nanotubes introduced into the abdominal cavity of mice show asbestos-like pathogenicity in a pilot study. Nature nanotechnology, 3(7), 423–428.

14. Prasad, S., Hogaboam, C. M., & Jarai, G. (2014). Deficient repair response of IPF fibroblasts in a co-culture model of epithelial injury and repair. Fibrogenesis & tissue repair, 7, 7.

15. Sharma, M., Nikota, J., Halappanavar, S., Castranova, V., Rothen-Rutishauser, B., & Clippinger, A. J. (2016). Predicting pulmonary fibrosis in humans after exposure to multi-walled carbon nanotubes (MWCNTs). Archives of toxicology, 90(7), 1605–1622.

16. Sundarakrishnan, A., Chen, Y., Black, L. D., Aldridge, B. B., & Kaplan, D. L. (2018). Engineered cell and tissue models of pulmonary fibrosis. Advanced drug delivery reviews, 129, 78–94.

17. Turner, M. D., Nedjai, B., Hurst, T., & Pennington, D. J. (2014). Cytokines and chemokines: At the crossroads of cell signalling and inflammatory disease. Biochimica et biophysica acta, 1843(11), 2563–2582.

18. Zuo, F., Kaminski, N., Eugui, E., Allard, J., Yakhini, Z., Ben-Dor, A., Lollini, L., Morris, D., Kim, Y., DeLustro, B., Sheppard, D., Pardo, A., Selman, M., & Heller, R. A. (2002). Gene expression analysis reveals matrilysin as a key regulator of pulmonary fibrosis in mice and humans. Proceedings of the National Academy of Sciences of the United States of America, 99(9), 6292–6297.