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Event: 914

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

Decrease, Apoptosis of ciliated epithelial cells

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

Decreased ciliated cell apoptosis

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
ciliated epithelial cell

Organ term

Organ term
lung

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
epithelial cell apoptotic process		decreased

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
Decreased lung function	KeyEvent	Karsta Luettich (send email)	Under development: Not open for comment. Do not cite	Under Development

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

Name
Cigarette smoke

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
human	Homo sapiens	Moderate	NCBI
mouse	Mus musculus	Moderate	NCBI
rat	Rattus norvegicus	Moderate	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
Adult	Moderate

Sex Applicability

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

Term	Evidence
Mixed	Moderate

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

Apoptosis, a main form of programmed cell death, is an essential part of tissue homeostasis during all life stages. Under normal conditions, apoptosis for example serves to eliminate damaged or infected cells and enable wound repair and resolution of inflammation (Elmore, 2007). In the airways, apoptosis occurs following exposure to cigarette smoke (Mebratu et al., 2011, Ning et al., 2013; Valencia-Gattas et al., 2016), hydrogen peroxide (Goldkorn et al., 1998), ozone (Triantaphyllopoulos et al., 2011); endotoxin (Tesfaigzi et al., 2000); infectious agents (Rajan et al., 2000; Monick et al., 2005) and ovalbumin sensitization (Truong-Tran et al., 2002; Reader et al., 2003; Takeyama et al., 2008; Liu et al., 2017). Ciliated cell apoptosis in lung epithelium is regulated by EGFR and PI3K, with both their activation resulting in decreased apoptosis (Tyner et al., 2006).

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). ?

Numerous apoptosis assays exist. They typically are based on the analysis of the characteristic features an apoptotic cell exhibits. For example, decreased DNA content, nuclear condensation and other morphological changes can be detected by flow cytometry (sub-G1 DNA content), Trypan Blue, or Hoechst staining. Electrophoresis or TUNEL assay can be used to determine the extent of DNA fragmentation. Annexin V staining can be used to confirm redistribution of phosphatidylserine from the inner plasma membrane to the extracellular surface. Additional propidium iodide staining would allow for a distinction between early and late apoptotic events. In addition, activation of caspases can be examined by caspase activity assays, and cytochrome c release from mitochondria during the early stages of apoptosis can be measured by ELISA. Finally, taking into consideration that the ratio between anti-apoptotic and pro-apoptotic proteins determines whether a cell lives or dies, examining the levels of BAX, BID, BAK, BIK, or BAD (pro-apoptotic) and Bcl-Xl and Bcl-2 (anti-apoptotic) by western blot and image analysis may also give an insight into cell fate (Bossy-Wetzel and Green, 2000; Oancea et al., 2006; Archana et al., 2013; Muppidi et al., 2004; Huerta et al., 2007; Loo, 2011; Kale et al., 2018). Inclusion of appropriate controls allows for quantitation of surviving or proliferating cell populations, as well as the extent and mode of cell death.

To support cell death in a specific subpopulation of the airway epithelium such as in ciliated epithelial cells would require identifying the cell type of interest first. This could be achieved by either staining the cell with a population-specific marker in situ, or by enriching the population of interest prior to monitoring cell proliferation and death. To the best of our knowledge, to date, only the double immunohistochemical/immunofluorescent staining of airway epithelial samples has been employed to demonstrate increased proliferation of goblet cells and decreased apoptosis of goblet or ciliated cells (Tyner et al., 2006; Takeyama et al., 2001; Tesfaigzi et al., 2004).

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

Ciliated cell apoptosis has been shown in a few mouse and human studies (Tyner et al., 2006; Martínez-Girón and Martínez-Torre, 2011). Epithelial cell apoptosis (presumably including ciliated cells) has been shown in numerous studies in mouse, rat, human (Monick et al., 2005; Hart et al., 1999; Matute-Bello et al., 1999; Rajan et al., 2000; Reader et al., 2003; Triantaphyllopoulos et al., 2011; Ning et al., 2013; Valencia-Gattas et al., 2016; Yang et al., 2013; Liu et al., 2017).

Evidence for Perturbation by Stressor

Cigarette smoke

Bik mRNA expression was significantly reduced in bronchial brushings and lung tissues of subjects with chronic bronchitis compared with nondiseased control subjects, and in C57BL/6 mice that were exposed to 250 mg/m³ of mainstream cigarette smoke for 6 h a day, 5 days a week, for 10 weeks. In addition, Bik mRNA and protein expression were significantly reduced in human airway epithelial cells differentiated at the air–liquid interface and treated with cigarette smoke extract (1,000 ng/ml total particulate matter) for 24 h at 5 days post-exposure (Mebratu et al., 2011).

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

Archana, M., Yogesh, T. L., & Kumaraswamy, K. L. (2013). Various methods available for detection of apoptotic cells-A review. Indian J. Cancer 50, 274.

Bossy-Wetzel, E., and Green, D.R. (2000). Assays for cytochrome c release from mitochondria during apoptosis. Methods Enzymol. 322, 235-242.

Elmore, S. (2007). Apoptosis: A Review of Programmed Cell Death. Toxicol. Pathol. 35, 495–516.

Goldkorn, T., Balaban, N., Matsukuma, K., Chea, V., Gould, R., Last, J., Chan, C., and Chavez, C. (1998). EGF-receptor phosphorylation and signaling are targeted by H2O2 redox stress. Am. J. Resp. Cell. Mol. Biol. 19, 786-798.

Hart, B.A., Lee, C.H., Shukla, G.S., Shukla, A., Osier, M., Eneman, J.D., and Chiu, J.F. (1999). Characterization of cadmium-induced apoptosis in rat lung epithelial cells: evidence for the participation of oxidant stress. Toxicology 133, 43–58.

Huerta, S., Goulet, E.J., Huerta-Yepez, S., and Livingston, E.H. (2007). Screening and detection of apoptosis. J. Surg. Res. 139, 143-156.

Kale, J., Osterlund, E.J., and Andrews, D.W. (2018). BCL-2 family proteins: changing partners in the dance towards death. Cell Death Differ. 25, 65-80.

Liu, Y., Pu, Y., Li, D., Zhou, L., & Wan, L. (2017). Azithromycin ameliorates airway remodeling via inhibiting airway epithelium apoptosis. Life Sci. 170, 1-8.

Loo, D.T. (2011). In situ detection of apoptosis by the TUNEL assay: an overview of techniques. Methods Mol. Biol. 682, 3-13.

Martínez-Girón, R., and Martínez-Torre, S. (2011). Apoptotic ciliated cells on sputum smear. Diagn. Cytopathol. 39, 941–942.

Matute-Bello, G., Liles, W.C., Steinberg, K.P., Kiener, P.A., Mongovin, S., Chi, E.Y., Jonas, M., and Martin, T.R. (1999). Soluble Fas ligand induces epithelial cell apoptosis in humans with acute lung injury (ARDS). J. Immunol. 163, 2217–2225.

Mebratu, Y.A., Schwalm, K., Smith, K.R., Schuyler, M., and Tesfaigzi, Y. (2011). Cigarette Smoke Suppresses Bik To Cause Epithelial Cell Hyperplasia and Mucous Cell Metaplasia. Am. J. Resp. Crit. Care Med. 183, 1531-1538.

Monick, M.M., Cameron, K., Staber, J., Powers, L.S., Yarovinsky, T.O., Koland, J.G., and Hunninghake, G.W. (2005). Activation of the Epidermal Growth Factor Receptor by Respiratory Syncytial Virus Results in Increased Inflammation and Delayed Apoptosis. J. Biol. Chem. 280, 2147–2158.

Muppidi, J., Porter, M., and Siegel, R.M. (2004). Measurement of apoptosis and other forms of cell death. Curr. Protoc. Immunol. 59, 3-17.

Ning, Y., Shang, Y., Huang, H., Zhang, J., Dong, Y., Xu, W., & Li, Q. (2013). Attenuation of cigarette smoke-induced airway mucus production by hydrogen-rich saline in rats. PLoS One 8, e83429.

Oancea, M., Mazumder, S., Crosby, M.E., and Almasan, A. (2006). Apoptosis assays. Methods Mol. Med. 129, 279-290.

Rajan, S., Cacalano, G., Bryan, R., Ratner, A. J., Sontich, C. U., van Heerckeren, A., et al. (2000). Pseudomonas aeruginosa induction of apoptosis in respiratory epithelial cells: analysis of the effects of cystic fibrosis transmembrane conductance regulator dysfunction and bacterial virulence factors. Am. J. Resp. Cell. Mol. Biol. 23, 304-312.

Reader, J.R., Tepper, J.S., Schelegle, E.S., Aldrich, M.C., Putney, L.F., Pfeiffer, J.W., and Hyde, D.M. (2003). Pathogenesis of mucous cell metaplasia in a murine asthma model. Am. J. Pathol. 162, 2069-2078.

Takeyama, K., Fahy, J., and Nadel, J. (2001). Relationship of epidermal growth factor receptors to goblet cell production in human bronchi. Am. J. Respir. Crit. Care Med. 163, 511-516.

Takeyama, K., Tamaoki, J., Kondo, M., Isono, K., and Nagai, A. (2008). Role of epidermal growth factor receptor in maintaining airway goblet cell hyperplasia in rats sensitized to allergen. Clin. Exp. Allergy 38, 857-865.

Tesfaigzi, Y., Fischer, M.J., Martin, A.J., and Seagrave, J. (2000). Bcl-2 in LPS- and allergen-induced hyperplastic mucous cells in airway epithelia of Brown Norway rats. Am. J. Physiol. Lung Cell. Mol. Physiol. 279, L1210-L1217.

Triantaphyllopoulos, K., Hussain, F., Pinart, M., Zhang, M., Li, F., Adcock, I., et al. (2011). A model of chronic inflammation and pulmonary emphysema after multiple ozone exposures in mice. Am. J. Physiol. Lung Cell. Mol. Physiol. 300, L691-L700.

Truong-Tran, A. Q., Ruffin, R. E., Foster, P. S., Koskinen, A. M., Coyle, P., Philcox, J. C., et al. (2002). Altered zinc homeostasis and caspase-3 activity in murine allergic airway inflammation. Am. J. Resp. Cell. Mol. Biol. 27, 286-296.

Tyner, J., Tyner, E., Ide, K., Pelletier, M., Roswit, W., Morton, J., Battaile, J., Patel, A., Patterson, G., Castro, M., et al. (2006). Blocking airway mucous cell metaplasia by inhibiting EGFR antiapoptosis and IL-13 transdifferentiation signals. J. Clin. Invest. 116, 309–321.

Valencia-Gattas M, Conner GE, Fregien NL (2016). Gefitinib, an EGFR Tyrosine Kinase inhibitor, Prevents Smoke-Mediated Ciliated Airway Epithelial Cell Loss and Promotes Their Recovery. Ahmad S, ed. PLoS ONE 11, e0160216.

Yang, Y.-X., Li, X.-L., Wang, L., Han, S.-Y., Zhang, Y.-R., Pratheeshkumar, P., Wang, X., Lu, J., Yin, Y.-Q., Sun, L.-J., et al. (2013). Anti-apoptotic proteins and catalase-dependent apoptosis resistance in nickel chloride-transformed human lung epithelial cells. Int. J. Oncol. 43, 936–946.