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Event: 405

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

irregularities, ovarian cycle

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
irregularities, ovarian cycle

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

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
ovulation cycle disrupted

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
Aromatase (Cyp19a1) reduction leading to reproductive toxicity AdverseOutcome Elise Grignard (send email) Open for citation & comment EAGMST Under Review

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
mice Mus sp. Low 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

Sex Applicability

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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

Biological state

The female ovarian cycle is the result of a balanced cooperation between several organs and is determined by a complex interaction of hormones. Ovarian cycle irregularities include disturbances in the ovarian cycle (e.g. longer cycle, persistent estrus) and/or ovulation problems (deferred ovulation or anovulation). The estrous cycle (also oestrous cycle) comprises the recurring physiologic changes that are induced by reproductive hormones in females. Estrous cycles start after sexual maturity in females and are interrupted by anestrous phases or pregnancies. During this cycle numerous well defined and sequential alterations in reproductive tract histology, physiology and cytology occur, initiated and regulated by the hypothalamic-pituitary-ovarian (HPO) axis. The central feature of the mammalian estrous cycle is the periodic maturation of eggs that will be released at ovulation and luteinisation of the follicles after ovulation to form corpora lutea. Adapted from www.oecd.org/chemicalsafety/testing/43754807.pdf Biological compartments

The cyclic changes that occur in the female reproductive tract are initiated and regulated by the hypothalamic-pituitary-ovarian (HPO) axis. Although folliculogenesis occurs independently of hormonal stimulation up until the formation of early tertiary follicles, the gonadotrophins luteinising hormone (LH) and follicle stimulating hormone (FSH) are essential for the completion of follicular maturation and development of mature preovulatory (Graafian) follicles. The oestrous cycle consists of four stages: prooestrus, oestrus, metoestrus (or dioestrus 1) and dioestrus (or dioestrus 2) orchestrated by hormones. Levels of LH and FSH begin to increase just after dioestrus. Both hormones are secreted by the same secretory cells (gonadotrophs) in the pars distalis of the anterior pituitary (adenohypophysis). FSH stimulates the development of the zona granulosa and triggers expression of LH receptors by granulosa cells. LH initiates the synthesis and secretion of androstenedione and, to a lesser extent, testosterone by the theca interna; these androgens are utilised by granulosa cells as substrates in the synthesis of estrogen. Pituitary release of gonadotrophins thus drives follicular maturation and secretion of estrogen during prooestrus. Gonadotrophin secretion by the anterior pituitary is regulated by luteinising hormone-releasing hormone (LHRH), produced by the hypothalamus. LHRH is transported along the axons of hypothalamic neurones to the median eminence where it is secreted into the hypothalamic-hypophyseal portal system and transported to the anterior pituitary. The hypothalamus secretes LHRH in rhythmic pulses; this pulsatility is essential for the normal activation of gonadotrophs and subsequent release of LH and FSH. Adapted from www.oecd.org/chemicalsafety/testing/43754807.pdf

Follicles that produce estrogens have sequestered pituitary FSH which in turn stimulates the aromatase reaction. Such follicles can undergo normal development and ovulation and contain eggs that readily resume meiosis when released. In the absence of an active local aromatase (i.e., no follicle-stimulating hormone), the follicles and oocytes become atretic and regress without ovulating. If aromatase is present, the estrogen and follicle stimulating hormone can further develop the follicular cells for normal luteal function after ovulation takes place (Ryan, 1982).

General role in biology

A sequential progression of interrelated physiological and behavioural cycles underlines the female's successful production of young. In many but not all species the first and most basic of these is estrous cycle, which is itself a combination of cycles.

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). ?

Methods that have been previously reviewed and approved by a recognized authority should be included in the Overview section above. All other methods, including those well established in the published literature, should be described here. Consider the following criteria when describing each method: 1. Is the assay fit for purpose? 2. Is the assay directly or indirectly (i.e. a surrogate) related to a key event relevant to the final adverse effect in question? 3. Is the assay repeatable? 4. Is the assay reproducible?

The pattern of events in the estrous cycle may provide a useful indicator of the normality of reproductive neuroendocrine and ovarian function in the nonpregnant female. It also provides a means to interpret hormonal, histologic, and morphologic measurements relative to stage of the cycle, and can be useful to monitor the status of mated females. Regular cyclicity is one of the key parameters in assessment of female reproductive function in rodents. Parameters assessed for cyclicity: - Number of cycling females - Number of females with regular cycles - Number of cycles - Estrous cycle length - Percentage of time spent in the various estrous cycle stages Estrous cyclicity provides a method for evaluating the endocrine disrupting activity of each test chemical under physiologic conditions where endogenous concentrations of estrogen vary. Abnormal cycles were defined as one or more estrous cycles in the 21-day period with prolonged estrus (≥3 days) and/or prolonged metestrus or diestrus (≥4 days) within a given cycle (Goldman, Murr, & Cooper, 2007).

Estrous cycle normality can be monitored in the rat and mouse by observing the changes in the vaginal smear cytology. Visual observation of the vagina is the quickest method, requires no special equipment, and is best used when only proestrus or estrus stages need to be identified. For details see: (Westwood, 2008), (Byers, Wiles, Dunn, & Taft, 2012) and OECD guidelines (www.oecd.org).

The observation that animals do not ovulate while exhibiting estrous cycles indicates that estrous cyclicity alone may not be a sufficient surrogate of healthy function of ovaries; the measurements of serum hormones and particularly FSH can contribute to more sensitivity indicators of healthy function of ovaries (Davis, Maronpot, & Heindel, 1994).

Monitoring of oestrus cyclicity is included in OECD test guidelines (Test No. 407: Repeated Dose 28-day Oral Toxicity Study in Rodents, 2008) [1], (Test No. 416: Two-Generation Reproduction Toxicity, 2001)[2] and (Test No. 443: Extended One-Generation Reproductive Toxicity Study, 2012) [3]and in USA EPA OCSPP 890.1450.

In vitro testing

The follicle culture models were developed for the in-vitro production of mature oocytes and used to study the process of folliculogenesis and oogenesis in vitro (Cortvrindt & Smitz, 2002). These in vitro cultures demonstrate near-identical effects to those found in vivo, therefore might be able to acquire a place in fertility testing, replacing some in-vivo studies for ovarian function and female gamete quality testing (Stefansdottir, Fowler, Powles-Glover, Anderson, & Spears, 2014).

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

The estrous cycle comprises the recurring physiologic changes that are induced by reproductive hormones in most mammalian females. Many of the mechanisms involved in the regulation of the reproductive axis are similar across species (particularly those mediated through the estrogen receptor), assessments of rodent estrous cyclicity can offer insight into potential adverse effects in humans (Goldman, Murr, & Cooper, 2007). While evaluations of vaginal cytology in the laboratory rodent can provide a valuable reflection of the integrity of the hypothalamic-pituitary-ovarian axis, other indices are more useful in humans to determine the functional status of the reproductive system (e.g. menses, basal body temperature, alterations in vaginal pH, cervical mucous viscosity, and blood hormone levels). Nevertheless, since many of the mechanisms involved in the regulation of the reproductive axis are similar across species (particularly those mediated through the estrogen receptor), assessments of rodent estrous cyclicity can offer insight into potential adverse effects in humans (Rasier, Toppari, Parent, & Bourguignon, 2006).

Regulatory Significance of the Adverse Outcome

An AO is a specialised KE that represents the end (an adverse outcome of regulatory significance) of an AOP. For KEs that are designated as an AO, one additional field of information (regulatory significance of the AO) should be completed, to the extent feasible. If the KE is being described is not an AO, simply indicate “not an AO” in this section.A key criterion for defining an AO is its relevance for regulatory decision-making (i.e., it corresponds to an accepted protection goal or common apical endpoint in an established regulatory guideline study). For example, in humans this may constitute increased risk of disease-related pathology in a particular organ or organ system in an individual or in either the entire or a specified subset of the population. In wildlife, this will most often be an outcome of demographic significance that has meaning in terms of estimates of population sustainability. Given this consideration, in addition to describing the biological state associated with the AO, how it can be measured, and its taxonomic, life stage, and sex applicability, it is useful to describe regulatory examples using this AO. More help

Chemicals may be found to interfere with reproductive function in the female rat. This interference is commonly expressed as a change in normal morphology of the reproductive tract or a disturbance in the duration of particular phases of the estrous cycle. This key event lies within the scope of testing for endocrine disrupting activity of chemicals and therefore for testing of female reproductive and developmental toxicity. Monitoring of oestrus cyclicity is included in OECD test guidelines (Test No. 407: Repeated Dose 28-day Oral Toxicity Study in Rodents, 2008), (Test No. 416: Two-Generation Reproduction Toxicity, 2001) and (Test No. 443: Extended One-Generation Reproductive Toxicity Study, 2012) and in USA EPA OCSPP 890.1450. While an evaluation of the estrous cycle in laboratory rodents can be a useful measure of the integrity of the hypothalamic-pituitary-ovarian reproductive axis, it can also serve as a way of insuring that animals exhibiting abnormal cycling patterns are excluded from a study prior to exposure to a test compound. When incorporated as an adjunct to other endpoint measures, a determination of a female's cycling status can contribute important information about the nature of a toxicant insult to the reproductive system. In doing so, it can help to integrate the data into a more comprehensive mechanistic portrait of the effect, and in terms of risk assessment, may provide some indication of a toxicant's impact on human reproductive physiology. Significant evidence that the estrous cycle (or menstrual cycle in primates) has been disrupted should be considered an adverse effect (OECD, 2008). Included should be evidence of abnormal cycle length or pattern, ovulation failure, or abnormal menstruation.

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

Byers, S. L., Wiles, M. V, Dunn, S. L., & Taft, R. A. (2012). Mouse estrous cycle identification tool and images. PloS One, 7(4), e35538. doi:10.1371/journal.pone.0035538

Cortvrindt, R. G., & Smitz, J. E. J. (2002). Follicle culture in reproductive toxicology: a tool for in-vitro testing of ovarian function? Human Reproduction Update, 8(3), 243–54.

Davis, B. J., Maronpot, R. R., & Heindel, J. J. (1994). Di-(2-ethylhexyl) phthalate suppresses estradiol and ovulation in cycling rats. Toxicology and Applied Pharmacology, 128(2), 216–23. doi:10.1006/taap.1994.1200

Goldman, J. M., Murr, A. S., & Cooper, R. L. (2007). The rodent estrous cycle: characterization of vaginal cytology and its utility in toxicological studies. Birth Defects Research. Part B, Developmental and Reproductive Toxicology, 80(2), 84–97. doi:10.1002/bdrb.20106

OECD. (2008). No 43: Guidance document on mammalian reproductive toxicity testing and assessment.

Rasier, G., Toppari, J., Parent, A.-S., & Bourguignon, J.-P. (2006). Female sexual maturation and reproduction after prepubertal exposure to estrogens and endocrine disrupting chemicals: a review of rodent and human data. Molecular and Cellular Endocrinology, 254-255, 187–201. doi:10.1016/j.mce.2006.04.002

Ryan, K. J. (1982). Biochemistry of aromatase: significance to female reproductive physiology. Cancer Research, 42(8 Suppl), 3342s–3344s.

Stefansdottir, A., Fowler, P. A., Powles-Glover, N., Anderson, R. A., & Spears, N. (2014). Use of ovary culture techniques in reproductive toxicology. Reproductive Toxicology (Elmsford, N.Y.), 49C, 117–135. doi:10.1016/j.reprotox.2014.08.001

Test No. 407: Repeated Dose 28-day Oral Toxicity Study in Rodents. (2008). OECD Publishing. doi:10.1787/9789264070684-en

Test No. 416: Two-Generation Reproduction Toxicity. (2001). OECD Publishing. doi:10.1787/9789264070868-en

Test No. 443: Extended One-Generation Reproductive Toxicity Study. (2012). OECD Publishing. doi:10.1787/9789264185371-en

Westwood, F. R. (2008). The female rat reproductive cycle: a practical histological guide to staging. Toxicologic Pathology, 36(3), 375–84. doi:10.1177/0192623308315665