This Key Event Relationship is licensed under the Creative Commons BY-SA license. This license allows reusers to distribute, remix, adapt, and build upon the material in any medium or format, so long as attribution is given to the creator. The license allows for commercial use. If you remix, adapt, or build upon the material, you must license the modified material under identical terms.

Relationship: 3717

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

Increased, E2 production in ovaries leads to Plasma E2, increased

Upstream event

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

Increased, E2 production in ovaries

Downstream event

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

Plasma E2, increased

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
Activation, estrogen receptor alpha leads to persistent vaginal cornification via increased kisspeptin release	adjacent	High		John Frisch (send email)	Under development: Not open for comment. Do not cite

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
mammals	mammals	Moderate	NCBI

Sex Applicability

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

Sex	Evidence
Female	High

Life Stage Applicability

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

Term	Evidence
Adult, reproductively mature	Moderate
Juvenile	Moderate

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

Production of estradiol (E2) by the ovaries has been well-established by the two-cell, two gonadotropin model of steroid biosynthesis (for review see Drummond 2006; Kimura et al. 2007; Palermo 2007; Beevors et al. 2024). Luteinizing hormone stimulates steroid production in theca cells, with follicle-stimulating hormone stimulates steroid production in granulosa cells.

Estradiol is a key signalling hormone, with increased production of estradiol in the ovary leading to increased secretion of estradiol into plasma.

Table 1: List of steroid synthesis enzymes with identifier of enzyme (Uniprot, 2025).

*Enzyme*	*Identifier*
Steroidogenic acute regulatory protein, mitochondrial (STAR)
Cholesterol side-chain cleavage enzyme, mitochondrial (CYP11A)	EC:1.14.15.6
3 beta-hydroxysteroid dehydrogenase (3B-HSD)	EC:1.1.1.145
Steroid 17-alpha-hydroxylase (CYP17A1)	EC:1.14.14.19
17-beta-hydroxysteroid dehydrogenase (17B-HSD)	EC:1.1.1.105
Aromatase (CYP19A1)	EC:1.14.14.14
3-oxo-5-alpha-steroid 4-dehydrogenase 2 (SRD5A2)	EC:1.3.1.22

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

This Key Event Relationship was part of an Environmental Protection Agency effort to develop AOPs that establish scientifically supported causal linkages between alternative endpoints measured using new approach methodologies (NAMs) and guideline apical endpoints measured in Tier 1 and Tier 2 test guidelines (U.S. EPA, 2024) employed by the Endocrine Disruptor Screening Program (EDSP). A series of key events that represent significant, measurable, milestones connecting molecular initiation to apical endpoints indicative of adversity were identified based on scientific review articles and empirical studies. Additionally, scientific evidence supporting the causal relationships between each pair of key events was assembled and evaluated. The present effort focused primarily on empirical studies with laboratory rodents and other mammals.

Empirical studies are focused on increased estradiol production in ovaries and resulting increased plasma estradiol, in support of development of AOP 623.

Authors of KER 3717 did a further evaluation of published peer-reviewed literature to provide additional evidence in support of the key event relationship. The literature used to support this KER began with the test guidelines and followed to primary, secondary, and/or tertiary works concerning the relevant underlying biology. In addition, search engines were used to target journal articles with term ‘Estradiol’ to locate representative empirical studies that support the key event relationship.

Evidence Supporting this KER

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

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

Increased estradiol production in gonads and resulting increased plasma estradiol have been studied in laboratory mammals by addition of hormones (Sashinda and Johnson 1976) and toxicants with endocrine disrupting properties (Li et al. 2018; Gan et al. 2024). Studies that link increases in steroidogenic enzymes in ovaries to increased estradiol levels in plasma have been useful in establishing plausibility (Li et al. 2018; Gan et al. 2024). Ovaries have been established as the primary source of estradiol in females by the two-cell, two gonadotropin model of steroid biosynthesis (Drummond 2006; Kimura et al. 2007; Palermo 2007; Beevors et al. 2024).

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

Species	Duration	Dose	Increased E2 production ovaries?	Increased plasma estradiol?	Summary	Citation
Rats (Rattus norvegicus)	24 hours	10 ug/L FSH and 1 ug/L LH at 2 hour intervals.	yes	yes	Female rats injected with FSH and LH had increased estradiol production in ovaries leading to increased plasma estradiol.	Sashinda and Johnson (1976)
Rats (Rattus norvegicus)	3 generations	1, 5 mg/kg BW Cadmium chloride	yes	yes	Female rats exposed to cadmium chloride had increased estradiol production in ovaries and statistically significant increased levels of steroidogenic enzymes (STAR, CYP11A1, 3B-HSD, CYP19A1) primarily at 5 mg/kg BW leading to statistically significant increased plasma estradiol at 5 mg/kg BW.	Li et al. (2018)
Mice (Mus musculus)	Until post natal day 55	30, 300, 3000 ug/kg/d triclosan	yes	yes	Female mice exposed to triclosan increased estradiol production in ovaries by increased statistically significant expression of steroidogenic enzyme genes (STAR at >= 300 ug/kg/d, CYP11A1 at >= 30 ug/kg/d, CYP17A1 at 3000 ug/kg/d, CYP19A1 at >= 300 ug/kg/d) leading to statistically significant increased plasma estradiol at >= 300 ug/kg/d.	Gan et al. (2024)

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

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

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

Response-response Relationship

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

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

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

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

Life Stage: Applies to adult, reproductively mature and juveniles.

Sex: Applies to females as specific to ovaries.

Taxonomic: Primarily studied in humans and laboratory rodents. Plausible for most mammals due to conserved hormone pathways regulating hypothalamus-pituitary-gonadal axis processes. Estradiol production in ovaries widespread among vertebrates, including mammals (Bondesson et al. 2015), birds (Hanlon et al. 2022), fish (Li et al. 2019), reptiles (Cruz-Cano et al. 2023), and amphibians (Bondesson et al. 2015), with corresponding increases in plasma estradiol resulting from release of estradiol from ovaries.

References

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

Beevors LI, Sundar S, Foster PA. 2024. Steroid metabolism and hormonal dynamics in normal and malignant ovaries. Essays in Biochemistry 68(4): 491-507.

Bondesson M, Hao R, Lin CY, Williams C, Gustafsson JA. 2015. Estrogen receptor signaling during vertebrate development. Biochimica et Biophysica Acta 1849(2): 142-151.

Cruz-Cano NB, Sanchez-Rivera UA, Alvarez-Rodriguez C, Cardenas-Leon M, Martinez-Torres M. 2023. Sex steroid receptors in the ovarian follicles of the lizard Sceloporus torquatus. Zygote. 31(4): 386-392.

Drummond AE. 2006. The role of steroids in follicular growth. Reproductive Biology and Endocrinology 4:16.

Gan H, Lan H, Hu Z, Zhu B, Sun L, Jiang Y, Wu L, Liu J, Ding Z, Ye X. 2024. Triclosan induces earlier puberty onset in female mice via interfering with L-type calcium channels and activating Pik3cd. Ecotoxicology and Environmental Safety 269: 115772.

Hanlon C, Ziezold CJ, Bedecarrats GY. 2022. The Diverse Roles of 17β-Estradiol in Non-Gonadal Tissues and Its Consequential Impact on Reproduction in Laying and Broiler Breeder Hens. Frontiers in Physiology 13: 942790.

Kimura S, Matsumoto T, Matsuyama R, Shiina H, Sato T, Takeyama K, Kato S. 2007. Androgen receptor function in folliculogenesis and its clinical implication in premature ovarian failure. Trends in Endocrinology and Metabolism 18(5): 183-189.

Li M, Sun L, Wang D. 2019. Roles of estrogens in fish sexual plasticity and sex differentiation. General and Comparative Endocrinology 277: 9-16.

Li Z, Li T, Leng Y, Chen S, Liu Q, Feng J, Chen H, Huang Y, Zhang Q. 2018. Hormonal changes and folliculogenesis in female offspring of rats exposed to cadmium during gestation and lactation. Environmental Pollution 238: 336-347.

Palermo R. 2007. Differential actions of FSH and LH during folliculogenesis. Reproductive BioMedicine Online 15(3): 326-337.

Sashida T, Johnson DC. 1976. Stimulation of the estrogen synthesizing system of the immature rat ovary by exogenous and endogenous gonadotropins. Steroids 27(4): 469-79.

U.S. Environmental Protection Agency. 2004. EDSP Test Guidelines and Guidance Document. https://www.epa.gov/test-guidelines-pesticides-and-toxic-substances/edsp-test-guidelines-and-guidance-document (retrieved 25 July 2025).

The UniProt Consortium. UniProt: the Universal Protein Knowledgebase in 2025. https://www.uniprot.org/ (retrieved 2 November 2025)..

Italics indicate edits from John Frisch February 2026. A full list of updates can be found in the Change Log on the View History page.