# Avian-Specific Evidence for an Estrogen Receptor Agonism Adverse Outcome Pathway Based on Chicken Embryos and LMH 3D Spheroids Exposed to Ethinylestradiol and Bisphenol A

## TL;DR

This study shows how estrogen receptor agonists like EE2 and BPA disrupt reproduction in chickens by altering gene expression and gonad development.

## Contribution

The study provides avian-specific evidence for an AOP linking estrogen receptor agonism to reduced fecundity in birds.

## Key findings

- EE2 and BPA increased plasma vitellogenin (VTG) and dysregulated estrogen-responsive genes in chicken embryos.
- Exposure to EE2 and BPA caused gonadal developmental impairments in male embryos, including oocyte-type cells and loss of testicular cords.
- LMH spheroids showed altered VTG concentrations and gene expression after exposure to EE2 and BPA.

## Abstract

Several adverse outcome pathways (AOPs) describe the
effects of
endocrine disrupting compounds on estrogen signaling. Substantial
data support an AOP related to estrogen receptor (ER) antagonism,
leading to decreased fecundity in fish. In this study, data were generated
for an ER agonism AOP leading to reduced fecundity in avian species
(AOP537). Chicken embryos and the chicken leghorn male hepatoma cell
line, LMH, were used to elucidate key events associated with estrogen
signaling following exposure to 17α-ethinylestradiol (EE2) and
bisphenol A (BPA). Embryos were exposed via egg injection. Viability
and hepatic estrogen-responsive gene expression data were collected
at midincubation (embryonic day [ED] 11). Changes in plasma vitellogenin
(VTG), gonad morphology and growth were evaluated prior to pipping
(ED20). Both chemicals dysregulated estrogen-responsive genes in hepatic
tissue and increased plasma VTG concentrations. In LMH spheroids,
EE2 and BPA altered estrogen-responsive genes and VTG concentrations
at 24 and 48 h, respectively. Gonadal histology revealed oocyte-type
cells and loss of testicular cords in male embryos exposed to EE2
and BPA. Overall, EE2 and BPA upregulated VTG mRNA expression, increased
plasma VTG, and caused impairments in gonadal development. These results
contribute avian-specific evidence to support an endocrine disruption
AOP describing the relationship between disrupted VTG synthesis and
impaired reproduction.