# Cytomolecular and In Vitro Toxicity Studies on Bisphenol A Effect on Ovine Granulosa Cells

## TL;DR

This study shows how Bisphenol A affects sheep ovarian cells, reducing their viability and hormone production at higher doses.

## Contribution

The study reveals BPA's biphasic effect on ovine granulosa cells, showing both estrogenic and suppressive effects at different concentrations.

## Key findings

- BPA reduced cell viability and metabolic activity at concentrations above 10 μm.
- Estrogen and progesterone levels increased at low BPA concentrations but decreased at higher doses.
- BPA altered gene expression related to apoptosis and steroidogenesis in a concentration-dependent manner.

## Abstract

Bisphenol A (BPA), an organic environment chemical, has extensive presence in consumer goods and food/feed items. The present study aimed to explore how BPA influenced the viability, proliferation, apoptosis, and steroidogenic activity of ovine ovarian granulosa cells (GCs). GCs were isolated from ovine ovaries obtained from a local abattoir and were cultured for 2, 3, and 6 days in the presence of varying BPA concentrations (0, 1, 10, 25, 50, and 100 μm). The proliferation and cytotoxicity of the GCs were evaluated using kit assays, and the 72‐h culture’s spent media were pooled to measure the hormone concentrations. qPCR was performed for the study of gene expression–related apoptosis and steroidogenesis. For further confirmation of viability and apoptosis, the trypan blue exclusion test and Hoechst staining were performed. The findings revealed that the metabolic activity was significantly reduced at 50 μm, while the cell viability dropped notably (p < 0.05) at concentrations of 10 μm and above. Hormonal analysis indicated a biphasic response: estrogen and progesterone levels were significantly elevated at lower BPA concentrations (1 μm) but reduced from 25 μm onwards. Cytotoxicity assessments showed marked changes in LDH and GST activity at 50 μm and increased MDA and ROS levels at 25 and 10 μm. However, total antioxidant activity (CUPRAC) remained unchanged compared to control samples. Gene expression analysis revealed a significant upregulation of ESR1, ESR2, PGR, and FSHR at 1 μm; BAX and CASP3 at 25 μm; and 17βHSD and BCL2 at 50 μm. Conversely, a significant downregulation was observed for 3βHSD1 at 1 μm, CYP19A1 at 50 μm, and StAR at 100 μm, with no notable changes in CYP11A1 and CYP17A1. Overall, the study demonstrated that BPA adversely affects GCs by disrupting their growth, steroidogenic function, and gene expression, exhibiting estrogenic effects at lower doses and suppressing hormone secretion at higher concentrations.