Effects of bisphenol A on apoptosis of ovarian preantral follicular granulosa cells and ovarian development in mice

Abstract

To investigate the effect of environmental estrogen bisphenol A (BPA) exposure on apoptosis of mouse ovarian preantral follicular granulosa cells and ovarian development and explore the underlying mechanism. Mouse ovarian preantral follicular granulosa cells were isolated from female ICR mice at postnatal day (PND) 10 and cultured in vitro. The cultured cells were treated with 0, 1, 10, 50, 100, 150, 200 and 500 μmol/L BPA, and the changes in cell proliferation, cell cycle, apoptosis and mitochondrial membrane potential were analyzed with CCK-8 method and flow cytometry. The protein expressions of Bcl-2, Bax, p53 and cyclin D1 in the treated cells were determined with Western blotting. Pregnant ICR mice were treated for a week with BPA at the concentration that produced significant effects on the preantral follicular granulosa cells, and the weight changes of the pregnant mice were recorded. The ovarian tissues of the offspring female mice were weighed at PND 10, 17, 21 and 42 followed by histological observation with HE staining and examination of Bcl-2 mRNA expression level with RT-qPCR. Compared with the control cells group, the isolated cells exposed to a low concentration of BPA (50 μmol/L) showed a significantly lowered apoptosis rate, increased mitochondrial membrane potential, and enhanced cellular proliferation (P < 0.05). Exposure to a higher BPA concentration at 200 μmol/L obviously enhanced cell apoptosis by reducing the mitochondrial membrane potential and repressed the cell proliferation (P < 0.05). BPA exposure at 50 μmol/L and 200 μmol/L produced opposite effects on the protein expressions of Bcl-2 (P < 0.01), Bax (P < 0.05) and p53 (P < 0.05) in mouse ovarian preantral follicular granulosa cells. BPA exposure at the doses of 10 and 35 mg/kg caused rapid weight increment of the pregnant mice and changes in ovarian index of the offspring female mice. In the offspring female mice, the changes in Bcl-2 mRNA expression in the ovarian tissue showed a similar pattern to that of ovarian index. Exposure of the pregnant mice to a high BPA concentration at 35 mg/kg resulted in accelerated follicular development into antral follicular stage in PND 21 offspring female mice. BPA can concentration-dependently regulate the function of ovarian preantral follicular granulosa cells in mice and potentially affects both the pregnant mice and the offspring female mice in light of early ovarian development.