Xenoestrogens Exposure At Environmental Relevant Doses Adversely Affects Ovine Follicular Development

RIVERA, O; VARAYOUD,J; RODRIGUEZ,H; DI MAURO, C; MUÑOZ DE TORO, MÓNICA; LUQUE, ENRIQUE H

Congreso; XXXVI Annual Conference of the Intenacional Embryo tranfer society; 2009

The exposure to endocrine disrupters may affect reproduction. Our hypothesis suggested that neonatal endocrine disrupter exposure affects ovarian ovine development. Female lambs were s.c. exposed from postnatal Day (PND) 1 to PND 14 to environmentally relevant doses of diethylstilbe- strol (DES; 5 μg kg-1, n = 11), bisphenol A (BPA; 50 μg kg-1, n = 11), or vehicle [control (C), n = 14]. On PND 30, the ovaries were weighed and paraffin-embedded. The whole ovaries were sectioned in sets of 4 adjacent 5-μm serial sections taken 200 μm apart. Follicular dynamics were established by histomorphological features on pricosirius-hematoxylin-stained sections (at least 10 sections/ovary). The total number of fol- licles/ovary was obtained considering the number of sections and section thickness (≥25 000 follicles/ovary). Follicles of different classes and multioocyte follicles were expressed as a percentage. Immunohistochemical expression of estrogen receptor alpha (ERa), androgen receptor (AR), Ki-67 (proliferation marker), and p27 (cyclin-dependent kinase inhibitor 1B) was evaluated. Proliferative index was obtained using Ki-67 expression in the granulose cells (GC) and theca cells of antral follicles. Antral follicles with ≤1% Ki-67 were considered atretic. Kruskal-Wallis and Dunn post-tests were used; P < 0.05 was accepted as significant. In the developing ovaries (PND 1, 5, and 10; n = 3 for each day), the stromal cells showed a high expression of ERa on PND 1, decreasing with age. Estrogen receptor beta (ERfi) was positive in oocytes, CG, and stromal cells, with a temporal pattern opposite to that of ERa. Androgen receptor was found in oocytes, GC, and stromal cells without changes along the studied ages. Results at PND 30 showed that both xenostrogens adversely affect ovarian development. The BPA treatment reduced the ovarian weight without changes in the total number of follicles. Lambs exposed to DES and BPA showed a reduction of the percentage of primordial follicles (C = 59.4 ± 2.9; DES = 36.9 ± 7.3; BPA = 49.8 ± 1.6) but an increase of transitory (C = 29.2 ± 1.8; DES =48.1 ± 4.4; BPA = 41.9 ± 2.5) and pri- mary (C = 1.5 ± 0.2; DES = 5.1 ± 2.0; BPA = 2.9 ± 0.5) follicles. In addition, xenoestrogen treatment increased the number of antral atretic follicles (C = 16.3%; DES = 27.2%; BPA = 49.1%) associated with an increased p27 expression. Besides that, BPA group showed a higher proliferation of GC (C = 12.3 ± 2.6; DES = 31.5 ± 5.3; BPA = 37.0 ± 6.8) and theca cells (C = 1.0 ± 0.3; DES = 7.6 ± 1.9; BPA = 15.1 ± 4.2) in antral follicles and an increased incidence of multioocyte follicles (C = 4.2 ± 1.1; DES = 8.1 ± 2.0; BPA = 18.2 ± 3.9). Our results show that an early exposure to environmentally relevant doses of BPA or DES disrupts the ovine ovarian development and suggest that these effects may be mediated through the early expression of ovarian ER. Xenoestrogen exposure could effect ovarian function with consequences on female fertility.