In mice, neonatal exposure to a synthetic estrogen, diethylstilbestrol (DES), induces polyovular follicles, which contain two or more oocytes per ovarian follicle. We reported previously that the estrogen receptor beta (ESR2) mediates DES signaling in polyovular follicle induction. However, the specific mechanism of polyovular follicle induction has not yet been clarified. Folliculogenesis in rodents begins soon after birth, accompanied by programmed oocyte death and germ cell loss. In this study, the effects of DES on oocyte death and on mRNA expression of genes thought to be involved in polyovular follicle induction were analyzed during a crucial period of folliculogenesis in the ovary of C57BL/6J, Fas(lpr/lpr) (lacking cell death receptor, FAS), and Esr2 knockout (Esr2 KO) mice. Neonatal DES exposure reduced programmed oocyte death in C57BL/6J mice; however, this reduction was not observed in Esr2 KO mice. In control Fas(lpr/lpr) mice, the oocyte apoptotic index was significantly lower than that in the control C57BL/6J mice. However, the polyovular follicle incidence in control 20-day-old Fas(lpr/lpr) mice was similar to that in the control C57BL/6J mice. Moreover, DES exposure changed mRNA expression of inhibin-alpha (Inha) in 2-day-old C57BL/6J mice. These results suggest that inhibition of oocyte death by DES through ESR2 may be one of the triggers for polyovular follicle induction. The FAS system is also involved in neonatal oocyte death; however, reduction of oocyte death is not sufficient for polyovular follicle induction. The combination of increased Inha mRNA and reduction of oocyte death in the ovaries of mice by DES through ESR2 might be correlated with polyovular follicle induction.

Natural and synthetic estrogens, Including diethylstilbestrol (DES), given during the critical period of newborn life Induce abnormalities in ovaries of mice. Induction of polyovular follicles (PFs) containing two or more oocytes in a follicle is one example. In this study, the involvement of estrogen receptor subtypes ERalpha and ERbeta in induction of PFs by neonatal treatment with DES was analyzed by using ERalpha knockout (alphaERKO) and ERbeta knockout (betaERKO) mice. Ovaries of mice injected with 3 microg DES for 5 days from the day of birth were examined histologically from 10 to 60 days of age, and the expression of genes involved in folliculogenesis was analyzed by real-time quantitative PCR. The PF Incidence (percent of PFs per 100 follicles greater than 50 microm in diameter) in the ovary of alphaERKO mice treated with DES was not different from that in the DES-treated wild-type mice. However, neonatal DES treatment did not increase the PF incidence in betaERKO mice, suggesting that PFs were induced by DES through ERbeta but not ERalpha. The expression of bone morphogenetic protein 15, growth differentiation factor 9, inhibin-alpha, Müllerian inhibiting substance, and other genes in the ovaries of DES-treated betaERKO mice was not different from that in the ovaries of DES-treated wild-type mice. These results indicate that ERbeta but not ERalpha is essential for DES to Induce PFs in mice.

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Abstract. Background/Aim: Neonatal diethylstilbestrol (DES) treatment induces polyovular follicles (PFs)