Inhibition of oocyte growth factors in vivo modulates ovarian folliculogenesis in neonatal and immature mice.

Growth differentiation factor-9 (GDF9) and bone morphogenetic protein-15 (BMP15) are among the key regulators transmitting the signaling between the oocyte and the surrounding granulosa cells. Previously, it has been shown that a recombinant BMP type II receptor ectodomain-Fc fusion protein (BMPR2ecd-Fc) is able to inhibit the actions of GDF9 and BMP15 in vitro. Here, we have produced bioactive BMPR2ecd-Fc, which was injected i.p. into neonatal mice. Early folliculogenesis was first studied by injecting mice five times with various doses of BMPR2ecd-Fc during the postnatal days 4-12. Folliculogenesis was affected dose dependently, as evidenced by a decreased mitogenesis of granulosa cells of the growing follicles. Furthermore, we also noticed a decrease in the number of secondary and tertiary follicles as well as an increase in the oocyte size. Electron microscopic analysis revealed that the ultrastructure of the granulosa cells of the primary follicles was not affected by the BMPR2ecd-Fc treatment. A second study was conducted to investigate whether a longer treatment with 12 injections during postnatal days 4-28 would inhibit folliculogenesis. Similar effects were observed in the two studies on the early follicular developmental stages. However, in the long-term study, later stages of folliculogenesis were not blocked but rather increased numbers of antral follicles, preovulatory follicles, and corpora lutea were found. We conclude that BMPR2ecd-Fc is a potent modulator of ovarian folliculogenesis in vivo, and thus, is a valuable tool for studying the physiology and downstream effects of oocyte-derived growth factors in vivo.

Regulation of membrane progestin receptors in the zebrafish ovary by gonadotropin, activin, TGF-beta and BMP-15.

Progestin hormones are vital for inducing oocyte maturation in fish by binding to membrane progestin receptors (mPRs). The aim of this study was to examine the expression and regulation of mPRalpha and mPRbeta in zebrafish follicles. First, defolliculated fully grown oocytes were subjected to immunofluorescent staining using anti-mPRalpha and mPRbeta antibodies, and their expression on the oocyte membrane was confirmed. Second, total protein was collected from zebrafish follicles and Western blotting revealed that the level of mPRalpha and mPRbeta increased with follicle development. We have previously shown that several members of the transforming growth factor-beta (TGF-beta) superfamily, including TGF-beta1, activin-A, and bone morphogenetic protein (BMP)-15, regulate oocyte maturation in zebrafish. Therefore, the third major focus of this study was to test if these growth factors, as well as gonadotropins, regulate the expression of mPRs. Overexpression of BMP-15 significantly reduced, while knockdown of BMP-15 increased, mPRbeta levels. However, mPRalpha expression level remained unchanged with BMP-15 overexpression or knockdown. Treatment of follicles with human chorionic gonadotropin (hCG) resulted in an increased in mPRbeta, but not mPRalpha, expression levels. Activin-A induced the expression of mPRalpha and mPRbeta in a dose- and time-dependent manner. On the other hand, TGF-beta1 treatment suppressed the expression of mPRbeta, but not mPRalpha. Taken together, these findings further support the role of mPRs in oocyte maturation and suggest that gonadotropins, BMP-15, activin-A, and TGF-beta1 exert their regulatory effects on oocyte maturation in part by regulating mPR expression.