Hedgehog signaling regulates the basement membrane remodeling during folliculogenesis in the neonatal mouse ovary.

In the mouse ovary, interactions between oocytes and somatic cells are essential for folliculogenesis and subsequent follicle development. The polyovular follicle (PF), which contains more than two oocytes in a follicle, can be induced in the neonatal mouse ovary when interactions between oocytes and somatic cells are disrupted by agents such as the potent synthetic estrogen diethylstilbestrol (DES) acting through estrogen receptor (ER) β. Hedgehog signaling is known to regulate granulosa cell proliferation, thecal cell differentiation, and follicle growth. To investigate the role of hedgehog signaling in the early folliculogenesis and in PF induction by DES, neonatal mouse ovaries were cultured with or without 10 μM cyclopamine (CPA), an inhibitor of hedgehog signaling, and grafted under the kidney capsule of adult ovariectomized host mice. The number and the incidence of PFs were significantly increased in organ-cultured ovaries post-grafting. Expression of procollagen type IV, alpha 1 (Col4a1) in organ-cultured ovaries was significantly reduced by CPA, but not by DES. The expression of two hedgehog ligands, Desert hedgehog (Dhh) and Indian hedgehog (Ihh), and a target gene, Hedgehog interacting protein (Hhip), was significantly increased by DES both in WT and ERβ KO mice. Therefore, we infer that DES can affect expression of those genes through ERα but not via suppression of hedgehog signaling. Thus, PFs are induced by DES or CPA, but the induction mechanism is different. Our results revealed an important role of hedgehog signaling in basement membrane remodeling during folliculogenesis even before thecal cell differentiation.