ABSTRACT
Bone morphogenetic protein 4 (BMP4) is essential for the development of primordial follicles, although its underlying mechanism remains largely unknown. By using cultured ovaries, the effects of BMP4 and the potential signal transduction pathways were investigated. Ovaries from 3-day-old female mouse pups were maintained in organ culture in the absence (control) or presence of BMP4 (100 ng/ml). At different culture time, the effects of BMP4 on primordial follicle growth and survival were assayed by follicle count and TUNEL labeling. The expression of phospho-SMAD1/5/8, Sohlh2, and c-kit were measured by immunohistochemistry, RT-PCR, and Western blotting. Immunohistochemistry was also performed to determine the expression pattern of BMP4, pSMAD1/5/8, Sohlh2, and c-kit in vivo during ovarian development. The results showed treatments of ovaries with BMP4 resulted in a significant (P < 0.05) increase on the primordial-to-primary follicle transition. The oocytes of primordial follicles treated with BMP4 were also less likely to undergo apoptosis. BMP4 enhanced the phosphorylation of SMAD1/5/8 and up-regulated the expression of Sohlh2 and c-kit in primordial follicles. During ovarian development in vivo, Sohlh2, and c-kit exhibited similar expression patterns to BMP4 and pSMAD1/5/8 in primordial follicles. The present studies suggest that BMP4/SMAD signaling pathway initiate primordial follicle growth and prevented oocyte apoptosis via up-regulation of Sohlh2 and c-kit.
Subject(s)
Basic Helix-Loop-Helix Transcription Factors/metabolism , Bone Morphogenetic Protein 4/metabolism , Ovarian Follicle/growth & development , Proto-Oncogene Proteins c-kit/metabolism , Smad Proteins/metabolism , Animals , Apoptosis/drug effects , Bone Morphogenetic Protein 4/pharmacology , Female , Humans , Mice , Oocytes/chemistry , Oocytes/drug effects , Oocytes/growth & development , Oocytes/metabolism , Ovarian Follicle/chemistry , Ovarian Follicle/drug effects , Ovarian Follicle/metabolism , Recombinant Proteins/pharmacology , Signal Transduction , Up-Regulation/drug effectsABSTRACT
The role of WNT/ß-catenin-signaling pathway is critical in mouse Sertoli cell maturation and tumorigenesis. This study aims to examine the effects of WNT/ß-catenin signaling on the cultured adult human Sertoli cells and the underlying molecular mechanisms. Glycogen synthase kinase 3ß (GSK-3ß) inhibitors, SB216763 and lithium chloride (LiCl), were used to activate WNT/ß-catenin-signaling pathway. 5-Bromo-2'-deoxyuridine (BrdU) incorporation assay and flow cytometry were used to analyze the proliferation and cell cycle of cultured human Sertoli cells, respectively. C-myc expression was accessed by immunofluorescence, real-time polymerase chain reaction and Western blot. The effects of c-myc on Sertoli cell proliferation were investigated by RNA interference technology and BrdU incorporation assay. The results showed activation of WNT/ß-catenin signaling stimulated human Sertoli cell proliferation. Obvious increases in c-myc messenger RNA and protein expression were observed after SB216763 and LiCl treatments. Knockdown of c-myc expression attenuated the ability of WNT/ß-catenin signaling to stimulate the proliferation of human Sertoli cells. WNT/ß-catenin signaling enhances human Sertoli cell proliferation via upregulation of c-myc expression.
