ABSTRACT
Some fibroblast growth factors (FGFs) affect ovarian follicle cell growth and/or differentiation. Whereas many FGFs activate several FGF receptors, FGF7 and FGF10 primarily activate only one, FGFR2B. As FGF7 is produced by bovine theca cells and acts on granulosa cells, we tested the hypothesis that FGF10 may also play a role in folliculogenesis in cattle. Reverse transcription-polymerase chain reaction demonstrated the presence of FGF10 mRNA in the oocytes and theca cells of the antral follicles, as well as in the preantral follicles. FGF10 protein was detected by immunohistochemistry in the oocytes of the preantral and antral follicles, and in the granulosa and theca cells of the antral follicles. FGF10 expression in theca cells changed during follicle development; mRNA abundance decreased with increasing follicular estradiol concentration in healthy follicles, and was lowest in highly atretic follicles. Culturing of granulosa cells in serum-free medium revealed FSH regulation of FGF10 receptor expression. The addition of FGF10 to cultured granulosa cells decreased the level of estradiol but did not alter cell proliferation. These data support a role for FGF10 in signaling to granulosa cells from theca cells and/or the oocyte.
Subject(s)
Fibroblast Growth Factor 10/metabolism , Ovarian Follicle/metabolism , Receptor, Fibroblast Growth Factor, Type 2/metabolism , Animals , Cattle , Female , Fibroblast Growth Factor 10/analysis , Fibroblast Growth Factor 10/genetics , Fibroblast Growth Factor 7/genetics , Fibroblast Growth Factor 7/metabolism , Follicle Stimulating Hormone/pharmacology , Granulosa Cells/drug effects , Granulosa Cells/metabolism , Insulin-Like Growth Factor I/pharmacology , Ovarian Follicle/chemistry , RNA, Messenger/analysis , RNA, Messenger/metabolism , Receptor, Fibroblast Growth Factor, Type 2/genetics , Theca Cells/drug effects , Theca Cells/metabolism , Tissue DistributionABSTRACT
In cattle, most evidence suggests that granulosa cells express LH receptors (LHR) after (or as) the follicle becomes dominant, however there is some suggestion that granulosa cells from smaller pre-dominant follicles may express several LHR mRNA splice variants. The objective of this study was to measure LHR expression in bovine follicles of defined size and steroidogenic ability, and in granulosa cells from small follicles (<6 mm diameter) undergoing differentiation in vitro. Semiquantitative RT-PCR demonstrated that LHR mRNA was undetectable in granulosa cells of follicles <7 mm diameter (nondominant follicles), and increased with follicle diameter in follicles >7 mm diameter. Splice variants with deletions of exon 10 and part of exon 11 were detected as previously described, and we detected a novel splice variant with a deletion of exon 3. Cultured granulosa cells contained LHR mRNA, but with significantly greater amounts of variants with deletions of exon 10 and/or exon 11 compared with cells from dominant follicles. FSH increased the abundance of some but not all LHR mRNA splice variants in cultured granulosa cells. The addition of LH to cultured cells did not increase progesterone secretion, despite the presence of LHR mRNA. Collectively, these data suggest that granulosa cells do not acquire functional LHR until follicle dominance occurs.
Subject(s)
Alternative Splicing , Cattle/genetics , Follicle Stimulating Hormone/pharmacology , Granulosa Cells/metabolism , Ovarian Follicle/cytology , Ovarian Follicle/drug effects , Receptors, LH/genetics , Animals , Cell Size , Cells, Cultured , Dose-Response Relationship, Drug , Female , Granulosa Cells/drug effects , Ovarian Follicle/metabolism , Protein Isoforms/genetics , Protein Isoforms/metabolism , RNA, Messenger , Receptors, LH/metabolismABSTRACT
Paracrine cell signaling is believed to be important for ovarian follicle development, and a role for some members of the fibroblast growth factor (FGF) family has been suggested. In the present study, we tested the hypothesis that FGF-8 and its cognate receptors (FGFR3c and FGFR4) are expressed in bovine antral follicles. RT-PCR was used to analyze bovine Fgf8, Fgfr3c and Fgfr4 mRNA levels in oocytes, and granulosa and theca cells. Fgf8 expression was detected in oocytes and in granulosa and theca cells; this expression pattern differs from that reported in rodents. Granulosa and theca cells, but not oocytes, expressed Fgfr3c, and expression in granulosa cells increased significantly with follicle estradiol content, a major indicator of follicle health. Fgfr4 expression was restricted to theca cells in the follicle, and decreased significantly with increasing follicle size. To investigate the potential regulation of Fgfr3c expression in the bovine granulosa, cells were cultured in serum-free medium with FSH or IGF-I; gene expression was upregulated by FSH but not by IGF-I. The FSH-responsive and developmentally regulated patterns of Fgfr3c mRNA expression suggest that this receptor is a potential mediator of paracrine signaling to granulosa cells during antral follicle growth in cattle.
