Expression of fibroblast growth factor-8 and regulation of cognate receptors, fibroblast growth factor receptor-3c and -4, in bovine antral follicles

Abstract

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. IntroductionAntral ovarian follicle growth in monovular species is regulated by a number of factors, the most well known of which are the gonadotropins. Follicles are considered to be follicle-stimulating hormone (FSH)-dependent until dominance occurs, after which they become luteinizing hormone-dependent (reviewed by Fortune et al. 2001, Ginther et al. 2001. It has also become clear that growth factors are key stimulatory/regulatory molecules. Several lines of evidence point to a critical role for members of the transforming growth factor-b (TGF-b) superfamily, especially growth/differentiation factor 9 and bone morphogenetic protein 15 (reviewed by Gilchrist et al. 2004, Juengel et al. 2004, Shimasaki et al. 2004.The fibroblast growth factor (FGF) family is emerging as a group of factors that are potentially important for follicle growth. For example, FGF-7 is expressed in theca cells, its receptor is expressed in granulosa cells (Parrott & Skinner 1998, Berisha et al. 2004, and FGF-7 stimulated bovine granulosa cell proliferation and inhibited steroidogenesis (Parrott & Skinner 1998). Another potentially interesting member of this family is FGF-8. Widely expressed in fetal tissues, this factor is predominantly expressed in the gonads of adult rodents and ruminants (MacArthur et al. 1995a, Buratini et al. 2005. Within the ovary, Fgf8 gene expression occurs only in the oocyte in adult mice (Valve et al. 1997), which suggests a potential role in signaling of follicular cells by the oocyte.There are five known FGF receptor (FGFR) genes (Kim et al. 2001, Sleeman et al. 2001, of which FGF-8 preferentially activates FGFR4 and the 'c' splice form of FGFR3 (Ornitz et al. 1996). mRNAs encoding Fgfr4 or Fgfr3c were not consiste...

Glial-derived neurotrophic factor promotes ovarian primordial follicle development and cell–cell interactions during folliculogenesis

“…FGF8 is expressed in antral follicles in rat and cow. It may also play a role in the development of ovarian and breast tumors (Valve et al 1997, Daphna-Iken et al 1998, Zammit et al 2002, Buratini et al 2005. STC1 was originally shown to have high expression levels in the ovary (Varghese et al 1998).…”

The roles of glial cell line-derived neurotrophic factor, brain-derived neurotrophic factor and nerve growth factor during the final stage of folliculogenesis: a focus on oocyte maturation

“…This expression profile suggests that FGF8 plays a paracrine role, signalling between the oocyte and follicular somatic cells during the final stages of follicular maturation (Valve et al 1997). However, it has been reported that FGF8 is expressed in oocytes, granulosa cells and theca cells within bovine antral follicles (Buratini et al 2005), suggesting an autocrine/paracrine mode of signalling. CTGF is a matrix-associated heparin binding protein that is regulated by gonadotrophins and is expressed in antral follicles and corpus lutea in the rat (Harlow et al 2007).…”

Expression of protease nexin-1 and plasminogen activators during follicular growth and the periovulatory period in cattle

“…These represent mature dominant growing, static and regressing follicles respectively (Price et al 1995). Cross-contamination of theca and granulosa cells was tested by detection of mRNA encoding cytochromes P450 aromatase (Cyp19) and 17a-hydroxylase (Cyp17) in each sample by PCR (Buratini et al 2005). Only granulosa cell samples negative for Cyp17, and only thecal samples negative for Cyp19 amplicons were included in the analysis (number of follicles used given above).…”

“…Briefly, for both theca and granulosa cells, total RNA (1 mg) was incubated with DNAse I (Invitrogen) and reverse transcribed with SuperScript II (Invitrogen) and oligo-d(T) primer (Buratini et al 2005). An aliquot (0.4 ml) of the cDNA template was amplified by PCR using 0.2 ml (2.5 U) Taq Polymerase (Amersham Pharmacia Biotech, Inc., Oakville, ON, Canada) in a 20 ml PCR buffer (Amersham Pharmacia Biotech) containing 0.1 mM dNTP mix, and 0.2 mM specific primers (Cao et al 2004).…”