Epidermal growth factor receptor activation by protein kinase C is necessary for FSH-induced meiotic resumption in porcine cumulus–oocyte complexes

Abstract

It is proved that epidermal growth factor (EGF)-like factors mediate gonadotropin-induced rodent oocyte maturation via EGF receptor (EGFR). However, the detail kinetics and signal pathway between FSH and EGF/EGFR is not clear in large animals. In the present study, we investigated the roles of EGFR and protein kinase C (PKC) in FSH-induced porcine oocyte meiotic resumption. Porcine cumulus-oocyte complexes were cultured in NCSU37 medium containing 10% porcine follicular fluid and germinal vesicle breakdown (meiotic resumption) was detected after different treatments. The results showed that EGF-like factor amphiregulin (AR) and EGFR mRNA were expressed in porcine cumulus cells, but not oocytes. FSH significantly induced AR mRNA expression with maximum at 4 h and activated EGFR phosphorylation at 8 h. AR (1-100 ng/ml) dose-dependently induced meiosis resumption of porcine oocyte. The specific EGFR inhibitor, AG1478, but not AG43 (the inactive analog of AG1478), completely blocked FSH, EGF, and AR-induced oocyte meiotic resumption; the inhibitory effect of AG1478 on FSH action gradually decreased when the inhibitor was added at 6 h or later and disappeared when it was added at 11 h; EGF reversed the inhibitory effect on FSH when AG1478 was added within 6 h. FSH triggered porcine oocyte meiotic resumption (at 20 h) later than that of EGF and AR (at 18 h). All these results supported that endogenously produced EGFR activator(s), possibly AR (maximum at 4 h) and EGFR activation (began at 6 h and finished within 11 h), in cumulus cells is necessary for FSHinduced porcine oocyte meiotic resumption (began at 18 h). Furthermore, PKC activator PMA mimicked but PKC inhibitor chelerythrine chloride inhibited FSH action, and AG1478 also suppressed PMA-induced porcine oocyte meiotic resumption. These data together suggested that EGFR activation, by PKC signal pathway, participates in FSH-induced porcine oocyte meiotic resumption.

Transcriptomic analysis of cyclic AMP response in bovine cumulus cells

“…We suppose that this could be due to either 1) an overactivation of the PKA pathway creating negative feedback at 6 h or 2) the FID treatment period of 6 h being too early to read any effect of ADAMTS1 stimulation, which had not yet trickled down to result in AREG activation. AREG is considered to be essential for transcription of EGF ligands (11). Interestingly, our findings corroborate an earlier observation that a forskolin exposure of Ͼ2 h resulted in downregulation of EGF ligands in cumulus cells (41).…”

Epidermal growth factor differentially regulates activin subunits in the zebrafish ovarian follicle cells via diverse signaling pathways

“…These four pathways were chosen also because they had been shown to regulate the functions of granulosa cells, cumulus cells and theca cells as well as final oocyte maturation. PKC pathway inhibited oocyte maturation in denuded oocytes but stimulated it in follicle-enclosed oocytes (Chen et al, 2008;Sun et al, 1999). In the frog, the PI3K pathway in the oocyte was activated during oocyte maturation (Ju et al, 2002).…”

Novel signaling mechanisms in the ovary during oocyte maturation and ovulation

“…It is now known that the EGF-like growth factors amphiregulin (AREG), epiregulin (EREG) and betacellulin (BTC), rather than EGF, are rapidly and transiently induced in the somatic cells of the preovulatory follicle by LH (Park et al 2004; Sekiguchi et al 2004; Ashkenazi et al 2005). LH/hCG induction of these EGF-like growth factors has been reported in multiple species (Fru et al 2007; Wang et al 2007; Chen et al 2008; Lindbloom et al 2008; Inoue et al 2009; Zamah et al 2010). The EGF-like growth factors are released from the cell surface as mature, soluble peptides by proteolytic cleavage of the ectodomain, and when added to cumulus-oocyte complexes or follicles in culture, activate the EGFR on the surfaces of the somatic cells to promote oocyte meiotic resumption and cumulus expansion.…”