Pre-maturation with cAMP modulators in conjunction with EGF-like peptides during in vitro maturation enhances mouse oocyte developmental competence

Abstract

Recent studies have independently shown that cyclic adenosine 3'5'-monophosphate (cAMP) modulation prior to in vitro maturation (IVM) and epidermal growth factor (EGF)-like peptide supplementation during IVM improve subsequent oocyte developmental outcomes. This study investigated the effects of an IVM system that incorporates these two concepts. Cumulus-oocyte complexes (COCs) were collected from pre-pubertal mice either 46 hr post-equine chorionic gonadotropin (eCG) (IVM) or post-eCG + post-human chorionic gonadotropin (hCG) stimulation (in vivo maturation; IVV). IVM COCs were treated with the cAMP modulators forskolin and IBMX for 1, 2, or 4 hr (pre-IVM phase) prior to IVM. COCs then underwent IVM with the EGF-like peptides amphiregulin or epiregulin, or with the common IVM stimulants follicle-stimulating hormone (FSH) or EGF. A pre-IVM phase increased the size of the subsequent blastocysts' inner-cell-mass compared to standard IVM, regardless of IVM treatment (P < 0.05). Unlike FSH or EGF, amphiregulin or epiregulin significantly increased blastocyst quality (trophectoderm and total cell numbers) and/or yield (P < 0.01) compared to standard IVM, and were the only treatments that produced blastocysts comparable to IVV-derived blastocysts. Forskolin acutely up-regulated EGF-like peptide mRNA expression after a 2-hr pre-IVM phase (P < 0.001), although EGF receptor and ERK1/2 activities were not significantly different than control. IVV-like levels of EGF-like peptide mRNA expression during IVM were maintained only by supplementing with EGF-like peptides and EGF, since expression levels induced by FSH were significantly lower in vitro than during IVV. However, EGF receptor and ERK1/2 phosphorylation levels were not significantly different across treatment groups. In conclusion, a pre-IVM phase in conjunction with IVM in the presence of EGF-like peptides endows high oocyte developmental competence, as evidenced by increased embryo yield and/or quality relative to FSH and EGF.