Richani D, Wang X, Zeng H T, Smitz J, Thompson J G, Gilchrist R B
Faculty of Health Sciences, Discipline of Obstetrics and Gynaecology, The Robinson Institute, Research Centre for Reproductive Health, School of Paediatrics and Reproductive Health, The University of Adelaide, Adelaide, Australia.
Mol Reprod Dev. 2014 May;81(5):422-35. doi: 10.1002/mrd.22307. Epub 2014 Feb 22.
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.
近期研究独立表明，体外成熟（IVM）前对环磷酸腺苷（cAMP）进行调控以及在IVM期间补充表皮生长因子（EGF）样肽可改善后续卵母细胞的发育结果。本研究调查了结合这两种理念的IVM系统的效果。从青春期前小鼠收集卵丘-卵母细胞复合体（COC），分别在马绒毛膜促性腺激素（eCG）注射后46小时进行IVM，或在eCG + 人绒毛膜促性腺激素（hCG）刺激后进行体内成熟（IVV）。IVM的COC在IVM前用cAMP调节剂福斯可林和异丁基甲基黄嘌呤处理1、2或4小时（IVM前期）。然后，COC用EGF样肽双调蛋白或表皮调节素，或用常用的IVM刺激剂促卵泡激素（FSH）或EGF进行IVM。与标准IVM相比，无论IVM处理如何，IVM前期均增加了后续囊胚内细胞团的大小（P < 0.05）。与FSH或EGF不同，与标准IVM相比，双调蛋白或表皮调节素显著提高了囊胚质量（滋养外胚层和总细胞数）和/或产量（P < 0.01），并且是唯一产生与IVV来源囊胚相当囊胚的处理方法。在2小时的IVM前期后，福斯可林急性上调了EGF样肽mRNA表达（P < 0.001），尽管EGF受体和ERK1/2活性与对照无显著差异。仅通过补充EGF样肽和EGF才能在IVM期间维持IVV样水平的EGF样肽mRNA表达，因为FSH诱导的表达水平在体外显著低于IVV期间。然而，各处理组间EGF受体和ERK1/2磷酸化水平无显著差异。总之，IVM前期结合存在EGF样肽的IVM赋予了高卵母细胞发育能力，相对于FSH和EGF，胚胎产量和/或质量增加证明了这一点。
