人未成熟卵母细胞体外成熟微环境对其发育潜能的影响及机制研究

In vitro maturation technique of human immature oocytes has great application value in reproductive medicine. However, the development potential of in vitro matured oocytes is still lower than those oocytes matured in vivo. The main reason for that was the inconsistent maturity of cytoplasm and nuclear. Based on the late development in reproductive biology and animal research, FSK/IBMX and/or EGF-like peptide were added into conventional IVM media for maturation of human immature oocytes. The developmental potential of the matured oocytes after fertilization in different IVM media was evaluated, and the cortical granules distribution, mitochondria membrane potential and spindle configuration were evaluated and compared by immunofluorescence technology and laser confocal microscopy. Mitochondrial DNA copies and maternal genes （Gdf9、Zar1、Mater、Dnmt1）mRNA assay through RT-PCR were also compared between the matured oocytes in different IVM media and in vivo matured oocytes. These results will help us find a useful method to evaluate the oocyte cytoplasm maturity and serve as an experimental evidences for a more efficient IVM protocol for immature oocytes.

未成熟卵母细胞体外成熟技术（IVM）在生殖医学领域具有重要的应用价值，而IVM后获得的成熟卵母细胞的发育潜能远远不及体内成熟的卵母细胞，核质成熟不同步是影响卵母细胞发育潜能的主要原因。基于最新的基础研究及动物实验的研究成果，本课题拟通过在IVM培养液中添加FSK/IBMX来延缓核成熟的进程和/或添加EGF样多肽改善卵母细胞成熟微环境的方法进行IVM培养，并以体内成熟卵母细胞的各项检测指标作为正常对照。根据卵母细胞受精后的发育潜能、胞质成熟相关超微结构改变的比较以及分子生物学的相关证据来判断FSK/IBMX和EGF样多肽在人卵IVM培养中的应用价值，本课题的研究结果有助于获得有效评价人卵母细胞胞浆成熟度的方法，并将为设计更高效的人未成熟卵母细胞的IVM方案提供实验依据。

未成熟卵母细胞体外成熟技术(IVM)在生殖医学领域具有重要的应用价值,而IVM后获得的成熟卵母细胞的发育潜能远远不及体内成熟的卵母细胞,核质成熟不同步是影响卵母细胞发育潜能的主要原因。基于最新的基础研究及动物实验的研究成果,本课题通过优化IVM培养液，通过添加EGF样多肽改善卵母细胞成熟微环境的方法进行IVM 培养,根据卵母细胞受精后的发育潜能、胞质成熟相关超微结构改变的比较来评价改良人卵IVM培养液的应用价值,发现优化的IVM培养液有助于未成熟卵母细胞的成熟，改善了卵母细胞成熟度，提高了卵母细胞体外发育潜能，获得了较传统受精液培养及传统IVM培养更好的培养效果，提高了优质囊胚形成率，本课题的研究结果帮助常规未成熟卵较多的体外助孕患者获得了更多的可利用胚胎，提高了治疗效率。通过发育潜能、超微结构及分子生物学的相关指标的分析对比，获得了有效评价卵母细胞体外成熟的方法，为设计更高效的人未成熟卵母细胞的IVM方案提供了实验及临床依据。