【作者】 岳顺利；

【导师】 严云勤；

【作者基本信息】 东北农业大学 ， 基础兽医学， 2005， 硕士

【摘要】 哺乳动物卵母细胞生长、发育和成熟的调节机制是当前生殖生物学及发育生物学的研究热点之一。在卵母细胞成熟过程中促性腺激素(尤其是FSH)起重要的作用。关于促性腺激素诱导卵母细胞恢复减数分裂的机制存在两个假说:一是促性腺激素诱导卵丘扩展使颗粒细胞与卵母细胞的缝隙连接中断,使抑制减数分裂的物质向卵母细胞的输入中断,导致减数分裂恢复;二是促性腺激素诱导卵丘细胞产生一些因子,克服抑制物质的作用而促使减数分裂的恢复。但是,迄今为止这些正因子的性质以及作用途径仍然有待研究。近年来,哺乳动物卵母细胞体外成熟技术已取得长足的进步。即便如此,某些动物卵母细胞体外成熟率仍然较低,仍存在许多问题没有解决。在人类及珍稀动物辅助生殖中卵母细胞的获得是十分困难的,因而最大限度的利用卵母细胞十分必要。有报道证明卵丘细胞产生的某些因子对卵母细胞的成熟有影响。因此我们以小鼠为实验动物模型,探索这些因子的作用,来提高裸卵和部分裸露的GV 期卵母细胞成熟和发育能力。哺乳动物卵母细胞正常发育是一系列复杂发育调节过程,通过缝隙连接和旁分泌协同作用卵母细胞最终完成发生。本研究应用缝隙连接解偶联剂结合共培养技术通过间接的手段探讨正因子进入途径及其对卵母细胞成熟的影响,从而为建立卵丘细胞共培养系统,提高卵母细胞成熟率和发育能力提供参考数据。本实验得到了如下的一些结果: 1、分离的卵丘细胞能够释放某些诱导DO 恢复减数分裂的因子,这些因子可以通过旁分泌发挥作用。2、分离的卵丘细胞可释放某些旁分泌因子,但是这些因子不能诱导完整的COCs 中的卵母细胞恢复减数分裂。3、FSH 刺激的卵丘细胞的产生某些因子,这些因子需要通过缝隙连接进入卵母细胞,并且这种机制在COCs 卵母细胞的减数分裂恢复中起主要作用。4、在FSH 作用下猪卵丘细胞可以产生提高小鼠PNO 和NO 的GVBD 比例的某些因子,但是它们不能提高PNO 和NO 的受精率和囊胚率。更多还原

【Abstract】 Regulation mechanism of growth, development and maturation of oocyte in mammal was one of foci of reproductive biology and developmental biology. Gonadotropins, especially FSH, played a important role in maturation of oocyte. There were two hypotheses on mechanism of meiotic resumption presently, one was cumulus oophorus expanded which induced by gonadotrophin breaked gap junctional communation between oocyte and granulosa cells, blocked the conduction of meiosis-inhibitiory signals from the outer cumulus cells to the oocyte, then meiotic resumption. The other was meiotic resumption by eliciting a maturation-inducing signal from the granulosa cells which overrided the inhibitory milieu of the follicle. However, property and regulation mechanism of the positive factor were still unknown. People made a great progress in IVM in mammal recently, even though maturation of oocyte in some species were still very low. Many problems about IVM were still remained. In assisted reproduction technology program of human and endangered species, oocyte obtaining was very difficult. It is nessary to use oocyte maximumly. Several studies reported culumus cells produced some positive factors which could influence maturation. So we use mouse as a model, to investigate the function of these factors, to improve maturation and development of PNO and NO at GV stage. Development of mammalian oocytes was a series of complex and developmentally regulated process. Gap junctional communication and paracrine signaling Interplay in those processes. Which kind of communication dominated in meiotic resumption was unknow. This study was to investigate the path way and the effect of the positive factors by using the gap junctional inhibitor and co-culture technique to provide basic data for maturation and development of oocyte in cumulus cell co-culture system. The results obtained were as follows: 1. Dissociated cumulus cells produced some meiosis-inducing factors which could act as a paracrine signal. 2. Dissociated cumulus cells produced some factors, but the factors could not induce meiotic resumption of oocyte in COCs. 3. The action of positive meiosis-inducing factors secreted by FSH-stimulated cumulus cells on oocyte depended on gap junctional communication which was dominant manner in meiosis resumption. 4. FSH-stimulated dissociated cumulus cells from swine produced some factors which could increase the percentage of GVBD of PNO and NO, but not improve percentage of fertilization and blastulae in mouse.更多还原