猪不同直径卵泡内卵母细胞胞质成熟及体外发育潜力差异原因的初探

Preliminary Study on the Mechnism Account for the Difference of Cytoplasmic Maturation and in vitro Developmental Potential of Porcine Oocytes from Different Diameter Follicles

试验根据直径将猪卵泡分为2组：G1组（4~7mm）和G2组（2~4mm）,对2组卵泡内获取的卵母细胞体外成熟率和发育潜能进行了比较,利用相对定量PCR检测了卵丘细胞中卵丘扩展相关基因Has2、Ptgs2、Ptx31及Pgr的表达水平,应用绝对定量PCR检测了成熟培养前后卵母细胞线粒体拷贝数,并利用5,5′-二巯基-2-硝基苯酸（DTNB）酶循环法检测了体外成熟培养过程中卵母细胞谷胱甘肽（GSH）的含量。结果显示,G1组和G2组卵母细胞体外成熟率分别为95.06%和68.19%,G1和G2组排出第一极体的成熟卵母细胞孤雌激活后的囊胚率分别为51.47%和29.44%,2组卵母细胞在体外成熟率和孤雌发育率上均差异显著（P〈0.05）。G1组卵丘细胞在体外成熟培养过程中的扩展程度明显高于G2组,G1组卵母细胞对应的卵丘细胞扩展相关基因Has2、Ptgs2、Ptx31、Pgr的表达水平高于G2组（Has2基因在卵母细胞成熟培养0、24h除外）;G1组卵母细胞线粒体数、谷胱甘肽含量均高于G2组。以上结果表明,大卵泡来源的卵母细胞体外成熟能力和发育潜力优于小卵泡来源的卵母细胞,这可能与卵母细胞成熟过程中卵丘扩展程度、卵丘扩展相关基因表达激活情况、卵胞质内谷胱甘肽含量和线粒体拷贝数有关。

We compared the in vitro maturation rate and parthenogenetic blastocyst rates of oocytes derived from follicles4 to 7mm（G1group）and 2to 4mm（G2group）in diameter.The cumulus expansion level,expansion relatived genes Has2,Ptgs2,Ptx31 and Pgr expression,GSH concentration and mitochondria copy number were also investigated.We found that the maturation rates of G1 and G2were 95.06%and 68.19%,respectively,and the parthenogenetic blastocyste rates of these two groups were 51.47%and 29.44%,respectively,there were significant differences in maturation rates and partehnogenetic blastocysts rates between these two groups（P〈0.05）.The expansion level of cumulus cells in G1 was clearly higher than that in G2.The expression of cumulus cell expansion related gene Has2,Ptgs2,Ptx31 and Pgrin G1 group were higher than that of G2group（except of Has2 gene in 0,24h）.Total GSH concentration and mitchondria numbers in G1 oocytes were higher than that in G2 oocytes.These results indicated that porcine oocytes from large follicles were better than these from small follicles in in vitro maturation and developmental potential,the cumulus cell expansion level,the expression of cumulus cell expansion related genes,concentration and mitochondria copy number might account for the difference.