小鼠孤雌胚2-细胞阻滞的研究

Studies on mouse parthenogenetic embryos 2-cell block

为探讨小鼠孤雌胚2-细胞阻滞的机制,本试验以昆明小鼠为研究对象,通过输卵管上皮细胞和垂体细胞作为饲养层培养小鼠孤雌胚胎,分析其作用机理。小鼠卵母细胞通过70mL/L乙醇激活5min,再用2mmol/L 6-DMAP、5μg/mL CB激活3h后,分别在不同饲养层的KSOM培养液中进行培养,观察并比较各培养条件下孤雌胚胎的发育情况。结果,培养至第24小时,各组无显著差异(P>0.05),都有较高卵裂率。培养至第48小时,用两种饲养层细胞培养的孤雌胚胎4-细胞～8-细胞发育效果好,与用单独一种饲养层细胞培养相比,差异显著(P<0.05),与对照组相比,差异极显著(P<0.01);发育至桑囊胚的比例为49.4%(42/85)(P<0.01)。结果表明,含有输卵管上皮细胞和垂体细胞的KSOM培养液可有效促进小鼠孤雌胚胎的发育并通过2-细胞阻滞(通过2-细胞阻滞率为74.1%),并可进一步提高其发育率(桑囊胚率为49.4%)。

In order to study the mechanism of mouse parthenogenetic embryos 2-cell block,we selected oviduct epithelial cells and pituitary cells for feeder layer to culture mouse parthenogenetic embryos. The mouse oocytes which were activated after dealing with 5 min in 70mL/L alcohol and 3 h in 2 mmol/L 6- DMAP and 5 g/mL CB, were cultured in KSOM medium containing different feeder layers. We observed and compared mouse parthenogenetic embryos developmental situation which were in various culture conditions. In result,the embryos had no significant difference in each group after 24 h culture,and all of them had high cleavage rate（P〉0. 05）. After being cultured to 48 h, the parthenogenetic embryos cultured in KSOM medium with two types of feeder cells had developmental effect of 4 to 8-cell,it was significantly different（P〈0.05） compared with single feeder layer cell and cotrol group（P〈0.01）. The ratio of embryos which can develop to Morula/blastocyst was 49.4%（42/85）（P〈0.01）. We inferred that KSOM medium containing oviduct epithelial cells and pituitary cells can effectively promote the development of mouse parthenogenetic embryos which can breakthrough 2-cell block（block rate to 74.1%）, and further improve embryos development（morula/blastocyst rate to 49.4 %）.