【作者】 雷安民； 敬晓棋； 严兴荣； 胥涛； 窦忠英；

【Author】 LEI An-min, JING Xiao-qi, YAN Xing-rong, XU Tao, DOU Zhong-ying(Shaanxi Sub-Center of National Stem Cell Engineering &Technology, Northwest Sci-Tech University of Agriculture &Forestry, Yangling, Shaanxi 712100, China)

【机构】 西北农林科技大学陕西省干细胞工程技术研究中心；

【摘要】 相同实验条件下,用三种不同培养液①（TCM199液内添加0.22mg/L丙酮酸钠,10nM Hepes,0.10iu HMG,1mg/mL17β-E₂,10%胎牛血清）②（SOF液内添加0.10 IU HMG,1mg/mL 17β-E₂,6mg/mLBSA）（CR₁液内添加0.10IU HMG,1mg/mL17β-E₂,6mg/mL BSA）对牛卵泡卵母细胞进行体外成熟的研究,①组进行了18批次实验,第一极体可见率为62.5%（303/485）;②组进行了9批次实验,第一极体可见率为52.5%（156/297）:③组进行了9批次实验,第一极体可见率为63.8%（192/301）。结果显示①②③三组之间第一极体可见率差异不显著。在此实验研究基础上,观察了培养液④（培养液①内添加转铁蛋白、EGF）和培养液⑤（培养液①内添加转铁蛋白、EGF、尿嘧啶）在牛卵泡卵母细胞体外成熟的效果。④组进行了20批次实验,第一极体可见率为63.6%（930/1462）;⑤组进行了6批次实验,第一极体可见率为67.9%（292/430）。结果显示①④⑤三组之间第一极体可见率差异不显著。显微操作结果显示,①组卵母细胞操作后的完整性较②③组好些,⑤组卵母细胞操作后的完整性较①④组好,⑤组卵母细胞成熟后,卵质膜柔韧性很好,操作过程几乎不发生质膜破损现象。添加丙酮酸钠与尿嘧啶的一个主要作用是保护卵母细胞中的线粒体功能,这一研究显示,对线粒体功能的保护尽管不能明显的提高第一极体可见率,但却可以提高卵质膜柔韧性,从而有利于卵母细胞的体外操作。更多还原

【Abstract】 In this study 3 groups of media, (1) TCM-199 supplemented with 0.22 mg/L sodium pyruvate, 10 nM Hepes, 0.10 IU HMG, 1 mg/mL17β-E2, and 10% fetal bovine serum, (2) SOF supplemented with 0.10 IU HMG, 1 mg/mL17β-E2 and 6 mg/mL BSA and (3) CR1 supplemented with 0.10 IU HMG, 1 mg/mL17B-E2 and 6 mg/mL BSA, were used for in vitro maturation of bovine oocytes under the same culture conditions. The ratios of visible PBl and repetition times in group (1), (2) and (3) were 62.5%(303/485), 52.5% (156/297) and 63.8% (192/301) and 18,9, and 9 respectively. The results showed that there were no significant differences in the proportion of visible PB1 among groups of (1), (2) and (3) ( P> 0.05). Based on the above experiment, we further investigated the effects on in vitro maturation of bovine oocytes by group(4) (group (1) supplemented with transferrin and EGF) and group (5) ((1) supplemented with transferrin , EGF and Uracil) . The rates of visible PBl were 63.6% (930/1462) in group (4) and 67.9% (292/420) in group (5), with 20 and 9 repetitions each. The results showed that there was no significant differences in the rates of visible PBl among group (1), (4) and (5). The micromanipulation results indicated that the integrality of oocytes in group (1) was better than that of (2) and (3), and that of group (5) was better than that of (1), (4) respectively. The flexibility of cytoplast in group (5) was ideal after in vitro maturation because the cell membrane seldom brokedown during the micromanipulation. One of major functions of adding sodium pyruvate and Urail was the protection of mitochondrial function of oocytes. The results of this study showed that the protection of mitochondrial function couldn’t improve the proportion of visible PBl, but can increase the cytoplast flexibility of the oocytes, thus being beneficial to the in vitro manipulation of bovine oocytes.更多还原