不同因素对奶牛分离精子ICSI效率的影响

Influence of Different Factors on the Efficiency of Bovine Sorted Sperm Injection

本研究旨在探讨不同的联合激活处理、精子预处理方法和单精子注射方式对奶牛分离精子ICSI效率的影响。借助显微操作仪将经不同预处理的奶牛分离X精子直接注入体外成熟22～24h的牛卵母细胞胞质内,注射过程采用回吸胞质和不回吸胞质2种处理,最后分别用3种激活方案对注射卵进行激活。结果表明:用CR1aa+Ionomycin+6-DMAP对ICSI注射卵进行激活后,卵裂率和囊胚率都高于A23187+CHX和7%ET+CHX两种联合激活组(63.74%vs61.09%、53.75%;28.54%vs17.68%、22.00%,P<0.05);采用percoll密度梯度离心法处理精子,卵裂率(77.10%vs62.19%,P<0.05)与囊胚率(26.95%vs22.82%,P>0.05)均高于上游法处理组;ICSI时回吸胞质获得的注射卵,其卵裂率和囊胚率均显著高于不回吸胞质处理组(63.10%,25.35%vs41.56%,19.40%,P<0.05)。结果表明,用percoll密度梯度离心法处理精子、注射时回吸胞质、用CR1aa+Iono-mycin+6-DMAP激活ICSI注射卵,可显著提高奶牛分离精子ICSI的效率。

This study was conducted to investigate the effects of different combined activation protocols,sperm preparing methods and injection methods on the efficiency of bovine ICSI.By the assistance of a micromanipulation system,bovine sorted X sperm,prepared by different pretreatments,was directly injected into an oocyte which was matured in vitro for 22-24 h,with or without performing cytoplasmic aspiration,then activated by three different combined activation protocols.The results showed that the cleavage and blastocyst rate of ICSI embryos activated by CR1aa ＋ Ionomycin ＋ 6-DMAP were higher than those activated by A23187 ＋ CHX and 7%ET ＋ CHX groups（63.74% vs 61.09%,53.75%;28.54% vs 17.68%,22.00%,P0.05）.After treating sperms with percoll density gradient centrifugation,the cleavage（77.10% vs 62.19%,P0.05）and blastocyst rate（26.95% vs 22.82%,P0.05）were both higher than that of the swim-up treatment group.The cleavage and blastocyst rate with performing cytoplasmic aspiration were both significantly higher than that of control during sorted sperm injection（63.10%,25.35% vs 41.56%,19.40%,P0.05）.The results indicated that treating sperms with percoll density gradient centrifugation,performing cytoplasmic aspiration and activating ICSI embryos by CR1aa ＋ Ionomycin ＋ 6-DMAP,can significantly improved the efficiency of bovine sorted sperm injection.