不同激活方法对猪卵胞质内单精子注射(ICSI)效果的影响

Effects of different methods on the activation and development of porcine oocytes after intracytoplasmic sperm injection(ICSI)

采用不同的注射方式结合人工激活方法处理猪卵母细胞,旨在探讨各处理方法对猪ICSI效果的影响。结果表明:假性注射+未激活处理组卵母细胞的激活率虽然高于无机械刺激+无激活处理对照组,但明显低于假性注射+人工激活处理组;精子注射卵母细胞经CaCl2(1.8pL,30mmol/L)、离子霉素(15μmol/L,40min)和电脉冲(场强0.4kV/cm、脉冲时程90μs、1次脉冲)激活处理后,其激活率无显著性差异(P>0.05),CaCl2处理组的受精率显著高于离子霉素处理组(P<0.05),并与电激活处理组无显著性差异(P>0.05);但是,在假性注射组和对照组中,CaCl2处理组的孤雌发育率极显著地低于离子霉素和电激活处理组(P<0.01);CaCl2处理组的卵裂率(P<0.01)和囊胚总细胞数(P<0.05)显著高于对照组。因此,单纯注射性机械刺激对猪卵母细胞的激活效果较差,有必要进行人工激活;ICSI卵母细胞经CaCl2溶液激活处理后,能够取得较好的ICSI效果,而不显著增加其孤雌发育率。

To explore effects of different methods on the activation and development of porcine oocytes after intracytoplasmic sperm injection（ICSI）,sperm injected,sham injection or non-injection oocytes were either activated by CaCl2（1.8 pL,30 mmol/L）,ionomycin（15 μmol/L,40 min） or electrical pulse（a single dc-pulse of 0.4 kV/cm for 90 μs）.Results were as follows： activation rates of sham injected and non-activated oocytes were higher than those of non-injected and non-activated oocytes,but obviously lower than those of sham injected and activated oocytes.Comparing the three different protocols of man-made activation,we found that there were no significant difference among those activation rates of the injected oocytes which were either activated by CaCl2,ionomycin or electrical pulse（P〉0.05）.Fertilization rates of sperm injected oocytes activated by CaCl2-injection were higher than those stimulated by ionomycin（P〈0.05）,but no difference with those stimulated by electrical pulse（P〉0.05）.While parthenogenetic development rates of sham injected and control oocytes activated by CaCl2-injection were significantly lower than those activated by ionomycin and electrical pulse（P〈0.01）.After being activated by CaCl2-injection,the sperm injected oocytes showed a higher rates of cleavage（P〈0.05） and total cell number of blastocysts（P〈0.05） compared to control ones.Those results indicated that the activate effect of porcine oocytes was poor when they were activated only by injection,in order to improve the activation rates of oocytes,it is necessary to carry out the man-made activation.The efficiency of ICSI of the porcine oocytes which were activated by CaCl2-injection was higher than that of the other two different activated protocols respectively.