不同融合参数对小鼠圆形精子细胞与卵母细胞融合胚胎发育的影响

Effects of different electrocution parameters on development of embryos fused by round spermatids and oocytes

目的利用手工克隆(handmade cloning,HMC)技术对小鼠圆形精子细胞与去透明带卵母细胞进行电融合,以研究电融合参数对融合胚胎发育潜能的影响。方法选取549枚卵母细胞与圆形精子细胞融合,根据电融合参数分为三组:A组(n=186)采用电场强度60V/mm、脉宽30μs、脉冲次数为2次的直流电脉冲;B组(n=188)采用电场强度80V/mm、脉宽20μs、脉冲次数为2次的直流电脉冲;C组(n=175)采用电场强度100V/mm、脉宽10μs、脉冲次数为2次的直流电脉冲。观察记录不同电融合参数下圆形精子细胞与无透明带卵母细胞的融合率、融合胚胎卵裂率以及囊胚形成率。结果(1)A组圆形精子细胞与去透明带卵母细胞的平均融合率为(84.87±3.84)%,与其他两组[(74.93±5.46)%、(73.60±5.48)%]相比平均融合率显著升高(P<0.05);(2)A组融合胚胎的平均卵裂率为(77.37±3.97)%,与B组、C组融合胚胎的卵裂率[(67.74±7.32)%、(66.17±9.87)%]相比显著升高(P<0.05);(3)A组的平均囊胚形成率为(41.28±6.50)%,显著高于C组[(31.03±6.09)%](P<0.05)。结论 A组采用电场强度60V/mm、脉宽30μs、脉冲次数为2的直流电脉冲是圆形精子细胞与去透明带卵母细胞融合的最佳方案,有利于提高胚胎的发育潜能。

Objective： To explore the effects of different electrofusion parameters on the developmental potential of embryos which were fused by round spermatids and zona-free oocytes with handmade cloning （HMC）. Methods： A total of 549 oocytes and round spermatids were fused by handmade cloning. The electric fusion parameters were divided into three groups according to the different electrofusion parameters. The electric field intensity was 60, 80 and 100V/mm in group A （n=186）, group B （n=188）and group C（n： 175）respectively. The pulse width was 30, 20, 10 μs of pulse width in group A, B, C respectively, and all number of direct current （DC） was 2. The electric fusion rates, cleavage rate and blastocyst formation rate of embryos fused by round spermatids and zona-free oocytes of mouse under the different electrofusion parameters were measured and recorded. Results： The results showed that the fusion rate of group A [（84.87 ± 3.84）%]was significantly higher than that of the other two groups [（74.93±5.46）%, （73.60±5.48）%, respectively] （P^0.05）. The cleavage rate of group A [（77.37±3.97%] was significantly higher than that of group B or group C [（67.74±7.32）G, （66.17 ± 9. 87）G, respectively] （P〈0.05）. The blastocyst rate of group A [（41.28±6.50） %] was significantly higher than that of group C [（31.03±6.09） %] （P%0.05）. Conclusions. The best electrofusion parameters are 60 v/ram and 30μsX2. It is beneficial to improve the developmental potential of embryo.