【摘要】 目的:研究冻存后的小鼠成熟卵母细胞复苏后培养不同时间对ICSI结果的影响,观察Ca2+载体霉素联合嘌呤霉素对ICSI受精失败的卵母细胞补救激活的有效性。方法:采用慢冻-快融程序化冷冻方法冻融小鼠成熟卵母细胞,复苏后卵母细胞分别培养不同时间(1h、2h、3h、4h、5h)后行ICSI,比较其受精和胚胎发育情况。冻融后受精失败的卵母细胞分为两组(A组:辅助激活,B组:不采取辅助激活),另外获取ICSI受精失败的新鲜卵母细胞(C组)采用和A组同样方法辅助激活,观察3组激活效果。结果:复苏的卵母细胞ICSI前培养3~4h后正常受精率、囊胚形成率明显高于培养1h、2h和5h实验组;A组激活率与B组相比,有明显差异(30.4%vs 6.7%,P<0.05);A组激活率和2PN2PB比例明显低于C组(30.4%vs 75.6%,P<0.05;21.4%vs 47.1%,P<0.05);A、C两组之间1PN2PB比例相比无统计学差异。结论:复苏的成熟卵母细胞ICSI前培养3~4 h后有助于卵母细胞结构的恢复,提高正常受精率和后期发育潜能。辅助激活在一定程度上可以挽救ICSI受精失败的卵母细胞;由于冻融损伤,复苏后ICSI受精失败的卵母细胞被辅助激活的能力明显下降。更多还原

【Abstract】 Objective:To investigate whether the culture time was associated with the results of the mouse oocytes intracytoplasmic sperm injection(ICSI) undergo cryopreservation protocol.And to attempt to remedy these mouse oocytes that fail to fertilization after ICSI by adopting assisted activation with calcium ionophore and puromycin.Method:After thawing,the survival oocytes with normal zona pellucida and cytoskeletal were randomly assigned to five groups by culture different time(1h,2h,3h,4h,5h) before inseminating.We compared the percentages of fertilization,embryo cleavage and blastocyst.Furthermore,this study assessed the effect of assisted activation with calcium ionophore and puromycin on mouse oocytes that fail to fertilization after ICSI.Unfertilized mouse oocytes without fragment were randomly assigned to two groups(group A:assisted activation,group B:without activation).Another fresh unfertilized oocytes(group C)were activated by the same activated method as group A.Results:For thawed oocytes,ICSI produced higher fertilization rate and blastocyst rate after culturing 3h and culturing 4h with respect to the results after culturing 1h and culturing 2h and 5h.For mouse oocytes that fail to fertilization after ICSI,the activation rate in group A was higher than the consequence of group B(30.4% vs 6.7%,P<0.05).Statistically decrease of activation rate and 2PN2PB rate were observed in group A than group C(30.4% vs 75.6%,P<0.05;21.4% vs 47.1%,P<0.05).No significant difference in 1PN2PB rate between group A and group C.Conculsion:After thawing,the 3h～4h incubation could permit restoration of the ultrastructure and improve the normal fertilization rate and embryo developing ability for mouse oocyte.Oocyte assisted activation with calcium ionophore and puromycin may be effective for mouse oocytes that fail to fertilize after ICSI.Because of irreversible injure during cryopreservation,the ability of been assisted activated was significant decreased for mouse thawed oocytes that failed to fertilize after ICSI compared to fresh unfertilized oocyte after ICSI.更多还原