冷冻后小鼠卵母细胞的透明带和质膜对体外受精的影响

Effects of the Membranes of Vitrificated Mouse Oocytes on in vitro Fertilization

以冷冻保存后的小鼠卵母细胞为实验材料,研究其膜对体外受精效果的影响。在室温(25±0.5)℃条件下,以10%乙二醇(EG)+10%二甲基亚砜(DMSO)为预处理液,EDFS30为玻璃化溶液。将卵母细胞于10%EG+10%DMSO溶液中预处理30s,然后再移入EDFS30中处理25s,以开放式拉长细管(OPS)为承载器投入液氮中,即两步法玻璃化冷冻保存。结果表明:冷冻卵母细胞受精后的卵裂率(46.67%)显著低于新鲜组的(86.06%)(P<0.05)。冷冻卵母细胞去透明带后质膜上的平均精子结合数(10.70)与对照组(10.81)无显著性差异(P>0.05),形成雌雄原核数也无显著性差异(2.49vs.2.59)(P>0.05)。冷冻卵母细胞透明带打孔后,其体外受精后卵裂率与新鲜组差异不显著(84.73%vs.91.19%)(P>0.05)。因此玻璃化冷冻保存小鼠卵母细胞透明带的变化是影响体外受精效果的主要因素之一。

To investigate the effect of the membranes of mouse oocyte on fertilization in vitro as a model using oocyte of mouse after vitrification. Under room temperature （25±0.5）℃, oocytes were vitrified in OPS by two steps vitification method with cryprotectant solutions-that is, 10%EG＋10%DMSO and EDFS30.0ocytes were firstly pretreated in 10% EG＋10% DMSO for 30 s, then exposed to EDFS30 for 25 s. The results showed that the cleavage rate of vitrificated oocytes were significantly lower （46.67% vs. 86.06%）（P 〈 0.05） than that of controls. The average number of sperm combined on the surface of the oocytes （10.70vs. 10.81） and the average number of pronuclears in oocytes （2.49 vs.2.59） were all similar （P 〉 0.05） between vitrificated and no-treated oocytes. After partial zona pellucida incision by piezo micromanipulation （ZIP）, the cleavage rate of the vitrificated oocytes was similar（84.73% vs.91.19%）（P 〉 0.05） with that of controls. In conclusion, the change of the zona pellucida of vitrificated mouse oocytes was one of the most important reasons to reduce the fetilization rate.