超促排卵后体外受精对小鼠囊胚发育的影响

Effect of in vitro fertilization following superovulation on the development of mouse blastocysts

目的观察超促排卵后体外受精对小鼠囊胚发育的影响。方法根据体外受精的关键步骤,包括超促排卵、体外受精和体外胚胎培养,设置自然交配组(n=8)、超促排自然交配组(n=8)、体外培养组(n=5)、超促排体外培养组(n=6)和体外受精组(n=15)。根据Gardner分级,观察各组E3.5的囊胚发育情况。比较超促排体外培养组和体外受精组Ⅴ级囊胚的孵出面积之比,分析体外受精组囊胚移植后的胎鼠妊娠率。统计学方法采用χ^(2)检验和Mann-Whitney非参数检验。结果体外受精组的Ⅰ~Ⅴ级囊胚构成比同其他4组显著不同(总体χ^(2)=188.200,P<0.001),体外受精组的E3.5囊胚主要以Ⅲ~Ⅴ级囊胚为主,比例达74.5%,且Ⅴ级囊胚占到51.2%,而其他4组主要以Ⅲ级和Ⅳ级囊胚为主。体外受精组Ⅴ级囊胚的孵出面积比例显著高于超促排体外培养组[0.11(0.05,0.18)与0.04(0.02,0.07),P<0.017],且体外受精组Ⅴ级囊胚的孵出面积呈现偏态分布。体外受精组的囊胚移植后,E12.5胎鼠的妊娠率显著低于体外培养组(45.3%与63.4%,χ^(2)=8.647,P<0.05)。此外,将体外受精组孵出面积<5%和5%~10%的囊胚分别进行移植,发现孵出面积<5%的囊胚移植后,E12.5胎鼠的妊娠率显著高于孵出面积5%~10%组(52.5%与29.7%,χ^(2)=7.578,P<0.05)。结论超促排卵后的体外受精可导致小鼠囊胚发育不同步,且孵出面积比例较大(5%~10%)的体外受精囊胚移植妊娠率较低。

Objective To observe the effect of in vitro fertilization(IVF)following superovulation on the development of mouse blastocysts.Method Based on the key procedures of IVF,including superovulation,IVF,in vitro embryo culture,five groups were set up:natural mating group(n=8),superovulation+natural mating group(n=8),in vitro culture group(n=5),superovulation+in vitro culture group(n=6),IVF group(n=15).According to Gardner's classification standard,the development status of mouse blastocysts was observed at E3.5.The ratio of hatched area of stageⅤblastocysts was compared between superovulation+in vitro culture group and IVF group,and the survival rate of fetuses after transferring of IVF mouse blastocysts was analyzed.The statistical methods were performed by Chi-square test and Mann-Whitney nonparametric test.Result The constitute ratios of different blastocyst stages(Ⅰ-Ⅴ)in IVF group significantly differed from other four groups(the globalχ^(2)=188.200,P<0.001).At E3.5,blastocysts in IVF group were mainly at the stageⅢ-Ⅴ,accounting for 74.5%,and the ratio of stageⅤblastocysts reached to 51.2%,and blastocysts in other four groups were mainly at the stageⅢandⅣ.The ratio of the hatched area of stageⅤblastocysts in IVF group was significantly higher than that in superovulation+in vitro culture group[0.11(0.05,0.18)vs 0.04(0.02,0.07),P<0.017],and the hatched area ratio of stageⅤblastocysts in IVF group showed skewed distribution.After blastocysts transferring,the survival rate of fetuses at E12.5 in IVF group was significantly lower compared to in vitro culture group(45.3%vs 63.4%,χ^(2)=8.647,P<0.05).Moreover,when transferring IVF blastocysts with different hatched area ratio(<5%vs 5%-10%)to uteruses,the data showed that the survival rate of fetuses at E12.5 from blastocysts(<5%hatched area)was robustly higher than blastocysts(5%-10%hatched area)(52.5%vs 29.7%,χ^(2)=7.578,P<0.05).Conclusion IVF following superovulation leads to asynchronous development of mouse blastocysts,and IVF blastocysts with large hatched area(5%-10%)have lower survival rate after transferring.