玻璃化冷冻及程序冷冻对人卵巢组织卵泡活性的影响

Effects of Vitrification and Program Freezing on the Follicular Viability of Human Ovarian Tissues

目的:比较玻璃化冷冻及程序冷冻对人卵巢组织卵泡活性的影响。方法:将15例人卵巢组织20 min内运至实验室,切割成1 mm×5 mm×5 mm大小的卵巢皮质片,随机分为新鲜组、玻璃化冷冻组及程序冷冻组。玻璃化冷冻组及程序冷冻组均采用优化的实验方法进行。新鲜及复苏组织体外培养后,在光镜及电镜下分析卵泡的形态正常率,采用激素释放试验及异体移植试验检测冷冻复苏的人卵巢组织的功能,并与新鲜组比较。结果:冷冻复苏组及新鲜对照组卵巢组织皮质内均可见形态正常和异常的始基及初级卵泡,新鲜对照组卵泡形态正常率显著高于玻璃化冷冻组和程序冷冻组(P<0.05);程序冷冻组形态正常率稍高于玻璃化冷冻组(P>0.05)。3组均可见超微结构正常及异常的原始卵泡、初级卵泡。卵巢皮质片体外培养后持续分泌雌二醇和孕酮。新鲜组激素分泌水平高于玻璃化冷冻组和程序冷冻组,但差异无统计学意义(P>0.05)。异体移植试验显示,程序冷冻组较玻璃化冷冻组具有更高的移植物回收数及卵泡密度。结论:由于程序冷冻人卵巢组织卵泡具有较高的发育潜能,因此其较玻璃化冷冻更有应用前景。

Objective：To compare the effect of vitrification and program freezing on the follicular viability of human ovarian tissues.Methods：Ovarian tissues from15 patients were transported to the laboratory within 20 min after excision.Pieces of ovarian tissues（1 mm×5 mm×5 mm）were randomly divided into fresh group,vitrification group and program freezing group.Optimal experimental method was used in vitrification group and program freezing group.The fresh and resuscitated tissues were cultivated in vitro,and the follicular morphology was analyzed by using light and electron microscope.The viability of ovarian tissues resuscitated from freezing was detected by hormones releasing assay and allograft assay,and then compared with that of fresh group.Results：Morphologically normal and abnormal primordial and primary follicles could be seen in resuscitated freezing group and fresh group.The rate of morphologically normal follicles in fresh group was significantly higher than those in vitrification group and program freezing group（P〈0.05）.The rate of morphologically normal follicles was slightly higher in program freezing group than that in vitrification group（P〉0.05）.Primordial and primary follicles with normal and abnormal ultrastructure could be seen in all the three groups.Ovarian cortical pieces secreted estradiol and progesterone continuously after in vitro culture.Hormone levels in fresh group were higher than those in vitrification group and program freezing group（P〉0.05）.Allograft assay showed that there were more graft recovery number and higher follicular density in program freezing group than those in vitrification group.Conclusions：Follicle has higher potentiality of development in program freezing human ovarian tissues,so program freezing is more promising than vitrification in application.