两种方法冻存胎儿卵巢组织后的活力判断

目的 比较常规慢速冻存法与快速冻存法保存胎儿卵巢组织的效果,为快速冻存法的应用提供依据.方法 以1.5 mol/L乙二醇与0.1 mol/L蔗糖作冷冻保护剂,分别采用常规慢速冻存法和三步快速冻存法保存1 mm3大小的胎儿卵巢组织,解冻后观察两组卵巢组织内的超微结构;以去势裸鼠为受体,将解冻后的胎儿卵巢皮质片移植于裸鼠的双肾被膜下,观察移植2周和8周后移植物的组织学结构、移植物存活率、卵泡密度及受体鼠血清E2水平.结果 与新鲜胎儿卵巢组织相比,慢速和快速冷冻后卵巢组织中形态正常的卵母细胞的比例都明显减少(P<0.05).移植后2周的两移植组均有卵泡存活,原始卵泡广泛分布于基质中;移植后8周的两移植组的卵泡数量都明显减少,但初级卵泡的数量相对增加.比较移植2周和8周后移植物的存活率、单位面积移植物内的原始卵泡或初级卵泡数及受体鼠血清雌激素水平,两移植组并无显著性差异.结论 三步快速冻存法可有效保存胎儿卵巢组织的生物活性,解冻后卵巢组织的形态学结构、雌激素分泌与常规慢速冻存组并无显著性差异.

Viability evaluation of human fetal ovarian tissue after frozen by two different freezing methods

Objective To compare the effects of slow-rate freezing method with that of rapid freezing method for cryopreservation of fetal ovarian tissue on viability of ovarian tissue and provide some data for the application of rapid freezing method.Methods Human fetal ovarian slices(1mm3 in size) were frozen with conventional slow cooling method and three-step rapid freezing method,respectively,using freezing solution containing 1.5mol/L ethylene glycol and 0.1mol/L sucrose.After thawed,electron microscopy was used to observe the ultrastructural changes in ovarian tissue.Ovariectomized female nude mice were chosen as recipients,two fetal ovarian tissue slices were separately inserted under kidney capsule of two sides in each mouse.The survival of ovarian grafts,histological structure,follicular density and serum E2 levels in recipients were detected after transplantation for two or eight weeks.Results Compared with that of fresh ovarian tissue,the number of oocytes with normal morphology in slow and rapid freezing groups was significantly decreased(P<0.05).Two weeks after transplantation,numerous follicles were found in both groups and primordial follicles were widely dispersed in the stroma.Eight weeks after transplantation,the number of follicles in both groups was reduced,but the number of primary follicles was relatively increased.There were no significant differences in the survival rate of ovarian grafts,follicular density and serum E2 level between the two groups.Conclusion The viability of fetal ovarian tissue can be well preserved by three-step rapid-freezing method and there are no differences in morphology and estrogen secretion of ovarian tissue after thawed,compared with the conventional slow freezing method.