【摘要】 目的探索不同卵巢组织条厚度对超低温冷冻效果的影响,同时评估卵巢组织的凋亡情况。方法卵巢组织取自10例手术的妇女。冷冻前行卵巢组织切片,按最小厚度随机分成1.0、0.5、0.25mm 3组。采用二甲基亚砜(DMSO)为冷冻保护剂的程序慢速冷冻法进行超低温冷冻保存。冻融后进行卵泡计数及各组间正常卵泡形态学分析比较,使用TdT介导的dUTP缺口末端标记技术(TUNEL)进行组织细胞凋亡检测。上述结果与新鲜卵巢组织进行比较。结果新鲜卵巢组中97.6%原始卵泡保持形态学正常,切片厚度为1.0、0.5、0.25 mm三组形态正常率分别为71.1%、73.9%和72.7%,与新鲜组比较,三实验组下降明显,差别有统计学意义(χ~2=15.66,P<0.001);而三冷冻组间相比差别无统计学意义(χ~2=0.153,P=0.926)。TUNEL原位杂交显示窦前卵泡颗粒细胞中未见凋亡阳性信号,窦卵泡中可见散在阳性信号细胞,卵巢间质细胞亦有散在凋亡信号。结论 0.5～1.0mm厚度卵巢组织为行超低温冷冻保存是较合理厚度,卵泡闭锁的机理值得进一步研究。更多还原

【Abstract】 Objective:To explore the effect of different thicknesses of ovarian tissue strips on the outcomes of human ovarian cryopreservation. Methods:The ovarian tissues were got from 10 women who underwent operations.The tissues were cut into fixed depths before frozen.In accordance with the minimum thickness,the strips were randomly divided into 3 groups:1.0 mm,0.5 mm and 0.25 mm,respectively.With dimethyl sulfoxide(DMSO) as the cryoprotectant,the ovarian tissues were cryopreserved by a slow-freezing procedure.After thawing, follicles were counted and the proportion of morphologically intact follicles in each group was analyzed.Also, the TUNEL was used for apoptosis detection.The results were also compared with those of the fresh ovarian tissues. Results:In the fresh tissues,97.6%of primordial follicles maintained normal morphology.Compared with that in the fresh group,the proportions of morphologically normal primordial follicles in three groups of frozen tissues(71.1%,73.9%and 72.7%in groups of 1 mm,0.5 mm and 0.25 mm,respectively) decreased significantly(χ~2=15.66,P<0.001).But the difference was not significant among three groups of the frozen tissues(χ~2=0.153,P = 0.926).The result of TUNEL showed that granulosa cells from preantral follicles had no positive signal.But there were scattered positive cells in the antral follicles,and also in the ovarian stroma in the frozen tissues. Conclusions:The reasonable thickness for the ovarian tissue cryopreservation is 0.5 - 1 mm.The mechanism of follicular atresia is worth further study.更多还原