梯度平衡法改善猪卵巢组织玻璃化保存效果

目的研究梯度平衡法对卵巢组织玻璃化冷冻后组织形态学及凋亡的影响。方法取新鲜猪卵巢切成小块,未进行任何处理的为对照组、采用梯度平衡法冷冻的为梯度平衡组、采用直接平衡法冷冻的为直接平衡组。HE染色观察3组组织形态,原位末端转移酶标记(TUNEL)技术检测细胞凋亡情况。结果与对照组相比,梯度平衡组和直接平衡组卵泡的正常率均下降(均P<0.05),但后两组比较无显著差异(P>0.05)。与对照组相比,梯度平衡组与直接平衡组的细胞凋亡率均显著增加(P<0.05),但梯度平衡组相对于直接平衡组,卵母细胞、颗粒细胞、基质细胞凋亡率(分别为14.8%vs.17.6%、13.8%vs.15.2%和15.7%vs.18.5%)均显著降低(P<0.05)。结论梯度平衡法可以减少玻璃化冻融卵巢组织过程中的细胞凋亡。

Gradient equilibrium method improves the vitrification preservation effect of porcine ovarian tissue

Objective:To evaluate the effect of gradient equilibrium method on the morphology and apoptosis of ovarian tissue after vitrification.Methods:Fresh pig ovaries were cut into small pieces.The ovarian tissue without any treatment was used as the control group,those frozen by the gradient equilibrium method were in the gradient equilibrium group,and those frozen by the direct equilibrium method were in the direct equilibrium group.The tissue morphology of three groups were observed by HE staining,and cell apoptosis was detected by in situ terminal transferase labeling(TUNEL)technique.Results:The normal rate of follicles in the gradient equilibrium group and the direct equilibrium group decreased compared with the control group(P<0.05),but there was no significant difference between these two groups(P>0.05).The apoptosis rate of the gradient equilibrium group and the direct equilibrium group were significantly increased compared with the control group(P<0.05).However,the apoptosis rates of oocytes,granulosa cells and stromal cells(14.8%vs.17.6%,13.8%vs.15.2%and 15.7%vs.18.5%respectively)in gradient equilibrium group were significantly lower compared with the direct equilibrium group(P<0.05).Conclusions:Gradient equilibrium method can reduce cell apoptosis in the process of vitrification and thawing of ovarian tissue.