海藻糖用于小鼠卵母细胞玻璃化冷冻的研究

Effect of trehalose on mouse oocyte vitrification

目的探讨海藻糖对小鼠卵母细胞玻璃化冷冻的影响。方法采用12只小鼠控制性超排卵获得的360枚成熟的卵母细胞,分别放入含1.0、0.5、0.3及0 mol/L的海藻糖中孵育3 h。部分卵母细胞进行碘化丙啶(PI)染色;320枚卵母细胞进行玻璃化冷冻和复苏,并进行体外受精,观察卵母细胞成活率、受精率、卵裂率及囊胚形成率。结果 0.3 mol/L海藻糖组复苏率、受精率与对照组无显著差异(92.59%vs.90.90%,70.67%vs.61.43%),但是卵裂率、囊胚形成率均高于对照组(83.02%vs.44.19%,72.72%vs.47.37%)(P<0.001);0.5 mol/L海藻糖组复苏率、受精率、囊胚形成率与对照组无显著差异,但是卵裂率高于对照组(65.22%vs.44.19%)(P<0.05);0.3 mol/L海藻糖组囊胚形成率高于0.5 mol/L海藻糖组(72.72%vs.50.00%)(P<0.001);1.0 mol/L海藻糖组卵母细胞复苏率、受精率、卵裂率、囊胚形成率均低于对照组低(P<0.001)。结论一定浓度范围的海藻糖对小鼠卵母细胞的玻璃化冷冻有保护作用,0.3 mol/L海藻糖浓度比较适合小鼠卵母细胞玻璃化冷冻。

Objective： To examine the potential effect of applying different concentration of extracellular trehalose on mouse oocyte vitrification. Methods： The recover rate of mouse oocytes was detected as a functional index of different extracellular trehalose concentrations. A total of 360 oocytes were obtained from 12 superstimulated mice and cultured for 3 hours in freezing medium added 0, 0.3, 0.5, and 1.0 mol/L trehalose. Forty oocytes were stained with propidium iodide （PI）. Three hundreds and twenty oocytes were vitrified, warmed and tested the developmental potential after in vitro fertilization and culture. Results： There was no PI staining positive cell among the four groups. The fertilization rate and recover rate in 0.3 mol/L trehalose group were not significantly different with control group（92.59% vs. 90.90%, 70.67% vs. 61.43%）（P〉0.05）. The cleavage rate and blastocyst rate in the 0.3 mol/L group were significantly higher than those in the control group（83.02% vs. 44.19%, 72.72% vs. 47.37%）（P〈0. 001）. The recover rate, fertilization rate and blastocyst rate in 0. 5 mol/L trehalose group were not significantly different with those in control group （P〉0.05）, but the cleavage rate in 0.5 mol/L group was significantly higher than that in control group （65.22% vs. 44.19%） （P〈0.05）. The blastocyst rate in 0.3 mol/L group was significantly higher than that in 0. 5 mol/L group（72.72% vs. 50.00%） （P〈0. 001）. Conclusions. Certain amount of trehalose had protective effect on the vitrification oocyte of mouse. A concentration of 0.3 mol/L trehalose was more suitable for oocytes vitrification in mouse.