两种玻璃化冻融方案对人成熟期卵母细胞的冷冻效果比较

Comparation of two vitrified frozen-thrawed procedures for cyropreservation of mature human oocytes

目的 探讨两种玻璃化冻融方案对人成熟期（MⅡ期）卵母细胞纺锤体、染色体的损伤程度.方法 收集体外受精胚胎移植（IVF-ET）中受精失败MⅡ卵母细胞,分别用A法（实验组A,胎牛血清配方＋梯度平衡方案）以及B法（实验组B,重组白蛋白血清方案）冻融,对照组为未冷冻卵母细胞.解冻后存活卵母细胞及对照组卵母细胞固定后进行免疫荧光染色,之后用激光扫描共聚焦显微镜观察纺锤体、染色体形态.结果 实验组A、B的存活率分别为95.8%（23/24）、96.7%（29/30）,差异无统计学意义（P〉0.05）.实验组A的纺锤体和染色体形态正常率为13%（3/23）和13%（3/23）与对照组[18.2%（6/33）、18.2%（6/33）]差异无统计学意义（P〉0.05）.实验组B的纺锤体和染色体正常率为0%（0/29）和0%（0/29）与对照组差异有统计学意义（P〈0.05）.A组纺锤体缺失率（Ⅳ型）为65.3%（15/23）,与对照组39.4%（13/33）差异无统计学意义（P〉0.05）.B组纺锤体缺失率（Ⅳ型）为79.3%（23/29）,与对照组39.4%（13/33）差异有统计学意义（P〈0.01）.结论 玻璃化冻融方案A对人MⅡ期卵母细胞的纺锤体、染色体损伤程度较小.

Objective To compare the effects of two vitrified frozen-thrawed procedures on spindle and chromosome configurations of mature （M Ⅱ ） human oocytes. Methods M Ⅱ oocytes which failed fertilization during in vitro fertilization and embryo transfer （IVF-ET） were collected and treated with either procedure A （cryopreservation agent with FBS formula gradient balancing,A group） or procedure B （cryopreservation agent with recombined serum albumin B group）. The same procedures were given to a group of un-cryopreserved oocytes （control group）. After thawing,surviving oocytes of study groups and fresh oocytes of control group were stained by immunofluorescence. Then the spindles and chromosomes of oocytes were observed under confocal laser scanning microscope. Results The survival rates of group A and B were 95.8% （23/24） and 96.7% （29/30） respectively,with no significant difference betw een the study groups （P〉0.05）.13% （3/23） of spindle and 13% （3/23） of chromosome configurations in the A group remained normal,comparable with the findings in control group[18.2%（6/33） and 18.2%（6/33）,respectively]（P〉0.05）,while normal rates of spindle and chromosome configurations in B group[0%（0/29） and 0%（0/29）]were significantly lower than those in control group （P〈0.05）. The rates with absent spindle in A group and control group were 65.3% （15/23） and 39.4% （13/33） respectively （P〉0.05）,while the rate with absent spindle in B group was 79.3% （23/29）,with significant difference as compared with control group （P 〈0.01 ）. Conclusion The vitrified frozen-thrawed procedure A is less harmful to the spindle and chromosome configurations of human M Ⅱ oocytes.