冻融液的温度对人未成熟卵母细胞玻璃化冻融效果的影响  

The effect of incubation temperature of vitrification and warming solutions on the viability and maturation of vitrified-thawed human premature oocytes

作　　者：龙晓林[1] 黄玉玲[1] 孙筱放[1] 范勇[1] 杜红姿[1] 石宇[1] 张文红[1] 李莉[1] 

机构地区：[1]广州医学院第三附属医院生殖中心,广东广州510150

出　　处：《生殖医学杂志》2010年第3期219-223,共5页Journal of Reproductive Medicine

基　　金：广东省卫生局基金资助项目(B2009110)

摘　　要：目的探讨冻融液的操作温度对未成熟人卵母细胞(GV、MI)玻璃化冻融效果的影响。方法收集卵胞浆内单精子注射(ICSI)患者的GV、MI卵母细胞,根据冻融液操作温度的不同分为A、B、C组。冻融后存活及未冷冻对照组(D组)卵母细胞体外成熟(IVM)培养,成熟的MII卵母细胞固定后进行免疫荧光染色,然后用Nikon CISI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GA、GB、GC卵母细胞冻融后的存活率和体外成熟率分别为100%、81.3%、68.8%和33.3%、83.3%、72.7%,仅GC组的存活率与对照比较存在显著性的差异(P<0.05);实验组中只有GB组获得了纺锤体(20%)和染色体(10%)形态正常卵母细胞。实验组MA、MB、MC卵母细胞冻融后的存活率分别为71.4%、100%、83.3%,各实验组与对照组比较无显著性差异(P>0.05);MA组体外成熟率与对照组比较存在极显著性的差异(0%比57.1%,P<0.01),MB、MC与对照组的体外成熟率比较无显著性的差异(66.7%、80%比57.1%,P>0.05);仅MC获得1个正常纺锤体和染色体形态卵子。结论冻融液的合适操作温度可以提高未成熟卵子的冻融效果。Objective： To investigate the viability and maturation of frozen-thawed human premature oocytes exposed to different temperatures of vitrification and warming solutions. Methods： The premature oocytes （GV and MI stages） were collected from our ICSI patients and exposed to different temperatures of vitrification solution before frozen and warming solutions （Group A, B,C） in the freezing/thawing procedures. The temperatures were as follows： Group A： equilibration solution at 37℃, vitrification solution at room temperature, warming solution at 37~C; Group B： both vitrification and warming solution at room temperature; Group C： both vitrification and warming solutions at 37℃. The frozen-thawed oocytes and the fresh oocytes （control, Group D） were cultured for in vitro maturation. The survival rate and maturation rate were compared between groups. The immunofluorescene-stained oocytes were examined under a confocal laser scanning microscope to check the spindle configuration and chromosome arrangement. Results： The survival rates and MII rates of GV stage oocytes in Group A, B, C were 100 %, 81.3 %, 68.8% and 33.3% ,83.3% ,72.7%, respectively, and only the survival rate of Group C was significantly lower than that of control （P 〈 0. 05）. The normal spindle and chromosome configuration were only observed in the oocytes of Group B, with the rates of 20% and 10%, respectively. For those frozenthawed oocytes of MI stage, the survival rates in Group A,B,C were 71. 4%, 100% and 83. 3%, not significantly different from that of control. The MII rates in Group A, B and C were 0%, 66. 7% and 80%, respectively, with the significantly lower MII rate in Group A than in other groups P〈0.01）. Only one oocyte in Group C was found with normal spindle and chromosome configurations. Conclusions： The right operation temperature of vitrification and warming solutions can improve the outcomes of the vitrified-thawed human premature oocytes.

关 键 词：玻璃化冷冻 未成熟卵母细胞 纺锤体 染色体 温度 

分 类 号：R[医药卫生]