小鼠囊胚玻璃化冷冻研究及超微结构观察

发布时间：2018-01-16 21:47

本文关键词：小鼠囊胚玻璃化冷冻研究及超微结构观察　出处：《昆明医学院》2006年硕士论文　论文类型：学位论文

【摘要】：目的研究由两种冷冻保护剂乙二醇(Ethylene Glycol，EG)和二甲亚砜(dimethylsuphoxide，DMSO)单独或联合配伍的改良玻璃化溶液对小鼠囊胚超快速冷冻保存的效果；比较聚蔗糖(Ficoll 70)的添加对超快速冷冻的作用；初步确定囊胚冷冻保存最佳玻璃化溶液及玻璃化冷冻的最佳囊胚发育阶段；通过透射电镜观察找出囊胚冷冻损伤的超微结构改变。方法 1、对6-8W龄昆明小白鼠促排卵，处死小鼠后获得小鼠初级囊胚和扩张囊胚。2、分别采用六组玻璃化溶液(ES35、EDS35、ESF10、ESF20、EDSF10和EDSF20)对小鼠初级囊胚和扩张囊胚进行玻璃化冷冻。3、透射电镜下观察新鲜扩张囊胚和各冷冻组冻融扩张囊胚的超微结构。结果 1、对于初级囊胚的存活率和孵出率均是EDSF10组最高，与新鲜对照组无显著性差异；ES35组最低与EDSF10组和新鲜对照组之间均存在显著性差异；2、对于扩张囊胚存活率各实验组间均无显著性差异，与新鲜对照组均有显著性差异；对于扩张囊胚的孵出率ES35组最低，EDSF10组最高，与新鲜对照组均存在显著差异。3、EG(?)DMSO对于囊胚的存活率和孵出率均有统计学意义。4、Ficoll70对于初级囊胚的存活率有统计学意义，浓度为10％或20％时的存活率均显著高于浓度为0％时，浓度为10％时存活率高于20％时，，但无统计学意义。5、除ES35组外各冷冻组，初级囊胚存活率显著高于扩张囊胚；EDS35组、ESF20组和EDSF20组的初级囊胚孵出率均显著高于扩张囊胚孵出率。6、透射电镜观察显示扩张囊胚冷冻损伤主要是细胞膜性结构和细胞骨架结构的损伤。结论 1、DMSO的添加对于改善玻璃化溶液具有重要作用，尤其是有助于初级囊
[Abstract]:Objective to study two kinds of cryopreservation agents, ethylene glycol Glycoline (EGG) and dimethyl sulfoxide dimethylsuphoxide (DMSO). Effects of modified vitrification solution alone or in combination with DMSO on ultrafast cryopreservation of mouse blastocysts; The effects of polysucrose Ficoll 70 on ultra-fast freezing were compared. The optimum vitrification solution for cryopreservation of blastocyst and the optimum stage of blastocyst development for vitrification were preliminarily determined. The ultrastructural changes of blastocyst cryopreservation injury were observed by transmission electron microscope (TEM). Methods 1. Primary blastocyst and expanded blastocyst were obtained from 6-8 W old Kunming mice. Six groups of vitrified solution (ES35) EDS35 were used respectively to obtain mouse primary blastocyst and dilated blastocyst. ESF10 ESF20 EDSF10 and EDSF20) vitrified mouse primary blastocyst and expanded blastocyst. The ultrastructure of fresh expanded blastocysts and freeze thawed blastocysts were observed under transmission electron microscope. Results 1. The survival rate and hatching rate of primary blastocysts in EDSF10 group were the highest, and there was no significant difference between the two groups. There was significant difference between ES35 group and EDSF10 group and fresh control group. 2. There was no significant difference in blastocyst survival rate between each experimental group and fresh control group. The hatching rate of expanded blastocysts in ES35 group was the highest, which was significantly different from that in fresh control group. DMSO has statistical significance for survival rate and hatching rate of blastocyst. 4 Ficoll70 has statistical significance for survival rate of primary blastocyst. The survival rate at the concentration of 10% or 20 was significantly higher than that at the concentration of 0, and the survival rate at the concentration of 10 was higher than that at the time of 20, but there was no statistical significance. 5. The survival rate of the freezing group was higher than that of the control group except the ES35 group. The survival rate of primary blastocyst was significantly higher than that of expanded blastocyst. The hatching rate of primary blastocysts in EDS35 group and EDSF20 group was significantly higher than that in expanded blastocyst group. Transmission electron microscopy (TEM) showed that the freeze injury of expanded blastocyst was mainly caused by membrane structure and cytoskeletal structure. Conclusion 1 the addition of DMSO plays an important role in improving vitrification solution, especially in primary sac.
【学位授予单位】：昆明医学院
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R321-33

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