冷冻载体对扩张期囊胚解冻结局的影响

背景：囊胚的冷冻复苏对人类辅助生殖技术的发展具有重要意义,合适的冷冻载体可以降低冷冻毒性提高冷冻效率,是改善囊胚冷冻复苏结局的研究热点之一。目的：观察不同冷冻载体对昆明小鼠扩张期囊胚玻璃化冷冻解冻结局的影响。方法：以新鲜昆明小鼠扩张期囊胚作为对照组,随机选取同期小鼠胚进行玻璃化冷冻解冻,冷冻前使用激光仪行人工皱缩,使用玻璃化法进行冷冻和解冻,按实验载体不同分为冷冻环组、闭合拉细麦管组和自制麦管叶片组;冷冻环组和自制麦管叶片组投入液氮前与冷冻保护剂接触时间〈40s,40-60s和60-90s;解冻后囊胚使用胚胎囊期培养基添加10%血清替代品进行微滴培养。比较解冻后组间囊胚丢失率、囊胚复苏率和培养24h囊胚孵出率。结果与结论：相比闭合拉细麦管组,冷冻环组和自制麦管叶片组的囊胚丢失率降低,胚胎复苏率增高（P〈0.05）,各冷冻组复苏囊胚培养24h孵出率均低于对照组（P〈0.05）。使用冷冻环载体和自制麦管叶片载体冷冻解冻囊胚,投入液氮前与冷冻保护剂接触时间90s内不同时间胚胎复苏率和孵出率的比较差异无显著性意义。结果说明冷冻环载体和自制麦管叶片载体用于昆明小鼠扩张期囊胚玻璃化解冻复苏的效果较好。

Effect of frozen carriers on the outcomes of blastocysts freezing-thawing in the expanding period

BACKGROUND： Blastocyst freezing-thawing has great significance to the development of the human assisted reproductive technology; favorable frozen carrier can reduce the frozen toxic and improve refrigeration efficiency, which has been considered as the hot spot in improving the outcome of blastocyst freezing-thawing OBJECTIVE： To discuss the effects of different frozen carriers on the outcomes of Kunming mice blastocyst vitrification freezing-thawing in expanding period. METHODS： The fresh Kunming mouse blastocysts in the expanding period were considered as the control group, and then the mouse embryo were randomly selected from the same period for the vitrification thawing, and before frozen, artificial shrinkage was performed by the laser line; the vitrification method was used for freezing and thawing, and the embryos were divided into cryoloop group, closed pulled straw group and self-made straw-leaf group according to different experiment carriers. The duration of contact cryoprotectant before input liquid nitrogen were 〈 40 seconds, 40-60 seconds and 60-90 seconds in cryoloop group and self-made straw-leaf group; after thawing, 10% serum replacement was added in to the blastocyst medium supplemented for the micro-drop culture in embryonic sac period. The effects of the vitrified mouse blastocysts were assessed by the lost rate, recovery rate and hatched rate after cultured for 24 hours. RESULTS AND CONCLUSION： Compared with closed pulled straw group, the lost rate of the blastocysts was reduced and the recovery rate of the embryo was increased in cryoloop group and self-made straw-leaf group （P 〈 0.05）, while the hatched rate after cultured for 24 hours in the freezing groups was lower than that in the control group （P 〈 0.05）. There was no significant difference of the recovery rate and hatched rate in cryoloop group and self-made straw-leaf group when the duration of contact cryoprotectant before input liquid nitrogen was 90 seconds. These findings indicate that cryoloop carrier and self-made straw-leaf carrier has a good effect on blastocyst vitrification freezing-thawing in the expanding period.