子宫内膜细胞共培养联合序贯成组培养对人早期冷冻胚胎发育的支持作用

Uterine endometrial cells coculture and sequential system support the development of early human discarded frozen embryos

目的探讨子宫内膜细胞体外共培养联合序贯成组培养对人早期废弃冷冻胚胎发育的影响。方法获取人子宫内膜细胞并冷冻保存。共培养前,解冻内膜细胞并培养至30%～50%汇合时解冻人早期废弃胚胎,胚胎冷冻年限为3～10年。2～3个胚胎一起转移至已更换胚胎培养液的内膜细胞上行成组共培养,待胚胎发育至8-细胞期更换为囊胚培养液,观察胚胎复苏存活、桑葚胚及囊胚形成情况。结果人子宫内膜细胞生长良好。采取内膜细胞联合序贯成组培养废弃胚胎33个,总体桑葚胚形成率45.5%,囊胚形成率27.2%,其中Ⅰ～Ⅱ级胚胎桑葚胚、囊胚形成率皆高于Ⅲ～Ⅳ胚胎,差异有统计学意义(P<0.05)。结论子宫内膜细胞共培养联合序贯成组培养对人早期冷冻时限较长的胚胎发育具有积极的支持作用,可为充分利用宝贵资源进行胚胎实验室质控、胚胎干细胞研究等领域提供支持性方案。

Objective To observe the effects of endometrial cells coculture and sequential system on early human embryonic development in vitro. Methods Human uterine endometrial cells were isolated and preserved in frozen status. Then the cells were cultured to 30%- 50 % cell fusion, and then coeultured with thawed early human frozen discarded embryos （frozen for 3 - 10 years） in fresh embryo culture media. When the embryos developed in 8-cell stage, blastula culture media was applied. The percentages of resuscitation, morulas formation and blastcysts hatching were observed. Results Human endometrial cells had viability in vitro culture. Coculture with endometrial cells could support the formation of morula （45.5% , P〈0.05） and blastoeyst （27. 2%, P〈0. 05）. Furthermore, classⅠ and Ⅱembryos had higher proficiency of morula （57.7% and 0.0 %, P〈0.05） and blastocyst （34. 6 % and 0. 0%, P〈 0.05） development compared with class Ⅲ-Ⅳ embryos. Conclusions Endometrial cells coculture and sequential system support the development of early human frozen discarded embryos （frozen for 3 - 10 years） in vitro, which can serve as an efficient protocol for human discarded embryo culture in vitro.