Effect Of Early Secretive Phase Human Endometrial Cells On Early Mouse Embryonic Development In Vitro

Objective Endometrial cells and mouse embryo are co-cultured to investigate the effect of human endometrial cells on early mouse embryonic development in vitro, so we can deduce the mechanism of coculture, to prepare for the study of human autologous endometrial epithelial cells and embryos coculture, furtherly to increase the pregnancy rate in patients with implantation failure by use of a co-culture system for human embryos development on autologous endometrial monolayer cells. Methods Endometrium specimens were obtained from volunteers undergoing IVF for the fallopian tube factor during the early secretive phase, then the tissue was minced and digested enzymatically. Firstly,the endometrial cells were cultured, achieved generation, cryopreserved, thawed and cryo-thawed, so we gained the post-cryo-thawed(Group A) and passage endometrial cells(Group B) monolayer. Secondly, immunohistochemistry was employed to express ER, PR in the endometrial cells.Thirdly,2-cell mouse embryos were collected after insemination for 24h,then distributed individually the 2-cell embryos to coculture (Group A and B) or medium culture alone(control group), observed every 24h and to grade embryos on the basis"of morphology, then selected some morula to detect cell apoptosis by the TUNEL. At last, to compare the number of embryos at the 4-8cell,morula,expanded,and hatching blastocyst after culture 24,48,72,96h,at the same time,to detect the apoptosis of 8-cell embryos and morula. Results 1. The cryo-thawed and passage human endometrial cells were applied to test inimmunohistochemistry, they were both employed to express ER, PR.2. After coculture and medium culture alone for 24h, some of the mouse embryos stopped in two-cell stage, but the number of embryos that stopped in two-cell stage in the co-culture group(A and B) were remarkably less than that in the control group(C), group A and group B were not different, the blocked embryos did not cleave after 24h.3. The rates of formation of four-to eight-cell,morula,blastocyst and hatching blastocyst were significantly higher(P< 0.005) in coculture group than those in control group,but those were similar between the group A and group B.4. The rates of formation of high quality embryos(grade A) were higher(A:63.29%, B:62.79%) in coculture group than those in control.There were no difference between the group A and group B.5. The percentage of eight-cell and morula with apoptotic blastomere(P<0.001=.as well as the number of apoptotic cells per blastomere were significantly lower in the coculture group than those in control group.6. Some morula were selected randomly in three groups to scatter, then counted the number of cells.The mean cells per morula were no difference between coculture group and control group.However the percentage of apoptotic cell of morula were.*significantly lower in the cocultured embryos than those in control group. Conclusion l.To coculture the mouse embryos with early secretive phase human endometrial cellscan improve the developmental block in 2-cell stage efficiently. 2.To coculture the mouse embryos with human endometrial cells can promote thedevelopment and enhance the quality in early stage of mouse embryos. 3.Coculture can reduce the incidence of apoptosis of mouse embryos.