A Role For TGF-β1 During Mouse Development Cultured In Vitro And Embryo Implantation And Its Related Mechanism

Purpose: 1 To investigate the role of TGF-β1 in in vitro culture of mouse embryo and embryo implantation by comparing the development status and implantation rate after blastocyste transfer between embryos cultured in M16 and M16 containing different level of TGF-β1.2 To study the immunoregulatory role for TGF-β1 during pregnant by comparing the IL-10/IFN-γin maternal-fetal interface between M16+TGF-β1 10ng/ml and control group(M16).3 To study the influence of mouse embryo culture media and TGF-β1 on embryo galectin-1 mRNA expression.Method: 1. 2-cell stage mouse embryo from superovulated mice were divided into four groups.Group A,B and C were cultured in Ml6 with different level of TGF-β1(lng/ml,10ng/ml and 50ng/ml respectively),and group D were contrals cultured with Ml6.The percentages of treated embryos reaching fixed stages(early blastocyst, expanding blastocyst and hatched blastocyst) were monitored when cultured for 68h and 92h respectively.The expanding blastocys cultured for 68h in Ml6+ TGF-β1 10ng/ml and in controls were transferred to pseudopregnant mouse. On the 6th day post transferring,the implantation rate of two groups were counted.2 On the 6th day post transferring,the embryo-placenta were removed from uterus and cultured for 24h.The supernate were collected and measured for IL 10/IFN-γby ELISA.3. Comparing the galectin-1 mRNA expression of embryos from in vivo and cultured in different media (M16,M16+ TGF-β1 1ng/ml and M16+ TGF-β1 50ng/ml).Results: 1.TGF-β1 can improve the development of mouse embryos cultured in vitro with more reaching blastocyst stage.When the TGF-β1 level was 10ng/ml,it is the most suitable media for embryos,in which the percentage of embryos reaching early, expanding, hatched stage after cultured for 68h or 92h were respectively 15.6%, 68.09%, 1.42% and 6.38%, 28.37%, 53.19%. When TGF-βl≥50ng/ml, the promoting effect decline.2. The implantation rate of embryos cultured in M16+TGF-β1 for 68h is higher than that of Ml6 significantly (35.42% vs 17.19%,p<0.05).3. The IFN-γlevel in supernate of M16+TGF-β1 group is lower than that of controls significantly (30.89±11.31pg/ml vs 43.23±18.09pg/ml, P<0.05)。4.Galectin-l mRNA expression level of embryos in vivo was higher than which cultured in Ml6 significantly. However the TGF-β1 did not influence the galectin-1 gene expression.Conclusion: Suitable level of TGF-β1 in mouse embryo in vitro culture system can effectively improve embryo implantation rate. The mechanism are improving the embryos quality and developing rate, and also decreasing the IFN-y production in maternal-fetal interface,which cause a Th2 bias. The embryo in vitro culture system may decrease the galectin-1 mRNA expression level, which maybe one of the reason for lower implantation rate and higher spontaneouse abortion rate for embryos cultured in vitro. However, TGF-β1 can not improve the galectin-1 gene expression.