생쥐 배아발달에 미치는 인슐린의 효과 = Effect of Insulin on Development of Mouse Preimplantion Embryos

다국어 초록 (Multilingual Abstract)

Although insulin is recognized as an important fetal growth factor, interaction with embryonic cells during the early development stages prior to the appearance of a functional pancreas has not been definde. But there is evidence that insulin binding to the cells of the preimplantation mouse embryos can be detected first at the morula stage, while autoradiographic studies have confirmed that insulin binding is receptor mediated. Other sutides have provided evidence that insulin is capable of stimulating glucose transport and protein synthesis in preimplantation mouse embryos. In this study, to investigate the effect of insulin on development of mouse preimplantation embryos, we performed the culture of mouse embryos from 1-cell zygote to blastocyst in modified Earle`s blanced salf solution(mEBSS), A chemically defined simple medium, supplemented with porcine insulin,hypoxanthine and/or ethylenediaminetetraacetic acid(EDTA), and the embryo development was scored at 24hours(2-cell)and96hours(morula,blastocyst)of culture. We also measured EC for blastocyst development form 2-cell enbryos inmEBSS supplemented with various concentrations of insulin, And the mouse embryos were stained to express the insulin receptor by immunohistochemical nethod usingan anti-insulin receptor immunoglobulinG.In addition to the immunohistochemical studies for localization of insulim receptor,experiments were undertaken to evaluate whether of not insulin exerts a stimulation of protein synthesis at early stages of mammalian enbryo genesis. The results were as follows: 1. The first cleavage rate of embryos derived from random-bred female mice was appoximately 90% in all experimental groups. In addition, although 40.6%and23.6% of 2-cell embryos developed to morula and blastocyst stage, respectively,in the control group, fewer than10% developed beyond 2-cell stage the hypoxanthine-supplemented medium(p<0.001)but significantly more embryos developed to morula(>80%)and blastocysts(>60%)in the presence of insulin of EDTA (p<0.001). 2. Insulin caused an increase of 20% in blastocyst development in the presence of EDTA(p<0.01)and dffectively reversed the 2-cell block by hypoxanthine(p<0.001). 3. The EC of insulin for induction of blastocyst development form 2-cell embryos was 13.1nM in the presence of0.45 bovine serum albumin. 4. Insulin receptor was expressed by immunohistochemical staining using an antiinsulin receptor immunoglobulin G at the blastocyst stage of mouse embryos. 5. Culture with insulin for 4hour caused an increase of13% in protein synthesis but it completely blocked the effect of insulin in both-supplemented medium at the mourla and blastocyst stage(P<0.05 and P<0.01, respectively). From the above results, we conclude that insulin may play a role as an important growth factor in embryogenesis early development of mouse preimplantation embryos by the insulin-receptor mediated interaction