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KMID : 0350519920450020597
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Journal of Catholic Medical College
1992 Volume.45 No. 2 p.597 ~ p.607
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Effect of Insulin on Cleavage and Protein Synthesis in Mouse Preimplantation Embryos
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Abstract
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Although insulin is recognized as an important fetal growth factor, its interaction with embryonic cells during the early develpmental stages prior to the appearance of a functional pancreas has not been defined.
But there is evidence that insulin binding to the cells of the preimplantation mouse embrys can be detected first at the morula stage, while autoradiographic studies have confirmed that insulin binding is recoptor mediated. Other studies 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 salt solution(mEBSS), a chemically defined
simple
medium, supplemented with porcine insulin, hypoxanthine and/or ethylenediaminetetraacetic acid(EDTA), and the embryo development was scored at 24 hours(2-cell) and 96 hours(morula, blastocyst) of culture, We also measured EC50 for blastocyst
develpment
from 2-cell embryos in mEBSS supplemented with various concentrations for blastocyst development from 2-cell embryos in mEBSS supplemented with various concentrations of insulin. And the mouse embryos were stained to express the insulin receptor
by
immunohistochemical method using an anti-insulin receptor immunoglobulin G. In addition to the immunohistochemical studies for localization of insulin receptor, receptor, experiments were undertaken to evaluate whether or not insulin exerts a
stimulation of protein synthesis at early stages of mammalian embryogenesis.
@ES The results were as follows:
@EN 1. The first clevage rate of embryos derived from random-bred female mice was approximately90% in all experimental groups. In addition, although 40.6% and 23.6% of 2-cellembryos developed to morula and blastocyst stage, respectively, in the
control
group, fewer than 10% developed beyond2-cell stage in the hypoxanthine-supplemented medium (P<0.001) but significantly more embryos developed to morulae(>80%) and blastocysts(>60%) in the presence of insulin or EDTA(P<0.001).
2. Insulin caused an increase of 20% in blastocyst develpment in the presence of EDTA (P<0.01) and effectively reversed the "2-cell block" by hypoxanthine(P<0.001).
3. The EC50 of insulin for induction of blastocyst develpment from 2-cell embryos was 13.1 nM in the presence of 0.4% bovine serum albumin.
4. Insulin receptor was expressed by immunohistochemical staining using an anti-insulin receptor immunoglobulin G at the blastocyst stage of mouse embryos.
5. Culture with insulin for 4 hour caused an increase of 13% in protein synthesis by 8-cell embryo(P>0.05), 64% by morula(P<0.05), and 99% by blastocyst(P<0.01) respectively compared with control.
6. Addition of anti-insulin antibody to control had no effect on protein synthesis but in completely blocked the effect of insulin in both su-pplemented medium at the morula 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 and early develpment of mouse preimplantation embryos by the insulin-receptor mediated interaction.
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KEYWORD
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