Effect Of Insulin And Glucose On Premi Plantation Development Of ICR Mouse In Vitro

【Objective】:Try tostudy the effects and roles of insulin and glucose on the development of ICR mouse embryos in virto.To provide the thertical and experimental bases for the optimization of the cutlture medium of embryos.【Methods】:Experiment 1:To observe the effects of insulin and glucose on the development of one-cell mouse embryos in virto.Embryos were divided into four groups:Control:group 1,embryos were cultured in CZB medium with no inlusin and glucose;Treatments:group2,3,4,group 2,embryos were cultured in CZB medium with 0.05μg/ml insulin,group 3,embryos were cultured in CZB medium with 3 mmol/L glucose,group 4,embryos were cultured in CZB medium with 0.05μg/ml insulin and 3 mmol/L glucose respectively.The embryos were cultured up to 120h at 37℃under 5% CO₂ in a sealed culture chamber and observed every 24h under a Nikon inverted microscope.The culture efficiency was evaluated by determining the proportion of embryos reaching two-cell,four-cell,morula,blastocyst,hatched blastocyst and counting its total cells in the blastocysts and hatched blastocysts stage.Experiment 2:To observe the effects of different concentrations of insulin and glucose on the development of one-cell mouse embryos in virto.This experiment were finished by two steps:The first step,embryos were divided into eight groups:Control:group 1,embryos were cultured in CZB medium with no inlusin and glucose;The left seven groups are treatments:group 2,embryos were cultured in CZB medium with 0.05μg/ml insulin,group 3,4,5,6,7,8,embryos were cultured in CZB medium with 0.05μg/ml insulin and 0.2,0.5,1,3,5,or 10 mmol/L glucose respectively.The second step,embryos were divided into nine groups:Control:group 1,embryos were cultured in CZB medium with no inlusin and glucose;The left eight groups are treatments:group 2,embryos were cultured in CZB medium with 0.5μg/ml insulin,group 3,4,5,6,7,8,embryos were cultured in CZB medium with 0.5μg/ml insulin and 0.2,0.5,1,3,5,or 10 mmol/L glucose respectively,group 9 is last step's group 7.Similarly,measuring the percentages of two-cell,four-cell,morula,blastocyst,hatched blastocyst and counting the total cell counts in the blastocysts and hatched blastocysts stage.【Results】:Experiment 1:There were no significant differences in the proportions of two-cell,four-cell and morula embryos between the treatments and the control(P＞0.05).Addition of glucose or glucose and insulin significantly increased blastocyst and hatched blastocyst development(P＜0.05).The all treatments significantly increased the total cell counts as compared with controls(P＜0.05).Culture with glucose and insulin significantly increased the proportions of hatched blastocyst and the total cell number as compared with others groups(P＜0.05).Experiment 2:The first step:There were no significant differences in the proportions of two-cell,four-cell and morula embryos between the treatments and the control(P＞0.05).Except group 2,other treatments significantly increased blastocyst and hatched blastocyst development(P＜0.05).The all treatments significantly increased the total cell counts as compared with controls(P＜0.05).Addition of 0.05μg/ml insulin and 5 mmol/L glucose significantly increased the proportions of hatched blastocyst and the total cell number as compared with others groups(P＜0.05);The second step:The first step:There were no significant differences in the proportions of two-cell,four-cell and morula embryos between the treatments and the control(P＞0.05).Except group 2,other treatments significantly increased blastocyst and hatched blastocyst development(P＜0.05).The all treatments significantly increased the total cell counts as compared with controls(P＜0.05).Group 6 and group 9 significantly increased hatched blastocyst development(P＜0.05),however culture with 0.05μg/ml insulin and 5 mmol/L glucose significantly increased total cell number as compared with others groups(P＜0.05). 【Conclusion】:(1) Insulin and glucose at the same time is presented at the same time in CZB medium could promote the development in vitro of the ICR embryos.(2).Addition of 0.05 insulin and 5 glucose in CZB medium are better than other group in embryos culture in vitro.