转染人源 Omentin-1、Vaspin 妊娠期糖尿病脂肪细胞胰岛素受体底物和磷脂酰肌醇3激酶表达变化

摘要

目的：观察转染人源网膜素1（Omentin-1）、内脏脂肪组织源性丝氨酸蛋白酶抑制剂（Vaspin）的妊娠期糖尿病（GDM）脂肪细胞胰岛素受体底物1／2（IRS-1／2）、磷脂酰肌醇3激酶（PI3K）表达变化。方法复苏、传代及诱导分化GDM前脂肪细胞。构建Omentin-1、Vaspin过表达载体，以3个不同过表达梯度（1．0、2．5、5．0μg）转染传代脂肪细胞，以无转染组为对照。采用实时荧光定量PCR法检测各组脂肪细胞Omentin-1、Vaspin、IRS-1／2、PI3K mRNA；采用Western blotting法检测各组脂肪细胞Omentin-1、Vaspin、IRS-1／2、PI3K蛋白及酪氨酸磷酸化IRS-1／2；采用[3H]-2-脱氧-D-葡萄糖摄取测定法测算各组脂肪细胞葡萄糖的摄取率。结果随Omentin-1表达增加，转染人源Omentin-1脂肪细胞中IRS-1、PI3K（P85a）mRNA及蛋白表达增加，IRS-2 mRNA及蛋白表达未发生明显变化， IRS-1酪氨酸磷酸化程度明显升高，IRS-2酪氨酸磷酸化程度未发生明显变化，葡萄糖摄取率上升。随Vaspin表达增加，转染人源Vaspin脂肪细胞IRS-1、IRS-2、PI3K（P85a）mRNA及蛋白表达均未出现明显变化，IRS-1、IRS-2酪氨酸磷酸化程度均未出现明显变化，葡萄糖摄取率变化不明显。结论转染人源Omentin-1的GDM脂肪细胞IRS-1和PI3K（P85a）表达升高，葡萄糖摄取率升高；转染人源Vaspin的GDM脂肪细胞无此变化。%Objective To observe the expression changes of insulin receptor substrate-1/2 ( IRS-1/2) and phosphati-dy inositol 3 kinase (PI3K) in gestational diabetes mellitus (GDM) adipocyte cells after anthropogenic Omentin-1, visceral adipose tissue-derived serine protease inhibitor ( Vaspin) transfection.Methods Recoveried, extended and differentiated GDM preadipocyte cells were prepared, and then we built Omentin-1, Vaspin overexpression carrier, and transfected the extended adipose cells with different overexpression levels (1.0, 2.5 and 5.0μg).Meanwhile, no transfection group was taken as the contrast.Using Q-PCR to detect the mRNA expression of Omentin-1, Vaspin, IRS-1/2 and PI3K (P85a), using Western blotting to detect protein expression of Omentin-1, Vaspin, IRS-1/2 and PI3K ( P85a) as well as IRS-1/2 phosphorylation levels, using [3H]-2-deoxidation-D-glucose uptake assay to detect the rates of glucose uptake in different transfection groups.Results With the increasing Omentin-1 expression, the mRNA and protein expression of IRS-1 and PI3K(P85a) were increased, but those of IRS-2 had no significant changes, IRS-1 phosphorylation increased obviously, IRS-2 phosphorylation had no significant changes, and the glucose uptake rate increased significantly.With the increasing Vaspin expression, all of the mRNA, protein and phosphorylation in IRS-1/2 and PI3K(P85a) had no significant changes, so did the glucose uptake rates.Conclusion In GDM adipocyte cells after anthropogenic Omentin-1 transfection, IRS-1, PI3K( P85a) and glucose uptake rates were increased, but no changes in those of GDM adipocyte cells after anthropogenic Vaspin transfection.