脂肪因子Omentin-1、Visfatin体外表达与老年2型糖尿病胰岛素抵抗的关系

Effect of Omentin-1 and Visfatin over expression in vitro on insulin signal transduction pathway of type 2 diabetes

目的探讨脂肪因子Omentin-1、Visfatin与老年2型糖尿病(T2DM)胰岛素抵抗(IR)的内在联系。方法复苏、传代及诱导分化老年人源T2DM前脂肪细胞,构建Omentin-1、Visfatin过表达载体,进行载体转化、培养和提取;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;Q-PCR检测Omentin-1、Visfatin、胰岛素受体底物(IRS)-1/2、磷脂酰肌醇-3激酶[PI3K(P85a)]mRNA表达,Western印迹检测Omentin-1、Visfatin、IRS-1、IRS-2、PI3K(P85a)蛋白表达及IRS-1/2酪氨酸磷酸化水平,[~3H]-2-脱氧-D-葡萄糖摄取测定细胞葡萄糖摄取率的变化。结果 (1)Omentin-1、Visfatin mRNA及蛋白表达随转染浓度梯度升高而表达量增加,所构建的Omentin-1、Visfatin过表达载体有效;(2)随Omentin-1表达增加,IRS-1及PI3K(P85a)mRNA和蛋白表达及IRS-1磷酸化程度均出现明显增加,IRS-2未出现明显变化;(3)随Visfatin表达增加,IRS-1/2、PI3K(P85a)mRNA、蛋白表达及IRS-1/2磷酸化程度未出现明显变化;(4)脂肪细胞的葡萄糖摄取率随Omentin-1表达增加而升高,而与Visfatin过表达浓度变化无明显关系。结论 Omentin-1表达可能通过某种途径激活了胰岛素受体后信号传导通路,导致了IRS-1和PI3K活化和表达,从而发挥其胰岛素增敏作用;Visfatin在老年T2DM血清中的高表达,很可能只是一种肥胖、T2DM糖脂代谢紊乱的补偿机制,与T2DM IR的发生没有必然联系。

Objective To investigate the relationship between Omentin-1, Visfatin and insulin resistance in type 2 diabetes （T2DM） from the classic insulin signaling pathways （PI3K pathway）.Methods T2DM preadipocyte cells were recoveried, extended and differentiated to build Omentin-1, Visfatin over expression carrier, and then were transformated, cultivated and the plasmids were extracted.Adipocytes were transfected with 3 different over expression levels （1.0 μg, 2.5 μg ,5.0 μg） and extended, with no transfection group was selected as control group.The expressions of Omentin-1, Visfatin, insulin receptor substrate （IRS）-1/2, phosphatidy inositol 3 kinase [PI3K （P85a）]mRNA were examined by Q-PCR, protein expression of Omentin-1,Visfatin, IRS-1, IRS-2, PI3K （P85a） and IRS-1/2 phosphorylation levels were detected by Western blot, glucose uptake rates in different concentration of plasmid transfection was detected with [3H]-2-deoxidation-D-glucose uptake assay.Results Omentin-1 and Visfatin mRNA and protein expression were increased with increasing of concentration of plasmid transfection, which indicated that the constructed over expression carrier of Omentin-1 and Visfatin were effective.With the Omentin-1 expression increasing, the mRNA and protein expression of IRS-1 and PI3K （P85a） increased obviously, so did the IRS-1 phosphorylation, but IRS-2 had no obvious changes.With the Visfatin expression increasing, all of the mRNA, protein and phosphorylation in IRS-1/2 and PI3K （P85a） had no obvious changes.Glucose uptake rate of adipocytes elevated with the increasing expression of Omentin-1 and had no obvious changes with Visfatin increasing expression.Conclusions The expression of Omentin-1 might activate signaling pathways of IRS-1 and PI3K （P85） by some way and lead to the excitation and expression of IRS-1 and PI3K indirectly, which plays a role of insulin sensitization.The high expression of Visfatin in T2DM serum may be probably a compensation mechanism of obesity and sugar lipid metabolic disorder of T2DM, there is no connection with the occurrence of insulin resistance.