3T3L1脂肪细胞对大鼠胰岛β细胞功能障碍的影响

目的 通过3T3L1脂肪细胞与大鼠原代胰岛细胞共培养,从细胞整体水平探讨脂肪细胞对胰岛β细胞功能的影响.方法 研究分为两组:(1)对照组:SD大鼠原代胰岛细胞组;(2)共培养组:SD大鼠原代胰岛细胞与3T3L1脂肪细胞共培养.对各组胰岛细胞观察以下指标:(1)胰岛素释放试验和细胞内胰岛素含量;(2)用RT-PCR检测葡萄糖转体2(GLUT2)、葡萄糖激酶(GCK)及内向整流钾离子通道6.2(Kit6.2)的mRNA表达水平;(3)用免疫沉淀和Western印迹检测胰岛素受体β(IR-β)、胰岛素受体底物-1(IRS-1)蛋白及其酪氨酸磷酸化程度.结果 (1)共培养组在低糖水平胰岛素分泌明显增多(0.79±0.35)ng·h-1·ml-1胰岛比(0.38±0.09)ng·h-1·ml-1胰岛,P=0.028],而高糖刺激时两组胰岛素分泌差异无统计学意义(P=0.760),共培养组刺激指数较对照组降低(1.57±0.61)比(2.84±0.92),P=0.04].两组的胞内胰岛素含量差异无统计学意义(P=0.102).(2)共培养组胰岛细胞GLUT2、GCK、Kil6.2的mRNA表达下调为0.27 ±0.11,(P=0.01)、0.32±0.24,(P=0.009)、0.41±0.09,(P=0.003).(3)共培养组胰岛细胞IR-β、IRS-1蛋白表达及其酪氨酸磷酸化程度均下降.结论 3T3L1脂肪细胞通过下调胰岛细胞葡萄糖刺激的胰岛素分泌(GSIS)的相关基因,及通过抑制其自身胰岛素信号通路,从而影响GSIS.脂肪细胞可能参与了2型糖尿病胰岛细胞功能障碍的发生.

3T3L1 adipocytes induce rat beta-cell dysfunction

Objective To investigate the integrated effects of adipocytes on rat beta-cells, differentiated 3T3L1 adipocytes and rat islet cells co-culture system was established. Methods There were two groups; control group ( SD rat islet cells) and co-culture group (islet cells and 3T3L1 adipocytes co-culture system). Islet cells were obtained for determination of (1) insulin secretion and insulin content; (2) mRNA expressions of GLUT2, GCK and Kir6. 2; (3) protein expressions of IR-β, IRS-1 and their tyrosine phosphorylation leveL Results (1) At low glucose, insulin secretion of co-culture group increased compared with that of control group (0.79±0.35) ng·h-1 · ml-1 islet vs (0.38±0.09 ) ng · h-1 · ml-1 · islet, P = 0. 028 . At high glucose, insulin secretion of those two groups was almost at the same level (P =0.760). Compared with control group (2. 84 ±0.92) , stimulation index (SI, insulin release at high glucose/ low glucose) of co-culture system decreased to (1. 57 ±0. 61, P =0.04). And the insulin content of the both groups was almost at the same level (P=0.102). (2) The mRNA of GCK, GLUT2 and Kir6. 2 in co-culture group downregulated to (0. 27 ±0. 11, P = 0. 01) , (0. 34 ±0. 24, P = 0. 009) and (0.41 ± 0. 09, P = 0. 003) compared with control group (mRNA = 1). ( 3) The protein levels of IR-β, IRS-1 and their tyrosine phosphorylation decreased in co-culture system. Conclusions 3T3L1 adipocytes are involved in beta-cell dysfunction, which may facilitate the development of type 2 diabetes. The effects may be mediated by multiple pathways, which include downregulation of GSIS related gene expressions and suppression of islet cell insulin signaling.