高浓度葡萄糖对胰岛β细胞株NIT-1细胞Irs2、FoxO1表达的影响

摘要

目的:探讨高浓度葡萄糖对胰岛β细胞胰岛素信号通路信号中胰岛素受体底物2(insulin reoeptor substance 2,Irs2)、叉头框转录因子1 (forkhead transcription factors O1,FoxO1)表达的影响.方法:胰岛β细胞株NIT-1细胞培养48 h后,随机分为不同葡萄糖浓度(5.6,11.1,16.7,22.5,27.6 mmol/L)组,继续培养3 d后终止,用MTT比色法检测细胞增殖,免疫荧光法检测细胞凋亡,放射免疫分析方法检测胰岛素水平,Real-Time PCR法检测Irs2、FoxO1 mRNA变化.结果:(1)在葡萄糖浓度为11.1 mmol/L时,NIT-1细胞增殖活性最强、细胞凋亡率最低,胰岛素分泌水平最高.随着葡萄糖浓度升高,细胞增殖活性和胰岛素分泌水平明显下降,细胞凋亡率逐渐增高.当葡萄糖浓度达27.6 mmol/L 时,细胞增殖活性和胰岛素分泌水平达最低,细胞凋亡率达到最高.(2)葡萄糖浓度为11.1 mmol/L时,Irs2 mRNA的转录达到最高水平,随着葡萄糖浓度的增高,Irs2 mRNA的转录水平呈现逐渐减少趋势;FoxO1基因的mRNA转录水平则随着葡萄糖浓度的增高呈不断增加状态.结论:在高浓度葡萄糖条件下,葡萄糖"毒性"作用通过下调Irs2 mRNA表达、增加核内的FoxO1 mRNA表达以调节胰岛β细胞的功能及增殖和凋亡,分子间均有其独立的调控机制,同时又相互关联.%Objective:To evaluate the effects of high concentration of glucose on insulin receptor substance 2 (Irs2) and forkhead transcription factors O1 (FoxO1) expression of pancreatic β cells. Methods: Pancreatic (3 cell lines NIT-1 cells were cultured 48 hours, then were randomly divided into 1 to 5 groups, were cultured with 10% FBS DMEM medium containing glucose of 5. 6 , 11. 1 , 16. 7, 22. 5 and 27. 6 mmol/L for 3 days. The expression of Irs2 and FoxO1 mRNA in the cells was analyzed using the real-time PCR. Cell proliferation was determined by MTT, cell apoptosis was observed by immunofluorescence staining, and insulin secretion was detected by radioimmunoassay. Results: Cell proliferation and insulin secretory function in mouse NIT-1 cells were the best at the concentration of 11. 1 mmol/L glucose, and cell apoptosis was the worst. With the increase of glucose concentration, cell proliferation and insulin secretion reduced obviously, the cell apoptosis increased gradually. Under the 27. 6 mmol/L glucose concentration, the proliferation rate and insulin secretion level were the lowest, the apoptosis rate was the best. Under the 11.1 mmol/L glucose concentration, Irs2 mRNA expression level was the best, with the increase of glucose concentration, it was decreased, but FoxOl mRNA expression level was increased. Conclusion: With high concentration of glucose, glucose toxicity can re-duce Irs2 mRNA expression, increase FoxO1 mRNA expression of nuclear, regulate functional and survival of pancreatic βcells. They have independent molecular regulatory mechanism but relate each other.