高糖经内质网应激途径诱导胰岛内皮细胞凋亡

Hyperglycemia induced islet endothelial cell line MS-1 apoptosis through endoplasmic reticulum stress pathway

目的:探讨高糖诱导胰岛微血管内皮细胞株MS-1凋亡及其可能机制。方法:体外培养MS-1细胞,用含有不同浓度葡萄糖(5.6、25.0和33.6 mmol/L)的培养液分组培养12、24 h。流式细胞术分析各组细胞凋亡率变化;四甲基偶氮唑盐(MTT)法检测各组细胞增殖率变化;RT-PCR分析GRP78、CHOP、caspase-3、caspase-12 mRNA表达变化情况;Western blot分析GRP78、CHOP蛋白表达变化情况。结果:与5.6 mmol/L组相比,培养12 h及24 h后,25.0和33.6 mmol/L组MS-1细胞凋亡率显著增加(P<0.05);而细胞增殖率显著下降(P<0.05),且呈浓度和时间依赖性。进一步研究表明,在高糖刺激12 h及24 h后,cas-pase-3、caspase-12 mRNA表达显著上调(P<0.05),CHOP mRNA和蛋白表达水平均显著上调(P<0.05);而GRP78 mRNA和蛋白表达水平在刺激12 h后显著上调,24 h后却显著下降(P<0.05)。结论:高糖可以促进胰岛内皮细胞凋亡增加,并呈浓度和时间依赖性升高,其机制可能与启动胰岛内皮细胞内质网应激有关。

Objective：To investigate the effect of hyperglycemia on the apoptosis and proliferation of islet endothelial cell line MS-1.Methods： MS-1 cells were treated with different concentrations of glucose（5.6,25.0 and 33.6 mmol/L） for different time（12 or 24 h）.The apoptotic cells were detected by flow cytometry,and the cell proliferation was tested using 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide（MTT） assay.The expressions of GRP78,CHOP,caspase-3,caspase-12 mRNA were examined by reverse transcription and polymerase chain reaction（RT-PCR）,and the expressions of GRP78,CHOP protein were examined by Western blot.Results： After treated with glucose for 12 h or 24 h,the apoptotic rates were obviously higher in 25.0 and 33.6 mmol/L group than in 5.6 mmol/L group（P 0.05）,while the cell proliferation was significantly lower in 25.0 and 33.6 mmol/L glucose treated group than that in 5.6 mmol/L glucose treated group（P 0.05）.The mRNA levels of CHOP,caspase-3,caspase-12 and the protein level of CHOP increased in a dose-dependent manner after treated with glucose（P 0.05）;while the mRNA and protein levels of GRP78 increased after treated for 12 h,and decreased at 24 h（P 0.05）.Conclusion： Hyperglycemia significantly promotes apoptosis and decreases proliferation of MS-1 cells,the mechanism may account for endoplasmic reticulum stress.