茶多酚对高糖时内皮细胞产生NO、NOS和ICAM-1的影响

The effect of tea polyphenols on the production of NO,NOS and ICAM-1 by endothelial cells under high glucose

目的:研究在高糖环境下人脐静脉内皮细胞(ECV304)产生一氧化氮(NO)量及其合酶和表达细胞间黏附分子1(intercellular adhesion molecule 1,ICAM鄄1)的变化,以及茶多酚(tea polyphenols,TPs)的干预作用。方法:培养ECV304,加入不同培养液,分为正常对照组、高糖组、茶多酚组、茶多酚干预组,以硝酸还原酶法测定培养液中NO的代谢产物NO2/NO3鄄(NOX-)含量,用分光光度法检测培养液中NOS活性,用逆转录鄄多聚酶链反应(RT鄄PCR)和细胞酶联免疫吸附法(ELISA)观察ICAM鄄1的基因和细胞表面蛋白的表达。结果:高糖组NO含量、NOS活性、ICAM鄄1的基因和蛋白表达较正常对照组显著增加,且ICAM鄄1的蛋白表达呈现一定的时间依赖性;茶多酚干预组显著抑制高糖时NO含量、NOS活性以及ICAM鄄1的基因和蛋白表达的上调。结论:高糖时内皮细胞NOS的激活、NO的大量释放以及ICAM鄄1mRNA和蛋白的表达上调,对糖尿病早期进展起一定作用,茶多酚对这种损伤具有明显的保护作用。

Objective: To investigate the effect of high glucose on the production of NO, NOS and ICAM-1 in human umbilical vein endothelial cells (ECV304), and to further study the intervention of tea polyphenols (TPs). Methods: ECV304 was cultured in DMEM medium with or without high glucose. The experiment was divided into four groups randomly: control group, high glucose group, TPs group and TPs treatment group. Metabolic production of NO, NO2-/NO3-(NOX-) and the activity of NOS in supernatant were detected by HNO3 reductase and spectrophotometry assay respectively. Meanwhile ICAM-1 mRNA expression in cells were measured by RT-PCR and protein levels on cell membrane was determined by ELISA. Results: The content of NO, the activity of NOS, ICAM-1 mRNA expression and protein level were found to be significantly increased in high glucose group compared with control group(P < 0.01), and ICAM-1 protein expression was time-dependent; compared with high glucose group, the content of NO, the activity of NOS, ICAM-1 mRNA expression and protein level were found to be markedly decreased in TPs treatment group(P < 0.01). Conclusion: Results suggest that the activation of NOS and NO release , the up-regulation of ICAM-1 mRNA and protein expression in endothelial cells under high glucose play an important role in the early phase of diabetes mellitus and TPs has a significantly protective effect.