高糖诱导人THP1细胞Toll样受体4信号通路变化及匹伐他汀的拮抗作用

Effects of high glucose on the expression and signal pathway of Toll-like receptor 4 in THP1 cells and the antagonism of pitavastatin

目的探讨高糖刺激下人单核/巨噬细胞系(Thp1)细胞表面Toll样受体4(TLR4)表达和信号通路活性变化,并探讨匹伐他汀对高糖引起该信号通路表达变化的作用及可能的机制。方法体外培养Thp1细胞,分为对照组(0mmol·L^(-1)葡萄糖)、低葡萄糖浓度组(5.5mmol·L^(-1)葡萄糖)、高葡萄糖浓度组(15、25 mmol·L^(-1)葡萄糖)、高糖(15mmol·L^(-1)葡萄糖)+匹伐他汀组(0.01、0.1、1μmol·L^(-1)匹伐他汀),各组细胞干预24 h后,Real-TimePCR方法检测各组细胞TLR4、MCP1、IL6基因表达水平,Western印迹法检测各组细胞TLR4、Myd88蛋白表达水平。结果随着葡萄糖浓度从5.5mmol·L^(-1)升高到25 mmol·L^(-1),Thp1细胞TLR4、MCP1、IL6基因表达及TLR4、Myd88蛋白水平逐渐增加。Thp1细胞在高糖刺激前给予匹伐他汀预孵1 h,TLR4、MCP1、IL6基因水平及TLR4、Myd88蛋白水平随匹伐他汀浓度的增加逐渐下降。结论高糖能剌激人Thp1细胞TLR4信号通路激活,呈剂量依赖性诱导TLR4、IL6、MCP1表达升高,且高糖激活的信号通路可能为Myd88依赖途径;匹伐他汀可以拮抗高糖诱导的TLR4信号通路激活,抑制炎症反应的放大。

Objective To investigate the effect of high glucose on the expression and signal pathway of Toll-like receptor 4 （TLR4） in Thpl cells and the antagonism of pitavastatin. Methods Thpl cells were cultured in vitro and treated with different concentration of glucose（0, 5.5, 15 and 25 mmol· L^-1）, or high glucose（15 rnmol · L^-1） ＋pitavastatin（0. 01,0. 1 and 1 laxnol · L^-1） for 24 h, respectively. Real-time PCR was used to detect the expression of TLR4, MCP1 and IL6 genes, Western blot was used to detect the protein levels of TLR4 and Myd88. Results TLR4, MCP1 and IL6 gene expression and the protein levels of TLR4, Myd88 gradually increased with the increase of glucose concentration. Afterintervention of pitavastatin, TLR4, MCP1 and IL6 gene expression and TLR4, Myd88 protein levels were decreased with the increase of pitavastatin concentration. Conclusion High glucose can activate the signaling pathway of TLR4 in Thpl celis and increase the expression of TLR4, IL6 and MCP1 genes in a dose-depending manner; and this activation might be through Myd88-dependent pathway. Pitavastatin can antagonize the activation of TLR4 signaling pathway induced by high glucose.