高糖环境对肝星状细胞TLR4表达及脂多糖诱导的炎症因子表达的影响

Effect of high glucose on toll-like receptor 4 expression and LPS-induced proinflammatory cytokine production in hepatic stellate cells in vitro

目的研究高糖环境对肝星状细胞Toll样受体4(TLR4)表达及脂多糖(LPS)诱导的炎症因子表达的影响。方法体外培养大鼠肝星状细胞株HSC-T6,采用高糖刺激,Western blot及RT-PCR检测细胞TLR4蛋白及mRNA表达;低糖或高糖培养基预处理肝星状细胞24 h后采用LPS(100 ng/ml)刺激,2 h后Western blot检测NF-κB核转位,24 h后RT-PCR检测MCP-1和IL-6mRNA表达,ELISA检测细胞上清两种炎症因子的浓度。结果高糖环境可上调肝星状细胞TLR4蛋白及mRNA表达(均为P<0.01),且呈明显的时效-量效关系。高糖可促进肝星状细胞NF-κB核转位(P<0.01),并促进MCP-1和IL-6表达及分泌(均为P<0.01)。高糖预处理可增强LPS诱导的肝星状细胞NF-κB核转位(P<0.01)、MCP-1和IL-6表达及分泌(均为P<0.01)。结论高糖可上调肝星状细胞TLR4的表达,并可促进脂多糖激活NF-κB而增加炎症因子表达及分泌。

Objective To investigate the effect of high glucose on the expressions of toll-like receptor 4 （TLR4） and proinflammatory cytokine production induced by lipopolysaccharide （LPS） in hepatic stellate cells in vitro. Methods Hepatic stellate cell line T6 was cultured in vitro and stimulated by high glucose. The mRNA and protein expression of TLR4 were detected by RT-PCR and Western blotting, respectively. After a 24-h pretreatment with high or low glucose, the cells were stimulated with LPS for 2 h, and Western blotting was used to detect the nuclear translocation of nuclear factor-loB （NF-κB）; at 24 h of LPS exposure, the cells were examined for MCP-1 and IL-6 mRNA and protein expression levels with RT-PCR and ELISA, respectively. Results High glucose significantly increased the mRNA and protein expressions of TLR4 （P〈0.01） in a time- and dose-dependent manner. High glucose promoted NF-κB nuclear translocation and significantly enhanced the expression and secretion of both MCP-1 and IL-6 （P〈0.01）. Pretreatment with high glucose significantly promoted LPS-induced NF-κB nuclear translocation （P〈0.01） and the mRNA expression and secretion of MCP-1 and IL-6. Conclusions High glucose can increase TLR4 mRNA and protein expressions in hepatic stellate cells and promote LPS-induced NF-κB activation and production of proinflammatory cytokines.