连翘对LPS作用的巨噬细胞TLR4蛋白表达的影响

Effects of forsythia suspense on TLR4 protein in macrophage induced by LPS

目的观察连翘对LPS诱导巨噬细胞(RAW264.7)16h后TLR4蛋白表达的影响,探讨连翘对抗内毒素作用及其可能机制。方法体外实验共分6组,分别为control组,LPS组,连翘高、中、低剂量组及多黏菌素B组;连翘高、中、低剂量组及多黏菌素B组分别加入连翘水煎液50mg/ml、25mg/ml、12.5mg/ml及多黏菌素B 10ug/ml培养0.5h后,再加入10ng/ml LPS,培养16h;Western blot法测各组细胞TLR4蛋白表达变化。结果control组RAW264.7细胞只表达少量TLR4,给予LPS刺激16h,TLR4表达明显升高(P<0.01),连翘高、中、低剂量组预处理均可明显降低TLR4的表达(P<0.01),并呈剂量依赖关系;连翘高剂量组与多黏菌素B组比较无显著性差异(P>0.05)。结论连翘预处理能抑制LPS诱导的巨噬细胞TLR4的表达可能是其抗内毒素作用的重要机制。

Objective To observe the effects of Forsythia suspense the expression of TLR4 in macrophage RAW264.7 induced by LPS and to explore the anti-endotoxin and immunity modulation function of AFS and its possible mechanisms . Methods Cultured RAW264.7 cells have been randomly divided into six groups：control group： without any treatment;LPS group：adding LPS at final concentration of 10ng/ml for 16h;high concentration of Forsythia suspense, median concentration of Forsythia suspense, low concentration of Forsythia suspense group： 30mins preconditioning of Forsythia suspense （50mg/ml, 25mg/ml, 12. 5mg/ml in final concentration） before adding LPS;polymyxin B group：30mins preconditioning of Ploy B （10ug/ml in final concentration） before adding LPS. The expression of TLR4 protein on RAW264.7 were measured by western blot. Results It was found that TLR4 protein had a basic expression in RAW264.7 cells under normal condition;and it was increased significantly after LPS acted on RAW264.7 cells than that of control group at 16h（P〈0.01） ;adding Forsythia suspense before the presence of LPS, the expressions of TLR4 protein in high concentration of Forsythia suspense,median concentration of Forsythia suspense,low concentration of Forsythia suspense group decreased remarkably than that of LPS group（ P〈0.01）, which was dose-dependent. There was no significant difference between high concentration of Forsythia suspense group and polymyxin B group（P〉0.05）. Conclusion It was verified that Forsythia suspensa had the effect of anti-endotoxin probably by inhabiting the up-regulation of TLR4 protein in macrophage induced by LPS.