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目的:探讨表没食子儿茶素-3-没食子酸酯(EGCG)对细菌脂多糖(LPS)诱导的人单核细胞株THP-1 Toll样受体4(TLR4)及下游信号转导通路的干预作用。方法:采用不同浓度的EGCG和LPS处理THP-1细胞,MTT检测细胞增殖能力;实时荧光定量RT-PCR(qRT-PCR)检测细胞TLR4和肿瘤坏死因子α(TNF-α)mRNA的表达;Western blotting检测TLR4以及下游信号分子胞外信号调节激酶(ERK)、c-Jun氨基末端激酶(JNK)、p38的蛋白水平变化;ELISA检测细胞培养上清TNF-α蛋白含量。结果:EGCG(20 mg·L-1)能降低LPS刺激的THP-1细胞TLR4(蛋白和mRNA)的表达(P<0.05);EGCG(5,10,20 mg·L-1)能抑制TLR4下游分子ERK,JNK,p38的磷酸化,并呈现一定的浓度依赖性(P<0.05);同时,EGCG(20 mg·L-1)能干预LPS刺激的细胞TNF-α(蛋白和mRNA)的表达(P<0.05)。结论:EGCG可通过干预LPS刺激的TLR4表达及下游分子ERK,JNK,p38磷酸化,抑制细胞TNF-α表达,这可能是EGCG的抗炎机制之一。
AIM: To investigate the effects of epigallocatechin-3-gallate (EGCG) on human monocytic THP-1 Toll-like receptor 4 (TLR4) and its downstream signal transduction pathway induced by lipopolysaccharide (LPS) effect. Methods: The THP-1 cells were treated with different concentrations of EGCG and LPS, and the proliferation of THP-1 cells was detected by MTT assay. The expressions of TLR4 and TNF-α mRNA were detected by real-time quantitative RT-PCR (qRT- The protein levels of TLR4, ERK, JNK and p38 were detected by Western blotting. The contents of TNF-α in cell culture supernatants were detected by ELISA. Results: EGCG (20 mg · L-1) decreased the expression of TLR4 (protein and mRNA) in THP-1 cells stimulated by LPS (5, 10 and 20 mg · L-1) (P <0.05); At the same time, EGCG (20 mg · L-1) could interfere with the LPS-stimulated cell TNF-α (protein and mRNA) phosphorylation of ERK, JNK and p38 in a concentration-dependent manner Expression (P <0.05). CONCLUSION: EGCG can inhibit the expression of TLR4 and down-regulate the phosphorylation of ERK, JNK and p38 in LPS-stimulated cells, which may be one of the anti-inflammatory mechanisms of EGCG.