摘要
目的探讨泽泻对脂多糖(Lipopolysaccharides,LPS)诱导RAW264.7细胞炎症反应的影响及作用机制。方法采用MTT法来检测泽泻对RAW264.7细胞的增殖活性的影响。Griess法检测细胞中NO的释放情况。实时定量PCR检测LPS诱导的RAW264.7细胞中环氧合酶-2(COX-2)、诱导型一氧化氮合酶(iNOS)、白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)和白细胞介素-10(IL-10)等的mRNA表达水平,采用Docking法将泽泻中主要有效成分与TLR1蛋白进行分子对接,检测结合能、结合位点、氢键的形成,采用荧光素酶报告基因检测泽泻对TNF-α诱导NF-κB活化的影响。结果泽泻在0~100μg/mL的范围内没有显著的细胞毒性。泽泻能显著降低NO的释放,泽泻能显著下调COX-2、iNOS、IL-1β、IL-6、TNF-α等炎症因子mRNA表达,Docking结果显示泽泻多个主要有效成分能够与TLR1多个位点结合并形成氢键,而对TNF-α诱导NF-κB活化无显著影响。结论泽泻具有抗炎活性,能够抑制LPS诱导的RAW264.7细胞所发生的炎症反应,可能与抑制TLR1有关,且表现出多成分协同作用的特点。
Objective To investigate the effect of Zexie on inflammatory response of RAW264.7 cells induced by lipopolysaccharide(LPS). Methods MTT was used to evaluate the viability of RAW264.7 cells after Zexie incubation. Griess was used to detect the secretion of NO by LPS induced RAW264.7 cells. The expression levels of mRNA of cyclooxygenase-2(COX-2), inducible nitric oxide synthase(iNOS), inter-leukin-1β(IL-1β) and interleukin-6(IL-6) were detected by real-time quantitative PCR(RT-PCR). Molecular docking the main effective components in Zexie with toll-like receptor 1(TLR1) protein and detection of binding energy, binding sites and the formation of hydrogen bonds. Using the luciferase report gene evaluate Zexie on the activity of NF-kappa B induced TNF-α. Results Zexie had no significant cytotoxicity in the range of 0-100 μg/mL, and could cause significant cell damage in the range of 150-400 μg/mL. Griess assay showed that Zexie could significantly reduce NO release, and 25 μg/mL had the most significant effect on reducing LPS-induced NO production. QPCR assay showed that Zexie could significantly down-regulate the expression of COX-2, iNOS, IL-1 beta and IL-6, and was dose-dependent on the down-regulation of iNOS, IL-1β and IL-6. Docking results show that the main effective components in Ze xie can binding with TLR1 in multiple site and form hydrogen bonding, and had no effect on TNF-α induced NF-κB activation. Conclusion Zexie has strong anti-inflammatory activity and can inhibit LPS-induced inflammation in RAW264.7 cells. It may be related to inhibition of TLR1, and it shows a synergistic effect of multiple components.
作者
谢治深
黄小虹
袁永
宋军营
张振强
王辉
徐江雁
XIE Zhi-shen;HUANG Xiao-hong;YUAN Yong;SONG Jun-ying;ZHANG Zhen-qiang;WANG Hui;XU Jiang-yan(Henan University of Chinese Medicine,Zhengzhou Henan 450046 China)
出处
《时珍国医国药》
CAS
CSCD
北大核心
2020年第7期1586-1590,共5页
Lishizhen Medicine and Materia Medica Research
基金
中国博士后基金(2018M642761)
河南省中医药科学研究专项课题(2018ZYD12,2018ZY1009)
河南省高等学校重点科研项目计划(19A520002,19A360021)
关键词
泽泻
炎症
TOLL样受体
脂多糖
Zexie
Inflammatory
RAW264.7 cells
Lipopolysaccharides
