青蒿琥酯对小鼠巨噬细胞RAW264.7中TLR4介导的炎症通路的抑制作用  

The inhibitory effects of artesunate on TLR4-mediated inflammation pathways in RAW264.7 cells

作　　者：匡梅[1] 岑彦艳[1] 覃容欣[1] 潘夕春[1] 周红[1] KUANG Mei;CEN Yanyan;QIN Rongxin;PAN Xichun;ZHOU Hong(Department of Pharmacology, College of Pharmacy, Army Medical University, Chongqing 400038, Chin)

机构地区：[1]第三军医大学药学系药理教研室,重庆400038

出　　处：《免疫学杂志》2018年第5期401-406,共6页Immunological Journal

基　　金：国家自然科学基金(81673495);重庆市基础与前沿重点课题(cstc2015jcyj BX0049);第三军医大学科技成果转化基金(2015XZH04)

摘　　要：目的观察青蒿琥酯(artesunate,AS)对脂多糖(lipopolysaccharide,LPS)刺激小鼠RAW264.7细胞Toll样受体4(Toll-like receptor 4,TLR4)介导炎症通路活化的影响,以探讨AS的抗炎作用机制。方法采用免疫荧光观察胞内TLR4表达和分布;免疫印迹检测TLR4及下游炎症通路关键分子的表达和活化;酶联免疫吸附法检测细胞培养上清中TNF-α、IL-6浓度。结果 AS对LPS诱导的TLR4表达及其在细胞中的聚集均有抑制作用;AS同时抑制TLR4衔接蛋白髓分化因子88和含TIR结构域干扰素诱导衔接蛋白表达,也抑制依赖于二者活化的肿瘤坏死因子受体相关因子6表达;对下游MAPK通路,AS抑制p38表达和磷酸化、JNK磷酸化,但对ERK1/2无显著影响;对下游NF-κB通路,AS下调抑制性-κBα(inhibitory-κBα,IκBα)的磷酸化,进而减少NF-κB亚基p50和p65活化入核的数量;最后,AS能够抑制LPS诱导的TNF-α和IL-6释放。结论 AS通过抑制LPS诱导的TLR4通路活化,减少致炎细胞因子的释放,从而发挥其抗炎作用。This study was performed to investigate the anti-inflammatory mechanism of artesunate（AS） by evaluating the influence of AS on the activation of Toll-like receptor 4（TLR4） signaling of RAW264.7 cells treated with lipopolysaccharide（LPS）. Immunofluorescence was used to observe the intracellular expression and distribution of TLR4; immunoblotting was used to detect the expression levels and activation of proteins involved in TLR4-mediated inflammatory pathways; while ELISA was used to detect the release of TNF-α and IL-6 from RAW264.7 cells. Data showed that AS inhibited both the expression and assembly of TLR4 within cells. AS also inhibited the expressions of adaptors, myeloid differentiation factor 88（My D88） and TIR domain-containing adaptor inducing interferon β（TRIF）, as well as TNF receptor-associated factor 6（TRAF6）. For downstream MAPK pathway, AS could inhibit the expression and phosphorylation of p38, and phosphorylation of JNK, but showed no influence on ERK1/2; For downstream NF-κB pathway, AS inhibited the phosphorylation of inhibitory κBα, thereby reduced the amounts of nuclear NF-κB subunits p50 and p65. Finally, AS significantly inhibited LPS-induced TNF-α and IL-6 release from RAW264.7. Taken together, AS can decrease LPS-induced inflammatory cytokine release from macrophages by inhibiting the activation of TLR4 signaling.

关 键 词：青蒿琥酯 LPS TLR4 炎症因子 

分 类 号：R931.6[医药卫生—生药学]