摘要
目的研究紫草素抑制由脂多糖(LPS)诱导的巨噬细胞炎症的机制。方法以淀粉培养基注射BALB/c小鼠腹腔,诱导并分离巨噬细胞,以APC标记F4/80染色,并用流式细胞仪检测分离巨噬细胞情况;用1 mg·L-1的LPS刺激上述分离的巨噬细胞,分组如下:DMSO溶剂对照组,LPS对照组,LPS+低剂量紫草素组(0.1μmol·L^(-1));LPS+中剂量紫草素组(1μmol·L^(-1));LPS+高剂量紫草素组(10μmol·L^(-1));通过ELISA和实时定量PCR(qRT-PCR)方法分别测定各处理组培养上清液以及细胞中肿瘤坏死因子-α(TNF-α)、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)的表达情况;Western blot分别测定Toll样受体4(TLR4)和核因子-κB(NF-κB)的p65亚基磷酸化表达情况。结果流式细胞检测可知,APC标记的F4/80染色巨噬细胞的纯度可达97.8%;ELISA和实时定量PCR结果显示,紫草素处理后可以抑制由LPS刺激细胞因子TNF-α、IL-1β和IL-6的表达(P<0.05),并增加IL-10的表达(P<0.05),且呈现出一定的浓度依赖性;Western blot结果显示,紫草素能下调由LPS刺激的TLR4的表达水平和NF-κB的p65亚基磷酸化表达。结论紫草素的抗炎机制可能是通过TLR4介导的信号通路活化NF-κB,抑制IL-1β、IL-6和TNF-α的分泌,并促进IL-10的分泌。
Objective Tostudy the roles of Shikonin inhibits LPS-induced macrophage inflammatory andanti-inflammatory effect ofperitoneal macrophages.Methods The peritoneal macrophages of BALB/c mouse were inducedby injectionintraperitoneal mediumand were stained with F4/80.The groupswere divided as follows:DMS0control group,1mg?L1LPS control group,LPS+0.1jjimol?L1Shikonin group,LPS+1jjimol?L1Shikonin group and LPS+10jjimol?L1Shikonin group.The levels of TNF-a,IL-1p,IL-6and IL-10were detected by ELISA and quantitative real-time PCR method.The expressions of TLR4and phosphorylation of p65(pp65)were measured by Western blot.Results After treatment with shikonin,the expression of T N F-a,IL-lp,IL-6and IL-10stimulated by LPSwere decreased(P<0.05),while the expression of IL-10showed the opposite effect(P<0.05).The expression level of TLR4and pp65in peritoneal macrophages stimulated with LPS were inhibited by Shikonin.Conclusion The anti-inflammatory mechanism of shikonin might be mediated by TLR4/N F-kB signaling pathway to inhibit IL-lp,IL-6and TNF-a and promote the secretion of IL-10.
作者
李叶静
谈弋
LI Yejing;TAN Yi(School of Pharmaceutical Sciences ^Wuhan University, Wuhan, Hubei 430060 , China;Department of Clinical Laboratory, Central Hospital, Enshi Tujia and Miao Autonomous Prefecture, Enshi, Hubei 445000, China)
出处
《安徽医药》
CAS
2017年第8期1384-1387,共4页
Anhui Medical and Pharmaceutical Journal