摘要
目的:观察表儿茶素对脂多糖(LPS)诱导小鼠单核巨噬细胞RAW264.7分泌炎症因子的影响,并探讨其作用机制。方法:用脂多糖(1 mg·L-1)刺激生长良好的RAW264.7细胞24 h建立体外细胞炎症模型,以噻唑蓝(MTT)比色法测定不同浓度表儿茶素对RAW 264.7细胞的毒性作用,Griess试剂法检测一氧化氮(NO)含量,酶联免疫吸附测定法(ELISA)检测细胞上清液中肿瘤坏死因子-α(TNF-α),白细胞介素-1(IL-1)和白细胞介素-6(IL-6)含量,蛋白免疫印迹法(Western blot)检测细胞一氧化氮合酶(i NOS)及丝裂原活化蛋白激酶(mitogen-activated protein kinases,MAPKs)磷酸化表达。结果:表儿茶素在100μmol·L-1时对RAW 264.7细胞无毒性作用。与空白组比较,LPS可明显诱导RAW264.7细胞分泌炎症因子NO,TNF-α,IL-1和IL-6(P<0.01);与LPS组比较,25~100μmol·L-1的表儿茶素可以明显降低LPS诱导的RAW 264.7细胞释放炎症因子NO,TNF-α,IL-1和IL-6(P<0.05,P<0.01),抑制i NOS蛋白及MAPKs亚族p38,细胞外信号调节激酶(extracellular signal regulated kinases,ERK)和c-jun氨基末端激酶(c-jun terminal kinase,JNK)蛋白磷酸化水平表达,并呈现浓度依赖关系。结论:表儿茶素可以抑制LPS诱导的RAW264.7细胞炎症反应,其抗炎作用可能与减少炎症因子NO,TNF-α,IL-1和IL-6,抑制i NOS蛋白表达及p38MAPK,ERK1/2和JNK的磷酸化水平有关。
Objective： To study the effects of epicatechin on inflammatory cytokines in lipopolysaccharide（ LPS）-induced macrophage model and its mechanism. Method： The in vitro inflammation model was established by stimulating RAW 264. 7 cells with LPS（ 1 mg·L^-1） for 24 h. The toxic effect of macrophages were detected by 3-（ 4,5-Dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide（ MTT） assay. The content of nitricoxide（ NO） was assayed by Griess reagent. Enzyme linked immunosorbent assay（ ELISA） were used to assay the content of inflammatory mediators,such as tumor necrosis factor-α（ TNF-α）,interleukin^-1（ IL^-1） and interleukin-6（ IL-6） in cell supernatant. The protein expressions of nitric oxide synthase（ iNOS） and the related proteins of mitogen-activated protein kinase（ MAPK） pathway were tested by Western blot. Result： The cell viability was not significantly affected by epicatechin at 100 μmol·L^-1. Compared with control group,LPS could significantly induce RAW 264. 7 cells to secrete inflammatory mediators,like NO,TNF-α,IL^-1 and IL-6（ P〈0. 01）. Compared with model group,25-100 μmol·L^-1of epicatechin in LPS-induced RAW 264. 7 cells greatly inhibited the release of inflammatory mediators,such as NO,TNF-α,IL^-1 and IL-6（ P〈0. 05,P〈0. 01）,and inhibit the expressions of iNOS,and the protein expression of p38 MAPK,ERK1 /2 and JNK in a dose dependent manner. Conclusion： Epicatechin can inhibit LPS-induced inflammatory response in RAW 264. 7 cells,and its anti-inflammatory effect may be related to reduction of inflammatory cytokines,like NO,TNF-α,IL^-1 and IL-6,inhibition of the gene expression of iNOS and phosphorylation of p38 MAPK,ERK1 /2 and JNK.
出处
《中国实验方剂学杂志》
CAS
CSCD
北大核心
2017年第4期159-163,共5页
Chinese Journal of Experimental Traditional Medical Formulae
基金
黑龙江省教育厅科学技术研究项目(12511369)
黑龙江八一农垦大学学成
引进人才科研启动计划课题项目(XDB2014-02)
