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目的观察五灵胶囊(Wuling capsules,WL)中部分化学成分对脂多糖(lipopolysaccharider,LPS)诱导大鼠原代枯否细胞(Kupffer cell,KC)表达ERK(extracellular signal-regulated kinase)和核因子(nuclear factor-kappa B,NF-κB)调节炎症因子和介质的作用。方法分离大鼠KC,采用60μg L 1 LPS诱导KC分泌炎症因子及NOS,ELISA测定TNF-α、IL-6、IL-8,比色法测定NOS,Western blot检测全蛋白ERK、p-ERK、NF-κBp50、NF-κBp65、p-NF-κBp65、p-IκK、p-IκB及核、浆蛋白NF-κBp65、p-NF-κBp65的变化。结果 WL明显下调LPS活化KC表达p-ERK、p-NF-κBp65、p-IκK、p-IκB蛋白,模拟WL混合成分(Mix)及6单体成分明显下调LPS活化KC表达p-ERK和p-NF-κBp65信号通路蛋白。各受试药物组均能降低核内p-NF-κBp65表达,减少p-NF-κBp65入核抑制炎性基因转录,降低活化KC过量分泌TNF-α、IL-6、IL-8和NOS。结论 WL中五味子醇甲、五味子乙素、隐丹参酮、丹参酮ⅡA、柴胡皂苷D是干预LPS诱导KC表达ERK、NF-κB信号通路蛋白,抑制炎性因子与介质基因转录而减少TNF-α、IL-6、IL-8和NOS分泌的有效成分。
Objective To observe the effect of partial chemical constituents of Wuling capsules (WL) on the expression of extracellular signal-regulated kinase (ERK) and nuclear factor (Kp) in primary Kupffer cells (KC) induced by lipopolysaccharide nuclear factor-kappa B, NF-κB) regulates inflammatory cytokines and mediators. Methods The KCs secreted by KC were induced by 60μg L 1 LPS, TNF-α, IL-6 and IL-8 were detected by ELISA, NOS was detected by ELISA, Western blot was used to detect ERK and p-ERK, The changes of NF-κBp50, NF-κBp65, p-NF-κBp65, p-IκK, p-IκB and nuclear, plasma protein NF-κBp65 and p-NF- Results WL significantly down-regulated the expressions of p-ERK, p-NF-κBp65, p-IκK and p-IκB proteins in LPS-activated KCs, and mimic WL mixed components and 6 monomer components downregulated p-ERK and p NF-κBp65 signaling pathway protein. Each test drug group could reduce the expression of p-NF-κBp65 in the nucleus, reduce the transcription of p-NF-κBp65, inhibit the transcription of inflammatory genes, and decrease the overproduction of TNF-α, IL-6, IL-8 and NOS by activated KC. Conclusion Schizandrin A, Schisandrin B, cryptotanshinone, tanshinone Ⅱ A and saikosaponin D in WL can reduce the expression of TNF-α and TNF-α in KCs by inducing the expression of ERK and NF-κB signaling pathways in KCs induced by LPS, IL-6, IL-8 and NOS secretion of the active ingredient.