桑枝总黄酮体外抗炎活性及机制研究

Effects of Total Flavones from Morus alba L. on Inflammatory Reaction of Macrophages

目的研究桑枝总黄酮的体外抗炎活性及其作用机制。方法 IFN-γ和LPS协同造成小鼠RAW264.7细胞炎症模型;Griess反应测定细胞上清液中NO产量;FRAP测定细胞总抗氧化能力;RT-PCR检测iNOS、COX-2、HO-1、IL-1β、IL-6、TNF-α mRNA表达;Western blot检测iNOS、p-ERK蛋白表达水平。结果桑枝总黄酮呈剂量依赖性抑制细胞上清液中NO含量并且无细胞毒性,提高细胞总抗氧化能力,下调iNOS、COX-2、IL-1β、IL-6 mRNA和iNOS、p-ERK蛋白的表达,上调HO-1 mRNA表达,但对TNF-α mRNA表达影响不大。结论桑枝总黄酮部分通过MAPK中的ERK信号转导通路抑制iNOS基因和蛋白的表达从而抑制NO的产量,提高细胞总抗氧化能力,同时下调COX-2、IL-1β、IL-6等炎症介质和上调抗炎介质HO-1的表达而发挥抗炎效果。

Objective To characterize anti-inflammative effect of total flavones fromMorus albaL（TFM） on RAW264.7 macrophages stimulated with interferon-γ（IFN-γ） / lipopolysaccharide（LPS）.Methods Using IFN-γplus LPS to stimulateRAW 264.7 macrophages as inflammatory cell model.Griess reaction was used for nitric oxide measurement,MTT assay wereused for cell proliferation and viability,RT-PCR was used for assaying mRNA expression and Western blot was used for protein expression.Results TFM dose-dependently inhibited both NO production and iNOS expression in IFN-γ＋LPS stimulated macrophages with out inference on cell viability.The mRNA expression of inflammatory cytokines such as IL-1β and IL-6 was significantly inhibited by TFM treatment.TFM suppressed gene expression of COX-2 while increased HO-1 expression.The protein expression of iNOS and p-ERK was inhibited by TFM.ConclusionOur study suggests that TFM may exert protective effects against inflammation damage through the mechanism of inhibiting excessive NO,reducing pro-inflammatory mediators such as COX-2,IL-1β,IL-6,and increasing HO-1 expression and enhancing total antioxidant ability.The inhibition of iNOS and COX-2 expression by TFM may be partially mediated through extracellular regulated protein kinases（ERK/MAPK）pathway.