丹皮酚抑制脂多糖诱导的小鼠BV2小胶质细胞NF-κB p65磷酸化及炎症因子表达

Paeonol inhibits the phosphorylation of NF-κB p65 and the expression of inflammatory cytokines in mouse BV2 microglia induced by lipopolysaccharide

目的研究丹皮酚对脂多糖(LPS)诱导的小鼠BV2小胶质细胞炎症反应的影响及其可能机制。方法体外培养小鼠BV2小胶质细胞,先给予不同浓度丹皮酚处理24 h,然后采用LPS刺激12 h激活BV2小胶质细胞。采用CCK-8法检测细胞活力,显微镜观察细胞形态变化,实时定量PCR检测BV2小胶质细胞肿瘤坏死因子α(TNF-α)、白细胞介素1β(IL-1β)、IL-12和IL-6 mRNA表达,Western blot法检测BV2小胶质细胞核因子κB p65(NF-κB p65)蛋白表达及其磷酸化水平。结果丹皮酚处理可提高LPS刺激后BV2小胶质细胞活力,抑制LPS诱导的BV2小胶质细胞变大变圆的过度活化状态。丹皮酚可浓度依赖性地降低LPS诱导的小胶质细胞TNF-α、IL-1β、IL-12和IL-6 mRNA表达水平;丹皮酚不影响NF-κB p65蛋白表达,但可浓度依赖性地降低LPS诱导的NF-κB p65磷酸化水平。结论丹皮酚可抑制LPS诱导的BV2小胶质细胞NF-κB p65磷酸化及其下游炎性细胞因子的转录表达。

Objective This study is aimed to investigate the effect of paeonol on inflammation of BV2 microglia induced by lipopolysaccharide(LPS),and the underlying mechanism.Methods Mouse BV2 microglia was cultured in vitro.BV2 microglia was pretreated with paeonol of different concentration for 24 hours,then stimulated by LPS for 12 hours.Cell viability was detected by CCK-8 assay.Morphological changes of microglia were monitored by microscopy.The mRNA expression of TNF-α,IL-1β,IL-12 and IL-6 by BV2 microglia was measured by real time quantitative-PCR.The protein expression of NF-κB p65 and phosphorylated NF-κB p65(p-NF-κB p65)was determined by Western blot analysis.Results Paeonol treatment improved cell viability,and inhibited over-activation of BV2 microglia challenged by LPS.Paeonol treatment concentration-dependently suppressed LPS induced mRNA expression of inflammatory cytokines including TNF-α,IL-1β,IL-6,and IL-12 by BV2 microglia.Phosphorylation of NF-κB p65,but not protein level of NF-κB p65,was suppressed by paeonol treatment in a concentration-dependent manner.Conclusion Paeonol inhibits LPS induced phosphorylation of NF-κB p65 and transcription of downstream proinflammatory cytokines in BV2 microglia.