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目的以LPS刺激N9小胶质细胞建立神经炎症模型,探索瑞香素对其的保护作用和机制,为瑞香素在神经退行性疾病的防治方面提供依据。方法用MTT法检测瑞香素对N9细胞的毒性作用;硝酸还原酶法测定瑞香素对LPS刺激N9细胞产生一氧化氮(NO)的抑制作用;一氧化氮合酶分型法检测瑞香素对LPS刺激N9细胞产生一氧化氮合酶(iNOS)酶活力的影响;Western Blot法检测iNOS、COX-2、TLR4蛋白的表达情况。结果 1～10μg.ml-1的瑞香素对炎症因子一氧化氮(NO)的产生有一定抑制作用,且能抑制一氧化氮合酶(iNOS)蛋白的表达,其作用机制可能与抑制Toll样受体4(TLR4)的表达有关。结论瑞香素1～10μg.ml-1剂量下可以抑制LPS诱导的小胶质细胞活化产生的炎症因子,其发挥药效的分子机制可能与其调控小胶质细胞TLR4信号转导通路有关。
Objective To establish neuroinflammation model by stimulating N9 microglial cells with LPS, and to explore the protective effect and mechanism of daphnetin on it, and to provide basis for daphnetin in the prevention and treatment of neurodegenerative diseases. Methods MTT assay was used to examine the toxic effect of daphnetin on N9 cells. Nitric acid reductase method was used to determine the inhibitory effect of daphnetin on nitric oxide (NO) production by LPS in N9 cells. Nitric oxide synthase Stimulated N9 cells produce nitric oxide synthase (iNOS) enzyme activity; Western Blot detection of iNOS, COX-2, TLR4 protein expression. Results Daphnetin at a concentration of 1 ~ 10μg.ml-1 inhibited the production of nitric oxide (NO) and inhibited the expression of nitric oxide synthase (iNOS), which may be related to the inhibition of Toll-like Receptor 4 (TLR4) expression. Conclusion Daphnetin can inhibit the inflammatory cytokines produced by LPS-induced microglial activation in the dose of 1 ~ 10μg.ml-1, and its molecular mechanism may be related to the regulation of TLR4 signal transduction pathway in microglia.