竹节参总皂苷通过NF-κB通路对LPS致RAW264.7细胞炎症的保护作用研究

Study on protective effect of total saponins of Panax japonicus on LPS-induced RAW264.7 cell inflammation through NF-κB pathway

目的:探讨竹节参总皂苷对脂多糖(LPS)刺激的RAW264.7巨噬细胞的抗炎作用。方法:噻唑蓝(MTT)法检测不同浓度的竹节参总皂苷对RAW264.7细胞活性的影响;一氧化氮(NO)试剂盒法检测竹节参总皂苷对LPS刺激RAW264.7细胞的NO释放量;酶联免疫吸附法(ELISA)检测肿瘤坏死因子-α(TNF-α)、白介素1-β(IL-1β)分泌;逆转录聚合酶链反应法(RT-PCR)检测诱导型一氧化氮合酶(iNOS),TNF-α,IL-1βmRNA的表达;免疫印迹法(Western blot)检测胞核内核转录因子-κB p65(NF-κB p65)蛋白表达。结果:竹节参总皂苷的安全用药范围为≤80 mg·L-1;与LPS模型组相比,竹节参总皂苷各剂量组(0.1,1,10,40 mg·L-1)均能显著降低LPS刺激下RAW264.7细胞NO,TNF-α和IL-1β分泌;竹节参总皂苷各剂量组(10,40 mg·L-1)均能抑制iNOS,TNF-α,IL-1βmRNA表达和NF-κB p65蛋白表达。结论:初步证实了竹节参总皂苷对LPS致RAW264.7巨噬细胞炎症的保护作用,作用机制可能与NF-κB通路有关。

Objective： To observe the anti-inflammatory effect of total saponins of Panax japonicus on LPS-induced RAW264.7 macrophages. Method： The effect of total saponins of P. japonicus of different concentrations on RAW264.7 cell viability was determined with the MTT method. The NO kit assay was adopted to detect the NO release of total saponins of P. japonicus to LPS-induced RAW264.7 cells. The enzyme linked immunosorbent assay （ELISA） was used to detect the secretion of tumor necrosis factor-α（TNF-α） and interleukin 1-β （IL-1β）. The reverse transeriptase-polymerase chain reaction （RT-PCR） was used to determine the expression of inducible nitric oxide synthase （iNOS）,TNF-α,IL-1β. The protein expression of nuclear transcription factor-κB p65 （NF-κB p65） was tested by Western blot. Result： The safe medication range of total saponins of P. japonicus was less than 80 mg·L^-1 Compared with the LPS model group, total saponins of P. japonicus high, middle and low dose groups （0.1, 1, 10, 40 mg·L^-1 could significantly reduce the secretion of NO, TNF-α, IL-1β of LPS-induced RAW264.7 cells, and inhibit the expressions of iNOS, TNF-α and IL-1β mRNA and the protein expression of NF-κB p65. Conclusion： This study preliminarily proves the protective effect of total saponins of P. japonicus on LPS-induced RAW264.7 macrophages. Its action mechanism may be related to NF-κB signal pathway.