滨蒿总黄酮体外抗呼吸道合胞病毒活性及其对先天免疫信号通路的影响

Anti-respiratory syncytial virus(RSV)activity and effects of the total flavonoids from Artemisia scoparia Waldst et Kit.on the innate immune signaling pathway in vitro

为探讨滨蒿总黄酮(ASTF)体外抗呼吸道合胞病毒(RSV)活性及其对RSV感染人肺上皮细胞A549天然免疫信号通路的影响,采用细胞病变评分法和Viral ToxGlo法检测ASTF体外抗RSV的作用,Western-blotting法检测先天免疫信号通路蛋白表达水平,流式微球技术(CBA)和ELISA检测炎性细胞因子水平。结果显示,ASTF在体外对RSV有明显的抑制作用,且随着浓度的增加,抗病毒作用增强,呈量效关系。进一步试验发现,与RSV对照组相比,500μg/mL ASTF可显著降低先天免疫信号通路中线粒体抗病毒信号传送蛋白(mitochondrial antiviral signaling protein,MAVs)、核因子κB抑制蛋白(inhibitor of NF-κB,IκBα)、蛋白激酶B(protein kinase B,AKT)、受体相互作用蛋白激酶3(receptor interacting proteins 3,RIP3)和TANK结合激酶1(TANK binding kinase 1,TBK1)的蛋白表达水平。157、500μg/mL ASTF对RSV感染A549细胞分泌的炎症因子IL-1β、IL-2、IL-6、IL-10和TNF-α的水平有明显的干预作用(P<0.05),表现出下调趋势。结果表明,ASTF通过先天免疫信号通路发挥免疫调节作用,从而达到抗RSV的效果。

To investigate the anti-respiratory syncytial virus(RSV) activity of total flavonoids from Artemisia scoparia Waldst et Kit.(ASTF) and its effects on the innate immune signaling pathway in human pulmonary alveolar epithelial cell line A549 infected by RSV in vitro.Cytopathic effect scoring method and Viral Tox Glo were used to detect the anti-RSV activity of ASTF in vitro.Western-blotting was exploited to detect the expression levels of proteins in the innate immune signaling pathway.Cytometric bead array(CBA) and enzyme-linked immunosorbent assay(ELISA) were used to measure the level of inflammatory cytokines.ASTF had an obvious inhibitory activity against RSV in vitro.Moreover,the anti-RSV effect increased in a dose-dependent manner with the concentration of ASTF.Further experiments found that compared with the RSV control group,500 μg/m L of ASTF could significantly reduce the protein expression levels of mitochondrial antiviral signaling protein(MAVs),inhibitor of nuclear factor-κB(IκBα),protein kinase B(AKT),receptor interacting proteins 3(RIP3) and TANK binding kinase 1(TBK1)in the innate immune signaling pathway.Furthermore,157 or 500 μg/m L of ASTF had an obvious intervention effect on the levels of inflammatory cytokines IL-1β,IL-2,IL-6,IL-10 and TNF-ɑ secreted by RSV-infected A549 cell,showing down-regulated trends.In conclusion,ASTF plays an immunomodulatory role via inhibiting the innate immune signaling pathway,so as to achieve the anti-RSV efficacy.