CAJ | 학술논문

目的：研究氧化苦参碱的抗流感病毒活性及其潜在机制。方法：培养MDCK和A549细胞，并设空白对照组(DMSO，0.5%)、病毒处理组、阳性对照组(利巴韦林，200μmol/L)和氧化苦参碱处理组(12.5、25、50、100 mol/L)，每组重复6孔。空斑抑制法测定氧化苦参碱抗病毒药效；构建荧光素酶报告质粒，并用Western blot和ELISA检测氧化苦参碱对流感病毒诱导的细胞Toll样受体4(TLR4)、髓样分化因子88(MyD88)和肿瘤坏死因子受体相关因子6(TRAF6)启动子转录、TLR4-Myd88-TRAF6-NF-κB信号通路活化及炎性因子释放的影响。结果：空斑抑制实验显示氧化苦参碱在12.5～100μmol/L范围内可显著降低流感病毒复制，其半数有效浓度(EC )为19.95μmol/L。启动子荧光素酶报告质粒检测显示12.5μmol/L氧化苦参碱可显著降低流感病毒诱50导的TLR4、MyD88和TRAF6转录(P＜0.05)；Western blot检测显示12.5μmol/L氧化苦参碱可显著降低流感病毒诱导的TLR4-Myd88-TRAF6-NF-κB通路的活化(P＜0.05)；ELISA检测显示12.5μmol/L氧化苦参碱可显著降低流感病毒诱导的IL-1β、IL-6、TNF-α、IL-8、IFN-β和IFN-γ释放(P＜0.05)。结论：氧化苦参碱具抗流感病毒活性，其潜在机制可能与其抑制流感病毒诱导的TLR4-Myd88-TRAF6-NF-κB信号通路活化有关。μ与病毒处理组比较，
목적：연구양화고삼감적항류감병독활성급기잠재궤제。방법：배양MDCK화A549세포，병설공백대조조(DMSO，0.5%)、병독처리조、양성대조조(리파위림，200μmol/L)화양화고삼감처리조(12.5、25、50、100 mol/L)，매조중복6공。공반억제법측정양화고삼감항병독약효；구건형광소매보고질립，병용Western blot화ELISA검측양화고삼감대류감병독유도적세포Toll양수체4(TLR4)、수양분화인자88(MyD88)화종류배사인자수체상관인자6(TRAF6)계동자전록、TLR4-Myd88-TRAF6-NF-κB신호통로활화급염성인자석방적영향。결과：공반억제실험현시양화고삼감재12.5～100μmol/L범위내가현저강저류감병독복제，기반수유효농도(EC )위19.95μmol/L。계동자형광소매보고질립검측현시12.5μmol/L양화고삼감가현저강저류감병독유50도적TLR4、MyD88화TRAF6전록(P＜0.05)；Western blot검측현시12.5μmol/L양화고삼감가현저강저류감병독유도적TLR4-Myd88-TRAF6-NF-κB통로적활화(P＜0.05)；ELISA검측현시12.5μmol/L양화고삼감가현저강저류감병독유도적IL-1β、IL-6、TNF-α、IL-8、IFN-β화IFN-γ석방(P＜0.05)。결론：양화고삼감구항류감병독활성，기잠재궤제가능여기억제류감병독유도적TLR4-Myd88-TRAF6-NF-κB신호통로활화유관。μ여병독처리조비교，
OBJECTIVE: To explore the effects and mechanisms of oxymatrine on anti-influenza A virus (IAV) activity. METHODS: MDCK and A549 cells were cultured and divided into 4 groups: blank control (DMSO, 0.5%), virus-treated group,positive group (ribavirin,200 μmol/L) and oxymatrine-treated group (12.5,25,50 and 100 μmol/L) ,with 6 repeats in each group. The anti-IAV activity of oxymatrine was determined by plaque inhibition assay. The influence of oxymatrine on the IAV-induced transcription of TLR4,MyD88 and TRAF6,the activation of TLR4-Myd88-TRAF6-NF-κB signal pathway,and the release of inflammatory cytokins were determined by luciferase reporter plasmid,Western blot and ELISA assays,respectively. RESULTS: Plaque inhibition assay showed that oxymatrine,at a range of 12.5-100μmol/L,could significantly inhibit the replication of IAV in vitro,and its EC50 was 19.95μmol/L. Luciferase reporter assay showed that oxymatrine,at the concentration of 12.5μmol/L,could significantly inhibit the transcription of TLR4,MyD88 and TRAF6 induced by IAV infection (compared with virus-treated group,P<0.05). Western blot assay showed that oxymatrine (12.5μmol/L) could significantly inhibit the IAV-induced activation of TLR4-Myd88-TRAF6-NF-κB pathway (compared with virus-treated group,P<0.05). ELISA assay showed that oxymatrine (12.5 μmol/L) could significantly inhibit the IAV-induced release of inflammatory cytokins (compared with virus-treated group,P<0.05). CONCLUSION: Oxymatrine possessed anti-IAV activity,and its mechanism may be related to its ability to inhibit the IAV-induced activation of TLR4-MyD88-TRAF6-NF-κB pathway.