氧化苦参碱抗流感病毒药效及机制研究

doi:10.3969/j.issn.1004-616x.2014.03.004

癌变·畸变·突变

氧化苦参碱抗流感病毒药效及机制研究

陈小璇，代剑平*，万倩英，朱丹霞，王革非，李康生

汕头大学医学院微生物与免疫学教研室，广东汕头 515041

收稿日期:2013-12-25 修回日期:2014-04-01 出版日期:2014-05-30 发布日期:2014-05-30
通讯作者: 代剑平，E-mail：daijpedu2008@163.com
作者简介:陈小璇(1984- )，女，广东省普宁市人，本科，助理实验师，研究方向：免疫药理学。E-mail：chen.x.x.85@163.com
基金资助:
国家自然科学基金项目(81374040)

The effects and mechanisms of oxymatrine on anti- influenza A virus activity

CHEN Xiao-xuan，DAI Jian-ping*，WAN Qian-ying，ZHU Dan-xia，WANG Ge-fei，LI Kang-sheng

Department of Microbiology and Immunology, Shantou University Medical College, Shantou 515041, Guangdong, China

Received:2013-12-25 Revised:2014-04-01 Online:2014-05-30 Published:2014-05-30

摘要/Abstract

摘要：

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，200 μmol/L)和氧化苦参碱处理组(12.5、25、50、100 μmol/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12.5～100 μmol/L范围内可显著降低流感病毒复制，其半数有效浓度(EC50)为19.95 μmol/L。与病毒处理组比较，启动子荧光素酶报告质粒检测显示12.5 μmol/L氧化苦参碱可显著降低流感病毒诱导的TLR4、MyD88和TRAF6转录(P＜0.05)；Western blot检测显示12.5 μmol/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12.5 μmol/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。

关键词: 氧化苦参碱, 流感病毒, TLR4信号通路, 炎性因子

Abstract:

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 12.5－100 μmol/L，could significantly inhibit the replication of IAV in vitro，and its EC50 was 19.95 μmol/L. Luciferase reporter assay showed that oxymatrine，at the concentration of 12.5 μmol/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 (12.5 μmol/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.

Key words: oxymatrine, influenza A virus, TLR4 pathway, inflammatory cytokines

陈小璇，代剑平*，万倩英，朱丹霞，王革非，李康生. 氧化苦参碱抗流感病毒药效及机制研究[J]. 癌变·畸变·突变, doi: 10.3969/j.issn.1004-616x.2014.03.004.

CHEN Xiao-xuan，DAI Jian-ping*，WAN Qian-ying，ZHU Dan-xia，WANG Ge-fei，LI Kang-sheng. The effects and mechanisms of oxymatrine on anti- influenza A virus activity[J]. Carcinogenesis, Teratogenesis & Mutagenesis, doi: 10.3969/j.issn.1004-616x.2014.03.004.

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氧化苦参碱抗流感病毒药效及机制研究

The effects and mechanisms of oxymatrine on anti- influenza A virus activity

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