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荆芥挥发油体外抗甲型流感病毒作用及机制的研究
引用本文:何婷,陈恬,曾南,汤奇,苟玲,刘金伟,彭希,于柳.荆芥挥发油体外抗甲型流感病毒作用及机制的研究[J].中药药理与临床,2012(3):51-55.
作者姓名:何婷  陈恬  曾南  汤奇  苟玲  刘金伟  彭希  于柳
作者单位:成都中医药大学药学院,中药资源系统研究与开发利用省部共建国家重点实验室培育基地;成都医学院病原生物学教研室
基金项目:国家自然科学基金项目(30973923);"2010年度高层次留学人才回国资助人选"项目;"2010年度留学回国人员科技活动项目择优资助"项目(国家人社部人社厅函[2010]411、412号)
摘    要:目的:观察荆芥挥发油及其主要成分胡薄荷酮与薄荷酮体外对甲型流感病毒(H1N1)增殖的影响,并探讨其TLR/IFN信号通路机制。方法:MTT法检测受试药对狗肾传代细胞(MDCK)的半数中毒浓度(TC50)及最大无毒浓度(TC0);血凝法测定甲型流感病毒鼠肺适应株A/PR/8/34(H1N1)对MDCK细胞的感染性;MTT法和细胞病变法(CPE)测定三种药物体外抑制流感病毒增殖的有效浓度(IC50)和治疗指数(TI)。Real-Time PCR法观察药物对H1N1感染MDCK细胞后8h细胞TLR7、IFN-βmRNA表达水平的影响。结果:荆芥挥发油、胡薄荷酮和薄荷酮对MDCK细胞的TC50分别为0.20mg/ml、0.36mg/ml与0.43mg/ml,TC0分别为0.10mg/ml、0.10mg/ml与0.25mg/ml。荆芥挥发油和胡薄荷酮3.1×10-3~0.1mg/ml浓度时能抑制流感病毒增殖,其IC50分别为1.7×10-3mg/ml与1.9×10-3mg/ml,TI分别为120.83、184.54,呈一定量效关系;薄荷酮对流感病毒染毒的MDCK细胞几乎无保护作用。荆芥挥发油3.1×10-3~0.10mg/ml浓度,胡薄荷酮1.3×10-2~0.10mg/ml及薄荷酮7.8×10-3~0.10mg/ml浓度时对流感病毒具有明显直接杀灭作用,其IC50分别为3.1×10-3、7.2×10-3与1.9×10-3mg/ml,TI分别为63.99、50.00和82.88。荆芥挥发油和胡薄荷酮0.1mg/ml浓度时均显著提高TLR7、IFN-βmRNA表达水平,0.25mg/ml薄荷酮显著提高TLR7mRNA表达水平。结论:荆芥挥发油与胡薄荷酮体外明显抑制甲型流感病毒(H1N1)在MDCK细胞中的增殖,薄荷酮作用不明显;三者均对流感病毒有显著直接杀灭作用。其抗病毒作用机制可能与其激活TLR7,诱导IFN-β高表达,激活病毒模式识别系统有关。

关 键 词:荆芥挥发油  胡薄荷酮  薄荷酮  MDCK细胞  甲型流感病毒  Toll受体  IFN-β

Study on the effect and mechanism of anti-influenza A virus of the volatile oil of Herba Schizonepetae Briq. in vitro
Institution:He Ting,Chen Tian1,Zeng Nan,Tang Qi,Gou Ling,Peng Xi,Liu Jinwei(Pharmacy College,Chengdu University of Traditional Chinese Medicine,et al.,Chengdu 610075;1 Department of Pathogenic Biology,Chengdu Medical College,Chengdu 610083)
Abstract:Objective :To study the anti-influenza A virus effect of the volatile oil of Herba Schizonepetae Briq.(VOHS),pulegone and menthone in vitro,and to explore its mechanism of TLR/IFN signaling pathway.Methods:The median toxic concentration(TC50)and maximum non-toxic concentration(TC0)of VOHS,pulegone and menthone on MDCK cells were detected by MTT assay.The transmissibility of influenza A virus(H1N1) stain adapted mouse lung on MDCK cells were detected by hemagglutinin method.The effect of VOHS,pulegone and menthone in H1N1-infected MDCK cells was evaluated via MTT assay and quantitative measurement of the changes in the Cytopathic effects(CPE).MDCK cells were infected with H1N1 in vitro,and the cells were harvested at 8h post-infection,then real-time RT-PCR was used to detect the expression of TLR7 and IFN-β mRNA.Results:It was found that the TC50 of VOHS and pulegone on MDCK cells were 1.68×10-3mg/ml and 1.94×10-3 respectively.VOHS and pulegone showed inhibition effect on virus proliferation after adsorption.Their IC50 were 1.68×10-3mg/ml and 1.94×10-3mg/ml respectively,and therapeutic index(TI)were 120.83 and 184.54 respectively,in dose dependence manner.However,menthone was not observed that had inhibition effect on virus proliferation in MDCK cells.After infection with H1N1,the expression of TLR7 and IFN-β mRNA were significantly increased in VOHS and pulegone group,the expression of TLR7 mRNA was significantly increased in Menthone group.Conclusion:This study showed that VOHS and pulegone had the effect of anti-influenza A virus(H1N1)in vitro,and VOHS,pulegone and menthone also displayed significant antiviral activity against H1N1 when mixed with virus directly.The mechanism of the anti-influenza A virus effect of VOHS,pulegone and menthone maybe related to increasing the expression of TLR7mRNA and its downstream products of IFN-β mRNA.
Keywords:volatile oil of Herba Schizonepetae Briq  (荆芥挥发油)  pulegone  menthone  MDCK cell  influenza A virus  Toll-like receptor  IFN-β
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