栀子苷抑制甲型流感病毒感染细胞中Toll样受体7/核因子-κB信号通路  

Antagonism of geniposide on Toll-like receptor 7/nuclear factor-κB signaling pathways in cells with influenza A virus infection

作　　者：张春晶[1] 于海涛[1] 

机构地区：[1]齐齐哈尔医学院生物遗传、生物化学与分子生物学教研室,161042

出　　处：《中华微生物学和免疫学杂志》2010年第8期749-754,共6页Chinese Journal of Microbiology and Immunology

基　　金：黑龙江省青年基金资助项目（QC2009C89）

摘　　要：目的 研究栀子苷对甲型流感病毒H3N2感染所致的Toll样受体7（Toll-like receptors7,TLR7）及其介导的细胞内转录因子核因子-κB（nuclear factor-kappa B,NF-κB）活性及前炎症细胞因子TNF-α和IL-6释放的影响,以探讨栀子苷干预甲型流感病毒作用的分子生物学机制.方法 甲型流感病毒H3N2作为刺激因素,利用双荧光素酶顺式报告系统和免疫荧光实验检测栀子苷对NF-κB转录活性及NF-κB核转位的影响.RT-PCR法进一步验证栀子苷对NF-κB上下游靶基因TLR7、TNF-α和IL-6 mRNA表达水平的影响.结果 通过NF-κB双萤光素酶报告系统检测发现,与正常对照组比较,病毒感染的细胞中NF-κB荧光素酶报告活性明显升高 与病毒损伤组比较,栀子苷治疗组NF-κB荧光素酶报告活性明显受到抑制.荧光倒置显微镜下观察NF-κB p65磷酸化水平及核转位变化结果显示,病毒感染的细胞中NF-κB磷酸化水平及人核率明显升高 与病毒损伤组比较,栀子苷治疗组NF-κB磷酸化水平及入核率明显受到抑制.RT-PCR结果显示栀子苷能明显抑制病毒诱导的TLR7、TNF-α和IL-6 mRNA的高表达.结论 栀子苷可拮抗甲型流感病毒H3N2感染细胞中TLR7介导的细胞内转录因子NF-κB信号转导通路的活化,并可抑制其下游靶基因TNF-α和IL-6 mRNA的高表达水平来发挥抗病毒感染的作用.Objective To observe how geniposide as an anti-inflammatory agent through inhibition Toll-like receptor 7/nuclear factor-κB signaling pathways activation, as well as TNF-α and IL-6 release infectioned by influenza virus. Methods Epithelial cells was exposed to human influenza viruses A/Gui/81/23（H3N2） infection for 2 h before treatment with geniposide for 24 h. NF-κB responsive element luciferase reportor gene was transfected and dual luciferase cis-reporting systems was used to assay the transcriptional activity of NF-κB under the stimulated circumstance of influenza virus infection. The phosphory level and nuclear transposition of NF-κB was observed by fluorescence inverted microscope. RT-PCR was used to detect the gene transcription level of TLR7, TNF-α and IL-6. Results The relative luciferase reporter assay of NF-κB was apparently improved by influenza virus infection. But geniposide significantly repressed the relative value of luciferase. The phosphorylation level and rate for nuclear transposition of NF-κB was apparently improved by influenza A virus infection observed by fluorescence inverted microscope. But geniposide significantly repressed the phosphorylation level and rate for nuclear transposition. RT-PCR showed upregulation of TLR7 and pre-inflammatory markers TNF-α and IL-6 in A549 cells infected by influenza virus, geniposide had a significant effect on the expression of TLR7 and inflammatory markers TNF-α and IL-6 after treated with influenza virus. Conclusion Geniposide as an antiinflammatory agent antagonized influenza A virus infection through inhibiting Toll-like receptor 7/nuclear factor-κB signaling pathways activation, as well as on the downregulation of the downstream inflammatory markers target gene expression TNF-α and IL-6.

关 键 词：栀子苷 甲型H3N2流感病毒 Toll样受体7/核因子-κB信号通路 炎症细胞因子 

分 类 号：R28[医药卫生—中药学]