摘要
目的研究柯萨奇B3病毒(coxsackievirus group B type3,CVB3)感染对固有免疫识别受体—黑色素瘤分化相关基因5(melanoma differentiation-associated gene-5,MDA-5)表达的影响及其意义。方法采用RT-PCR扩增CVB3正负链以确证感染成功,CVB3感染正常HeLa细胞后用RT-PCR的方法检测MDA-5和IFN-β的表达变化,RT-PCR检测CVB3感染IFN-α处理过细胞MDA-5的表达情况,将细胞分为对照组(转染空载体)和MDA-5组(转染MDA-5表达质粒pEF-BOS-MDA-5),将空载体或pEF-BOS-MDA-5质粒与IFN-β启动子荧光报告质粒和作为内参照的pc MV-lacZ质粒瞬时转染进两组细胞,CVB3感染8h后收集细胞检测IFN-β启动子活性变化。结果与未感染组相比,CVB3感染下调固有免疫识别受体MDA-5及IFN-β的表达;IFN-α可明显上调MDA-5的表达,而CVB3感染抑制IFN-α的这种上调作用;CVB3感染下调MDA-5介导的IFN-β启动子的活化。结论CVB3感染能下调固有免疫识别受体MDA-5的表达并抑制MDA-5对IFN-β启动子的活化,从而下调IFN-β的表达,这可能是病毒性心肌炎中病毒逃逸机体免疫应答的机制之一。
Objective To detect an innate immune recognition receptor melanoma differentiationassociated gene5 （MDA-5） expression after coxsacklevirus group B type 3 （CVB3） infection and explore its significance. Methods The infection was confirmed with the evidence of the existence of positive and negative strands of CVB3. Normal HeLa cells were infected with CVB3; subsequently MDA-5 and IFN-β expression was analyzed by RT-PCR. HeLa cells were primed with IFN-α and then infected with CVB3. Cells were harvested and RT-PCR was conducted to analyze MDA-5 expression. Cells were divided into two groups： control （transfected with vector） and MDA-5 （transfected with pEF-BOS-MDA-5）. Each group was transfected with vector or pEF-BOS-MDA-5, IFN-β promoter luciferase reporter plasmid and pcMV-lacZ as internal control. Then the cells were mock infected or CVB3 infected for 8h. Then the IFN-β promoter luciferase activity was analyzed. Results CVB3 infection down-regulated MDA-5 and IFN-β expression. IFN-α up-regulated MDA-5 expression obviously, which could be inhibited by CVB3 infection. Additionally, CVB3 infection inhibited MDA- 5-mediated IFN-β promoter activation. expression, inhibited MDA-5-mediated Conclusions CVB3 infection down-regulated MDA-5 IFN-β promoter activation and then alleviated IFN-β expression, which might be a mechanism to escape host immune response during viral myocarditis.
出处
《复旦学报（医学版）》
CAS
CSCD
北大核心
2008年第4期477-482,共6页
Fudan University Journal of Medical Sciences
基金
上海市科委产学研项目(04DZ11601)
上海市医学领军人才基金(LJ06011)
教育部留学回国基金(FGF120809)
