埃可病毒6型对恒河猴树突状细胞的感染研究

目的：肠道病毒作为小核糖核酸病毒的成员之一，其病原特性已得到了广泛的研究，为进一步探讨其尚未完全清楚的病理学机理，利用来源于恒河猴的不同免疫细胞，对埃可病毒6型（Echovirus 6，Echo 6）毒株与树突状细胞的作用关系及其特征做了初步分析。方法：利用流式细胞仪及抗不同树突状细胞表面特征分子的抗体，从恒河猴外周血及骨髓分离未成熟树突状细胞，经体外诱导培养后，以Echo 6病毒在感染复数为0.1的条件下感染树突状细胞，并于感染后不同时间点收集细胞，检测病毒载量，绘制病毒增殖曲线，同时对上清中相关细胞因子含量做出动态检测。结果：源于恒河猴外周血PBMC分选出的树突状细胞以成熟期树突状细胞（CD11⁺/CD83⁺）居多；而源于骨髓的细胞经粒细胞-巨噬细胞集落刺激因子（granulocyte-macrophage colony stimulating factor，GM-CSF）和重组人白细胞介素-4（recombinant human interleukin-4，rhIL-4）刺激培养后诱导分化的树突状细胞则以未成熟细胞为主。Echo 6型病毒分别对未成熟树突状细胞和成熟树突状细胞进行感染后显示，病毒在未成熟树突状细胞中有明显增殖的趋势而非在成熟树突状细胞中增殖。进一步检测病毒感染这些细胞后效应分子的表达情况发现，感染了Echo 6型病毒的未成熟树突状细胞出现肿瘤坏死因子（tumor necrosis factor-α，TNF-α）和白细胞介素-6（interleukin-6，IL-6）转录表达水平的明显上升，同时这些细胞的表面标记呈现CD11⁺/CD83⁺趋势。结论：Echo 6型病毒能够感染未成熟树突状细胞并在其中出现明显增殖趋势，并在上调细胞TNF-α和rhIL-6表达水平的同时，促进这些细胞转变为成熟树突状细胞。

Objective:As a member of picornavirus family,the pathogenic characterisitcs of enterovirus have been extensively studied. In order to further discuss its pathological mechanism,which was not completely clear,the relationship between Echovirus type 6 (Echovirus 6,Echo 6) and rhesus dendritic cells and its characteristics were preliminarily analyzed using different immune cells derived from rhesus monkeys. Methods:Immature dendritic cells (imDCs) were isolated from the peripheral blood and bone marrow of rhesus monkeys using specific antibodies against the surface molecules of dendritic cells by flow cytometry. After induction culture in vitro,dendritic cells were infected with Echo 6 at low multiplicity of infection (moi=0.1). The samples were obtained at different time points after infection. Then,the viral loads were measured and the virus proliferation curve was drawn. In addition,the levels of cytokines in the supernatant were dynamically detected. Results:Dendritic cells derived from PBMC of peripheral blood of rhesus monkeys were mostly mature dendritic cells (mDCs,CD11⁺/CD83⁺). Whereas,dendritic cells derived from bone marrow induced by granulocyte-macrophage colony stimulating factor (GM-CSF) and recombinant human interleukin-4 (rhIL-4) in vitro were mainly immature (imDCs). The imDCs and mDCs were infected by Echo 6 in vitro. The virus significantly proliferated in imDCs rather than in mDCs. In addition,the expression levels of effect molecules such as tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6) were significantly increased in Echo6-infected imDCs. Moreover,the surface markers of these imDCs showed a trend of CD11⁺/CD83⁺. Conclusion:Echo 6 can infect imDCs,have a trend of obvious proliferation in these cells,and promote the transformation of imDCs mDCs while up-regulating the expression levels of TNF-αand rhIL-6.

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