HFRS患者汉滩病毒特异性CTL克隆靶细胞的建立及意义

目的:建立肾综合征出血热(HFRS)患者汉滩病毒(HTNV)核衣壳蛋白(NP)特异性CTL克隆的靶细胞.方法:采用Ficoll密度梯度离心法分离HFRS患者外周血单个核细胞(PBMC),并用EB病毒(EBV)转化B淋巴细胞,建立B淋巴母细胞样细胞系(B-LCL).然后,用含HTNVS基因的重组痘苗病毒(rVV)转染B-LCL,间接免疫荧光(IFA)检测NP的表达,并用HTNV特异性T细胞克隆对转染rVV的B-LCL进行细胞毒杀伤实验.结果:B淋巴细胞经EBV感染4wk左右,可形成B-LCL.HTNV-S基因在B-LCL中能有效表达NP,且能被HTNV特异性T细胞克隆识别.结论:成功建立了HFRS患者永生化B-LCL.含HTNVS的基因重组痘苗病毒转染的B-LCL可用作HTNV特异性CTL克隆的靶细胞.

Establishment and significance of target cells of Hantaan virus-specific CTL clone in patients with hemorrhagic fever with renal syndrome

AIM: To establish the target cells of CTL clone which is specific for the nucleocapsid protein (NP) of Hantaan virus (HTNV) in patients with hemorrhagic fever with renal syndrome (HFRS). METHODS: Peripheral blood mononuclear cells (PBMCs) of patients with HFRS, separated by Ficoll Hypaque density gradient centrifugation and infected by EB virus (EBV), were used to establish immortalized B LCLs. The immortalized B LCLs were infected by recombinant vaccinia viruses harboring HTNV S gene (rVV). Expression of NP was detected with indirect immunofluorescent assay (IFA) and the recognition of NP by HTNV specific T cell clones was performed by cytotoxicity assays. RESULTS: B lymphocytes became immortalized B LCLs after 4 weeks. NP was effectively expressed in B LCLs by rVV and was recognized by HTNV specific T cell clones. CONCLUSION: Immortalized B LCLs are established successfully. B LCLs infected by rVV can be used as target cells of HTNV specific CTL clones.