丙型肝炎病毒（HCV）F蛋白（ARFP）是近年来报道的HCV核心抗原（core）基因的核糖体读码框移蛋白，其功能尚未充分阐明。上海巴斯德研究所肿瘤病毒研究组邓强副研究员等与上海瑞金医院张欣欣教授合作，应用合成F蛋白15肽对MHC-II人源化转基因小鼠（HLA-DR1以及HLA-DP4）进行免疫筛选，通过DNA免疫和脾细胞增殖实验，鉴定获得HLA-DR1和-DP4限制的3种F蛋白特异性T细胞表位；进一步通过临床合作研究，应用上述鉴定的T细胞表位，对62例慢性丙型肝炎病人外周血淋巴细胞进行体外扩增和干扰素γ的细胞内染色分析，首次证实HCV慢性感染过程中存在F特异的CD4+T细胞应答。
相对于免疫机能正常小鼠实验模型，HCV慢性感染病人中F特异的T细胞应答显著减弱，反映了病毒持续性感染过程中T细胞功能损伤或耐受的重要特征。研究人员同时对慢性丙肝病人中F蛋白特异的抗体应答和治疗特征进行了深入分析。
该项工作充分体现了上海巴斯德所研究人员与临床研究人员在医学转化型研究方面的共同努力。
这一研究发表在12月6日国际学术期刊PLoS ONE上。
该课题获国家自然科学基金、中科院知识创新工程以及十一五国家重大科技专项经费支持。（生物谷Bioon.com）
生物谷推荐原文出处：
PLoS ONE doi:10.1371/journal.pone.0014237
Characterization of the Specific CD4+ T Cell Response against the F Protein during Chronic Hepatitis C Virus Infection
De-Yong Gao1,2, Gen-Di Jin1, Bi-Lian Yao1, Dong-Hua Zhang1, Lei-Lei Gu1, Zhi-Meng Lu1, Qiming Gong1, Yu-Chun Lone3, Qiang Deng4*, Xin-Xin Zhang1*
1 Department of Infectious Diseases, Ruijin Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China, 2 Songjiang Hospital Affiliated to Shanghai First People's Hospital, Shanghai Jiaotong University, Shanghai, China, 3 INSERM U 542, H?pital Paul Brousse, Villejuif, France, 4 Unit of Tumor Virology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China
Abstract
Background
The hepatitis C virus (HCV) Alternate Reading Frame Protein (ARFP or F protein) presents a double-frame shift product of the HCV core gene. We and others have previously reported that the specific antibodies against the F protein could be raised in the sera of HCV chronically infected patients. However, the specific CD4+ T cell responses against the F protein during HCV infection and the pathological implications remained unclear. In the current study, we screened the MHC class II-presenting epitopes of the F protein through HLA-transgenic mouse models and eventually validated the specific CD4+ T cell responses in HCV chronically infected patients.
Methodology
DNA vaccination in HLA-DR1 and-DP4 transgenic mouse models, proliferation assay to test the F protein specific T cell response, genotyping of Chronic HCV patients and testing the F-peptide stimulated T cell response in the peripheral blood mononuclear cell (PBMC) by in vitro expansion and interferon (IFN)- γ intracellular staining.
Principal Findings
At least three peptides within HCV F protein were identified as HLA-DR or HLA-DP4 presenting epitopes by the proliferation assays in mouse models. Further study with human PBMCs evidenced the specific CD4+ T cell responses against HCV F protein as well in patients chronically infected with HCV.
Conclusion
The current study provided the evidence for the first time that HCV F protein could elicit specific CD4+ T cell response, which may provide an insight into the immunopathogenesis during HCV chronic infection.
