Mustafa A S, Shaban F A, Al-Attiyah R, Abal A T, El-Shamy A M, Andersen P, Oftung F
Department of Microbiology; Department of Medicine, Kuwait University, Safat; Chest Diseases Hospital, Kuwait.
Scand J Immunol. 2003 Feb;57(2):125-34. doi: 10.1046/j.1365-3083.2003.01204.x.
We have used a synthetic-peptide approach to map epitope regions of the Mycobacterium tuberculosis ESAT-6 antigen recognized by human T cells in relation to major histocompatibility complex (MHC) restriction. ESAT-6-specific CD4+ T-cell lines were established by stimulating peripheral blood mononuclear cells from 25 HLA-DR-typed tuberculosis patients with complete antigen in vitro. The established T-cell lines were then screened for proliferation and interferon-gamma (IFN-gamma) secretion in response to eight overlapping 20-mer peptides covering the ESAT-6 sequence. The response of the T-cell lines to ESAT-6 and peptides from a human leucocyte antigen (HLA)-heterogeneous group of donors suggested the presence of multiple epitopes and promiscuous recognition of the antigen. Analysis of antigen and peptide recognition in the presence of anti-HLA class I and class II antibodies suggested that the T-cell lines recognized ESAT-6 in association with HLA-DR and -DQ molecules. Furthermore, testing of selected T-cell lines with ESAT-6 and the peptides in the presence of autologous and allogeneic HLA-DR- and -DQ-typed antigen-presenting cells identified HLA-DR2, -DR52 and -DQ2 amongst the HLA molecules involved in the presentation of ESAT-6 and its peptides to human Th1 cells. In addition, the T-cell lines were cytotoxic for monocytes and macrophages pulsed with ESAT-6 and peptides. In conclusion, the recognition of ESAT-6 by IFN-gamma-secreting and cytotoxic CD4+ T cells in association with frequently expressed HLA class II molecules supports the application of this antigen to either specific diagnosis or subunit vaccine design.
我们采用了合成肽方法来绘制结核分枝杆菌ESAT-6抗原的表位区域，这些表位区域可被人类T细胞识别，并与主要组织相容性复合体（MHC）限制相关。通过在体外使用完整抗原刺激25名经HLA-DR分型的结核病患者的外周血单核细胞，建立了ESAT-6特异性CD4+ T细胞系。然后，针对覆盖ESAT-6序列的8个重叠的20肽进行筛选，检测所建立的T细胞系的增殖和干扰素-γ（IFN-γ）分泌情况。T细胞系对ESAT-6以及来自人类白细胞抗原（HLA）异质供体组的肽的反应表明存在多个表位，并且该抗原具有混杂识别性。在存在抗HLA I类和II类抗体的情况下对抗原和肽识别情况进行分析，结果表明T细胞系识别与HLA-DR和-DQ分子相关联的ESAT-6。此外，在自体和同种异体HLA-DR和-DQ分型的抗原呈递细胞存在的情况下，用ESAT-6和肽对选定的T细胞系进行检测，结果在参与将ESAT-6及其肽呈递给人类Th1细胞的HLA分子中鉴定出HLA-DR2、-DR52和-DQ2。另外，T细胞系对用ESAT-6和肽脉冲处理的单核细胞和巨噬细胞具有细胞毒性。总之，分泌IFN-γ的细胞毒性CD4+ T细胞对ESAT-6的识别以及与频繁表达的HLA II类分子的关联，支持了将该抗原应用于特异性诊断或亚单位疫苗设计。
