Wehler Thomas C, Karg Michael, Distler Eva, Konur Abdo, Nonn Marion, Meyer Ralf G, Huber Christoph, Hartwig Udo F, Herr Wolfgang
Department of Medicine III - Hematology and Oncology, Johannes Gutenberg-University of Mainz, Mainz, Germany.
J Immunol Methods. 2008 Nov 30;339(1):23-37. doi: 10.1016/j.jim.2008.07.017. Epub 2008 Aug 27.
Current methods for the detection and isolation of antigen-specific CD4(+) and CD8(+) T cells require the availability of peptide/MHC multimers or are restricted to cells that produce cytokines after antigen contact. Here we show that de novo cell surface expression of the TNF-receptor family member CD137 (4-1BB) identifies recently activated, but not resting, human CD4(+) and CD8(+) memory T cells. Maximum CD137 expression level is uniformly observed in both T-cell subsets at 24h after stimulation with antigen. In experiments with CMV and EBV-reactive T cells, we confirmed the specificity of CD137 expression by co-staining with peptide/HLA tetramers. Substantial proportions of CD137(+) T cells did not produce IFN-gamma, suggesting that CD137 detects a broader repertoire of antigen-specific T cells. Activated CD137(+) T cells could be easily purified by MACS and expanded in vitro thereafter. This CD137-based enrichment method was capable of isolating 2-fold higher numbers of anti-viral CD4(+) and CD8(+) T cells compared to the IFN-gamma secretion assay. In conclusion, antigen-triggered CD137 expression allows the rapid detection and sorting of virus-reactive CD4(+) and CD8(+) T cells. The CD137 assay is most attractive for the simultaneous targeting of anti-viral T helper and effector cells in monitoring studies and adoptive immunotherapy trials.
目前用于检测和分离抗原特异性CD4(+)和CD8(+) T细胞的方法需要肽/MHC多聚体，或者仅限于抗原接触后产生细胞因子的细胞。在此我们表明，TNF受体家族成员CD137（4-1BB）的从头细胞表面表达可识别最近激活而非静止的人类CD4(+)和CD8(+)记忆T细胞。在用抗原刺激后24小时，在两个T细胞亚群中均均匀观察到最大CD137表达水平。在针对巨细胞病毒（CMV）和EB病毒（EBV）反应性T细胞的实验中，我们通过与肽/HLA四聚体共染色证实了CD137表达的特异性。相当比例的CD137(+) T细胞不产生IFN-γ，这表明CD137可检测到更广泛的抗原特异性T细胞库。激活的CD137(+) T细胞可通过磁珠分选（MACS）轻松纯化，随后在体外扩增。与IFN-γ分泌检测相比，这种基于CD137的富集方法能够分离出数量多两倍的抗病毒CD4(+)和CD8(+) T细胞。总之，抗原触发的CD137表达可实现病毒反应性CD4(+)和CD8(+) T细胞的快速检测和分选。在监测研究和过继性免疫治疗试验中，CD137检测对于同时靶向抗病毒辅助性T细胞和效应细胞极具吸引力。
