Ex vivo analysis of human memory CD4 T cells specific for hepatitis C virus using MHC class II tetramers.
Ex vivo analysis of human memory CD4 T cells specific for hepatitis C virus using MHC class II tetramers.
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DOI：
10.1172/jci18509
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发表时间：
2003-09
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通讯作者：
C. Day;Nilufer P. Seth;M. Lucas;H. Appel;L. Gauthier;G. Lauer;G. Robbins;Z. Szczepiorkowski;D. Casson;R. Chung;Shannon Bell;G. Harcourt;B. Walker;P. Klenerman;K. Wucherpfennig
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作者：
C. Day;Nilufer P. Seth;M. Lucas;H. Appel;L. Gauthier;G. Lauer;G. Robbins;Z. Szczepiorkowski;D. Casson;R. Chung;Shannon Bell;G. Harcourt;B. Walker;P. Klenerman;K. Wucherpfennig
遏制丙型肝炎病毒（HCV）和其他慢性人类病毒感染与病毒特异性CD4 T细胞的持久性有关，但尚未实现循环CD4 T细胞的体外表征。为了进一步确定这些细胞的表型和功能，我们开发了一种基于细胞肽交换机制的四聚体形式MHC II类/肽复合物的新方法。HLA-DR分子作为带有共价连接的CLIP肽的前体表达，该前体可以在连接体切割后与病毒肽有效交换。在自发解决HCV病毒血症的受试者中，而不是慢性进行性感染的受试者中，HCV四聚体标记的细胞可以通过磁珠捕获分离，尽管循环CD4 T细胞的频率非常低（1:12 00至1:11 1000）。这些T细胞表达一组表面受体（CCR7+CD45RA-CD27+），表明它们对次级淋巴结构具有监视功能，并且由于它们利用了有限的Vbeta库，因此经历了显著的体内选择。这些研究证明了临床结果与针对这种病毒的循环CD4 T细胞的存在之间的关系。此外，他们还表明，记忆性CD4 T细胞的罕见群体可以在人类疾病中进行体外研究。
Containment of hepatitis C virus (HCV) and other chronic human viral infections is associated with persistence of virus-specific CD4 T cells, but ex vivo characterization of circulating CD4 T cells has not been achieved. To further define the phenotype and function of these cells, we developed a novel approach for the generation of tetrameric forms of MHC class II/peptide complexes that is based on the cellular peptide-exchange mechanism. HLA-DR molecules were expressed as precursors with a covalently linked CLIP peptide, which could be efficiently exchanged with viral peptides following linker cleavage. In subjects who spontaneously resolved HCV viremia, but not in those with chronic progressive infection, HCV tetramer-labeled cells could be isolated by magnetic bead capture despite very low frequencies (1:1,200 to 1:111,000) among circulating CD4 T cells. These T cells expressed a set of surface receptors (CCR7+CD45RA-CD27+) indicative of a surveillance function for secondary lymphoid structures and had undergone significant in vivo selection since they utilized a restricted Vbeta repertoire. These studies demonstrate a relationship between clinical outcome and the presence of circulating CD4 T cells directed against this virus. Moreover, they show that rare populations of memory CD4 T cells can be studied ex vivo in human diseases.