Day Cheryl L, Seth Nilufer P, Lucas Michaela, Appel Heiner, Gauthier Laurent, Lauer Georg M, Robbins Gregory K, Szczepiorkowski Zbigniew M, Casson Deborah R, Chung Raymond T, Bell Shannon, Harcourt Gillian, Walker Bruce D, Klenerman Paul, Wucherpfennig Kai W
Howard Hughes Medical Institute, Partners AIDS Research Center, Boston, Massachusetts, USA.
J Clin Invest. 2003 Sep;112(6):831-42. doi: 10.1172/JCI18509.
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.
丙型肝炎病毒（HCV）及其他慢性人类病毒感染的控制与病毒特异性CD4 T细胞的持续存在有关，但尚未实现对循环CD4 T细胞的体外特性分析。为了进一步明确这些细胞的表型和功能，我们基于细胞肽交换机制开发了一种生成MHC II类/肽复合物四聚体形式的新方法。HLA-DR分子以前体形式表达，带有共价连接的CLIP肽，连接子裂解后可与病毒肽有效交换。在自发清除HCV病毒血症的受试者中，而非慢性进行性感染的受试者中，尽管循环CD4 T细胞中的频率非常低（1:1200至1:111000），但通过磁珠捕获仍可分离出HCV四聚体标记的细胞。这些T细胞表达了一组表面受体（CCR7 + CD45RA - CD27 +），表明其对次级淋巴结构具有监视功能，并且由于它们利用有限的Vβ库，因此在体内经历了显著的选择。这些研究证明了临床结果与针对该病毒的循环CD4 T细胞的存在之间的关系。此外，它们表明可以在体外对人类疾病中罕见的记忆CD4 T细胞群体进行研究。
