Extensive Replicative Capacity of Human Central Memory T Cells

摘要

To characterize the replicative capacity of human central memory(T_CM)CD4 T cells,we have developed a defined culture system optimized for the ex vivo expansion of Ag-specific CD4~+ T cells.Artificial APCs(aAPCs)consisting of magnetic beads coated with Abs to HLA class II and a costimulatory Ab to CD28 were prepared;peptide-charged HLA class II tetramers were then loaded on the beads to provide Ag specificity.Influenza-specific DR*0401 CD4 T_CM were isolated from the peripheral blood of normal donors by flow cytometry.Peptide-loaded aAPC were not sufficient to induce resting CD4 T_CM to proliferate.In contrast,we found that the beads efficiently promoted the growth of previously activated CD4 T_CM cells,yielding cultures with >80% Ag-specific CD4 cells after two stimulations.Further stimulation with peptide-loaded aAPC increased purity to >99% Ag-specific T cells.After in vitro culture for 3-12 wk,the flu-specific CD4 T_CM had surface markers that were generally consistent with an effector phenotype described for CD8 T cells,except for the maintenance of CD28 expression.The T_CM were capable of 20-40 mean population doublings in vitro,and the expanded cells produced IFN-gamma,IL-2,and TNF-alpha in response to Ag,and a subset of cells also secreted IL-4 with PMA/ionomycin treatment.In conclusion,aAPCs expand T_CM that have extensive replicative capacity,and have potential applications in adoptive immunotherapy as well as for studying the biology of human MHC class II-restricted T cells.