Functionally Mature CD4 and CD8 TCRαβ Cells Are Generated in OP9-DL1 Cultures from Human CD34+ Hematopoietic Cells

Human CD34(+) hematopoietic precursor cells cultured on delta-like ligand 1 expressing OP9 (OP9-DL1) stromal cells differentiate to T lineage cells. The nature of the T cells generated in these cultures has not been studied in detail. Since these cultures do not contain thymic epithelial cells which are the main cell type mediating positive selection in vivo, generation of conventional helper CD4(+) and cytotoxic CD8(+) TCR alpha beta cells is not expected. Phenotypically mature CD27(+)CD1(-) TCR gamma delta as well as TCR alpha beta cells were generated in OP9-DL1 cultures. CD8 and few mature CD4 single-positive TCR alpha beta cells were observed. Mature CD8 single-positive cells consisted of two subpopulations: one expressing mainly CD8 alpha beta and one expressing CD8 alpha alpha dimers. TCR alpha beta CD8 alpha alpha and TCR gamma delta cells both expressed the IL2R beta receptor constitutively and proliferated on IL-15, a characteristic of unconventional T cells. CD8 alpha beta(+) and CD4(+) TCR alpha beta cells were unresponsive to IL-15, but could be expanded upon TCR stimulation as mature CD8 alpha beta(+) and CD4(+) T cells. These T cells had the characteristics of conventional T cells: CD4(+) cells expressed ThPOK, CD40L, and high levels of IL-2 and EL-4; CD8(+) cells expressed Eomes, Runx3, and high levels of granzyme, perforin, and IFN-gamma. Induction of murine or human MHC class I expression on OP9-DL1 cells had no influence on the differentiation of mature CD8(+) cells. Similarly, the presence of dendritic cells was not required for the generation of mature CD4(+) or CD8(+) T cells. These data suggest that positive selection of these cells is induced by interaction between T precursor cells. The Journal of Immunology, 2009, 183: 4859-4870.