Hematopoietic precursor/stem cells (HPCs) are progenitors to all functional immune cell lineages. We established a novel stromal cell line expressing (Delta-like 1) DL1, interleukin-7 (IL-7), and FMS-like tyrosine kinase 3 Ligand (Flt3-L) and demonstrated efficient differentiation and expansion of early T cells from adult CD34+ HPCs. However, continuous IL-7 signaling led to the production of immature CD8+ T lymphocytes at the expense of CD4+ T lymphocytes. It is known that the developing double positive T cells shut down IL-7 mediated signaling and require TCR signaling for survival and positive selection. We hypothesize that IL-7 inhibits the TCR signaling event required for positive selection. We found that by removal of IL-7, followed with CD3/CD28 stimulation, the early T cells were able to differentiate into mature single positive CD4 T cells expressing CD3, CD28, and TCRαβ. This can also be demonstrated by bypassing TCR signaling through stimulation with PMA/ionomycin. When further stimulated, these in vitro derived CD4+ T cells differentiated into various T helper subtypes and produced effector cytokines including IL-2, IFN-γ, and IL-17. This novel culture system can greatly facilitate the study of late T cell development and has potential for immunotherapeutic applications.
Supported by Vectorite Biomedica Inc. and Yongling Foundation.