Hematopoietic stem cells: c-mpl and TNF receptor function

Sammanfattning: Hematopoietic stem cells (HSC) are crucial to life. Daily and throughout the life span of an individual they are responsible for replacing approximately 10(12) mature blood cells. Mature cells are, for example, required for transportation of oxygen, defense against infections and to stop bleeding. Thus, our aim with this study was to characterize these rare murine (Lin-Sca1+c-kit+) and human (CD34+CD38-) bone marrow (BM) HSC with respect to their viability, proliferation, differentiation and expansion in response to cytokines. When these studies were initiated thrombopoietin (Tpo) was postulated to act primarily as a megakaryocyte lineage-specific factor. However, we demonstrated that Tpo directly and synergistically enhanced multilineage growth and differentiation from both Lin-Sca1+c-kit+ and CD34+CD38- BM cells, when combined with cytokines like c-kit ligand (KL), flt3 ligand (FL), and the interleukins 3 and 11. In addition, Tpo, more than any other early-acting cytokines, efficiently promoted viability of single CD34+CD38- cells in the absence of cell division, further implicating that Tpo directly acted on candidate stem cells. Interestingly, a direct comparison of human long-term culture-initiating cells (LTC-IC) and the more primitive extended LTC-IC (ELTC-IC), demonstrated distinct and stringent requirements for their ex vivo expansion. Specifically, ELTC-IC were potently expanded in serum-free medium in the presence of high concentrations of KL+FL+Tpo for 12 days. We also observed that Tpo-induced clonal growth of human CD34+CD38- progenitor cells was almost completely abrogated by tumor necrosis factor-a (TNF-a) and transforming growth factor-b. Additionally, we demonstrated in mice that functional signaling through the Fas and TNF receptors severely compromised HSC short- and long-term multilineage repopulating activity.