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As an early acting growth factor, flt-3 ligand (FL) promotes the ex vivo expansion of hematopoietic stem and progenitor cells. The effect and mechanism of FL on the development of the megakaryocytic lineage remain unclear. In this study, we compared the effects of FL and stem cell factor (SCF) in combination with other megakaryocyte-promoting cytokines on the differentiation and proliferation of megakaryocytic progenitors and investigated the expression of flt-3 receptors on megakaryocytic cell lines. In liquid cultures of enriched CD34+ cells from human umbilical cord blood for 14 days, FL plus thrombopoietin (TPO), interleukin-3 (IL-3), and IL-6 promoted the expansion of nucleated cells, CD34+ cells, CD34+CD38– cells, and megakaryocyte colony-forming units (CFU-MK) by 300 ± 115-, 23.8 ± 11.3-, 33.9 ±28.6-, and 584 ± 220-fold, respectively. Replacing FL with SCF significantly decreased the yield of all cell types. Using murine bone marrow (BM) cells, we demonstrated that FL at a range of 0–100 ng/ml had no significant mitogenic effect on CFU-MK formation. TPO increased CFU-MK (p &lt; 0.001) but the effect was not significantly modified by FL. While one human acute lymphoblastic leukemia sample expressed high levels of flt-3 receptor, the four megakaryocytic cell lines (Meg-01, CHRF-288-11, M-07e, and Dami) did not show any positive expression. Our data suggest that the present cytokine combination and expansion conditions provide an effective and potentially useful system for the clinical expansion of cord blood for bone marrow transplantation (BMT). FL alone did not stimulate megakaryocytopoiesis, possibly due to the lack of receptor expression on megakaryocytes. The effect of FL in augmenting the expansion of CFU-MK in liquid culture might be due to the early action of FL at the pluripotent stem cell stage.

Effects of Flt-3 Ligand in Combination with TPO on the Expansion of Megakaryocytic Progenitors