The FLT3+ fraction within the bone marrow lem-sca+kit+ stem cell compartment efficiently reconstitutes B and T lymphopoiesis but lacks ability to long-term reconstitute myelopoiesis

The cytokine tyrosine kinase receptor flt3 and its ligand (FL) have been demonstrated to play an important role in early hematopoiesis. Although less clear, studies in flt3 deficient mice and the ability of FL to support expansion of candidate human stem cells, have implicated that flt3/FL might also play a specific role in regulating the hematopoietic stem cell (HSC) pool. Lin-Sca+kit+ cells represent only 0.05% of the bone marrow (BM) cells but contain virtually all its long-term reconstituting activity. Flow cytometric analysis revealed that approximately 40%-50% of cells in the BM Lin-Sca+kit+ HSC compartment coexpressed flt3. Utilizing flow cytometric cell sorting Lin-Sca+kit+flt3+ (LSKF+) and Lin-Sca+kit-i-fU3- (LSKF-) cells were obtained from Ly5.1+ mice at high purity and investigated for their ability to reconstitute hematopoiesis in lethally irradiated recipients (Ly5.2+). Transplantation of as few as 750-1000 LSKF+ as well as LSKF- cells in competition with 200,000 unfractionated BM cells (Ly5.2+) resulted in high levels of reconstitution (20%-40%) by 4 weeks. However, whereas LSKF- cells reconstituted both lymphopoiesis and myelopoiesis, LSKF+ cells efficiently reconstituted lymphopoiesis but not myeloid cell production. Furthermore, whereas LSKF- cells supported long-term reconstititution of all blood lineages, LSKF+ cells showed lymphoid-restricted rapidly declining reconstitution, although B and T cell production could be observed as late as 4 to 6 months following transplantation. Despite of their inability to reconstitute myelopoiesis in vivo, LSKF-t-cells showed high proliferative and myeloid potential in vitro. When the whole LSK population was cultured in vitro in FL for 5 days, no long-term in vivo reconstituting activity could be recovered, lending further support to the lack of flt3 expression on BM HSC. In contrast, LSK cells incubated in the presence of the ligand for c-kit showed maintained long-term reconstituting activity. Thus, whereas long-term repopulating HSC in murine BM express high levels of c-kit. they appear to lack constitutive expression of flt3. Rather, acquisition of flt3 expression in the HSC pool appears to be associated with ability to efficiently reconstitute B and T lymphopoiesis, but with loss of long-term multilineage reconstituting potential.