Fms-related tyrosine kinase 3 expression discriminates hematopoietic stem cells subpopulations with differing engraftment-potential: identifying the most potent combination.

Background: Fms-related tyrosine kinase 3 (Flt3)-ligand (FL) promotes the proliferation, differentiation, development, and mobilization of hematopoietic cells. We previously found that FL-mobilized hematopoietic stem cells (HSC) engraft efficiently, whereas FL-expanded bone marrow HSC do not. The function of FL-mobilized c-Kit(+) Sca-1(+)Lin(-)(KSL) subpopulations has not been systematically evaluated. A precise definition of the repopulating ability is needed to define which HSC subpopulations are critical for long-term chimerism and tolerance induction. FL significantly mobilized c-Kit(hi) and c-Kit(lo) Sca-1(+)Lin(-) cells into peripheral blood (PB). Here, we evaluated the influence of Flt3 expression on long-term repopulating ability of HSC subpopulations.

Methods: c-Kit(hi) or c-Kit(lo) KSL cells were sorted from PB of FL-treated green fluorescent protein-positive donors. The function of these cells was evaluated using competitive reconstitution assays, colony-forming units spleen, and colony forming cell assays. The function of c-Kit(hi) CD34(-)Flt3(-) KSL, c-Kit CD34(+)Flt3(-) KSL, c-Kit(hi) CD34(+)Flt3(+) KSL were investigated in an in vivo transplantation model.

Results: Only FL-mobilized PB c-Kit(hi) KSL cells exhibited high spleen colony-forming unit activity, generated high numbers of both lymphoid and myeloid colonies in vitro, and rescued ablated recipients. FL-mobilization expanded both c-Kit(hi) CD34(+)Flt3(-) cells (short-term HSC) and c-Kit(hi) CD34(-)Flt3(-) KSL cells (long-term HSC). There was a significant decrease in c-Kit CD34Flt3 KSL late multipotent progenitors in PB. A combination of c-Kit(hi) CD34Flt3 and c-Kit CD34(+)Flt3(-) KSL cells offered the most effective rescue of ablated recipients.

Conclusions: These data suggest that engraftment of purified HSC is influenced by both short- and long-term repopulating populations and that Flt3 expression may be useful for selecting the most critical HSC subpopulations for transplantation.