Human CD34+ hematopoietic stem cells capable of multilineage engrafting NOD/SCID mice express flt3: distinct flt3 and c-kit expression and response patterns on mouse and candidate human hematopoietic stem cells

Abstract

The cytokine tyrosine kinase receptors c-kit and flt3 are expressed and function in early mouse and human hematopoiesis. Through its ability to promote ex vivo expansion and oncoretroviral transduction of primitive human hematopoietic progenitors, the flt3 ligand (FL) has emerged as a key stimulator of candidate human hematopoietic stem cells (HSCs). However, recent studies in the mouse suggest that though it is present on shortterm repopulating cells, flt3 is not expressed on bone marrow long-term reconstituting HSCs, the ultimate target for the development of cell replacement and gene therapy. Herein we demonstrate that though only a fraction of human adult bone marrow and cord blood CD34 ؉ longterm culture-initiating cells (LTC-ICs) express flt3, most cord blood lymphomyeloid HSCs capable of in vivo reconstituting nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice are flt3 ؉ . The striking difference in flt3 and c-kit expression on mouse and candidate human HSCs translated into a corresponding difference in flt3 and c-kit function because FL was more efficient than SCF at supporting the survival of candidate human HSCs. In contrast, SCF is far superior to FL as a viability factor for mouse HSCs. Thus, the present data provide compelling evidence for a contrasting expression and response pattern of flt3 and c-kit on mouse and human HSCs. IntroductionAll blood cells arise from a common hematopoietic stem cell (HSC), 1 which, because of its self-renewal capacity, can maintain hematopoiesis throughout the lifespan of an individual. 2,3 The molecular mechanisms regulating HSC fate decisions between self-renewal and lineage commitment remain largely unknown. 2 However, evidence supports that blood-lineage maturation and HSC self-renewal are regulated in part by cytokines interacting with their specific surface receptors. Stem cell factor (SCF, or c-kit ligand) and thrombopoietin (TPO) have been shown to act directly on HSCs, 4-9 and gene-targeting studies have demonstrated that SCF and TPO play important roles in sustaining HSC number and function, in contrast to other cytokines. 10-13 Flt3 (or flk2) is, like c-kit, a cytokine tyrosine kinase receptor expressed primarily at very early stages of hematopoiesis. [14][15][16][17][18] Mice deficient in the expression of flk2 or flt3 ligand (FL) show deficient lymphopoiesis. 19,20 FL has also been implicated as a key and potent stimulator of candidate mouse and human HSCs and is frequently used to promote HSC ex vivo expansion and oncoretroviral-mediated gene transfer. [21][22][23][24][25] However, it has not been established whether this reflects a role of flt3 in HSC generation, maintenance, homing, engraftment, or expansion after transplantation.Surprisingly, recent studies revealed that mouse adult long-term reconstituting HSCs (LTR-HSCs) do not express flt3 during steady-state hematopoiesis and that the up regulation of flt3 expression within the HSC compartment is accompanied by loss of self-renewal capacity. 26,27 In addition, SCF, but not FL, eff...

FLT3: ITDoes matter in leukemia

“…11,12 However, a recent study showed that in human CD34-positive cells, the FLT3-positive fraction has the active long-term reconstituting activity. 31 The accumulated data clearly point to FLT3 as an important receptor in early hematopoiesis.…”

Hematopoietic developmental pathways: on cellular basis

“…Around 40-80% of hCD34 þ bone marrow and cord blood cells express hFlt3 (Rappold et al, 1997;Sitnicka et al, 2003). Although a fraction of both the hFlt3 þ and the hFlt3 À populations gave rise to multilineage colonies containing all myelo-erythroid components, the hFlt3 þ hCD34 þ and hFlt3 À hCD34 þ populations predominantly formed GM and erythroid colonies, respectively (Rappold et al, 1997;Gotze et al, 1998;Sitnicka et al, 2003), and cells with NOD-SCID reconstitution activity reside in the hCD34 þ hFlt3 þ fraction (Sitnicka et al, 2003). Our recent experiments have revealed that hFlt3 is expressed in the entire human bone marrow and cord blood hCD34 þ hCD38 À population, and purified hFlt3 þ hCD34 þ hCD38 À cells could reconstitute multilineage cells for a long term in the NOD-SCID/ IL-2Rgnull xenogeneic transplantation system (unpublished data).…”

Internal tandem duplications of the FLT3 gene are present in leukemia stem cells

“…2,3 The receptor tyrosine kinase FLT3 is expressed in CD34 ϩ hematopoietic stem/progenitor cells and plays an important role in normal hematopoiesis. [4][5][6][7] FLT3 is also expressed on the leukemic blasts in the majority of cases of acute leukemia, although its expression is no longer tightly coupled to CD34 expression. [8][9][10][11] Internal tandem duplication mutations of FLT3 (FLT3/ITD mutations) occur in approximately 23% of patients with AML and are associated with an increased relapse rate and reduced overall survival.…”