Modulation of haematopoietic progenitor development by Flt-3 ligand

The Flt-3 receptor is expressed in primitive haematopoietic cells and its l igand exerts proliferative effects on these cells in vitro in synergy with other cytokines, To increase our knowledge of the functional properties of the human Flt-3 ligand (FL) as relating to in vitro expansion of haematopoi etic stem cells, the effects on murine haematopoiesis of FL alone or in com bination with other growth factors were studied. Analysis of Flk-2/Flt-3 mR NA expression indicated that Flk-2/Flt-3 was preferentially expressed in pr imitive haematopoietic cell populations. To examine the expression of the F lk-2/Flt-3 receptor on megakaryocyte progenitors (CFU-Meg), Flk-2/Flt-3 pos itive and negative CD34(+) populations were separated from human bone marro w and cultured in a plasma clot culture system, CFU-Meg colonies were found in the Flk-2/Flt-3 negative fraction. Myeloid (CFU-GM) derived colonies ap peared in the presence of FL alone. Neither FL+IL-3 nor FL+IL-3+IL-6 had an y effect on the generation of megakaryocyte colonies (CFU-MX), due to the l ack of FL, receptor expression on megakaryocyte progenitors. Bone marrow ce lls remaining after 5-fluorouracil (5-FU) treatment of mice represent a ver y primitive population of progenitors enriched for reconstituting stem cell s. This cell population expressed FL receptors, as revealed by RT-PCR analy sis. Addition of FL alone did not enhance the replication of such cells in liquid cultures as compared to controls. However, a significantly greater g eneration of myeloid progenitors (CFU-GM) in clonogenic assays was observed in the presence of FL+IL-3, FL+GM-CSF or FL+CSF-1, In addition, the effect s of FL on in vitro expansion of murine haematopoietic stem cells were stud ied using lineage-negative (lin(-)) Sca-1 positive (Sca-1(+)) c-kit positiv e (c-kit(+)) marrow cells from 5-FU treated mice, FL enhanced the survival of primitive murine lin- Sca-1(+) c-kit(+) cells. FL and IL-6 were able to significantly expand murine progenitor stem cells in vitro and promote thei r survival. These studies strongly suggest that FL significantly and select ively enhanced the generation of myeloid progenitors in vitro and increased myeloid progenitor responsiveness to later acting growth factors. In addit ion, FL synergized with IL-6 to support in vitro expansion of haematopoieti c progenitors and promoted the survival of lin(-) Sca-1(+) c-kit+ cells, (C ) 1999 Academic Press.