Organ-selective homing defines engraftment kinetics of murine hematopoietic stem cells and is compromised by ex vivo expansion

Hematopoietic reconstitution of ablated recipients requires that intravenou sly (IV) transplanted stem and progenitor cells "home" to organs that suppo rt their proliferation and differentiation. To examine the possible relatio nship between homing properties and subsequent engraftment potential, murin e bone marrow (BM) cells were labeled with fluorescent PKH26 dye and inject ed into lethally irradiated hosts. PKH26(+) cells homing to marrow or splee n were then isolated by fluorescence-activated cell sorting and assayed for in vitro colony-forming cells (CFCs). Progenitors accumulated rapidly in t he spleen, but declined to only 6% of input numbers after 24 hours. Althoug h egress from this organ was accompanied by a simultaneous accumulation of CFCs in the BM (plateauing at 6% to 8% of input after 3 hours), spleen cell s remained enriched in donor CFCs compared with marrow during this time. To determine whether this differential homing of clonogenic cells to the marr ow and spleen influenced their contribution to short-term or long-term hema topoiesis in vivo, PKH26(+) cells were sorted from each organ 3 hours after transplantation and injected into lethally irradiated Ly-5 congenic mice. Cells that had homed initially to the spleen regenerated circulating leukoc ytes (20% of normal counts) approximately 2 weeks faster than cells that ha d homed to the marrow, or PKH26-labeled cells that had not been selected by a prior homing step. Both primary (17 weeks) and secondary (10 weeks) reci pients of "spleen-homed" cells also contained approximately 50% higher numb ers of CFCs per femur than recipients of "BM-homed" cells. To examine wheth er progenitor homing was altered upon ex vivo expansion, highly enriched Sc a-1(+)c-kit(+)Lin(-) cells were cultured for 9 days in serum-free medium co ntaining interleukin (IL)-6, IL-11, granulocyte colony-stimulating factor, stem cell factor, flk-2/flt3 ligand, and thrombopoietin. Expanded cells wer e then stained with PKH26 and assayed as above. Strikingly, CFCs generated in vitro exhibited a 10-fold reduction in homing capacity compared with fre sh progenitors. These studies demonstrate that clonogenic cells with differ ential homing properties contribute variably to early and late hematopoiesi s in vivo. The dramatic decline in the homing capacity of progenitors gener ated in vitro underscores critical qualitative changes that may compromise their biologic function and potential clinical utility, despite their effic ient numerical expansion. (C) 1999 by The American Society of Hematology.