The ex vivo expansion of feline marrow cells leads to increased numbers of BFU-E and CFU-GM but a loss of reconstituting ability.
Some studies in mice suggest that hematopoietic stem cells can be maintained and possibly expanded ex vivo. As there is a paucity of data from larger animals, we have studied hematologic reconstitution following autologous marrow transplantation in cats. Transplantation of very low density marrow cells (<1.050 g/ml), termed "1050 cells," at 2 x 10(5) cells/kg leads to rapid hematopoietic recovery (granulocytes >200/microl by day 20+/-2 and platelets >50 x 10(3)/microl by day 21+/-3). Recovery rates are comparable when 1-2 x 10(7) nucleated marrow cells/kg are infused, suggesting that reconstituting cells are enriched 50- to 100-fold in the 1050 cell preparation. To explore if the numbers of reconstituting cells could be expanded ex vivo, 1050 cells were cultured in the presence of 5 ng/ml recombinant human interleukin 1beta, 10 ng/ml recombinant canine (rc)G-CSF, 2 U/ml rHu erythropoietin, and 5 ng/ml rc stem cell factor. Maximum numbers of BFU-E and colony-forming units-granulocyte/macrophage (CFU-GM) were generated at day 6. However, when 10(6) 1050 cells/kg (5x that needed for hematologic recovery) were cultured for six days and all resulting cells infused into irradiated donor animals, two of nine (22%) engrafted. Even when flt3 ligand (100 ng/ml) was added to cultures, only two of five animals (40%) engrafted (p = NS versus studies without flt3 ligand). These data confirm that BFU-E and CFU-GM provide inaccurate estimates of reconstituting cells and demonstrate that the number or function of feline reconstituting cells is impaired by in vitro culture with cytokines.
Department of Medicine, University of Washington, Seattle 98195-7710, USA.
J L
JL
Abkowitz
M R
MR
Taboada
K M
KM
Sabo
G H
GH
Shelton
D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology* / Q000187:drug effects; D002415:Cats; D002452:Cell Count; D002478:Cells, Cultured; D002498:Centrifugation; D003114:Colony-Forming Units Assay; D015672:Erythroid Precursor Cells / Q000166:cytology*; D006098:Granulocytes / Q000166:cytology*; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000166:cytology; D006801:Humans; D008264:Macrophages / Q000166:cytology*; D008565:Membrane Proteins / Q000494:pharmacology
D008565:Membrane Proteins; C084881:flt3 ligand protein