Stem cell factor prevents Fas-mediated apoptosis of human erythroid precursor cells with Src-family kinase dependency

Objective. The Pas ligand (Fas-L) expressed on mature erythroblasts may induce apoptosis of more immature erythroid cells that express Pas, whereas stem cell factor (SCF) may prevent Fas-mediated cell death in hematopoietic progenitor cells, The manner in which SCF prevents Fas-mediated cell death still is unclear. Given the essential role of SCF and the potentially important involvement of the Fas/Fas-L system in the development of erythrocytes, we studied mechanisms related to SCF prevention of Fas-mediated apoptosis, Materials and Methods. We used primary cultured human erythroid colony-forming cells (ECFC) derived from CD34(+) cells and enriched glycophorin A positive (GPA(+)) c-kit(+) cells in ECFC, Apoptosis of ECFC was induced by an Fas-L mimetic monoclonal antibody CH11. DNA fragmentation and the activation of caspase-3 and caspase-8 were measured using commercially available kits, Characterization of expanded cells was performed using multiparameter flow cytometry, Lyn kinase activity was measured by enolase kinase assays, Results. SCP inhibited the CH11-induced DNA fragmentation of ECFC as well as enriched GPA(+) c-kit(+) cells in ECFC, but not those of GPA(+) c-kit(-) cells. SCF also inhibited the activation of caspase-3 and caspase-8, without downregulation of the surface expression of Fas, suggesting that SCF prevents apoptosis through uncoupling of Pas ligation from subsequent caspase activation, PP2, a specific inhibitor of Src-family kinases, antagonized the effects of SCF in preventing Pas-mediated apoptosis. Conclusions. We propose that SCF prevents Fas-mediated apoptosis of erythroid progenitor cells in a manner dependent on the activity of Src-family tyrosine kinases, We also identified active Lyn in erythroid cells, These data suggest the presence of a novel Src-family-dependent function of SCF in the development of erythrocytes. (C) 2001 International Society for Experimental Hematology. Published by Elsevier Science Inc.