The reduction of in vitro radiation-induced Fas-related apoptosis in CD34(+) progenitor cells by SCF, FLT-3 ligand, TPO, and IL-3 in combination resulted in CD34(+) cell proliferation and differentiation

Recovery from radiation-induced (RT) bone marrow aplasia depends on appropr iate cytokine support. The early effects of exogenous cytokines at the hema topoietic stem and progenitor cell (HSPC) level following irradiation are s till largely unknown, especially those of survival factors such as stem cel l factor (SCF) and Flt-3 ligand (FL). This study was aimed at A) clarifying Fas/Fas-Ligand (Fas-L) implication in RT apoptosis of CD34(+) cells and B) assessing the capacity of a combination of cytokines to mitigate RI apopto sis in HSPCs in vitro, We showed that most of in vitro gamma-irradiated CD3 4(+) HSPCs incubated in a medium devoid of cytokines underwent progressive apoptosis-related changes from 6 h (i.e., decreased CD34 antigen expression , Annexin V binding); then Fas/Fas-L coexpression occurred from 10 h on, A strong DNA fragmentation, as assessed by TUNEL assay and propidium iodide s taining, was observed at 24 h, Within a 2.5- to 6-Gy dose range, the RI apo ptotic process finally led to 97% CD34(+) cell death within 48 h with a com plete loss of functionality. Unirradiated cells incubated in the same condi tions displayed a significantly reduced apoptotic pattern, The early additi on of a combination of SCP, FL, thrombopoietin, and interleukin 3 (4F) afte r tell irradiation prevented 15% (2.5 Gy) and 12% (4 Gy) of HSPCs, respecti vely, from RI apoptosis, whereas these cytokines used as single factors wer e inefficient, Furthermore, irradiated HSPCs (2.5 Gy) incubated with 4F in a serum-free culture system for seven days proliferated, giving rise to an increase in the number of total cells (x 5.6-fold) and CD34(+) cells (x 4.2 -fold) and to megakaryocytic and granulomonocytic precursors, These results show that the prevention of apoptosis in in vitro irradiated HSPCs depends on an early combination cytokine support, These data suggest that the earl y therapeutic administration of anti-apoptotic cytokines may be critical fo r preserving functional HSPCs from in vivo radiation damage.