EXPRESSION OF FAS CD95 AND BCL-2 BY PRIMITIVE HEMATOPOIETIC PROGENITORS FRESHLY ISOLATED FROM HUMAN FETAL LIVER/

The cell-surface expression and the functional status of the CD95/Fas antigen on primitive hematopoietic progenitors PHPs) freshly isolated from human fetal liver (FL) were studied. PHPs were phenotypically def ined as CD34(++)CD38(-/+) cells. The most immature subfractions of PHP s, CD34(++)CD38(-) and CD34(++)CD38(+) FL cells, expressed CD95, where as the more mature CD34(++)CD38(++) and CD34(+)CD38(++) FL cells displ ayed low CD95 expression. Combinations of cytokines, such as kit ligan d (KL) + interleukin-3 or KL + granulocyte-macrophage colony-stimulati ng factor (GM-CSF) upregulated the expression of CD95 on PHPs upon in vitro culture. Tumor necrosis factor-alpha (TNF-alpha) and interferon- gamma (IFN-gamma) further increased the CD95 expression induced by KL + GM-CSF. The hematopoietic potential of sorted CD34(++)lineage (lin)( -) CD95(+) versus CD34(++)lin(-)CD95(-) FL cells was compared by colon y-forming unit-culture (CFU-C) assays performed in serum-deprived medi um. Lin(+) cells were composed of erythrocytes, monocytes, T cells, B cells, and natural killer cells. Our results indicated that both CD95( -) and CD95(+) subsets contained pluripotent progenitors, generating m yeloid and erythroid progenitors. The functional status of CD95 and th e effects of TNF-alpha and IFN-gamma, cytokines known to induce CD95-m ediated apoptosis, were analyzed by incubation of PHPs in the presence of anti-CD95 monoclonal antibodies (MoAbs). The effect of anti-CD95 M oAbs was measured by viable cell counting, flow cytometry, and CFU-C a ssays. A decrease of CFU-C numbers was observed in the presence of ant i-CD95 MoAbs and TNF-alpha and/or IFN-gamma. However, whereas growth f actor deprivation induced apoptosis of PHPs, cross-linking of CD95 did not lead to apoptosis of PHPs measured by flow cytometry and viable c ell counting. The correlation of increased intracytoplasmic levels of bcl-2 with high levels of cell-surface CD34 and the presence of CD95 o n fresh FL cells suggests that bcl-2 may be involved in protecting aga inst CD95-mediated apoptosis of FL PHPs. (C) 1996 by The American Soci ety of Hematology.