Fas Ligand Is Present in Human Erythroid Colony-Forming Cells and Interacts With Fas Induced by Interferon g to Produce Erythroid Cell Apoptosis

Interferon g (IFNg) inhibits the growth and differentiation of highly purified human erythroid colony-forming cells (ECFCs) and induces erythroblast apoptosis. These effects are doseand time-dependent. Because the cell surface receptor known as Fas (APO-1; CD95) triggers programmed cell death after activation by its ligand and because incubation of human ECFCs with IFNg produces apoptosis, we have investigated the expression and function of Fas and Fas ligand (FasL) in highly purified human ECFCs before and after incubation with IFNg in vitro. Only a small percentage of normal human ECFCs express Fas and this is present at a low level as detected by Northern blotting for the Fas mRNA and flow cytometric analysis of Fas protein using a specific mouse monoclonal antibody. The addition of IFNg markedly increased the percentage of cells expressing Fas on the surface of the ECFCs as well as the intensity of Fas expression. Fas mRNA was increased by 6 hours, whereas Fas antigen on the cell surface increased by 24 hours, with a plateau at 72 hours. This increase correlated with the inhibitory effect of IFNg on ECFC proliferation. CH-11 anti-Fas antibody, which mimics the action of the natural FasL, greatly enhanced IFNgmediated suppression of cell growth and production of apoptosis, indicating that Fas is functional. Expression of FasL was also demonstrated in normal ECFCs by reverse transcriptase-polymerase chain reaction and flow cytometric analysis with specific monoclonal antibody. FasL was constitutively expressed among erythroid progenitors as they matured from day 5 to day 8 and IFNg treatment did not change this expression. Apoptosis induced by IFNg was greatly reduced by the NOK-2 antihuman FasL antibody and an engineered soluble FasL receptor, Fas-Fc, suggesting that Fas-FasL interactions among the ECFCs produce the erythroid inhibitory effects and apoptosis initiated by IFNg. r 1998 by The American Society of Hematology.