Influence of FADD constructs on the differentiation pathway. (A-B) U937 cells were infected with lentiviral constructs encoding either EGFP alone (T or ○) or wild-type FADD (WT or ▪) or mutated FADD in which the death effector domain has been partly deleted (DN or •), then selected on EGFP expression by cell sorting. (A) Expression of FADD (24 kDa) and FADD-DN (17 kDa) was analyzed by immunoblotting. HSC70 indicates loading control. Molecular weights are in kilodaltons. Cells were either left untreated or treated with 100 ng/mL CH11 Fas antibody plus 0.8 μg/mL CHX for 6 hours before measuring the percentage of apoptotic cells after Hoechst 33352 staining of the nuclear chromatin. (B) Cells were exposed to 20 nM TPA for indicated times before measuring CD11b expression by flow cytometry. (C) Monocytes were infected with lentiviral constructs encoding either EGFP alone (▪) or wild-type FADD (□) and treated for 4 days as in Figure 1 to induce their differentiation into macrophages (Mac) or dendritic cells (DC) before flow cytometry analysis of the cell-surface expression of CD71 or CD1a. Results are normalized to EGFP-infected monocytes. Results are the mean ± SD of at least 3 independent experiments. **P < .01; ***P < .005; NS indicates not significant.