Caspase-mediated RIP1 and FLIP cleavage in cells undergoing macrophagic differentiation. (A) Peripheral-blood monocytes (Mo) were treated as in Figure 1 for indicated times (days) to trigger their differentiation into macrophages (Mac) or dendritic cells (DC) before analyzing the expression of indicated proteins by immunoblotting. (B) U937 cells transfected with either an empty vector (Co) or a mutated caspase-8–expressing vector (C8-DN) were treated with 20 nM TPA for indicated times (hours) before immunoblot analysis of indicated proteins. (C-D) U937 cells transduced with a lentivirus encoding an empty vector (ΔMCS or ▪) or RIP1 mutated on the caspase cleavage site (RIPm or □) were exposed to TPA as in panel B before (C) immunoblot analysis of indicated proteins. RIP1 cleavage fragment was normalized to FADD expression (fold increase). *Cleavage fragment. (D) CD11b expression measured by FACS analysis. One representative of at least 3 independent experiments is shown, or mean ± SD of 3 independent experiments. **P < .01.