The caspase inhibitor z-VAD-fmk induces a switch from differentiation to death.

(A,B) U937 cells were exposed to 20 nM TPA in the presence of indicated concentrations of z-VAD-fmk for 48 hours (A) or 100 μM z-VAD-fmk for indicated times (B, ●) before counting the percentage of cells stained with trypan blue (○ indicates U937 cells exposed to 100 μM z-VAD-fmk in the absence of TPA). (C) U937 cells were either left untreated or treated with TPA (20 nM, 48 hours), z-VAD-fmk (100 μM, 48 hours), or the TPA/z-VAD-fmk combination. The percentages of cells labeled with annexin V–FITC (left panels) and propidium iodide (PI; right panels) were measured by flow cytometry. (D) Western blot analysis of poly(ADP-ribose)polymerase (PARP) in cells treated as above or with etoposide (VP16; 50 μM, 6 hours). HSC70 expression was used for checking loading. (E) Radical oxygen species measured by flow cytometry in cells exposed for indicated times to 20 nM TPA, in the absence (white bars) and presence (black bars) of 100 μM z-VAD-fmk. NAC indicates cotreatment with 25 mM N-acetyl-cysteine for 48 hours. Mean ± SD of 3 experiments in triplicate (A,B,E) or 1 representative of 3 independent experiments (C,D).