High concentrations of z-VAD-fmk augment TNFα-stimulated neutrophil apoptosis. Human neutrophils were incubated in the presence of (A) z-VAD-fmk, (B) Boc-D-fmk, (C) z-IETD-fmk (caspase 8 inhibitor), or (D) Ac-LEHD-cmk (caspase 9 inhibitor) at the concentrations shown or vehicle control for 30 minutes prior to the addition of 200 U/mL TNFα (▴) or PBS (▪). The cells were then incubated for 6 hours and percent apoptosis determined by annexin V-FITC staining. (E) A representative experiment showing z-VAD-fmk inhibition of TNFα-induced apoptosis at low inhibitor concentrations (30 μM) and the augmentation of cell death at the higher concentrations (300 μM). ^*P < .05 (significant inhibition of TNFα-stimulated apoptosis), #P < .05 (significant augmentation of TNFα-stimulated apoptosis). All values are mean ± SEM of n = 3-5 separate experiments, each performed in triplicate. Numbers in quadrants indicate percentage of cells.