Broad-spectrum caspase inhibitors but not a cathepsin L–selective inhibitor sensitize neutrophils to TNF-α–induced cell death.

(A) Cleavage of the caspase-3–preferred fluorogenic substrate, Ac-DEVD-AMC, was measured (see “Materials and methods”) in whole-cell lysates prepared from neutrophils preincubated with or without zVAD (100 μM) for 1 hour and then stimulated with or without TNF-α (10 ng/mL) for 2 additional hours. Results shown represent means ± SEM of 3 experiments. *P < .05 compared with unstimulated neutrophils at 2 hours. (B) Neutrophils were preincubated with zVAD (100 μM) or Boc-D (100 μM) or the cathepsin L–selective inhibitor zFF (100 μM) for 1 hour before the addition of TNF-α (10 ng/mL) or an equivalent volume of buffered saline at time 0. Cellular binding of annexin V–FITC (AV⁺) was subsequently assayed at 6 hours with flow cytometry. Results are means ± SEM of 3 independent experiments. *P < .05 compared with neutrophils stimulated with TNF-α alone.