PHAGOCYTES Caspase-Mediated Proteolysis and Activation of Protein Kinase Cd Plays a Central Role in Neutrophil Apoptosis

Neutrophils undergo constitutive apoptosis when aged ex vivo. Recent studies have indicated roles for Fas/CD95 and the nicotinamide adenine dinucleotide phosphate (NADPH)oxidase system in this process. We have investigated the role of protein kinase C (PKC) in neutrophil death. We show that there is proteolysis and activation of the novel isoform PKCd in aged neutrophils and that this process is accelerated by the addition of an agonistic Fas antibody. PKCd proteolysis occurs before the onset of any detectable features of apoptosis and pharmacologic inhibition of this enzyme inhibits neutrophil apoptosis. PKCd cleavage and activation is dependent on caspase-8/FADD-like interleukin-1b converting enzyme (FLICE)–mediated processing of caspase-3/ CPP32. Neutrophil survival is prolonged by the addition of broad spectrum (BD.fmk) or caspase-8 targeted (zIETD.fmk) peptide caspase inhibitors. Inhibition of PKCd does not prevent apoptosis triggered by factor withdrawal in immature hematopoietic cells, including normal human CD341 progenitors indicating that within a given lineage, the mechanisms of apoptosis may be differentiation-stage–specific. Ex vivo aging of neutrophils leads to the increasing production of reactive oxygen species and this is attenuated in cells treated with either caspase or PKCd inhibitors. Proteolytically activated PKCd acts as a molecular link between the Fas/CD95 receptor and the NADPH-oxidase system and plays a central role in regulating the process of neutrophil apoptosis. r 1999 by The American Society of Hematology.