Reactive ,-unsaturated aldehydes are major components of common environmental pollutants and are products of lipid oxidation. While these aldehydes have been demonstrated to induce apoptotic cell death in various cell types, we recently observed that the ,-unsaturated aldehyde acrolein (ACR) can inhibit constitutive apoptosis of polymorphonuclear neutrophils, and thus potentially contribute to chronic inflammation. The present study was designed to investigate the biochemical mechanisms by which two representative ,-unsaturated aldehydes, ACR and 4-hydroxynonenal (HNE), regulate neutrophil apoptosis. While low concentrations of either aldehyde (< 10 µM) mildly promoted apoptosis in neutrophils (reflected by increased phosphatidylserine exposure, caspase-3 activation, and mitochondrial cytochrome c release), higher concentrations prevented critical features of apoptosis (caspase-3 activation, phosphatidylserine exposure), and caused delayed neutrophil cell death with characteristics of necrosis/oncosis. Inhibition of caspase-3 activation by either aldehyde occurred in spite of increases in mitochondrial cytochrome c release, and occurred in close association with depletion of cellular GSH and with cysteine modifications within caspase-3. However, pro-caspase-3 processing was also prevented, due to inhibited activation of caspases -9 and -8 under similar conditions, suggesting that ACR (and to a lesser extent HNE) can inhibit both intrinsic (mitochonchria-dependent) and extrinsic mechanisms of neutrophil apoptosis at initial stages. Collectively, our results indicate that ,-unsaturated aldehydes can inhibit constitutive neutrophil apoptosis by common mechanisms, involving changes in cellular GSH status resulting in reduced activation of initiator caspases as well as inactivation of caspase-3 by modification of its critical cysteine residue.