ROS regulate neutrophil apoptosis and CD95 clustering. (A) Apoptosis was delayed by inhibitors of oxygen radical formation. Neutrophils were cultured with 100 μM N-acetylcysteine or 10 μM desferrioxamine for 18 hours prior to assessment of apoptosis by cell morphology. Generation of ceramide and clustering of CD95 on aging neutrophils depends on the generation of hydroxyl radicals. Neutrophils were cultured with and without desferrioxamine, stained for ceramide (B) or CD95 and cells with clustered CD95 enumerated (C). (D) Intracellular levels of GSH were reduced within 3 to 7 hours of neutrophil culture to one third of the original level. Data for panels A, C, and D are mean ± SD of 3 separate experiments. *P < .05.