PEROXYNITRITE INDUCES APOPTOSIS OF HL-60 CELLS BY ACTIVATION OF A CASPASE-3 FAMILY PROTEASE

Apoptosis is an active process critical for the homeostasis of organis ms. Enzymes of the caspase family are responsible for executing this p rocess. We have previously shown that peroxynitrite (ONOO-), a biologi cal product generated from the interaction of nitric oxide and superox ide, induces apoptosis of HL-60 cells. The aim of this study was to el ucidate the mechanisms involved in the execution process of peroxynitr ite-induced apoptosis. Proteolytic cleavage of poly(ADP-ribose) polyme rase, an indication of caspase-3 family protease activation and an ear ly biochemical event accompanying apoptosis, was observed in a time-de pendent manner during peroxynitrite-induced apoptosis of HL-60 cells. Activation of caspase-3 during peroxynitrite-induced apoptosis was sub stantiated by monitoring proteolysis of the caspase-3 proenzyme and by measuring caspase-3 activity with a fluorogenic substrate. Furthermor e, pretreatment of HL-60 cells with N-acetyl-Asp-Glu-Val-Asp-aldehyde, a specific inhibitor of caspase-3, but not N-acetyl-Tyr-Val-Ala-Asp-a ldehyde, a specific inhibitor of caspase-1, decreased peroxynitrite-in duced apoptosis. These results suggest that the activation of a caspas e-3 family protease is essential for initiating the execution process of peroxynitrite-induced apoptosis of HL-60 cells.