Reactive Oxygen Species Participate in Peroxynitrite-Induced Apoptosis in HL-60 Cells

Abstract

Peroxynitrite (ONOO−) is a physiological product generated by the interaction of superoxide (O.−2) and nitric oxide (.NO). We have previously shown that peroxynitrite induces apoptosis in HL-60 cells. In the present study, we demonstrated that peroxynitrite generates reactive oxygen species (ROS) in HL-60 cells. Brief exposure of HL-60 cells to ONOO−induced elevation of lucigenin chemiluminescence, indicating generation of superoxide anion. Exogenous superoxide dismutase (SOD), a scavenger of O.−2, fully abolished the chemiluminescence response, further supporting this notion. Following O.−2generation, the accumulation of hydrogen peroxide (H2O2) was observed. The addition of SOD exacerbated but that of catalase attenuated peroxynitrite-induced DNA fragmentation, suggesting that this H2O2production contributes to the apoptotic process. In addition, pre-treatment of HL-60 cells with N-acetyl-L-cysteine (15 mM), a ROS scavenger, fully scavenged peroxynitrite-elicited ROS generation and effectively inhibited ONOO−-induced apoptosis, further enforcing this hypothesis. In summary, our results suggest that ONOO−-stimulated ROS formation may serve as a mechanism for the propagation of peroxynitrite-mediated apoptotic cell death in an intact cell system.