The role of poly (ADP-ribose) polymerase in necrosis and apoptotic-like delayed neuronal death after acute hydrogen peroxide injury

Reactive oxygen species are postulated to be the cause of, or mediators of, neuronal death associated with neurodegenerative diseases, brain trauma, epilepsy, hypoxia, hyperoxia, and ischemia/reperfusion injury. DNA is a major macromolecular target of these ROS, and ROS-induced DNA strand breaks activate poly (ADP-ribose) polymerase-1 (PARP-1). Upon activation PARP-1 uses NAD + as a substrate to catalyze the transfer of ADPribose subunits to a host of nuclear proteins. In the face of extensive DNA strand breaks, PARP-1 activation can lead to depletion of intracellular NAD(P)(H) pools, large decreases in ATP, and can threaten cell survival. Accordingly, studies show that lack of PARP-1 activity after acute oxidative injury leads to notable increases in cell survival. The current study was undertaken to determine the role of H2O2-induced PARP-1 activation in the two distinct types of cell death, apoptosis and necrosis, in differentiated PC12 cells---a model of sympathetic neurons. The results revealed that H2O 2 exposure resulted in an immediate synthesis of poly ADP-ribose and decreases in intracellular NAD(P)(H) and ATP. Addition of the chemical PARP inhibitor 3-aminobenzamide (AB) prior to H2O2 exposure blocked the synthesis of poly ADP-ribose and helped maintain intracellular NAD(P)(H) and ATP levels. H2O2 injury was characterized by an immediate, necrotic cell death 2 hours after injury and a delayed apoptotic-like death 12--24 hours after injury. This apoptotic-like death was characterized by apoptotic membrane changes and apoptotic DNA fragmentation but was not associated with measurable caspase-3 activity. AB delayed cell death beyond 24 hours and increased cell survival by ∼25%. This increase in cell survival was accompanied by significantly decreased necrosis and the apoptotic-like death associated with H2O2 exposure. AB also restored caspase-3 which could be attributed to the activation of the upstream activator of caspase-3, caspase-9. Taken together, these results suggest that the maintenance of intracellular ATP levels associated with PARP-1 inhibition shifts cell death from necrosis to apoptosis and from apoptosis to cell survival. Furthermore, the shift from necrosis to apoptosis may be explained, in part, by the energy-dependent activation of caspase-9.