Activation of phospholipase D involved in both injury and survival in A549 alveolar epithelial cells exposed to H2O2

To determine the role of the phospholipase D (PLD) pathway in injury and survival of alveolar epithelial cells, A549 cells were exposed to H₂O₂ (500 μM) which resulted in time-dependent injury and bi-phasic increase of PLD activity at 5 min and at 3 h, respectively. n-Butanol (0.5%) inhibited PLD activation, attenuated cell injury at 5 min of H₂O₂ exposure, but enhanced injury at 3 h of exposure. This activation was inhibited by treatment with catalase (500 units/ml). Exogenous phosphatidic acid mimicked the effects of PLD activation, and diphenyliodonium (NADPH oxidase inhibitor) reversed the decline in cell viability induced by H₂O₂ exposure. Propranolol (phosphatidic acid phospholydrolase inhibitor) and quinacrine (phospholipase A2 inhibitor) had weak effects on H₂O₂-induced PLD activation but reversed H₂O₂-induced injury. We speculate that PLD activation at the initiation of H₂O₂ exposure predominantly results in NAPDH oxidase activation, which mediates A549 cell injury, but turns to mediating cell survival as the H₂O₂ attack continues, which might be mainly due to the accumulation of intracellular phosphatidic acid.