Activators of protein kinase C selectively mediate cellular cytotoxicity to hypoxic cells and not aerobic cells

Purpose: By understanding the signal transduction pathways through which a cell responds to changes in environmental oxygen levels, we may be able to therapeutically exploit this response by manipulating these pathways. Materials and Methods: The human adenocarcinoma cell line A549 was exposed to varying durations of hypoxia alone and then plated for survival, or treated with PKC activating agents for 1 h before plating for survival. Western blots were used to determine the kinetics of PKC epsilon and phospholipase C induction. Results: The level of hypoxic killing was directly related to the time of exposure and inversely related to the level of oxygen in the environment. Exposure of the cells to protein kinase C (PKC) activators for 1 h after chronic hypoxic exposure increased cell killing by at least an additional three logs beyond that found for hypoxia alone. Treatment of cells with an inactive phorbol ester 4α-phorbol-12,13-didecanoate (PDA) resulted in no increase in hypoxic cell killing, even at the highest concentrations of PDA which produced no detectable toxicity under normal aerobic conditions. Using inhibitors of phospholipases A₂ and C, we were able to completely inhibit the additional hypoxic cell killing induced by TPA, but not the uninduced hypoxic cell killing. Conclusion: These studies suggest that accumulation of phospholipid breakdown products may be responsible for TPA induced cell killing, and that hypoxic cells differ from aerobic cells in their ability to tolerate these products.