Tumor necrosis factor-α increases Mn-SOD expression: protection against oxidant injury

Antioxidant enzymes, including superoxide dismutase, are important for protecting the lung against O₂ injury. Manganese superoxide dismutase (Mn-SOD) is a superoxide anion (O₂^-·) scavenger located in the mitochondria, a primary site of O₂^-· production during hyperoxia. We studied the effects of tumor necrosis factor (TNF-α), a macrophage-derived cytokine, on Mn-SOD expression in human pulmonary adenocarcinoma cells. TNF-α significantly increased Mn-SOD activity and mRNA in a dose- and time-dependent manner. Mn-SOD activity was increased 3-fold and mRNA 20-fold after a 48-h incubation with TNF-α (25 ng/ml). To examine the mechanism of this increase, cells were incubated for 48 h with TNF-α (25 ng/ml) with or without cycloheximide (10 μM) or actinomycin D (10 μg/ml). Actinomycin D blocked the induction of Mn-SOD mRNA by TNF-α, but cycloheximide did not. These findings suggest that the effect of TNF-α requires gene transcription but not synthesis of new protein intermediates. To test the hypothesis that increased Mn-SOD protects against oxidative injury, pulmonary adenocarcinoma cells were incubated in TNF-α (25 ng/ml) for 48 h and then exposed to paraquat (PQ⁺), an intracellular O₂^-· generator. Cells pretreated with TNF-α had significantly improved survival in PQ⁺ compared with controls. At the LD₅₀ (6 μM) for control cells, 95% of TNF-α-treated cells survived, 85% at the LD₇₅ (10 μM), and 77% at the LD₉₀ (14 μM). Our results suggest that the induction of Mn-SOD by TNF-α in pulmonary adenocarcinoma cells is pretranslationally mediated and that increasing Mn-SOD activity with TNF-α confers protection against O₂ radicals.