Mn-Superoxide Dismutase Overexpression Enhances G2Accumulation and Radioresistance in Human Oral Squamous Carcinoma Cells

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Abstract
This study investigates the hypothesis that Mn-superoxide dismutase (MnSOD) influences cancer cell radiosensitivity by regulating the G2-checkpoint pathway. Human oral squamous carcinoma cells (SCC25) stably overexpressing MnSOD were irradiated (6 Gy) and assayed for cell survival, cell-cycle phase distributions, and bromodeoxyuridine (BrdU) pulse-chase flow-cytometric measurements of cell-cycle phase transits. Electron paramagnetic resonance (EPR) spectroscopy was used to measure steady-state levels of oxygencentered free radicals. Glutathione and glutathione disulfide levels were used as indicators of changes in the intracellular redox state. MnSOD overexpression increased radioresistance threefold to fourfold; this increase was associated with twofold to threefold increases in radiation-induced G2 accumulation. BrdU pulsechase and flow-cytometric measurements of the percentage of G1 and relative movement showed no significant changes in G1 and S transits; however, the percentage of G2 cells and BrdU-positive cells showed delayed G2+M transits in MnSOD-overexpressing irradiated cells. The steady-state levels of oxygen-centered free radicals were not significantly different in vector compared with MnSOD-overexpressing cells, suggesting that the free radical generation is essentially similar. MnSOD overexpression did prevent radiation-induced decreases in total glutathione content, which correlated with radioresistance and enhanced G2 accumulation. These results support the hypothesis that a "metabolic redox-response" to IR exposure regulates radiosensitivity by altering radiation-induced G2 accumulation.