Abstract
Reactive oxygen species (ROS) act as a mediator of ionizing radiation-induced cellular damage. Previous studies have indicated that MnSOD (SOD2) plays a critical role in protection against ionizing radiation in mammalian cells. In our study, we focused on superoxide dismutase (SOD2) and glutaredoxin (Grx2) proteins. It is well known that SOD eliminate superoxide anion, Grx reduces the oxidized macromolecules, and both SOD and Grx are conserved from bacteria to human. Therefore, SOD and Grx are essential enzymes in cellular homeostasis. In this study, we constructed two types of stable HeLa cell lines overexpressing SOD2, HeLa S3/SOD2 and T-REx HeLa/SOD2, to elucidate the mechanisms underlying the protection against radiation by SOD2. SOD2 overexpression in mitochondria enhanced the survival of HeLa S3 and T-REx HeLa cells following γ-irradiation. The levels of γH2AX significantly decreased in both cells compared with those in the control cells. MitoSoxTM Red assay revealed that both lines of SOD2-expressing cells showed suppression of the superoxide generation in mitochondria. Flow cytometry with a fluorescent probe revealed that the cellular levels of ROS increased in HeLa S3 cells during post-irradiation incubation, but the increase was markedly attenuated in HeLa S3/SOD2 cells. Furthermore, we examined whether and how cellular sensitivity to ionizing radiation is modulated by the overexpression of mitochondrial Grx (Grx2) in the cultured human cells. As the results of overexpression of SOD2 and Grx2 in T-REx HeLa cells, morphological alterations of mitochondria and levels of mitochondrial superoxide, DNA double-stranded breaks, protein oxidation and OXR1 expression were suppressed. These results suggest that antioxidant enzymes in mitochondria play important roles in various cellular responses to ionizing radiation.
