The proposed studies are based on the hypothesis that cellular and organ aging are, to a significant degree, the consequences of oxygen free radical-mediated mitochondrial aging or impairment resulting from the accumulation of mitochondrial DNA mutations. Despite much indirect evidence, there is, as yet, no direct proof of the mitochondrial hypothesis of aging. However, the ability to modulate the activity of Mn superoxide dismutase (MnSOD) within mitochondria by genetic means makes it possible to assess the role of oxygen free radicals generated by the mitochondria in the aging process. To achieve this modulation of MnSOD activity, mutant mice completely lacking in MnSOD and mice with a low level of MnSOD expression from a transgene bred into the MnSOD deficient stock will be used. The finding that fetal fibroblasts lacking MnSOD are sensitive to atmospheric oxygen and have a shorter replicative life span than normal cells indicates that intramitochondrial superoxide levels can be affected by alteration of MnSOD activity. Mutant mice lacking MnSOD quickly develop a dilated cardiomyopathy and metabolic acidosis and die within 10 days after birth. However, in Specific Aim I, chimeras composed of cells completely lacking MnSOD and wild type cells will be formed, and the survival of the MnSOD-deficient cells exposed to a high level of superoxide will be followed in several organs and in the brain. In this manner it will be possible to assess the vulnerability of different types of cells and of different regions of the brain to free- radical induced mitochondrial damage.
In Specific Aim II, the effects of chronically increased exposure to intra-mitochondrial superoxide will be studied in appropriately constructed transgenic animals with 10-30% of normal MnSOD activity. The longevity of these animals will determined, and the long term effects on mitochondrial DNA and the electron transport system will be assessed. In addition, behavioral studies will be carried out to determine whether any changes observed in the brain are correlated with alterations in behavior. These studies represent a new and innovative approach to the direct examination of the role of oxygen free radicals in producing mitochondrial mutations and dysfunction and, in turn, organ and organismal aging. Evidence that mitochondrial free radicals are indeed involved in the aging process would constitute a basis for searching for therapeutic approaches to the modulation of oxygen free radical levels within mitochondria.
| Huang, Ting-Ting; Raineri, Ines; Eggerding, Faye et al. (2002) Transgenic and mutant mice for oxygen free radical studies. Methods Enzymol 349:191-213 |
| Friedman, J S; Rebel, V I; Derby, R et al. (2001) Absence of mitochondrial superoxide dismutase results in a murine hemolytic anemia responsive to therapy with a catalytic antioxidant. J Exp Med 193:925-34 |
