The objective of this study was to determine the effects of aging and exercise training on SIRT1 activity and to identify a pathway linking SIRT1 to antioxidant response and cell cycle regulation in rats. SIRT1 is a NAD(+)-dependent deacetylase involved in the oxidative stress response and aging. The effects of aging and of moderate and prolonged exercise training in rats are unknown. We measured SIRT1 activity in heart and adipose tissue of young (6 months old), sedentary old (24 months), and trained old (24 months) rats using an assay kit. Peroxidative damage was determined by measuring levels of thiobarbituric reactive substances (TBARS) and the protein-aldehyde adduct 4-hydroxynonenal (4-HNE). MnSOD, catalase, and FOXO3a levels were evaluated by Western blot, and GADD45a, cyclin D(2), and FOXO3a mRNA by RT-PCR. Aging significantly reduced SIRT1 activity in heart, but not in adipose tissue, increased TBARS and 4-HNE and decreased Mn-SOD and catalase expression in both heart and adipose tissue. Aging did not affect FOXO3a protein expression in the heart or FOXO3a mRNA in adipose tissue. Exercise training significantly increased FOXO3a protein in the heart and FOXO3a mRNA in adipose tissue of aged rats. It also significantly increased Mn-SOD and catalase levels in both heart and adipose tissue. The exercise-induced increase in SIRT1 activity in the heart caused a decrease in cyclin D(2) and an increase in GADD45a mRNA expression. There was a similar decrease in cyclin D(2), and no changes in GADD45a mRNA expression in adipose tissue. We concluded that exercise training, which significantly increases SIRT1 activity, could counteract age-related systems impairment.

No abstract

Renal toxicity produced by paraquat involves the generation of reactive oxygen species (ROS) which can overwhelm antioxidant defences, leading to oxidant injury. However, there are conflicting reports regarding the activity and/or expression of antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) during oxidative stress injury. This study investigated the activity and expression of these enzymes in a renal epithelial cell line following exposure to paraquat. Confluent NRK-52E cells were incubated with increasing concentrations of paraquat (1-100mM) for up to 24 hours. Renal cell death was determined by measurement of lactate dehydrogenase release. Oxidant damage was determined via measurement of malondialdehyde formation and DNA strand breaks. The effects of paraquat on DNA and de novo protein synthesis were determined using radio-labelled thymidine and leucine respectively. ROS generation (superoxide anion and hydroxyl radical formation) was measured using nitrobluetetrazolium and deoxyribose assays. Antioxidant enzyme activities and expression were measured using established biochemical assays and Western blot analysis. Exposure of confluent NRK-52E cells to paraquat resulted in significant cell death involving increased lipid peroxidation, DNA damage and inhibition of DNA and de novo protein synthesis. Renal cell injury and death were secondary to increased ROS generation. Incubation with paraquat reduced SOD and CAT activities; in contrast, GSH-Px activity increased significantly. Although SOD expression was significantly reduced, catalase expression was unaffected. These results indicate that paraquat mediates renal toxicity via oxidative stress involving both an increase in ROS generation and reductions in SOD and CAT activities with a concomitant reduction in SOD expression.