Nitration of tryptophan 372 in succinyl-CoA:3-ketoacid CoA transferase during aging in rat heart mitochondria.

The main objective of this study was to test the hypothesis that in vivo post-translational modifications in proteins, induced by the endogenously generated reactive oxygen and nitrogen molecules, can alter protein function and thereby have an effect on metabolic pathways during the aging process. Succinyl-CoA:3-ketoacid coenzyme A transferase (SCOT), the mitochondrial enzyme involved in the breakdown of ketone bodies in the extrahepatic tissues, was identified in rat heart to undergo age-associated increase in a novel, nitro-hydroxy, addition to tryptophan 372, located in close proximity ( approximately 10 A) of the enzyme active site. Between 4 and 24 months of age, the molar content of nitration was more than doubled while specific enzyme activity increased significantly. The amount of SCOT protein, however, remained unchanged. In vitro treatment of heart mitochondrial soluble proteins with relatively low concentrations of peroxynitrite enhanced the nitration as well as specific activity of SCOT. Results of this study identify tryptophan to be a specific target of nitration in vivo, for the first time. We hypothesize that increases in tryptophan nitration of SCOT and catalytic activity constitute a plausible mechanism for the age-related metabolic shift toward enhanced ketone body consumption as an alternative source of energy supply in the heart.

Effect of coenzyme Q10 intake on endogenous coenzyme Q content, mitochondrial electron transport chain, antioxidative defenses, and life span of mice.

The main purpose of this study was to determine whether intake of coenzyme Q10, which can potentially act as both an antioxidant and a prooxidant, has an impact on indicators of oxidative stress and the aging process. Mice were fed diets providing daily supplements of 0, 93, or 371 mg CoQ10 /kg body weight, starting at 3.5 months of age. Effects on mitochondrial superoxide generation, activities of oxidoreductases, protein oxidative damage, glutathione redox state, and life span of male mice were determined. Amounts of CoQ9 and CoQ10, measured after 3.5 or 17.5 months of intake, in homogenates and mitochondria of liver, heart, kidney, skeletal muscle, and brain increased with the dosage and duration of CoQ10 intake in all the tissues except brain. Activities of mitochondrial electron transport chain oxidoreductases, rates of mitochondrial O2-* generation, state 3 respiration, carbonyl content, glutathione redox state of tissues, and activities of superoxide dismutase, catalase, and glutathione peroxidase, determined at 19 or 25 months of age, were unaffected by CoQ10 administration. Life span studies, conducted on 50 mice in each group, showed that CoQ10 administration had no effect on mortality. Altogether, the results indicated that contrary to the historical view, supplemental intake of CoQ10 elevates the endogenous content of both CoQ9 and CoQ10, but has no discernable effect on the main antioxidant defenses or prooxidant generation in most tissues, and has no impact on the life span of mice.

Effect of age and caloric restriction on coenzyme Q and alpha-tocopherol levels in the rat.

Alterations in the amount of coenzyme Q and alpha-tocopherol during aging and in response to 40% reduction in caloric intake were determined in homogenates and mitochondria of liver, heart and kidney of the rat. A comparison among 4-, 19- and 28-month-old ad libitum fed (AL) rats indicated an age-related loss in the amount of CoQ9 and alpha-tocopherol in mitochondria of all the three tissues. Depletion of alpha-tocopherol, but not of CoQ, was also detectable in tissue homogenates, apparently due to the preferential sequestration of CoQ in the mitochondrial fraction. Comparison of 19-month-old AL and calorically restricted (CR) rats indicated that CR elevates the level of mitochondrial CoQ, but greatly diminishes the alpha-tocopherol content. Activity of DT-diaphorase, a quinone reductase, increased with age as well as in response to CR. Altogether, results are interpreted to suggest that the widely observed age-related increase in mitochondrial oxidative damage may be associated with depletion of CoQ and alpha-tocopherol, which are known to act in tandem to prevent oxidative damage to membranes.

Coenzyme Q intake elevates the mitochondrial and tissue levels of Coenzyme Q and alpha-tocopherol in young mice.

The main objective of this study was to resolve the issue of whether the amounts of Coenzyme Q (CoQ), which is endogenously synthesized in cells, can be elevated in tissues and mitochondria of young mice by dietary supplementation with CoQ10. The prevalent view is that the uptake of exogenous CoQ by tissues other than plasma and liver either does not occur or is quite minimal. Mice, 6 mo of age, were fed 0, 148 or 654 mg CoQ10/(kg body x d) in their diets for 11 wk. CoQ10 intake enhanced both CoQ9 and CoQ10 homologues in the plasma, and in homogenates and mitochondria of liver, heart and skeletal muscle. CoQ was elevated in brain mitochondria, but not in the brain homogenate. The uptake of exogenous CoQ was higher in mitochondria of heart and skeletal muscle than those in liver. CoQ10 administration also elevated the alpha-tocopherol concentration in tissue homogenates and their mitochondria, thereby providing an in vivo indication of the "sparing" effect of CoQ on alpha-tocopherol. Results of this study demonstrate that, contrary to the historical view, both total and mitochondrial CoQ concentrations in the heart and skeletal muscle and in the mitochondria of brain of young mice can be augmented by dietary supplementation. Furthermore, CoQ intake enhances the antioxidative potential of tissues by elevating the endogenous amounts of alpha-tocopherol.

Effects of age and caloric restriction on glutathione redox state in mice.

The main purpose of this study was to determine whether the aging process in the mouse is associated with a pro-oxidizing shift in the redox state of glutathione and whether restriction of caloric intake, which results in the extension of life span, retards such a shift. Amounts of reduced and oxidized forms of glutathione (GSH and GSSG, respectively) and protein-glutathione mixed disulfides (protein-SSG) were measured in homogenates and mitochondria of liver, kidney, heart, brain, eye, and testis of 4, 10, 22, and 26 month old ad libitum-fed (AL) mice and 22 month old mice fed a diet containing 40% fewer calories than the AL group from the age of 4 months. The concentrations of GSH, GSSG, and protein-SSG vary greatly (approximately 10-, 30-, and 9-fold, respectively) from one tissue to another. During aging, the ratios of GSH:GSSG in mitochondria and tissue homogenates decreased, primarily due to elevations in GSSG content, while the protein-SSG content increased significantly. Glutathione redox potential in mitochondria became less negative, i.e., more pro-oxidizing, as the animal aged. Caloric restriction (CR) lowered the GSSG and protein-SSG content. Results suggest that the aging process in the mouse is associated with a gradual pro-oxidizing shift in the glutathione redox state and that CR attenuates this shift.

Effects of coenzyme Q(10) administration on its tissue concentrations, mitochondrial oxidant generation, and oxidative stress in the rat.

Coenzyme Q (CoQ(10)) is a component of the mitochondrial electron transport chain and also a constituent of various cellular membranes. It acts as an important in vivo antioxidant, but is also a primary source of O(2)(-*)/H(2)O(2) generation in cells. CoQ has been widely advocated to be a beneficial dietary adjuvant. However, it remains controversial whether oral administration of CoQ can significantly enhance its tissue levels and/or can modulate the level of oxidative stress in vivo. The objective of this study was to determine the effect of dietary CoQ supplementation on its content in various tissues and their mitochondria, and the resultant effect on the in vivo level of oxidative stress. Rats were administered CoQ(10) (150 mg/kg/d) in their diets for 4 and 13 weeks; thereafter, the amounts of CoQ(10) and CoQ(9) were determined by HPLC in the plasma, homogenates of the liver, kidney, heart, skeletal muscle, brain, and mitochondria of these tissues. Administration of CoQ(10) increased plasma and mitochondria levels of CoQ(10) as well as its predominant homologue CoQ(9). Generally, the magnitude of the increases was greater after 13 weeks than 4 weeks. The level of antioxidative defense enzymes in liver and skeletal muscle homogenates and the rate of hydrogen peroxide generation in heart, brain, and skeletal muscle mitochondria were not affected by CoQ supplementation. However, a reductive shift in plasma aminothiol status and a decrease in skeletal muscle mitochondrial protein carbonyls were apparent after 13 weeks of supplementation. Thus, CoQ supplementation resulted in an elevation of CoQ homologues in tissues and their mitochondria, a selective decrease in protein oxidative damage, and an increase in antioxidative potential in the rat.