Effect of aging, caloric restriction, and uncoupling protein 3 (UCP3) on mitochondrial proton leak in mice

Mitochondrial proton leak may modulate reactive oxygen species (ROS) production and play a role in aging. The purpose of this study was to determine proton leak across the life span in skeletal mitochondria from calorie-restricted and UCP2/3 overexpressing mice. Proton leak in isolated mitochondria and markers of oxidative stress in whole tissue were measured in female C57BL/6J mice fed ad-libitum (WT-Control) or a 30% calorie-restricted (WT-CR) diet, and in mice overexpressing UCP2 and UCP3 (Positive-TG), their non-overexpressing littermates (Negative-TG) and UCP3 knockout mice (UCP3KO). Proton leak in WT-CR mice was lower than that of control mice at 8 and 26 months of age. The Positive-TG mice had greater proton leak than the Negative-TG and UCP3KO mice at 8 months of age, but this difference disappeared by 19 and 26 months. Lipid peroxidation was generally lower in WT-CR vs. WT-Control mice and UCP3KO mice had greater concentrations of T-BARS (thiobarbituric acid reactive substances, a measure of lipid peroxidation) than did Positive-TG and Negative-TG. The results of this study indicate that sustained increases in muscle mitochondrial proton leak are not responsible for alterations in life span with calorie restriction or UCP3 overexpression in mice. However, UCP3 may contribute to the actions of CR through mechanisms distinct from increasing basal proton leak.     

Altered redox homeostasis in human diabetes saliva

J Oral Pathol Med (2012) 41 : 235–241 Background: Oxidative stress has been implicated in the pathogenesis of diabetes mellitus (DM). Levels of 8‐hydroxy‐2′‐deoxyguanosine (8‐OHdG), 8‐epi‐prostaglandin‐F_2α (8‐epi‐PGF2α), and total protein carbonyls were measured to assess whether DM is associated with altered salivary redox homeostasis. Methods: A total of 215 patients with diabetes and 481 healthy controls were recruited from the Department of Endocrinology at the Jewish General Hospital in Montreal. Levels of oxidative biomarkers were assayed using enzyme‐linked immunosorbent assay (ELISA) in whole unstimulated saliva. Associations of the redox data with exposure to insulin, metformin and dietary control were assessed by logistic regression analyses. Results: We observed (i) significantly higher mean levels of 8‐OHdG and protein carbonyls in whole unstimulated saliva of patients with diabetes compared to controls, (ii) higher mean levels of protein carbonyls in type 1 diabetes as well as higher mean levels of 8‐OHdG and protein carbonyls in type 2 diabetes compared to controls, (iii) elevated levels of protein carbonyls in diet‐controlled patients and in patients with diabetes on insulin and metformin, (iv) elevated levels of 8‐OHdG in patients on metformin, and (v) significant associations between subjects with DM and salivary 8‐OHdG and protein carbonyls. Conclusion: DM is associated with increased oxidative modification of salivary DNA and proteins. Salivary redox homeostasis is perturbed in DM and may inform on the presence of the disease and efficacy of therapeutic interventions.     

Occupational exposure characterization in professional sprayers: clinical utility of oxidative stress biomarkers

The impact of involuntary exposure to pesticides was studied in a group of professional sprayers (S) (25 ± 5 years old) exposed to various agrochemicals for about 10 years. The results were compared with a group of non exposed people (C). S group showed hematological, renal, pancreatic and hepatic biomarkers within the reference values established for the general population, including cholinesterase activity. In spite of that, all the biochemical tests were statistically different compared to C. On the other hand, oxidative stress biomarkers (OSB) such as plasma tocopherol and the total reducing ability of plasma were significantly decreased, while protein carbonyls, thiobarbituric acid-reactive substances, total glutathione and the sum of nitrites and nitrates were increased in the exposed group. Results demonstrated that screening laboratory tests could not be fully sensitive in detecting sub-clinical exposure to pesticides, and also suggest that OSB could be validated and included in health surveillance protocols.     

Oxidative stress and severe walking disability among older women

Background

Oxidative stress has been implicated in sarcopenia and the loss of muscle strength with aging, but the relationship between oxidative stress and decrease in muscle strength and physical performance has not been well characterized. Serum protein carbonyls are markers of oxidative damage to proteins and are caused by oxidative stress.

Methods

Serum protein carbonyls were measured at baseline and compared with a decrease in walking speed and development of severe walking disability (inability to walk or walking speed <0.4 m/sec) over 36 months of follow-up in 545 moderately to severely disabled women, aged ≥65 years, living in the community in Baltimore, Maryland (the Women’s Health and Aging Study I).

Results

After adjusting for age, body mass index, smoking, and chronic diseases, log_e protein carbonyls (nmol/mg) were associated with a decrease in walking speed over 36 months (P = .002). During follow-up, 154 women (28.2%) developed severe walking disability. After adjusting for the same potential confounders, log_e protein carbonyls were associated with incident severe walking disability (hazards ratio 1.42, 95% confidence interval, 1.02-1.98, P = .037).

Conclusion

High oxidative stress, as indicated by oxidative damage to proteins, is an independent predictor of decrease in walking speed and progression to severe walking disability among older women living in the community.     

Protective effect of N-acetyl-l-cysteine against disulfiram-induced oxidative stress and apoptosis in V79 cells

This work investigated the effect of N-acetyl-l-cysteine (NAC) on disulfiram (DSF) induced oxidative stress in Chinese hamster fibroblast cells (V79). An increase in oxidative stress induced by DSF was observed up to a 200 μM concentration. It was evidenced by a statistically significant increase of both GSH_t and GSSG levels, as well as elevated protein carbonyl (PC) content. There was no increase in lipid peroxidation (measured as TBARS). DSF increased CAT activity, but did not change SOD1 and SOD2 activities. Analysis of GSH related enzymes showed that DSF significantly increased GR activity, did not change Se-dependent GPx, but statistically significantly decreased non-Se-dependent GPx activity. DSF showed also pro-apoptotic activity. NAC alone did not produce any significant changes, besides an increase of GSH_t level, in any of the variables measured. However, pre-treatment of cells with NAC ameliorated DSF-induced changes. NAC pre-treatment restored the viability of DSF-treated cells evaluated by Trypan blue exclusion assay and MTT test, GSSG level, and protein carbonyl content to the control values as well as it reduced pro-apoptotic activity of DSF. The increase of CAT and GR activity was not reversed. Activity of both GPx was significantly increased compared to their values after DSF treatment. In conclusion, oxidative properties are at least partially attributable to the cellular effects of disulfiram and mechanisms induced by NAC pre-treatment may lower or even abolish the observed effects. These observations illustrate the importance of the initial cellular redox state in terms of cell response to disulfiram exposure.     

Sperm protein carbonylation

The cryopreservation of sperm is a well established technique that plays an essential role in dissemination of elite germplasm of livestock. Despite having numerous advantages, the cryopreservation induces certain stresses on sperm including structural and functional damages leading to impaired sperm quality and fertility, which might be associated with production of reactive oxygen species (ROS). In addition, the ROS upon reacting with sperm lipids, DNA and proteins may lead to a cascade of sperm damages. The sperm membrane contains a rich amount of polyunsaturated fatty acids, which increases their susceptibility to oxidative stress induced damages, leading to formation of secondary products. These secondary products result in oxidation of sperm proteins via carbonylation. The carbonylation could lead to disturbances in specific proteins that are involved in capacitation. The present review deals with sperm protein carbonylation.     

Long term smoking with age builds up excessive oxidative stress in bronchoalveolar lavage fluid

BACKGROUND: Epithelial lining fluid plays a critical role in protecting the lung from oxidative stress, in which the oxidised status may change by ageing, smoking history, and pulmonary emphysema. METHODS: Bronchoalveolar lavage (BAL) was performed on 109 young and older subjects with various smoking histories. The protein carbonyls, total and oxidised glutathione were examined in BAL fluid. RESULTS: By Western blot analysis, the major carbonylated protein in the BAL fluid was sized at 68 kDa, corresponding to albumin. The amount of carbonylated albumin per mg total albumin in BAL fluid was four times higher in older current smokers and three times higher in older former smokers than in age matched non-smokers (p<0.0001, p=0.0003, respectively), but not in young smokers. Total glutathione in BAL fluid was significantly increased both in young (p=0.006) and older current smokers (p=0.0003) compared with age matched non-smokers. In contrast, the ratio of oxidised to total glutathione was significantly raised (72%) only in older current smokers compared with the other groups. There was no significant difference in these parameters between older smokers with and without mild emphysema. CONCLUSIONS: Oxidised glutathione associated with excessive protein carbonylation accumulates in the lung of older smokers with long term smoking histories even in the absence of lung diseases, but they are not significantly enhanced in smokers with mild emphysema.     

Protein and lipid oxidative damage and complex I content are lower in the brain of budgerigar and canaries than in mice. Relation to aging rate

What are the mechanisms determining the rate of animal aging? Of the two major classes of endothermic animals, bird species are strikingly long-lived compared to mammals of similar body size and metabolic rate. Thus, they are ideal models to identify longevity-related characteristics not linked to body size or low metabolic rates. Since oxidative stress seems to be related to the basic aging process, we measured specific markers of different kinds of oxidative damage to proteins, like glutamic and aminoadipic semialdehydes (GSA and AASA, specific protein carbonyls), Nɛ-(carboxyethyl)lysine (CEL), Nɛ-(carboxymethyl)lysine (CML), and Nɛ-(malondialdehyde)lysine (MDAL), as well as mitochondrial Complex I content and amino acid and membrane fatty acyl composition, in the brain of short-lived mice (maximum life span [MLSP] 3.5 years) compared with those of long-lived budgerigar ‘parakeets’ (MLSP, 21 years) and canaries (MLSP, 24 years). The brains of both bird species had significantly lower levels of compounds formed as a result of oxidative (GSA and AASA), glycoxidative (CEL and CML), and lipoxidative (CML and MDAL) protein modifications, as well as a lower levels of mitochondrial complex I protein. Although it is known that fatty acid unsaturation is lower in many tissues of long-lived compared to short-lived mammals, this is not true in the particular case of brain. In agreement with this, we also found that the brain tissue of bugerigars and canaries contains no fewer double bonds than that of mice. Amino acid composition analyses revealed that bird proteins have a significantly lower content of His, Leu and Phe, as well as, interestingly, of methionine, whereas Asp, Glu, Ala, Val, and Lys contents were higher than in the mammals. These results, together with those previously described in other tissues of pigeons (MLSP, 35 years) compared to rats (MLSP, 4 years), indicate that oxidative damage to proteins, lipids and mitochondrial DNA are lower in birds (very long-lived species) than in short-lived mammals of similar body size. The lower degree of oxidative modification of bird brain proteins was not due to decreases in the target amino acids (lysine for CEL, CML, MDAL, and AASA; and arg and pro for GSA), since these were present in bird brain proteins at higher or similar levels than in those of mice. These results are consistent with the possibility that decreases in oxidative protein modification are caused at least in part by the low rate of mitochondrial oxygen radical generation in these birds, as in all long-lived homeothermic vertebrates investigated so far.Key words: Aminoadipic semialdehyde, complex I, glutamic semialdehyde, lipid peroxidation, Maillard reaction, malondialdehyde, maximum life span, Nɛ-(carboxyethyl)lysine, Nɛ-(carboxymethyl)lysine, Nɛ-(malondialdehyde)lysine, peroxidizability index, protein carbonyls, protein oxidation