Reactive Carbonyl Derivatives Research Articles

The alteration in peripheral neutrophils of patients with chronic kidney disease

Abstract Recent findings have demonstrated the impaired functions of neutrophils of patients with chronic renal failure. The purpose of our research was to study oxidative modified proteins, as well as the histone spectrum in neutrophils drawn from patients with chronic kidney disease, and to estimate the ability of neutrophils to form spontaneous neutrophil extracellular traps. In this work, we have assumed that metabolic alteration in neutrophils may develop at early stages of chronic kidney disease. Materials and methods: Neutrophils obtained from patients with various stages of chronic kidney disease and degrees of chronic renal failure were used. As control, blood samples obtained from 32 healthy individuals was employed. In the examined neutrophils, advanced oxidation protein products, protein reactive carbonyl derivatives, as well as nucleosomal histones were detected. The ability of neutrophils to form spontaneous neutrophil extracellular traps was estimated. Our results have demonstrated an increase of nucleosomal histones in neutrophils of all patients with chronic kidney disease. Moreover, our work fixes the rate of growth of intracellular advanced oxidation protein products and the decreasing of protein reactive carbonyl derivatives in neutrophils from patients with chronic kidney disease. Furthermore, we demonstrate the presence of the neutrophils with altered oxidative status and the decomposition of the histone spectrum in the circulation of patients with chronic kidney disease.

P43 - The modified proteins in erythrocytes at hemodialysis patients

The main goal of our research was to study the modified proteins and activity of integral band 3 protein in erythrocytes of patients with end-stage renal failure during hemodialysis. Depending on the starting nosological variant 2 groups were formed. The first group included 27 patients with end-stage renal failurein the outcome of chronic pyelonephritis, the second – 20 patients with end-stage renal failure, developed on a background of chronic glomerulonephritis. The control group consisted of 15 healthy subjects. The patients of 1-th and 2-nd groups were treated with hemodialysis. The concentration of reactive protein carbonyl derivates, membrane-binding hemoglobin were detected in erythrocytes. Activity of band 3 protein was detected following the protocol of I. Mindukshev et al. (2010). Comparison the results obtained was performed using non-parametric Mann-Whitney U-test (for independent variables).Before hemodialysis the significant increasing of concentration of reactive protein carbonyl derivates and membrane-binding hemoglobin in erythrocytes of both group patients in comparison with healthy ones was observed. During hemodialysis the further augmentation of modified proteins in erythrocytes took place.Different changes in the activity of the protein band 3 in erythrocytes of patients during hemodialysis were found. The similar type of protein band 3 activity changing was determined in erythrocytes of 1-th group patients. There was positive pair correlation between concentration of the reactive protein carbonyl derivates and protein band 3 activity (r= 0,31, p<0,05). Two different types of protein band 3 activity changing were determined in erythrocytes of 2-th group patients. Thus, the differences in modified proteins concentration and protein band 3 activity that occurred in the erythrocytes of patients during hemodialysis were established.

P64 - Myeloperoxidase activity and the oxidized proteins in blood neutrophils of patients with pneumonia

The main purpose of our investigation was to study myeloperoxidase activity and concentration of oxidized proteins in blood neutrophils of patients with ambulant pneumonia and secondary pneumonia which has arisen on a background of chronic obstructive pulmonary disease (COPD). Patients were divided into 2 groups. 17 patients with ambulant pneumonia moderate severity and respiratory insufficiency of grade 2 were included in the 1-st group. 20 COPD patients with secondary pneumonia moderate severity and with respiratory insufficiency of grade 2 were included in the 2-nd group. The control group consisted of 15 healthy subjects. The reactive protein carbonyl derivates, advanced oxidation protein products (AOPP) and myeloperoxidase activity were detected in neutrophils.In neutrophils of 1-st group patients the augmentation of reactive protein carbonyl derivates was observed in comparison with healthy ones. In neutrophils of 2-nd group patients the slight decrease of reactive protein carbonyl derivates was observed in comparison with healthy ones (by 17%). In neutrophils of 2-nd group patients the significant increasing AOPP in comparison with healthy ones (p <0.01) and 1 group patients (p <0.05) was fixed. Myeloperoxidase activity was higher in neutrophils of 1-th group patients in comparison with healthy ones. In neutrophils of 2-nd group patients myeloperoxidase activity was higher in comparison with the same of 1 group patients (by 67%, p <0.05). Our results showed the different direction of oxidized proteins formation neutrophils of patients with primary and secondary pneumonia. Besides that the varied degree of myeloperoxidase activity was fixed. Our results require more detailed understanding because they can reflect peculiar mechanisms of pneumonia development and determine the characteristics of their progression.

Interdependency of selected metabolic variables in an animal model of metabolic syndrome.

In the present study, the correlation between the percentage of glycated hemoglobin, taken as representative of changes in glucose homeostasis, and selected variables was investigated. Rats were treated for 8 weeks with diets containing 64% starch and 5% sunflower oil or containing 64% D-fructose mixed with: 5% sunflower oil; 3.4% sunflower oil and 1.6% salmon oil; or 3.4% sunflower oil and 1.6% safflower oil. Positive correlations were found between glycated hemoglobin and plasma albumin, urea, creatinine, phospholipids, triglycerides and total cholesterol, liver cholesterol, triglyceride and phospholipid content, and the plasma, liver, heart, kidney, soleus muscle and visceral adipose tissue content of thiobarbituric acid reactive substances, carbonyl derivatives and hydroperoxides. Inversely, negative correlations were observed between glycated hemoglobin and plasma calcium, iron and HDL-cholesterol concentrations, liver, heart, kidney, soleus muscle and visceral adipose tissue superoxide dismutase and catalase activity; as well as plasma, liver, heart, kidney, soleus muscle and visceral adipose tissue nitric oxide content. Only the liver glucokinase activity and liver, heart, kidney, soleus muscle and visceral adipose tissue glutathione reductase activity failed to display a significant correlation with glycated hemoglobin. These findings confirm the hypothesis that there is a close association between glucose homeostasis and other variables when considering the effects of long-chain polyunsaturated ω3 and ω6 fatty acids in rats with fructose-induced metabolic syndrome.

ATP content, oxidative stress and motility of beluga (Huso huso) semen: effect of short-term storage.

An effective technique for short-term storage of semen is essential when processing multiple sperm samples and when semen must be transported from collection sites to hatcheries for the fertilization of ova or to laboratories for cryopreservation. In this experiment, beluga (Huso huso) sperm were used to evaluate the effects of short-term storage on several quality parameters (i.e. motility, adenosine triphosphate (ATP) content and oxidative stress indices). Sperm cells exhibited > 50% motility during 3 days of storage with an average total duration of sperm motility varying from 13.33 ± 5.77 to 278.33 ± 25.65 s, and no motile spermatozoa were recorded after 9 days of storage. The levels of oxidative stress indices (thiobarbituric acid reactive substances and carbonyl derivatives of proteins) and antioxidant activity (superoxide dismutase) increased significantly after 3 days of storage. The ATP content also decreased significantly after 2 days of storage. The results of this study can be used to develop effective reproduction management and cryopreservation protocols for this endangered fish.

Biomarkers of oxidative stress and antioxidant defences as indicators of different disinfectants exposure in the heart of rainbow trout (Oncorhynchus MykissWalbaum)

The aim of this study was to evaluate the impacts of different disinfectants' treatment using in aquaculture on the oxidative stress biomarkers such as thiobarbituric acid reactive substrates (TBARS) and carbonyl derivatives of protein oxidative modification, as well as antioxidant defences [superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR) and glutathione peroxidase (GPx) activity] and total antioxidant capacity (TAC) in the heart tissue of rainbow trout (Oncorhynchus mykiss). In the disinfectants exposure, fish were treated to chloramine-T (final concentration 10 g m−3), chloride dioxide (5 g m−3), formalin (200 mL m−3) and CIP disinfectant based on peracetic acid and hydrogen peroxide (16 mL m−3) for 20 min and repeated three times every 3 days. Both chlorine dioxide and formalin treatment was indicated by a significant increase in the level of heart TBARS levels and carbonyl derivatives content and decreased SOD activity. Tissue oxidative stress biomarkers were unchanged upon chloramine-T or CIP disinfectant exposure. Increased oxidative stress could modify antioxidant defences, principally causing increased CAT activity in the heart tissue of formalin- or ClO2−-exposed fish. The correlation between oxidative stress biomarkers and GPx activity indicates that enzymes related to glutathione metabolism were responsible to formalin or ClO2−-induced oxidative stress. Hence, TBARS, carbonyl derivatives and antioxidant defences could be used as biomarkers in evaluating the toxicity of formalin and chlorine dioxide using as disinfectants to trout.

Antiepileptic Drugs Affect Protein, Lipid and DNA Oxidative Damage and Antioxidant Defense in Patients with Epilepsy

Summary Background: To get more insight into the effects of the most widely used antiepileptic drugs (AEDs) on the prooxidant/ antioxidant balance in epilepsy, a comparative analysis of the byproducts of oxidative damage and antioxidant de fense mechanisms was performed in patients with epilepsy treated with lamotrigine, carbamazepine and valproic acid. Methods: Byproducts of oxidative damage to proteins (reactive carbonyl derivatives, RCD and protein thiol groups, PSH), lipids (urinary isoprostanes, 8-epi-PGF2a) and DNA (urinary 8-hydroxy-2’-deoxyguanosine, 8-OHdG), as well as the activities of antioxidant enzymes superoxide dismutase (SOD) and glutathione peroxidase (GPX) were measured in 60 patients with newly diagnosed seizure (at illness onset and after 6 months of treatment with lamotrigine, carbamazepine or valproic acid) and in 20 healthy controls. Results: In patients with epilepsy, RCD, urinary 8-epi-PGF2a and 8-OHdG, together with SOD and GPX activities were significantly increased, while P-SH were only slightly decreased. After 6 months of treatment with AEDs, a decrease was observed in RCD, urinary 8-epi-PGF2a and 8-OHdG to values slightly higher or similar to the control, while P-SH remained unchanged. A decrease was also observed in SOD and GPX activities, although they remained significantly in creased compared to controls. Conclusions: The results of this study have shown that treatments with lamotrigine, carbamazepine and valproic acid affect the prooxidant/antioxidant balance in patients with epilepsy.

Markers of Oxidative Damage and Antioxidant Enzyme Activities as Predictors of Morbidity and Mortality in Patients With Chronic Heart Failure

BackgroundAlthough the majority of previous findings unequivocally confirmed the existence of systemic oxidative stress in chronic heart failure (CHF) patients, data on prognostic potential of biomarkers of oxidative lipid and protein damage are limited. We aimed to address the relation of oxidative stress markers to severity and prognosis in CHF secondary to ischemic cardiomyopathy. Methods and ResultsPlasma malondialdehyde (MDA), protein thiol groups (P-SH), reactive carbonyl derivatives (RCD), together with glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) activities were determined in 120 CHF patients and 69 healthy controls. Increased lipid peroxidation (MDA) and oxidation of plasma proteins (RCD; P-SH) s well as downregulated GSH-Px activity were found in CHF patients compared with controls. Significant correlation was obtained only for RCD content and remodeling indices (LVEDV: r = 0.469, P = .008; LVESV: r = 0.452; P = .011). Cox regression analysis demonstrated only MDA (HR = 3.33; CI: 1.55–7.12; P = .002) as independent predictor of death, whereas SOD was associated with unstable angina pectoris (HR = 2.09; CI: 1.16–3.78; P = .011). ConclusionsIn the course of CHF progression, carbonyl stress is implicated in the LV remodeling. Malondialdehyde level might be a useful parameter for monitoring and planning management of CHF patients.

Byproducts of protein, lipid and DNA oxidative damage and antioxidant enzyme activities in seizure

Purpose To get more insight into molecular mechanisms underlying oxidative stress and its role in different types of seizure, in this study, oxidative byproducts of proteins, lipids and DNA, as well as, antioxidant enzyme activities were studied in adult patients with epilepsy. Methods Study was performed in 60 patients with epilepsy and in 25 healthy controls. Plasma protein reactive carbonyl derivatives (RCD) and protein thiol groups (P-SH), byproducts of oxidative protein damage, as well as antioxidant enzyme activities, superoxide dismutase (SOD) and glutathione peroxidase (GPX) were studied spectrophotometrically. Urinary 8-epi-prostaglandin F 2α (8-epi-PGF 2α) and 8-hydroxy-2′-deoxyguanosine (8-OHdG), representative byproducts of lipid and DNA oxidative damage, respectively, were determined by enzyme immunoassay. Results RCD levels were significantly increased ( p = 0.001), while P-SH content was decreased in patients with first seizure ( p = 0.052) compared to controls, independently of the seizure type. Urinary 8-epi-PGF 2α and 8-OHdG were significantly increased in patients with epilepsy ( p = 0.001 and p = 0.001). Rise in 8-epi-PGF 2α was more pronounced in patients with generalized tonic–clonic seizure (GTCS) compared to those with partial seizure (PS). Both SOD and GPX activity were significantly increased in epileptic patients compared to controls ( p = 0.001 and p = 0.001), but only SOD activity was significantly higher in patients with GTCS than in those with PS. Conclusions Data on enhanced protein, lipid and DNA oxidation, together with upregulated antioxidant enzyme activities, confirm the existence of systemic oxidative stress in patients with epilepsy. It might be speculated that post-translational modification to existing functional proteins, particularly alterations to ion channels, might be at least partially responsible for acute early changes in neuronal networks.

Oxidative stress parameters as possible urine markers in patients with diabetic nephropathy

Objective Reactive oxygen species play a crucial role in the pathogenesis of diabetic nephropathy (DN). The present study was performed to assess oxidative stress parameters—thiobarbituric acid reactive substances (TBARS), reactive carbonyl derivates (RCDs), and total sulfhydryl groups (TSHGs)—in serum and urine of patients with DN. Methods All parameters were determined in patients with type 2 and type 1 diabetes mellitus and microalbuminuria (DMT2-MIA, DMT1-MIA, respectively) and patients with type 2 diabetes mellitus and macroalbuminuria (DMT2-MAA) compared to healthy controls. Results Serum and urine TBARS levels were higher in all patients with DN and microalbiminuria compared to the control group. RCD levels significantly increased in serum of patients with DMT2 relative to the controls as well as in urine of patients with DMT2-MAA and DMT1-MIA. In all groups of patients, TSHGs decreased in serum but not in urine of patients with DMT2-MAA. Conclusion Urine TBARS, RCDs, and TSHGs could be proposed as possible markers for oxidative damage of kidney in DN.

Multivitamin-Mineral Supplementation Prevents Lipid Peroxidation during “The Marathon des Sables”

Objective: We investigated the effect of a moderate mutivitamin and mineral supplementation containing mainly vitamin C (150.0 mg.day−1), vitamin E (24.0 mg.day−1) and β-carotene (4.8 mg.day−1) prior to and during an extreme running competition -the Marathon des Sables (MDS)- that consisted of six long races in the desert.Methods: Seventeen athletes participated in our double blind, placebo-controlled study. Blood samples were collected prior to the supplementation i.e. three weeks before the competition (D-21), two days prior to the MDS (D-2), after the third race (D3) and at the end of the competition (D7). Erythrocyte antioxidant enzyme activity (glutathione peroxidase (GPx), superoxide dismutase (SOD)), erythrocyte glutathione level (GSH), plasma non-enzymatic antioxidant status (uric acid, vitamin C, α-tocopherol, retinol, β-carotene), markers of plasma lipid peroxidation (thiobarbituric reactive substances (TBARS)), reactive carbonyl derivatives (RCD) and membrane damage (creatine kinase and lactate dehydrogenase activities) were measured.Results: In both groups, GSH levels, uric acid levels and membrane damage significantly increased during the competition while SOD activity significantly decreased. In Supplemented group, plasma α-tocopherol, β-carotene and retinol levels significantly increased after three weeks of supplementing. In contrast to Placebo group, α-tocopherol, vitamin C and retinol levels were significantly affected by the competition in Supplemented group. Moreover, no increase in TBARS was observed in Supplemented group during the competition, whereas TBARS significantly increased at D3 in the placebo group.Conclusion: The moderate multivitamin-mineral supplementation prevented the transient increase in TBARS levels during this extreme competition.

Circulating leukocyte heat shock protein 70 (HSP70) and oxidative stress markers in rats after a bout of exhaustive exercise

A novel method to measure oxidative stress resulting from exhaustive exercise in rats is presented. In this new procedure we evaluated the erythrocyte antioxidant enzymes, catalase (CAT) and glutathione reductase (GR), the plasma oxidative attack markers, reactive carbonyl derivatives (RCD) and thiobarbituric reactive substances (TBARS). Muscular tissue damage was evaluated by monitoring plasma creatine kinase (CK) and plasma taurine (Tau) concentrations. Also, we monitored total sulphydryl groups (TSG) and uric acid (UA), and the level of the 70 kDa heat shock protein (HSP70) in leukocytes as a marker of oxidative stress. In the study we found a correspondence between erythrocyte CAT and GR activities and leukocyte HSP70 levels, principally 3 h after the acute exercise, and this suggested an integrated mechanism of antioxidant defense. The increase in levels of plasma Tau was coincident with the increasing plasma levels of CK and TBARS, principally after two hours of exercise. Thus tissue damage occurred before the expression of any anti-oxidant system markers and the monitoring of Tau, CK or TBARS may be important for the estimation of oxidative stress during exhaustive exercise. Furthermore, the integrated analyses could be of value in a clinical setting to quantify the extent of oxidative stress risk and reduce the need to perform muscle biopsies as a tool of clinical evaluation.

Byproducts of oxidative protein damage and antioxidant enzyme activities in plasma of patients with different degrees of essential hypertension

Despite evidence that essential hypertension (EH) is a state of increased oxidative stress, the data on oxidative protein modifications is lacking. Besides, the role of extracellular antioxidant enzymes in EH has not been systematically studied. Study was performed in 45 subjects with EH and 25 normotensive controls. Patients were divided into three groups according to the 2003 ESH/ESC guidelines (grade 1-3). Plasma protein reactive carbonyl derivatives (RCD) and SH-groups (as byproducts of oxidative protein damage) as well as antioxidant enzyme activities superoxide dismutase (SOD), glutathione peroxidase (GPX) and catalase were studied spectrophotometrically and correlated with blood pressure (BP). RCD levels were increased in EH patients compared to controls and correlated significantly with both systolic blood pressure (SBP) (r = 0.495, P<0.01) and diastolic blood pressure (DBP) (r = 0.534, P<0.01). Plasma SH-groups content was significantly lower in all patients with EH, with no correlation with BP. SOD and catalase activity in patients with grade 1 EH were similar to that of controls. Patients with grade 2 and 3 of EH had lower SOD and catalase activity. However, significant correlation with SBP and DBP was observed for catalase only (r = -0.331; P<0.05 and r = -0.365; P<0.05, respectively). EH patients exhibited higher plasma GPX activity compared to those in controls, and it correlated with SBP (r = 0.328; P<0.05). The results presented show that increased oxidative protein damage is present in all grades of EH. In mild hypertension extracellular antioxidant enzyme activities are not decreased, suggesting they are probably not critical in early EH, but could be important in moderate to severe EH.

Protein oxidation in Type 2 diabetic patients on hemodialysis

Background Oxidative stress is considered to be a unifying link between diabetes mellitus (DM) and its complications, including nephropathy. There have been many reports on increased production of oxidants and decreased level of antioxidants in diabetic patients. The dialysis procedure contributes to oxidative stress. An increase in oxidative stress may contribute to the development of oxidative protein damage in diabetic patients. Our aim was to reveal the effects of diabetes and hemodialysis (HD) on oxidative modifications of plasma proteins. Methods We measured reactive carbonyl derivates (PCO), protein thiol (P-SH), and reduced glutathione (GSH) levels in Type 2 diabetic (DM) and diabetic hemodialysed patients (DHD) and in healthy control participants. Results Protein carbonyl (PCO) content increased significantly in all patient groups relative to the controls. The dialysis procedure caused an additional increase in PCO levels in DHD patients before and after dialysis compared with the level in DM patients. There was a significant decrease in P-SH levels in DHD patients compared with the level in healthy participants and DM patients. There was no significant difference in the whole blood GSH levels between the DM patients and control participants. It was significantly higher in DHD patients in comparison to the DM patients. Conclusions We conclude that PCO level increases in DM patients, and this increase is more profound in DHD patients, indicating that both diabetes and dialysis contribute to increased protein oxidation. The low P-SH level in DHD patients, but not in DM patients, suggests that dialysis is responsible for this decrease. We propose plasma PCO derivate as a novel specific marker for oxidative protein damage.

The effects of moderate-, strenuous- and over-training on oxidative stress markers, DNA repair, and memory, in rat brain

We have tested the hypothesis that training with moderate- (MT), strenuous- (ST), or over- (OT) load can cause alterations in memory, lipid peroxidation, protein oxidation, DNA damage, activity of 8-oxoG-DNA glycosylase (OGG1) and brain-derived neurotrophic factor (BDNF), in rat brain. Rat memory was assessed by a passive avoidance test and the ST and OT group demonstrated improved memory. The content of BDNF was increased only in the OT group. The oxidative damage of lipids and DNA, as measured by thiobarbituric acid reactive substances (TBARS), and 8-hydroxydeoxyguanosine (8-OHdG), did not change significantly with exercise. Similarly, the activity of DNA repair enzyme, 8-oxoguanine DNA glycosylase (OGG1), was not altered with exercise training. On the other hand, the content of reactive carbonyl derivatives (RCDs) decreased in all groups and the decrease reached significance levels in the ST and OT groups. The activity of the proteasome complex increased in the brain of OT. The findings of this study imply that over-training does not induce oxidative stress in the brain and does not cause loss of memory. The improved memory was associated with enhanced BDNF content.

Carbonyl stress in chronic renal failure: the effect of haemodialysis

Oxidative stress has been defined as a loss of balance between free radical production and the antioxidant systems. There have been many reports of increased production of oxidants and decreased levels of antioxidants in chronic renal failure (CRF) patients. An increase in oxidative stress may contribute to the development of oxidative protein damage in CRF. Our aim was to reveal oxidative modifications of plasma proteins by measuring 2,4-dinitrophenylhydrazine reactive carbonyl derivates (PCO), protein thiol (P-SH) and reduced glutathione (GSH) in predialytic uraemic and haemodialysed (HD) patients before and after dialysis. We included 20 predialytic uraemic patients, 20 HD patients and 20 healthy volunteers in our study. PCO concentration in predialytic uraemic patients increased compared with the concentration of the control group and this increase was more profound in HD patients. P-SH concentrations were significantly decreased in haemodialytic patients compared with those of controls. GSH level was higher in HD patients (both before and after dialysis). Increased PCO and decreased P-SH concentrations in all patient groups in comparison to the control subjects indicate increased protein oxidation. Our data in ESRD patients propose plasma protein carbonyl derivates and thiol concentrations as novel specific markers for oxidative protein damage.

Mechanical ventilation-induced oxidative stress in the diaphragm.

Prolonged mechanical ventilation (MV) results in oxidative damage in the diaphragm; however, it is unclear whether this MV-induced oxidative injury occurs rapidly or develops slowly over time. Furthermore, it is unknown whether both soluble (cytosolic) and insoluble (myofibrillar) proteins are equally susceptible to oxidation during MV. These experiments tested two hypotheses: 1). MV-induced oxidative injury in the diaphragm occurs within the first 6 h after the initiation of MV; and 2). MV is associated with oxidative modification of both soluble and insoluble proteins. Adult Sprague-Dawley rats were randomly divided into one of seven experimental groups: 1) control (n = 8); 2) 3-h MV (n = 8); 3). 6-h MV (n = 6); 4). 18-h MV (n = 8); 5). 3-h anesthesia-spontaneous breathing (n = 8); 6). 6-h anesthesia-spontaneous breathing (n = 6); and 7). 18-h anesthesia-spontaneous breathing (n = 8). Markers of oxidative injury in the diaphragm included the measurement of reactive (protein) carbonyl derivatives (RCD) and total lipid hydroperoxides. Three hours of MV did not result in oxidative injury in the diaphragm. In contrast, both 6 and 18 h of MV promoted oxidative injury in the diaphragm, as indicated by increases in both protein RCD and lipid hydroperoxides. Electrophoretic separation of soluble and insoluble proteins indicated that the MV-induced accumulation of RCD was limited to insoluble proteins with molecular masses of approximately 200, 120, 80, and 40 kDa. We conclude that MV results in a rapid onset of oxidative injury in the diaphragm and that insoluble proteins are primary targets of MV-induced protein oxidation.

Exercise training decreases DNA damage and increases DNA repair and resistance against oxidative stress of proteins in aged rat skeletal muscle.

Regular physical exercise retards a number of age-associated disorders, in spite of the paradox that free radical generation is significantly enhanced with exercise. Eight weeks of treadmill running resulted in nearly a 40% increase in maximal oxygen uptake in both middle-aged (20-month-old) and aged (30-month-old) rats. The age-associated increase in 8-hydroxy-2'-deoxyguanosine (8-OHdG) content was significantly attenuated in gastrocnemius muscle by exercise. The 8-OHdG repair, as measured by the excision of 32P-labeled damaged oligonucleotide, increased in muscle of exercising animals. The reactive carbonyl derivatives (RCD) of proteins did not increase with aging. However, when the muscle homogenate was exposed to a mixture of 1 mM iron sulfate and 50 mM ascorbic acid, the muscle of old control animals accumulated more RCD than that of the trained or adult groups. The chymotrypsin-like activity of proteasome complex increased in muscle of old trained rats. We suggest that regular exercise-induced adaptation attenuates the age-associated increase in 8-OHdG levels, and increases the activity of DNA repair and resistance against oxidative stress in proteins.

Effect of aging and late onset dietary restriction on antioxidant enzymes and proteasome activities, and protein carbonylation of rat skeletal muscle and tendon

Many studies have shown that lifelong dietary restriction (DR) can retard aging processes. Very few reports, however, are found that examined the effect of late onset DR on biochemical parameters in aging animals [Goto, S., Takahashi, R., Araki, S., Nakamoto, H., 2002b. Dietary restriction initiated in late adulthood can reverse age-related alterations of protein and protein metabolism. Ann. NY Acad. Sci. 959, 50–56]. We studied the effect of every-other-day feeding, initiated at the age of 26.5 months and continued for 3.5 months, on antioxidant enzymes, protein carbonyls, and proteasomes of the gastrocnemius muscle and tendon in rats. Age-related increase in the activity and content of Cu, Zn-SOD and the content of Mn-SOD was attenuated by the DR in both tissues. The same was true for glutathione peroxidase and catalase activities. Significant increase with age in protein reactive carbonyl derivatives (RCD) in the tendon was noted that was partially reversed by the DR. No significant change of RCD, however, was observed in the skeletal muscle. The age-related and DR-induced changes of the RCD in the tendon appeared to be associated with proteasome activity that decreases with age and increases by the DR. It is suggested that the late onset DR can have beneficial effects on the locomotive functions by reducing age-associated potentially detrimental oxidative protein damage in the tendon.

Regular Training Modulates the Accumulation of Reactive Carbonyl Derivatives in Mitochondrial and Cytosolic Fractions of Rat Skeletal Muscle

The oxygen flux into the mitochondria of skeletal muscle increases with exercise. However, the extent of oxidative damage to mitochondrial proteins of skeletal muscle has only been estimated. We studied the alteration of reactive carbonyl derivatives (RCD) in mitochondrial and cytosolic fractions of skeletal muscle following 9 weeks of swimming training in rats. The RCD content of mitochondria was significantly elevated compared with the cytosolic fraction of both control and exercised animals. Accumulation of RCD in muscle mitochondria of the exercised group was also significantly elevated (P < 0.05). On the other hand, alteration of the accumulation of RCD was not apparent in the cytosolic fraction of skeletal muscle. The activity of proteasome complex, however, was increased in the cytosolic fraction of exercised muscle (P < 0.05). The data suggest that mitochondria of skeletal muscle accumulate significantly larger amounts of RCD than the cytosolic fraction and the tendency of the accumulation varies in cell fractions. Exercise training increases the accumulation of protein damage in mitochondria of skeletal muscle but cytosolic proteins are protected by increased activity of proteasome complex and possibly by other antioxidant enzymes.