Cranberry fruit has been reported to have high antioxidant effectiveness that is potentially linked to its richness in diversified polyphenolic content. The aim of the present study was to determine the role of cranberry polyphenolic fractions in oxidative stress (OxS), inflammation and mitochondrial functions using intestinal Caco-2/15 cells. The combination of HPLC and UltraPerformance LC®-tandem quadrupole (UPLC-TQD) techniques allowed us to characterize the profile of low, medium and high molecular mass polyphenolic compounds in cranberry extracts. The medium molecular mass fraction was enriched with flavonoids and procyanidin dimers whereas procyanidin oligomers (DP > 4) were the dominant class of polyphenols in the high molecular mass fraction. Pre-incubation of Caco-2/15 cells with these cranberry extracts prevented iron/ascorbate-mediated lipid peroxidation and counteracted lipopolysaccharide-mediated inflammation as evidenced by the decrease in pro-inflammatory cytokines (TNF-α and interleukin-6), cyclo-oxygenase-2 and prostaglandin E2. Cranberry polyphenols (CP) fractions limited both nuclear factor κB activation and Nrf2 down-regulation. Consistently, cranberry procyanidins alleviated OxS-dependent mitochondrial dysfunctions as shown by the rise in ATP production and the up-regulation of Bcl-2, as well as the decline of protein expression of cytochrome c and apoptotic-inducing factor. These mitochondrial effects were associated with a significant stimulation of peroxisome-proliferator-activated receptor γ co-activator-1-α, a central inducing factor of mitochondrial biogenesis and transcriptional co-activator of numerous downstream mediators. Finally, cranberry procyanidins forestalled the effect of iron/ascorbate on the protein expression of mitochondrial transcription factors (mtTFA, mtTFB1, mtTFB2). Our findings provide evidence for the capacity of CP to reduce intestinal OxS and inflammation while improving mitochondrial dysfunction.

The role of oxidative stress in ischaemic heart disease has been thoroughly investigated in humans. Increased levels of ROS (reactive oxygen species) and RNS (reactive nitrogen species) have been demonstrated during ischaemia and post-ischaemic reperfusion in humans. Depending on their concentrations, these reactive species can act either as benevolent molecules that promote cell survival (at low-to-moderate concentrations) or can induce irreversible cellular damage and death (at high concentrations). Although high ROS levels can induce NF-κB (nuclear factor κB) activation, inflammation, apoptosis or necrosis, low-to-moderate levels can enhance the antioxidant response, via Nrf2 (nuclear factor-erythroid 2-related factor 2) activation. However, a clear definition of these concentration thresholds remains to be established. Although a number of experimental studies have demonstrated that oxidative stress plays a major role in heart ischaemia/reperfusion pathophysiology, controlled clinical trials have failed to prove the efficacy of antioxidants in acute or long-term treatments of ischaemic heart disease. Oral doses of vitamin C are not sufficient to promote ROS scavenging and only down-regulate their production via NADPH oxidase, a biological effect shared by vitamin E to abrogate oxidative stress. However, infusion of vitamin C at doses high enough to achieve plasma levels of 10 mmol/l should prevent superoxide production and the pathophysiological cascade of deleterious heart effects. In turn, n −3 PUFA (polyunsaturated fatty acid) exposure leads to enhanced activity of antioxidant enzymes. In the present review, we present evidence to support the molecular basis for a novel pharmacological strategy using these antioxidant vitamins plus n −3 PUFAs for cardioprotection in clinical settings, such as post-operative atrial fibrillation, percutaneous coronary intervention following acute myocardial infarction and other events that are associated with ischaemia/reperfusion.

Understanding the role of oxidative injury will allow for therapy with agents that scavenge ROS (reactive oxygen species) and antioxidants in the management of several diseases related to free radical damage. The majority of free radicals are generated by mitochondria as a consequence of the mitochondrial cycle, whereas free radical accumulation is limited by the action of a variety of antioxidant processes that reside in every cell. In the present review, we provide an overview of the mitochondrial generation of ROS and discuss the role of ROS in the regulation of endothelial and adipocyte function. Moreover, we also discuss recent findings on the role of ROS in sepsis, cerebral ataxia and stroke. These results provide avenues for the therapeutic potential of antioxidants in a variety of diseases.

MDs (mitochondrial diseases) are a clinically heterogeneous group of disorders characterized by impairment of the respiratory chain function with altered oxidative phosphorylation. We tested the hypothesis that the function of vascular endothelium is affected by increased oxidative stress in MDs. A total of 12 patients with MDs and pair-matched controls were studied. Endothelial function was assessed by measuring FMD (flow-mediated vasodilation) of brachial and common femoral arteries. The test was repeated after vitamin C (500 mg, twice a day) and E (400 mg, once a day) supplementation for 30 days and 90 days after vitamin withdrawal. FMD was reduced in patients compared with controls [AUC/τ (time-averaged area under the curve) for the brachial artery, 1.05±0.24 compared with 4.19±0.59% respectively, P <0.001; AUC/τ for the femoral artery, 0.98±0.19 compared with 2.36±0.29% respectively, P =0.001; values are means±S.E.M.] and correlated (brachial artery) with plasma lactate ( r =−0.63, P <0.01). Urinary 8-iso-PGF 2α (8-iso-prostaglandin F 2α ) was higher in patients than controls (505.6±85.9 compared with 302.5±38.7 pg/mg of creatinine; P <0.05) and correlated with plasma lactate ( r =0.70, P <0.05). Immunohistochemical analysis showed 8-iso-PGF 2α staining in MD-affected striated muscle cells and in blood vessels in muscle biopsies of patients. Antioxidant vitamins transiently restored FMD in patients [ΔAUC/τ (change in AUC/τ) for the brachial artery, +1.38±0.49%, P <0.05; ΔAUC/τ for the femoral artery, +0.98±0.24%, P <0.01] but had no effect on FMD in controls (brachial artery, −1.3±0.63%; and common femoral artery, −0.58±0.30%), thus abolishing the differences between patients and controls. The results of the present study indicate that oxidative stress is increased and is, at least partly, responsible for endothelial dysfunction in MDs.

Intra-uterine growth restriction is an independent risk factor for adult psychiatric and cardiovascular diseases. In humans, intra-uterine growth restriction is associated with increased placental and fetal oxidative stress, as well as down-regulation of placental 11β-HSD (11β-hydroxysteroid dehydrogenase). Decreased placental 11β-HSD activity increases fetal exposure to maternal glucocorticoids, further increasing fetal oxidative stress. To explore the developmental origins of co-morbid hypertension and anxiety disorders, we increased fetal glucocorticoid exposure by administering the 11β-HSD inhibitor CBX (carbenoxolone; 12 mg·kg −1 of body weight·day −1 ) during the final week of murine gestation. We hypothesized that maternal antioxidant (tempol throughout pregnancy) would block glucocorticoid-programmed anxiety, vascular dysfunction and hypertension. Anxiety-related behaviour (conditioned fear) and the haemodynamic response to stress were measured in adult mice. Maternal CBX administration significantly increased conditioned fear responses of adult females. Among the offspring of CBX-injected dams, maternal tempol markedly attenuated the behavioural and cardiovascular responses to psychological stress. Compared with offspring of undisturbed dams, male offspring of dams that received daily third trimester saline injections had increased stress-evoked pressure responses that were blocked by maternal tempol. In contrast, tempol did not block CBX-induced aortic dysfunction in female mice (measured by myography and lucigenin-enhanced chemiluminescence). We conclude that maternal stress and exaggerated fetal glucocorticoid exposure enhance sex-specific stress responses, as well as alterations in aortic reactivity. Because concurrent tempol attenuated conditioned fear and stress reactivity even among the offspring of saline-injected dams, we speculate that antenatal stressors programme offspring stress reactivity in a cycle that may be broken by antenatal antioxidant therapy.

Exercise may contribute to the maintenance of vascular function via enhanced liberation and action of bone-marrow-derived progenitor cells. Activity related changes in oxidative stress may also influence the number and function of these cells. In the present study, we sought to determine (i) whether adaptations in reactive hyperaemic FBF (forearm blood flow) response associated with long-term endurance exercise and short-term detraining were related to resting putative progenitor cell number and function, and (ii) whether oxidative stress affected these factors. Participants included men with a history of more than 30 years of moderate-to-high-intensity exercise (HI group) and healthy low-active age- and BMI (body mass index)-matched control subjects (LO group). Vascular reactive hyperaemic FBF response, resting CD34 + and CD34 + /VEGFR2 + (vascular endothelial growth factor receptor 2 + ] cell number, CFU-EC (colony-forming unit-endothelial cell) count and CFU-EC senescence were evaluated. Oxidative stress measures included OxLDL (oxidized low-density lipoprotein) and TAC (total antioxidant capacity). These measures were assessed following 10 days of detraining in the HI group. The HI group had greater peak reactive hyperaemic FBF responses compared with the LO group, despite no difference in resting CD34 + cell number, CD34 + /VEGFR2 + cell number, CFU-EC colonies or CFU-EC senescence. With detraining in the HI group, CD34 + cells declined 44%, and the percentage change in CD34 + /VEGFR2 + cells was positively correlated with the change in FBF response to reactive hyperaemia. The percentage change in CD34 + /VEGFR2 + cells and the percentage change in EPC (endothelial progenitor cell) senescence with detraining were related to the percentage change in TAC. These results reveal that changes in reactive hyperaemic FBF are closely related to activity dependent dynamic changes in CD34 + /VEGFR2 + cell number, which may be influenced by alterations in oxidative stress.

Exposure to an adverse intrauterine environment is recognized as an important risk factor for the development of cardiovascular disease later in life. Although oxidative stress has been proposed as a mechanism for the fetal programming phenotype, the role of mitochondrial O 2 •− (superoxide radical) production has not been explored. To determine whether mitochondrial ROS (reactive oxygen species) production is altered by in utero programming, pregnant ewes were given a 48-h dexamethasone (dexamethasone-exposed, 0.28 mg·kg −1 of body weight·day −1 ) or saline (control) infusion at 27–28 days gestation (term=145 days). Intact left ventricular mitochondria and freeze-thaw mitochondrial membranes were studied from offspring at 4-months of age. AmplexRed was used to measure H 2 O 2 production. Activities of the antioxidant enzymes Mn-SOD (manganese superoxide dismutase), GPx (glutathione peroxidase) and catalase were measured. Compared with controls, a significant increase in Complex I H 2 O 2 production was found in intact mitochondria from dexamethasone-exposed animals. The treatment differences in Complex I-driven H 2 O 2 production were not seen in mitochondrial membranes. Consistent changes in H 2 O 2 production from Complex III in programmed animals were not found. Despite the increase in H 2 O 2 production in intact mitochondria from programmed animals, dexamethasone exposure significantly increased mitochondrial catalase activity, whereas Mn-SOD and GPx activities were unchanged. The results of the present study point to an increase in the rate of release of H 2 O 2 from programmed mitochondria despite an increase in catalase activity. Greater mitochondrial H 2 O 2 release into the cell may play a role in the development of adult disease following exposure to an adverse intrauterine environment.

Both antioxidant supplementation and exercise training have been identified as interventions which may reduce oxidative stress and thus improve cardiovascular health, but the interaction of these interventions on arterial BP (blood pressure) and vascular function has not been studied in older humans. Thus in six older (71±2 years) mildly hypertensive men, arterial BP was evaluated non-invasively at rest and during small muscle mass (knee-extensor) exercise with and without a pharmacological dose of oral antioxidants (vitamins C and E, and α-lipoic acid). The efficacy of the antioxidant intervention to decrease the plasma free radical concentration was verified via EPR (electron paramagnetic resonance) spectroscopy, while changes in endothelial function in response to exercise training and antioxidant administration were evaluated via FMD (flow-mediated vasodilation). Subjects were re-evaluated after a 6-week aerobic exercise training programme. Prior to training, acute antioxidant administration did not change resting arterial BP or FMD. Six weeks of knee-extensor exercise training reduced systolic BP (from 150±8 mmHg at pre-training to 138±3 mmHg at post-training) and diastolic BP (from 91±5 mmHg at pre-training to 79±3 mmHg at post-training), and improved FMD (1.5±1 to 4.9±1% for pre- and post-training respectively). However, antioxidant administration after exercise training negated these improvements, returning subjects to a hypertensive state and blunting training-induced improvements in FMD. In conclusion, the paradoxical effects of these interventions suggest a need for caution when exercise and acute antioxidant supplementation are combined in elderly mildly hypertensive individuals.

Plasma levels of HDL (high-density lipoprotein)-cholesterol are strongly and inversely correlated with atherosclerotic cardiovascular disease. Both clinical and epidemiological studies have reported an inverse and independent association between serum HDL-cholesterol levels and CHD (coronary heart disease) risk. The cardioprotective effects of HDLs have been attributed to several mechanisms, including their involvement in the reverse cholesterol transport pathway. HDLs also have antioxidant, anti-inflammatory and antithrombotic properties and promote endothelial repair, all of which are likely to contribute to their ability to prevent CHD. The first part of this review summarizes what is known about the origins and metabolism of HDL. We then focus on the anti-inflammatory and antioxidant properties of HDL and discuss why these characteristics are cardioprotective.

The oxidation hypothesis for CHD (coronary heart disease) is attractive; however, the almost universal failure of antioxidant vitamin supplementation as a CVD (cardiovascular disease) risk modifier challenges the oxidation hypothesis, at least as a concept that easily ‘translates’ into clinical benefit for the population. At the same time, quality prospective data on lipid or protein oxidation markers as predictors of vascular events are sparse. In the present issue of Clinical Science , Woodward and co-workers provide much needed prospective data examining the relationship between markers of oxidative damage and CHD outcome in a general population. Despite noting the expected associations between several established CHD risk factors and CHD events, no significant link was observed between measured oxidation markers and CHD risk, a finding which further challenges the oxidation hypothesis for CHD.

The role of oxidative damage in the aetiology of coronary disease remains controversial, as clinical trials investigating the effect of antioxidants have not generally been positive. In the present study, 227 coronary cases, identified from a cohort study, were matched, by age and gender, with 420 controls in a nested case-control design. Stored plasma samples were analysed for F 2 -isoprostanes by stable isotope dilution MS, and specifically oxidized forms of apoA-I (apolipoprotein A-I) by HPLC of HDL (high-density lipoprotein). Median values of F 2 -isoprostanes were higher in plasma samples that contained oxidized apoA-I compared with samples with undetectable oxidized apoA-I (1542 compared with 1165 pmol/l). F 2 -Isoprostanes were significantly correlated with variants of non-oxidized apoA-II ( r =−0.15) and were associated with HDL-cholesterol ( P <0.0001). F 2 -Isoprostanes in cases (median, 1146 pmol/l) were not different from controls (1250 pmol/l); the odds ratio (95% confidence interval) for a 1 S.D. increase in F 2 -isoprostanes was 1.08 (0.91–1.29). Similarly, there was no independent association between the presence of oxidized apoA-I, detected in approx. 20% of the samples, and coronary risk. In conclusion, we found no evidence of associations between markers of lipid (F 2 -isoprostanes) and protein (oxidized apoA-I) oxidation and the risk of fatal or non-fatal coronary heart disease in a general population. This may be due to a true lack of association or insufficient power.

Liver cirrhosis is characterized by increased IHR (intrahepatic resistance) and lipid peroxidation, and decreased antioxidative defence. The present study investigates the effects of administration for 1 month of the antioxidant UDCA (ursodeoxycholic acid) in BDL (bile-duct-ligated) cirrhotic rats. Splanchnic haemodynamics, IHR, hepatic levels of TBARS (thiobarbituric acid-reacting substances), GSH (glutathione), SOD (superoxide dismutase) activity, nitrite, PIIINP (N-terminal propeptide of type III procollagen) and collagen deposition, histological examination of liver, mRNA expression of PIIIP-α 1 (type III procollagen) and TGF-β1 (transforming growth factor-β 1 ), protein expression of TXS (thromboxane synthase) and iNOS (inducible NO synthase), and TXA 2 (thromboxane A 2 ) production in liver perfusates were measured. The results showed that portal pressure and IHR, hepatic levels of PIIINP, hepatic collagen deposition, mRNA expression of PIIIP-α 1 and TGF-β 1 , protein expression of iNOS and TXS, and production of TXA 2 in liver perfusates were significantly decreased in UDCA-treated BDL rats. The increased levels of hepatic GSH and SOD activity and decreased levels of TBARS and nitrite were also observed in UDCA-treated BDL rats. In UDCA-treated BDL rats, the reduction in portal pressure resulted from a decrease in IHR, which mostly acted through the suppression of hepatic TXA 2 production and lipid peroxidation, and an increase in antioxidative defence, leading to the prevention of hepatic fibrosis.

Oxidative stress mediates cell injury during ischaemia/reperfusion. On the other hand, experimental findings suggest that ROS (reactive oxygen species) induce processes leading to ischaemic preconditioning. The extent and source of oxidative stress and its effect on antioxidant status in the human liver during intermittent ischaemia and reperfusion remains ill-defined. Therefore the aim of the present study was to investigate the occurrence of oxidative stress in humans undergoing liver resection. Liver biopsies, and arterial and hepatic venous blood samples were taken from ten patients undergoing hepatectomy with an intermittent Pringle manoeuvre. Plasma MDA (malondialdehyde) and hepatic GSSG levels were measured as markers of oxidative stress and plasma uric acid as a marker of xanthine oxidase activity. In addition, changes in hepatosplanchnic consumption of plasma antioxidants and hepatic levels of carotenoids and glutathione (GSH) were measured. After ischaemia, hepatosplanchnic release of MDA and increased hepatic GSSG levels were found. This was accompanied by the release of uric acid, reflecting xanthine oxidase activity. During reperfusion, ongoing oxidative stress was observed by further increases in hepatic GSSG content and hepatosplanchnic MDA release. Uric acid release was minimal during reperfusion. A gradual decrease in plasma antioxidant capacity and net hepatosplanchnic antioxidant uptake was observed upon prolonged cumulative ischaemia. Oxidative stress occurs during hepatic ischaemia in man mainly due to xanthine oxidase activity. Interestingly, the gradual decline in plasma antioxidant capacity and net hepatosplanchnic antioxidant uptake during prolonged cumulative ischaemia, preserved both hydrophilic and lipophilic hepatic antioxidant levels. Decreasing plasma levels and net hepatosplanchnic uptake of plasma antioxidants may warrant antioxidant supplementation, although it should be clarified to what extent limitation of oxidative stress compromises ROS-dependent pathways of ischaemic preconditioning.

Current strategies to lower the incidence of ISR (in-stent restenosis) following PCI (percutaneous coronary intervention) are aimed at modifying arterial healing after stent injury. This can impair endothelial recovery and render the vessel prone to acute thrombosis. As early restoration of endothelial integrity inhibits neointimal growth and thrombosis, alternative approaches which encourage this process may provide a more effective long-term result after PCI. Oxidative stress is enhanced after PCI and participates in the regulation of endothelial regeneration and neointimal growth. Moreover, evidence suggests antioxidants improve re-endothelialization and inhibit ISR. By promoting, rather than blocking, the healing process, antioxidant and other therapies may offer an alternative or additional approach over the antiproliferative approaches common to many current devices.

Free radical production is increased during diabetes. Serum albumin is a major antioxidant agent, and structural modification of albumin induced by glucose or free radicals impairs its antioxidant properties. Therefore the aim of the present study was to compare the antioxidant capacities and structural changes in albumin in patients with T2DM (Type 2 diabetes mellitus) treated with MET (metformin) or SU (sulfonylureas) and in healthy control subjects. Structural changes in albumin were studied by fluorescence quenching in the presence of acrylamide. Albumin thiols and fructosamines, reflecting oxidized and glycation-induced changes in serum albumin respectively, were assessed. Structural changes in albumin were demonstrated by a significant decrease in fluorescence quenching in patients with T2DM, with patients treated with MET exhibiting a significant difference in the conformation of albumin compared with patients treated with SU. Oxidation, resulting in a significant decrease in thiol groups and plasma total antioxidant capacity, and glycation, associated with a significant increase in fructosamines, were both found when comparing healthy control subjects with patients with T2DM. When patients treated with MET were compared with those treated with SU, oxidative stress and glycation were found to be significantly lower in MET-treated patients. In conclusion, patients with T2DM have a decrease in the antioxidant properties of serum albumin which may aggravate oxidative stress and, thus, contribute to vascular and metabolic morbidities. Moreover, a significant protection of albumin was found in patients with T2DM treated with MET.

In the present study, the effect of vitamin E (α-tocopherol) on mice skeletal muscle mitochondrial dysfunction and oxidative damage induced by an in vivo acute and severe hypobaric hypoxic insult (48 h at a barometric pressure equivalent to 8500 m) has been investigated. Male mice ( n =24) were randomly divided into the following four groups ( n =6): control (C), hypoxia (H), vitamin E (VE; 60 mg/kg of body weight intraperitoneally, three times/week for 3 weeks) and hypoxia+VE (HVE). A significant increase in mitochondrial protein CGs (carbonyl groups) was found in the H group compared with the C group. Confirming previous observations from our group, hypoxia induced mitochondrial dysfunction, as identified by altered respiratory parameters. Hypoxia exposure increased Bax content and decreased the Bcl-2/Bax ratio, whereas Bcl-2 remained unchanged. Inner and outer mitochondrial membrane integrity were significantly affected by hypoxia exposure; however, vitamin E treatment attenuated the effect of hypoxia on mitochondrial oxidative phosphorylation and on the levels of CGs. Vitamin E supplementation also prevented the Bax and Bcl-2/Bax ratio impairments caused by hypoxia, as well as the decrease in inner and outer mitochondrial membrane integrity. In conclusion, the results suggest that vitamin E prevents the loss of mitochondrial integrity and function, as well as the increase in Bax content, which suggests that mitochondria are involved in increased cell death induced by severe hypobaric hypoxia in mice skeletal muscle.

Previous studies have postulated the association between oxidative stress and Type 2 diabetes. Considering the long pre-diabetic period with IGR (impaired glucose regulation) and its high risk of developing diabetes, to test this hypothesis, we have investigated oxidative stress pathways and DNA damage in patients with IGR and newly diagnosed Type 2 diabetes. The study population consisted of 92 subjects with NGT (normal glucose tolerance), 78 patients with IGR and 113 patients with newly diagnosed diabetes. Plasma MDA (malondialdehyde) and TAC (total antioxidative capacity) status, erythrocyte GSH content and SOD (superoxide dismutase) activity were determined. A comet assay was employed to evaluate DNA damage. Compared with subjects with NGT, patients with IGR had reduced erythrocyte SOD activity. Patients with diabetes had a higher plasma MDA concentration, but a lower plasma TAC level and erythrocyte SOD activity, than the NGT group. Correlation analysis revealed a strong positive association between IR (insulin resistance) and MDA concentration, but negative correlations with TAC status and SOD activity. With respect to β-cell function, a positive association with TAC status and an inverse correlation with GSH respectively, were observed. The comet assay revealed slight DNA damage in patients with IGR, which was increased in patients with diabetes. Significant correlations were observed between DNA damage and hyperglycaemia, IR and β-cell dysfunction. In conclusion, the results of the present study suggest that hyperglycaemia in an IGR state caused the predominance of oxidative stress over antioxidative defence systems, leading to oxidative DNA damage, which possibly contributed to pancreatic β-cell dysfunction, IR and more pronounced hyperglycaemia. This vicious circle finally induced the deterioration to diabetes.

Engaging in regular physical activity reduces the risk of developing CVD (cardiovascular disease), but it is not certain to what degree this may be due to the anti-inflammatory effects of exercise. Following acute exercise, there is a transient increase in circulating levels of anti-inflammatory cytokines, whereas chronic exercise reduces basal levels of pro-inflammatory cytokines. Exercise training also induces the expression of antioxidant and anti-inflammatory mediators in the vascular wall that may directly inhibit the development of atherosclerosis. Limited studies in humans and more comprehensive assessments in animal models have confirmed that exercise is atheroprotective and helped identify a number of the mechanisms to explain these effects. This review explores the relationship between systemic and vascular wall inflammation and the role that the anti-inflammatory effects of exercise have on the development and progression of CVD.

Maximal exercise in normoxia results in oxidative stress due to an increase in free radical production. However, the effect of a single bout of moderate aerobic exercise performed in either relative or absolute normobaric hypoxia on free radical production and lipid peroxidation remains unknown. To examine this, we randomly matched {according to their normobaric normoxic V ̇ O 2peak [peak V ̇ O 2 (oxygen uptake)]} and assigned 30 male subjects to a normoxia ( n =10), a hypoxia relative ( n =10) or a hypoxia absolute ( n =10) group. Each group was required to exercise on a cycle ergometer at 55% of V ̇ O 2peak for 2 h double-blinded to either a normoxic or hypoxic condition [ F i O 2 (inspired fraction of O 2 )=0.21 and 0.16 respectively]. ESR (electron spin resonance) spectroscopy in conjunction with ex vivo spin trapping was utilized for the direct detection of free radical species. The main findings show that moderate intensity exercise increased plasma-volume-corrected free radical and lipid hydroperoxide concentration (pooled rest compared with exercise data, P <0.05); however, there were no selective differences between groups (state×group interaction, P >0.05). The delta change in free radical concentration was moderately correlated with systemic V ̇ O 2 ( r 2 =0.48, P <0.05). The hyperfine coupling constants recorded from the ESR spectra [a N =13.8 Gauss, and a H β =1.9 Gauss; where 1 Gauss=10 −4 T (telsa)] are suggestive of oxygen-centred free radical species formed via the decomposition of lipid hydroperoxides. Peripheral leucocyte and neutrophil cells and total CK (creatine kinase) activity all increased following sustained exercise (pooled rest compared with exercise data, P <0.05), but no selective differences were observed between groups (state×group interaction, P >0.05). We conclude that a single bout of moderate aerobic exercise increases secondary free radical species. There is also evidence of exercise-induced muscle damage, possibly caused by the increase in free radical generation.

The present study examined if free radicals and associated inflammatory sequelae influenced metabolic biomarkers involved in the neuro-endocrinological regulation of energy homoeostasis at high altitude. Sixteen mountaineers (11 males/five females) were matched for physical fitness and caloric intake and assigned in a double-blind manner to either antioxidant ( n =8) or placebo ( n =8) supplementation, which was enforced for 7 days at sea level and during an 11-day ascent to 4780 m. Enteral prophylaxis incorporated a daily bolus dose of 1 g of L -ascorbate, 400 international units of D , L -α-tocopherol acetate and 600 mg of α-lipoic acid. EPR (electron paramagnetic resonance) spectroscopic detection of PBN (α-phenyl- tert -butylnitrone) adducts confirmed an increase in the venous concentration of carbon-centred radicals at high altitude in the placebo group, whereas a decrease was observed in the antioxidant group ( P <0.05 compared with that at sea level). EPR detection of DMSO/A˙ − (DMSO-supplemented ascorbate free radical) demonstrated that the increase in carbon-centred radicals at high altitude was associated with a decrease in ascorbate ( r 2 =0.63; P <0.05). Ascent to high altitude (pooled placebo+antioxidant groups) also increased the expression of pro-inflammatory cytokines ( P <0.05 compared with that at sea level) and biomarkers of skeletal tissue damage ( P <0.05). Despite a general decrease in leptin, insulin and glucose at high altitude (pooled placebo+antioxidant groups; P <0.05 compared with that at sea level), persistent anorexia resulted in a selective loss of body fat ( P <0.05). In conclusion, antioxidant prophylaxis decreased the concentration of carbon-centred radicals at high altitude ( P <0.05 compared with the placebo group), but did not influence markers of inflammation, appetite-related peptides, ad libitum nutrient intake or body composition. Thus free radicals do not appear to be involved in the inflammatory response and subsequent control of eating behaviour at high altitude.

Elevated circulating levels of NEFAs (non-esterified fatty acids) are associated with states of insulin resistance and increased risk of vascular disease. Previous animal and human studies have demonstrated NEFA-induced endothelial dysfunction of large conduit arteries, reversible by the antioxidant ascorbic acid. We therefore investigated the effect of NEFAs on carbachol-induced endothelium-dependent vasodilation of rat resistance arteries in vitro using the technique of wire myography. In addition, we investigated the effect of co-incubation of NEFAs and ascorbic acid. Cumulative concentration–response curves to carbachol (endothelium-dependent vasodilation) and SNAP ( S -nitroso- N -acetyl- DL -penicillamine; endothelium-independent vasodilation) were constructed. Those to carbachol were repeated following a 30 min incubation with either oleic acid (10 −4 M) or palmitic acid (10 −4 M), demonstrating significant impairment of endothelium-dependent vasodilation with both [ P <0.05, comparison of pD 2 values (the negative log concentration of agonist required to effect a 50% response)]. A cumulative concentration–response curve to carbachol was repeated following co-incubation with palmitic acid (10 −4 M) and the antioxidant ascorbic acid (10 −5 M), demonstrating an abolition of the previously observed endothelial dysfunction induced by palmitic acid. There was no impairment of vasodilation to SNAP following NEFA incubation. We conclude that NEFAs directly impair endothelial function in rat resistance arteries via an increase in oxidative stress at the vascular endothelium.

Physical activity is associated with beneficial changes in serum lipids, but exhaustive exercise has been suggested to increase oxidative stress. To test the effect of ascorbate (vitamin C) on serum lipids and the metabolism of urate, which is the most important intrinsic antioxidant, during exhaustive exercise, we performed a randomized, blinded, placebo-controlled study on eight male well-trained athletes. subjects were randomly allocated to either a group given 1000 mg of ascorbate daily ( n =4) or a placebo group ( n =4). Fasting serum lipids and urate concentrations were measured before and after 3 weeks of training. Although serum low-density lipoprotein (LDL)-cholesterol levels decreased and high-density lipoprotein (HDL)-cholesterol levels increased significantly in the ascorbate group after the 3 weeks of training, serum LDL-cholesterol levels increased and HDL-cholesterol levels decreased significantly in the placebo group. Furthermore, serum urate levels were elevated significantly in the placebo group; however, these levels did not change in the ascorbate group. When compared with the placebo group, significantly higher serum HDL-cholesterol and lower serum LDL-cholesterol and urate levels were observed in the ascorbate group after training. In conclusion, our results suggested that ascorbate may contribute to the desirable changes in serum lipids during exhaustive training and suggest the significant association between ascorbate and urate under intense training.

Uric acid (UA) possesses free-radical-scavenging properties, and systemic administration is known to increase serum antioxidant capacity. However, it is not known whether this protects against oxidative stress. The effects of raising UA concentration were studied during acute aerobic physical exercise in healthy subjects, as a model of oxidative stress characterized by increased circulating 8-iso-prostaglandin F 2α (8-iso-PGF 2α ) concentrations. Twenty healthy subjects were recruited to a randomized double-blind placebo-controlled crossover study, and underwent systemic administration of 0.5 g of UA in 250 ml of 0.1% lithium carbonate/4% dextrose vehicle or vehicle alone as control. subjects performed high-intensity aerobic exercise for 20 min to induce oxidative stress. Plasma 8-iso-PGF 2α concentrations were determined at baseline, after exercise and after recovery for 20 min. A single bout of high-intensity exercise caused a significant increase in plasma 8-iso-PGF 2α concentrations from 35.0±4.7 pg/ml to 45.6±6.7 pg/ml ( P <0.01). UA administration raised serum urate concentration from 293±16 to 487±16 μmol/l ( P <0.001), accompanied by increased serum antioxidant capacity from 1786±39 to 1899±45 μmol/l ( P <0.01). UA administration abolished the exercise-induced elevation of plasma 8-iso-PGF 2α concentrations. High UA concentrations are associated with increased serum antioxidant capacity and reduced oxidative stress during acute physical exercise in healthy subjects. These findings indicate that the antioxidant properties of UA are of biological importance in vivo .

Methionine loading seems to be accompanied by increased oxidative stress and damage. However, it is not known how this oxidative stress is generated. We performed the present crossover study to further elucidate the effects of methionine loading on oxidative stress in the blood of healthy volunteers, and to examine possible preventative effects of N -acetylcysteine (NAC) administration. A total of 18 healthy subjects were given two oral methionine loads of 100 mg/kg body weight, 4 weeks apart, one without NAC (Met group), and one in combination with supplementation with 2×900 mg doses of NAC (Met+NAC group). Blood samples were collected before and 2, 4, 8 and 24 h after methionine loading for measurements of thiol levels, protein carbonyls, lipid peroxidation, cellular fibronectin and ferric reducing ability of plasma (FRAP; i.e. antioxidant capacity). After methionine loading, whole-blood levels of free and oxidized cysteine and homocysteine were increased in both groups. Furthermore, the total plasma levels of homocysteine were higher, whereas those of cysteine were lower, after methionine loading in both groups. Lower levels of oxidized homocysteine and a higher free/oxidized ratio were found in the Met+NAC group compared with the Met group. Although the antioxidant capacity decreased after methionine loading, no major changes over time were found for protein carbonyls or cellular fibronectin in either group. Our results suggest that methionine loading may initiate the generation of reactive oxygen species by the (auto)-oxidation of homocysteine. In addition, supplementation with NAC seems to be able to partially prevent excessive increases in the levels of homocysteine in plasma and of oxidized homocysteine in whole blood, and might thereby contribute to the prevention of oxidative stress.

1. Albumin is often administered intravenously to critically ill patients as a volume expander, to combat hypoalbuminaemia, and to decrease hyperbilirubinaemia. There is, however, an ongoing debate concerning the therapeutic benefit of the former which is an expensive form of treatment. 2. Albumin has several biological functions, in particular as a ligand binder. It also acts as an extracellular transition metal ion-binding and radical-scavenging antioxidant. These functions are influenced by the presence of an exposed thiol group (cys 34) on the surface of the albumin molecule. 3. The ability of infused albumin to influence the plasma thiol pool, and hence antioxidant potential, was investigated in patients with sepsis syndrome. 4. Plasma thiol levels rose rapidly after albumin infusion and remained elevated even after plasma albumin levels had declined significantly, due to interstitial leakage. Data are suggestive of some form of thiol exchange in the plasma of these patients between albumin and molecules containing oxidized thiol groups. 5. Administration of albumin to patients with sepsis syndrome leads to a sustained increase in plasma thiols. Thiols have several important antioxidant functions, and thiol repletion in these patients, who are known to suffer from oxidative stress, may have beneficial antioxidant effects. Antioxidant repletion may represent an important facet of clinically administered albumin.

1. A new method has been developed for measuring the total antioxidant capacity of body fluids and drug solutions, based on the absorbance of the ABTS*+ radical cation. 2. An automated method for use on a centrifugal analyser, as well as a manual method, is described. 3. The procedure has been applied to physiological antioxidant compounds and radical-scavenging drugs, and an antioxidant ranking was established based on their reactivity relative to a 1.0 mmol/l Trolox standard. 4. The Trolox equivalent antioxidant capacity of plasma from an adult reference population has been measured, and the method optimized and validated. 5. The method has been applied to investigate the total plasma antioxidant capacity of neonates and how this may be compromised in prematurity.

1. The copper-containing protein caeruloplasmin has several oxidase activities. 2. Its ability to catalyse the oxidation of ferrous ions to the ferric state (ferroxidase activity) makes it an important antioxidant in vivo . 3. Recent reports have suggested that oral supplementation with vitamin C can inhibit the oxidase activities of caeruloplasmin. 4. As expected, damage to DNA and membrane lipids was stimulated by mixtures of iron salt and ascorbate, and this damage could be inhibited by caeruloplasmin provided the molar ratio of ascorbate to caeruloplasmin was kept sufficiently low. 5. When the molar ratio of ascorbate to caeruloplasmin was greater than 200 substantial loss of ferroxidase antioxidant activity occurred. 6. It is unlikely, however, that oral supplementation with vitamin C can raise plasma levels sufficiently to inhibit caeruloplasmin activity in vivo.

1. The burst of damaging oxygen free-radicals at the time of reperfusion is one of the crucial factors affecting skin flap survival after an ischaemic interval. In these experiments the efficacy of the antioxidant and free-radical scavenger N -acetylcysteine in improving the survival of ischaemic rabbit epigastric skin flaps was tested. 2. At the time of reperfusion flaps were given: (1) balanced salt solution by intravenous whole-body administration, (2) N -acetylcysteine (200 mg/kg) by intravenous whole-body administration, (3) balanced salt solution by intra-arterial infusion into the flap, (4) N -acetylcysteine (20 mg/kg) by intra-arterial infusion into the flap, or (5) N -acetylcysteine (200 mg/kg) by intra-arterial infusion into the flap. Flap survival at 1 week, and tissue levels of parameters related to free-radical production, blood levels of thromboxane B 2 and peripheral resistance during reperfusion were determined. 3. Compared with controls (groups 1 and 3) which had flap survival rates (expressed as percentage surface area surviving) of 27.1% and 31.6%, respectively, N -acetylcysteine treatment in group 2 (55.2%) and group 4 (51.9%) resulted in significant ( P < 0.05) improvements in flap survival. The survival rate in group 5 (37.7%) was not significantly better than that of the controls. 4. N -Acetylcysteine significantly reduced parameters related to free-radical production in the skin flap after 30 min of reperfusion, determined as tissue levels of malonyldialdehyde and protein oxidation products. There was also a significant decrease in peripheral resistance when low-dose N -acetylcysteine (group 4) was infused intra-arterially into the flap. The systemic levels of thromboxane B 2 , a prostanoid promoting thrombosis, were not significantly altered by administration of N -acetylcysteine. 5. N -Acetylcysteine proved to be a successful therapeutic agent for the salvage of experimental ischaemic rabbit skin flaps, when used as a low-dose intra-arterial or high-dose intravenous treatment. The possible clinical benefits in humans have yet to be demonstrated.