Inter-individual variability in redox and performance responses after antioxidant supplementation: A randomized double blind crossover study.

AIM: We aimed to investigate the inter-individual variability in redox and physiological responses of antioxidant-deficient subjects after antioxidant supplementation. METHODS: Two hundred individuals were sorted by plasma vitamin C levels. A low vitamin C group (n = 22) and a control group (n = 22) were compared in terms of oxidative stress and performance. Subsequently, the low vitamin C group received for 30 days vitamin C (1 g) or placebo, in randomized, double-blind, crossover fashion, and the effects were examined through a mixed-effects model, while individual responses were calculated. RESULTS: The low vitamin C group exhibited lower vitamin C (-25 µmol/L; 95%CI[-31.7, -18.3]; p < 0.001), higher F2 -isoprostanes (+17.1 pg/mL; 95%CI[6.5, 27.7]; p = 0.002), impaired VO2max (-8.2 mL/kg/min; 95%CI[-12.8, -3.6]; p < 0.001) and lower isometric peak torque (-41.5 Nm; 95%CI[-61.8, -21.2]; p < 0.001) compared to the control group. Regarding antioxidant supplementation, a significant treatment effect was found in vitamin C (+11.6 µmol/L; 95%CI[6.8, 17.1], p < 0.001), F2 -isoprostanes (-13.7 pg/mL; 95%CI[-18.9, -8.4], p < 0.001), VO2max (+5.4 mL/kg/min; 95%CI[2.7, 8.2], p = 0.001) and isometric peak torque (+18.7; 95%CI[11.8, 25.7 Nm], p < 0.001). The standard deviation for individual responses (SDir) was greater than the smallest worthwhile change (SWC) for all variables indicating meaningful inter-individual variability. When a minimal clinically important difference (MCID) was set, inter-individual variability remained for VO2max , but not for isometric peak torque. CONCLUSION: The proportion of response was generally high after supplementation (82.9%-95.3%); however, a few participants did not benefit from the treatment. This underlines the potential need for personalized nutritional interventions in an exercise physiology context.

Dietary Cysteine Intake is Associated with Blood Glutathione Levels and Isometric Strength.

Glutathione is the most abundant cellular antioxidant and regulates redox homeostasis. Healthy individuals with certain antioxidant inadequacies/deficiencies exhibit impairments in physiological functions. The aim was to investigate whether low levels of dietary cysteine intake are associated with a) lower erythrocyte glutathione, b) increased plasma F2-isoprostanes, and c) impaired muscle function. Towards this aim, we recorded the dietary intake of the three amino acids that synthesize glutathione (i. e., glutamic acid, cysteine, and glycine) in forty-one healthy individuals, and subsequently measured erythrocyte glutathione levels. Maximal isometric strength and fatigue index were also assessed using an electronic handgrip dynamometer. Our findings indicate that dietary cysteine intake was positively correlated with glutathione levels (r=0.765, p<0.001). In addition, glutathione levels were negatively correlated with F2-isoprostanes (r=- 0.311, p=0.048). An interesting finding was that glutathione levels and cysteine intake were positively correlated with maximal handgrip strength (r=0.416, p=0.007 and r=0.343, p=0.028, respectively). In conclusion, glutathione concentration is associated with cysteine intake, while adequate cysteine levels were important for optimal redox status and muscle function. This highlights the importance of proper nutritional intake and biochemical screening with the goal of personalized nutrition.

Antioxidant supplementation, redox deficiencies and exercise performance: A falsification design.

The aim of the present study was to validate the idea of personalized redox supplementation by subjecting individuals to targeted and non-targeted antioxidant supplementation schemes. Seventy-three volunteers were screened for plasma vitamin C and erythrocyte glutathione levels. Three groups were formed: i) the "low vitamin C″ group (12 individuals with the lowest vitamin C levels; Low VitC), ii) the "low glutathione" group (12 individuals with the lowest glutathione levels; Low GSH) and iii) a control group (12 individuals with moderate vitamin C and glutathione levels). The three groups received 1 g of vitamin C or 1.2 g of NAC daily for 30 days in a crossover design with a wash-out period of 30 days. Both antioxidant treatments reduced the increased resting systemic oxidative stress levels, assessed via urine F2-isoprostanes, in the Low VitC and Low GSH groups (P < .05). A significant group × time interaction (P < .05) was found for VO2max and isometric peak torque after both treatments, with the Low VitC and Low GSH groups exhibiting improved performance only after the targeted treatment (vitamin C and NAC, respectively). A significant group × time interaction (P < .05) was found for fatigue index after NAC treatment, but not after vitamin C treatment. No interaction was found for the Wingate test after both treatments. Most of the evidence verifies the idea that antioxidant supplementation increases performance when a particular deficiency is reversed. This indicates that the presence of oxidative stress per se does not rationalize the use of antioxidants and emphasizes the need to identify "responsive" phenotypes.

Chronic administration of plasma from exercised rats to sedentary rats does not induce redox and metabolic adaptations.

The present study aimed to investigate whether endurance exercise-induced changes in blood plasma composition may lead to adaptations in erythrocytes, skeletal muscle and liver. Forty sedentary rats were randomly distributed into two groups: a group that was injected with pooled plasma from rats that swam until exhaustion and a group that was injected with the pooled plasma from resting rats (intravenous administration at a dose of 2 mL/kg body weight for 21 days). Total antioxidant capacity, malondialdehyde and protein carbonyls were higher in the plasma collected from the exercised rats compared to the plasma from the resting rats. Νo significant difference was found in blood and tissue redox biomarkers and in tissue metabolic markers between rats that received the "exercised" or the "non-exercised" plasma (P > 0.05). Our results demonstrate that plasma injections from exercised rats to sedentary rats do not induce redox or metabolic adaptations in erythrocytes, skeletal muscle and liver.

Antioxidants in Personalized Nutrition and Exercise.

The present review highlights the idea that antioxidant supplementation can be optimized when tailored to the precise antioxidant status of each individual. A novel methodologic approach involving personalized nutrition, the mechanisms by which antioxidant status regulates human metabolism and performance, and similarities between antioxidants and other nutritional supplements are described. The usefulness of higher-level phenotypes for data-driven personalized treatments is also explained. We conclude that personally tailored antioxidant interventions based on specific antioxidant inadequacies or deficiencies could result in improved exercise performance accompanied by consistent alterations in redox profile.

N-acetylcysteine supplementation increases exercise performance and reduces oxidative stress only in individuals with low levels of glutathione.

Most of the evidence indicates that chronic antioxidant supplementation induces negative effects in healthy individuals. However, it is currently unknown whether specific redox deficiencies exist and whether targeted antioxidant interventions in deficient individuals can induce positive effects. We hypothesized that the effectiveness of antioxidant supplements to decrease oxidative stress and promote exercise performance depends on the redox status of the individuals that receive the antioxidant treatment. To this aim, we investigated whether N-acetylcysteine (NAC) supplementation would enhance exercise performance by increasing glutathione concentration and by reducing oxidative stress only in individuals with low resting levels of glutathione. We screened 100 individuals for glutathione levels and formed three groups with low, moderate and high levels (N = 36, 12 per group). After by-passing the regression to the mean artifact, by performing a second glutathione measurement, the individuals were supplemented with NAC (2 × 600mg, twice daily, for 30 days) or placebo using a double-blind cross-over design. We performed three whole-body performance tests (VO2max, time trial and Wingate), measured two systemic oxidative stress biomarkers (F2-isoprostanes and protein carbonyls) and assessed glutathione-dependent redox metabolism in erythrocytes (glutathione, glutathione peroxidase, glutathione reductase, superoxide dismutase, catalase and NADPH). The low glutathione group improved after NAC supplementation in VO2max, time trial and Wingate by 13.6%, 15.4% and 11.4%, respectively. Thirty days of NAC supplementation were sufficient to restore baseline glutathione concentration, reduce systemic oxidative stress and improve erythrocyte glutathione metabolism in the low glutathione group. On the contrary, the 30-day supplementation period did not affect performance and redox state of the moderate and high glutathione groups, although few both beneficial and detrimental effects in performance were observed. In conclusion, individuals with low glutathione levels were linked with decreased physical performance, increased oxidative stress and impaired redox metabolism of erythrocytes. NAC supplementation restored both performance and redox homeostasis.

Iron Supplementation Effects on Redox Status following Aseptic Skeletal Muscle Trauma in Adults and Children.

Exercise-induced skeletal muscle microtrauma is characterized by loss of muscle cell integrity, marked aseptic inflammatory response, and oxidative stress. We examined if iron supplementation would alter redox status after eccentric exercise. In a randomized, double blind crossover study, that was conducted in two cycles, healthy adults (n = 14) and children (n = 11) received daily either 37 mg of elemental iron or placebo for 3 weeks prior to and up to 72 h after an acute eccentric exercise bout. Blood was drawn at baseline, before exercise, and 72 h after exercise for the assessment of iron status, creatine kinase activity (CK), and redox status. Iron supplementation at rest increased iron concentration and transferrin saturation (p < 0.01). In adults, CK activity increased at 72 h after exercise, while no changes occurred in children. Iron supplementation increased TBARS at 72 h after exercise in both adults and children; no changes occurred under placebo condition. Eccentric exercise decreased bilirubin concentration at 72 h in all groups. Iron supplementation can alter redox responses after muscle-damaging exercise in both adults and children. This could be of great importance not only for healthy exercising individuals, but also in clinical conditions which are characterized by skeletal muscle injury and inflammation, yet iron supplementation is crucial for maintaining iron homeostasis. This study was registered at Clinicaltrials.gov Identifier: NCT02374619.

Plasma from exercised rats administered to sedentary rats induces systemic and tissue inflammation.

Recent studies have consistently supported the active role of blood in mediating biochemical and physiological tissue adaptations. However, no study has investigated the possible contribution of circulating factors in an exercise setting. The aim of the study was to investigate the role of circulating factors in exercise adaptations by chronically administering to sedentary animals blood plasma collected from acutely exercised animals. Phase 1: Blood plasma was collected from rats that swam to exhaustion and from sedentary rats. Phase 2: Other rats were divided into two groups (n = 20 per group): the first group involved rats that were injected intravenously with blood plasma originating from rats that previously swam to exhaustion, the second group consisted of rats that were injected intravenously with blood plasma originating from sedentary rats. Tail-vein injections (2 mL/kg) were performed daily for 21 consecutive days. Inflammatory markers (C-reactive protein, interleukins-1α, 2, 6, 8, 10 and tumor necrosis factor-a) were measured in blood plasma, muscle, and adipose tissue. Sedentary rats administered with plasma from exercised rats had significantly higher levels in all inflammatory markers measured in blood, skeletal muscle and adipose tissue, compared to the sedentary rats administered with resting plasma. Our data demonstrate that administration of "exercised" blood to sedentary rats induced inflammation in plasma, muscle and adipose tissue. Exercise adaptations are not solely due to intrinsic processes in muscle or adipose tissue. Blood factors also play a crucial role in mediating signals for tissue adaptations.

Passive smoking reduces and vitamin C increases exercise-induced oxidative stress: does this make passive smoking an anti-oxidant and vitamin C a pro-oxidant stimulus?

The current interpretative framework states that, for a certain experimental treatment (usually a chemical substance) to be classified as "anti-oxidant", it must possess the property of reducing (or even nullifying) exercise-induced oxidative stress. The aim of the study was to compare side by side, in the same experimental setup, redox biomarkers responses to an identical acute eccentric exercise session, before and after chronic passive smoking (considered a pro-oxidant stimulus) or vitamin C supplementation (considered an anti-oxidant stimulus). Twenty men were randomly assigned into either passive smoking or vitamin C group. All participants performed two acute eccentric exercise sessions, one before and one after either exposure to passive smoking or vitamin C supplementation for 12 days. Vitamin C, oxidant biomarkers (F2-isoprostanes and protein carbonyls) and the non-enzymatic antioxidant (glutathione) were measured, before and after passive smoking, vitamin C supplementation or exercise. It was found that chronic exposure to passive smoking increased the level of F2-isoprostanes and decreased the level of glutathione at rest, resulting in minimal increase or absence of oxidative stress after exercise. Conversely, chronic supplementation with vitamin C decreased the level of F2-isoprostanes and increased the level of glutathione at rest, resulting in marked exercise-induced oxidative stress. Contrary to the current scientific consensus, our results show that, when a pro-oxidant stimulus is chronically delivered, it is more likely that oxidative stress induced by subsequent exercise is decreased and not increased. Reversely, it is more likely to find greater exercise-induced oxidative stress after previous exposure to an anti-oxidant stimulus. We believe that the proposed framework will be a useful tool to reach more pragmatic explanations of redox biology phenomena.

Adipocytokine levels in children: effects of fatness and training.

To investigate the effects of obesity and exercise training on plasma adipocytokines a sample of 42 children (lean = 24, %BF = 17.8 ± 7.5%; obese = 18; %BF = 29.1 ± 9.3%; mean age = 12.4 ± 1.9 yrs), were divided into 4 age-matched for activity groups: lean inactive (n = 11), obese inactive (n = 9), lean active (n = 13) and obese active (n = 9). Active children participated in swimming training (≥1 year, ≥3 times/week, ≥1 h per session, covering a distance of 10,000-12,000 m per week).Obese individuals demonstrated greater visfatin levels (3.3 ± 1.3 ng/ml) than their lean counterparts (2.6 ± 1.1 ng/ml; p = .06) whereas adiponectin was significantly lower in obese children (3.8 ± 1.9) than their lean counterparts (5.9 ± 2.7; p £ .05). Insulin and HOMA values were significantly greater in obese compared with lean children (p £ .05). Within obese individuals, active individuals had significantly lower visfatin levels (2.8 ± 1.2 ng/ml) compared with their inactive counterparts (3.8 ± 1.2 ng/ml; p £ .05). Resistin levels were comparable between groups (p > .05). Childhood obesity elevates visfatin and lowers adiponectin levels whereas exercise training could reduce visfatin levels in obese children.

Exercise as a model to study redox homeostasis in blood: the effect of protocol and sampling point.

Twenty males ran either on a level treadmill (nonmuscle-damaging condition) or on a downhill treadmill (muscle-damaging condition). Blood and urine samples were collected before and after exercise (immediately after, 1h, 4h, 24h, 48h, and 96h). The following assays were performed: F(2)-isoprostanes in urine, protein carbonyls in plasma, glutathione, superoxide dismutase, glutathione peroxidase, and catalase in erythrocytes. The main finding was that monophasic redox responses were detected after nonmuscle-damaging exercise compared to the biphasic responses detected after muscle-damaging exercise. Based on these findings, muscle-damaging exercise may be a more appropriate experimental model to induce physiological oxidative stress.

No effect of antioxidant supplementation on muscle performance and blood redox status adaptations to eccentric training.

BACKGROUND: It was recently reported that antioxidant supplementation decreases training efficiency and prevents cellular adaptations to chronic exercise. OBJECTIVE: This study aimed to investigate the effects of vitamin C and vitamin E supplementation on muscle performance, blood and muscle redox status biomarkers, and hemolysis in trained and untrained men after acute and chronic exercise. A specific type of exercise was applied (eccentric) to produce long-lasting and extensive changes in redox status biomarkers and to examine more easily the potential effects of antioxidant supplementation. DESIGN: In a double-blinded fashion, men received either a daily oral supplement of vitamin C and vitamin E (n = 14) or placebo (n = 14) for 11 wk (started 4 wk before the pretraining exercise testing and continued until the posttraining exercise testing). After baseline testing, the subjects performed an eccentric exercise session 2 times/wk for 4 wk. Before and after the chronic eccentric exercise, the subjects underwent one session of acute eccentric exercise, physiologic measurements were performed, and blood samples and muscle biopsy samples (from 4 men) were collected. RESULTS: The results failed to support any effect of antioxidant supplementation. Eccentric exercise similarly modified muscle damage and performance, blood redox status biomarkers, and hemolysis in both the supplemented and nonsupplemented groups. This occurred despite the fact that eccentric exercise induced marked changes in muscle damage and performance and in redox status after exercise. CONCLUSION: The complete lack of any effect on the physiologic and biochemical outcome measures used raises questions about the validity of using oral antioxidant supplementation as a redox modulator of muscle and redox status in healthy humans.

Ergogenic and antioxidant effects of spirulina supplementation in humans.

PURPOSE: Spirulina is a popular nutritional supplement that is accompanied by claiMSS for antioxidant and performance-enhancing effects. Therefore, the aim of the present study was to examine the effect of spirulina supplementation on (i) exercise performance, (ii) substrate metabolism, and (iii) blood redox status both at rest and after exercise. METHODS: Nine moderately trained males took part in a double-blind, placebo-controlled, counterbalanced crossover study. Each subject received either spirulina (6 g x d(-1)) or placebo for 4 wk. Each subject ran on a treadmill at an intensity corresponding to 70%-75% of their VO2max for 2 h and then at 95% VO2max to exhaustion. Exercise performance and respiratory quotient during exercise were measured after both placebo and spirulina supplementation. Blood samples were drawn before, immediately after, and at 1, 24, and 48 h after exercise. Reduced glutathione (GSH), oxidized glutathione (GSSG), GSH/GSSG, thiobarbituric acid-reactive substances (TBARS), protein carbonyls, catalase activity, and total antioxidant capacity (TAC) were determined. RESULTS: Time to fatigue after the 2-h run was significantly longer after spirulina supplementation (2.05 +/- 0.68 vs 2.70 +/- 0.79 min). Ingestion of spirulina significantly decreased carbohydrate oxidation rate by 10.3% and increased fat oxidation rate by 10.9% during the 2-h run compared with the placebo trial. GSH levels were higher after the spirulina supplementation compared with placebo at rest and 24 h after exercise. TBARS levels increased after exercise after placebo but not after spirulina supplementation. Protein carbonyls, catalase, and TAC levels increased similarly immediately after and 1 h after exercise in both groups. CONCLUSIONS: Spirulina supplementation induced a significant increase in exercise performance, fat oxidation, and GSH concentration and attenuated the exercise-induced increase in lipid peroxidation.

Uniform and prolonged changes in blood oxidative stress after muscle-damaging exercise.

BACKGROUND: The effect of eccentric exercise on the time-course changes in several indices of muscle damage and blood oxidative stress was examined. MATERIALS AND METHODS: Isometric torque, delayed-onset muscle soreness, creatine kinase, reduced glutathione (GSH), oxidized glutathione (GSSG), thiobarbituric-acid reactive substances (TBARS), protein carbonyls, catalase, uric acid, bilirubin and total antioxidant capacity (TAC) in blood were measured pre-, 24 h, 48 h and 72 h post-eccentric exercise of knee extensors in ten females. RESULTS: The concentration of all oxidative stress indices changed significantly in a way indicating increased oxidative stress in the blood (GSH and GSH/GSSG, decreased, whereas GSSG, TBARS, protein carbonyls, catalase, uric acid, bilirubin and TAC increased) peaking, in all but TBARS, at 48 h and returning toward baseline afterwards. CONCLUSION: We believe that muscle-damaging exercise should be viewed as a different challenge compared to non-muscle-damaging exercise with regard to its effects on blood oxidative stress.

Decreased blood oxidative stress after repeated muscle-damaging exercise.

PURPOSE: To examine the effect of repeated muscle-damaging exercise on the time-course changes in several indices of muscle damage, and to compare them with changes in blood oxidative stress indices. METHODS: Twelve females underwent an isokinetic exercise session consisting of 75 lengthening knee flexions, which was repeated after 3 wk. Isometric torque, range of movement (ROM), delayed-onset muscle soreness (DOMS), creatine kinase (CK), reduced glutathione (GSH), oxidized glutathione (GSSG), thiobarbituric acid-reactive substances (TBARS), protein carbonyls, catalase, uric acid, bilirubin, and total antioxidant capacity (TAC) in blood were measured before, immediately after, and 1, 2, 3, 4, and 7 d after lengthening contractions. RESULTS: All muscle damage indices (torque, ROM, DOMS, and CK) changed significantly after exercise. The concentration of all oxidative stress indices changed significantly in a way indicating increased oxidative stress in the blood (GSH and GSH/GSSG decreased, whereas GSSG, TBARS, protein carbonyls, catalase, uric acid, bilirubin, and TAC increased), peaking in all but bilirubin at 3 d and returning to baseline values by 7 d after exercise. The repeated bout of lengthening contractions induced significantly less changes in indices of muscle damage and blood oxidative stress than the first bout. In general, from the increasing or decreasing area under the curve calculated for each oxidative stress index, the second bout produced 1.8- to 6.1-fold less changes in oxidative stress than after the first bout. CONCLUSION: A repeated bout of lengthening contractions attenuated muscle damage and blood oxidative stress compared with the first bout.