Unlocking peak performance: The role of Nrf2 in enhancing exercise outcomes and training adaptation in humans.

Since the discovery of the nuclear factor erythroid-derived 2-like 2 (Nrf2) transcription factor thirty years ago, it has been shown that it regulates more than 250 genes involved in a multitude of biological processes, including redox balance, mitochondrial biogenesis, metabolism, detoxification, cytoprotection, inflammation, immunity, autophagy, cell differentiation, and xenobiotic metabolism. In skeletal muscle, Nrf2 signalling is primarily activated in response to perturbation of redox balance by reactive oxygen species or electrophiles. Initial investigations into human skeletal muscle Nrf2 responses to exercise, dating back roughly a decade, have consistently indicated that exercise-induced ROS production stimulates Nrf2 signalling. Notably, recent studies employing Nrf2 knockout mice have revealed impaired skeletal muscle contractile function characterised by reduced force output and increased fatigue susceptibility compared to wild-type counterparts. These deficiencies partially stem from diminished basal mitochondrial respiratory capacity and an impaired capacity to upregulate specific mitochondrial proteins in response to training, findings corroborated by inducible muscle-specific Nrf2 knockout models. In humans, baseline Nrf2 expression in skeletal muscle correlates with maximal oxygen uptake and high-intensity exercise performance. This manuscript delves into the mechanisms underpinning Nrf2 signalling in response to acute exercise in human skeletal muscle, highlighting the involvement of ROS, antioxidants and Keap1/Nrf2 signalling in exercise performance. Furthermore, it explores Nrf2's role in mediating adaptations to chronic exercise and its impact on overall exercise performance. Additionally, the influence of diet and certain supplements on basal Nrf2 expression and its role in modulating acute and chronic exercise responses are briefly addressed.

Endurance Training Improves Leg Proton Release and Decreases Potassium Release During High-Intensity Exercise in Normoxia and Hypobaric Hypoxia.

AIM: To assess the impact of endurance training on skeletal muscle release of H+ and K+. METHODS: Nine participants performed one-legged knee extension endurance training at moderate and high intensities (70%-85% of Wpeak), three to four sessions·week-1 for 6 weeks. Post-training, the trained and untrained (control) leg performed two-legged knee extension at low, moderate, and high intensities (40%, 62%, and 83% of Wpeak) in normoxia and hypoxia (~4000 m). The legs were exercised simultaneously to ensure identical arterial inflow concentrations of ions and metabolites, and identical power output was controlled by visual feedback. Leg blood flow was measured (ultrasound Doppler), and acid-base variables, lactate- and K+ concentrations were assessed in arterial and femoral venous blood to study K+ and H+ release. Ion transporter abundances were assessed in muscle biopsies. RESULTS: Lactate-dependent H+ release was similar in hypoxia to normoxia (p = 0.168) and was lower in the trained than the control leg at low-moderate intensities (p = 0.060-0.006) but similar during high-intensity exercise. Lactate-independent and total H+ releases were higher in hypoxia (p < 0.05) and increased more with power output in the trained leg (leg-by-power output interactions: p = 0.02). K+ release was similar at low intensity but lower in the trained leg during high-intensity exercise in normoxia (p = 0.024) and hypoxia (p = 0.007). The trained leg had higher abundances of Na+/H+ exchanger 1 (p = 0.047) and Na+/K+ pump subunit α (p = 0.036). CONCLUSION: Moderate- to high-intensity endurance training increases lactate-independent H+ release and reduces K+ release during high-intensity exercise, coinciding with increased Na+/H+ exchanger 1 and Na+/K+ pump subunit α muscle abundances.

Physiological and molecular predictors of cycling sprint performance.

The study aimed to identify novel muscle phenotypic factors that could determine sprint performance using linear regression models including the lean mass of the lower extremities (LLM), myosin heavy chain composition (MHC), and proteins and enzymes implicated in glycolytic and aerobic energy generation (citrate synthase, OXPHOS proteins), oxygen transport and diffusion (myoglobin), ROS sensing (Nrf2/Keap1), antioxidant enzymes, and proteins implicated in calcium handling. For this purpose, body composition (dual-energy X-ray absorptiometry) and sprint performance (isokinetic 30-s Wingate test: peak and mean power output, Wpeak and Wmean ) were measured in young physically active adults (51 males and 10 females), from which a resting muscle biopsy was obtained from the musculus vastus lateralis. Although females had a higher percentage of MHC I, SERCA2, pSer16 /Thr17 -phospholamban, and Calsequestrin 2 protein expressions (all p < 0.05), and 18.4% lower phosphofructokinase 1 protein expression than males (p < 0.05), both sexes had similar sprint performance when it was normalized to body weight or LLM. Multiple regression analysis showed that Wpeak could be predicted from LLM, SDHB, Keap1, and MHC II % (R 2 = 0.62, p < 0.001), each variable contributing to explain 46.4%, 6.3%, 4.4%, and 4.3% of the variance in Wpeak , respectively. LLM and MHC II % explained 67.5% and 2.1% of the variance in Wmean , respectively (R 2 = 0.70, p < 0.001). The present investigation shows that SDHB and Keap1, in addition to MHC II %, are relevant determinants of peak power output during sprinting.

A Mango Leaf Extract (Zynamite®) Combined with Quercetin Has Exercise-Mimetic Properties in Human Skeletal Muscle.

Zynamite PX®, a mango leaf extract combined with quercetin, enhances exercise performance by unknown molecular mechanisms. Twenty-five volunteers were assigned to a control (17 males) or supplementation group (8 males, receiving 140 mg of Zynamite® + 140 mg quercetin/8 h for 2 days). Then, they performed incremental exercise to exhaustion (IE) followed by occlusion of the circulation in one leg for 60 s. Afterwards, the cuff was released, and a 30 s sprint was performed, followed by 90 s circulatory occlusion (same leg). Vastus lateralis muscle biopsies were obtained at baseline, 20 s after IE (occluded leg) and 10 s after Wingate (occluded leg), and bilaterally at 90 s and 30 min post exercise. Compared to the controls, the Zynamite PX® group showed increased basal protein expression of Thr287-CaMKIIδD (2-fold, p = 0.007) and Ser9-GSK3ß (1.3-fold, p = 0.005) and a non-significant increase of total NRF2 (1.7-fold, p = 0.099) and Ser40-NRF2 (1.2-fold, p = 0.061). In the controls, there was upregulation with exercise and recovery of total NRF2, catalase, glutathione reductase, and Thr287-CaMKIIδD (1.2-2.9-fold, all p < 0.05), which was not observed in the Zynamite PX® group. In conclusion, Zynamite PX® elicits muscle signaling changes in resting skeletal muscle resembling those described for exercise training and partly abrogates the stress kinases responses to exercise as observed in trained muscles.

Pulmonary ventilation and gas exchange during prolonged exercise in humans: Influence of dehydration, hyperthermia and sympathoadrenal activity.

NEW FINDINGS: What is the central question of the study? Ventilation increases during prolonged intense exercise, but the impact of dehydration and hyperthermia, with associated blunting of pulmonary circulation, and independent influences of dehydration, hyperthermia and sympathoadrenal discharge on ventilatory and pulmonary gas exchange responses remain unclear. What is the main finding and its importance? Dehydration and hyperthermia led to hyperventilation and compensatory adjustments in pulmonary CO2 and O2 exchange, such that CO2 output increased and O2 uptake remained unchanged despite the blunted circulation. Isolated hyperthermia and adrenaline infusion, but not isolated dehydration, increased ventilation to levels similar to combined dehydration and hyperthermia. Hyperthermia is the main stimulus increasing ventilation during prolonged intense exercise, partly via sympathoadrenal activation. ABSTRACT: The mechanisms driving hyperthermic hyperventilation during exercise are unclear. In a series of retrospective analyses, we evaluated the impact of combined versus isolated dehydration and hyperthermia and the effects of sympathoadrenal discharge on ventilation and pulmonary gas exchange during prolonged intense exercise. In the first study, endurance-trained males performed two submaximal cycling exercise trials in the heat. On day 1, participants cycled until volitional exhaustion (135 ± 11 min) while experiencing progressive dehydration and hyperthermia. On day 2, participants maintained euhydration and core temperature (Tc ) during a time-matched exercise (control). At rest and during the first 20 min of exercise, pulmonary ventilation ( V ̇ E ${\skew2\dot V_{\rm{E}}}$ ), arterial blood gases, CO2 output and O2 uptake were similar in both trials. At 135 ± 11 min, however, V ̇ E ${\skew2\dot V_{\rm{E}}}$ was elevated with dehydration and hyperthermia, and this was accompanied by lower arterial partial pressure of CO2 , higher breathing frequency, arterial partial pressure of O2 , arteriovenous CO2 and O2 differences, and elevated CO2 output and unchanged O2 uptake despite a reduced pulmonary circulation. The increased V ̇ E ${\skew2\dot V_{\rm{E}}}$ was closely related to the rise in Tc and circulating catecholamines (R2  ≥ 0.818, P ≤ 0.034). In three additional studies in different participants, hyperthermia independently increased V ̇ E ${\skew2\dot V_{\rm{E}}}$ to an extent similar to combined dehydration and hyperthermia, whereas prevention of hyperthermia in dehydrated individuals restored V ̇ E ${\skew2\dot V_{\rm{E}}}$ to control levels. Furthermore, adrenaline infusion during exercise elevated both Tc and V ̇ E ${\skew2\dot V_{\rm{E}}}$ . These findings indicate that: (1) adjustments in pulmonary gas exchange limit homeostatic disturbances in the face of a blunted pulmonary circulation; (2) hyperthermia is the main stimulus increasing ventilation during prolonged intense exercise; and (3) sympathoadrenal activation might partly mediate the hyperthermic hyperventilation.

Angiotensin-Converting Enzyme 2 (SARS-CoV-2 receptor) expression in human skeletal muscle.

The study aimed to determine the levels of skeletal muscle angiotensin-converting enzyme 2 (ACE2, the SARS-CoV-2 receptor) protein expression in men and women and assess whether ACE2 expression in skeletal muscle is associated with cardiorespiratory fitness and adiposity. The level of ACE2 in vastus lateralis muscle biopsies collected in previous studies from 170 men (age: 19-65 years, weight: 56-137 kg, BMI: 23-44) and 69 women (age: 18-55 years, weight: 41-126 kg, BMI: 22-39) was analyzed in duplicate by western blot. VO2 max was determined by ergospirometry and body composition by DXA. ACE2 protein expression was 1.8-fold higher in women than men (p = 0.001, n = 239). This sex difference disappeared after accounting for the percentage of body fat (fat %), VO2 max per kg of legs lean mass (VO2 max-LLM) and age (p = 0.47). Multiple regression analysis showed that the fat % (ß = 0.47) is the main predictor of the variability in ACE2 protein expression in skeletal muscle, explaining 5.2% of the variance. VO2 max-LLM had also predictive value (ß = 0.09). There was a significant fat % by VO2 max-LLM interaction, such that for subjects with low fat %, VO2 max-LLM was positively associated with ACE2 expression while as fat % increased the slope of the positive association between VO2 max-LLM and ACE2 was reduced. In conclusion, women express higher amounts of ACE2 in their skeletal muscles than men. This sexual dimorphism is mainly explained by sex differences in fat % and cardiorespiratory fitness. The percentage of body fat is the main predictor of the variability in ACE2 protein expression in human skeletal muscle.

Contribution of oxygen extraction fraction to maximal oxygen uptake in healthy young men.

We analysed the importance of systemic and peripheral arteriovenous O2 difference ( a-v¯O2 difference and a-vf O2 difference, respectively) and O2 extraction fraction for maximal oxygen uptake ( V˙O2max ). Fick law of diffusion and the Piiper and Scheid model were applied to investigate whether diffusion versus perfusion limitations vary with V˙O2max . Articles (n = 17) publishing individual data (n = 154) on V˙O2max , maximal cardiac output ( Q˙max ; indicator-dilution or the Fick method), a-v¯O2 difference (catheters or the Fick equation) and systemic O2 extraction fraction were identified. For the peripheral responses, group-mean data (articles: n = 27; subjects: n = 234) on leg blood flow (LBF; thermodilution), a-vf O2 difference and O2 extraction fraction (arterial and femoral venous catheters) were obtained. Q˙max and two-LBF increased linearly by 4.9-6.0 L · min-1 per 1 L · min-1 increase in V˙O2max (R2  = .73 and R2  = .67, respectively; both P < .001). The a-v¯O2 difference increased from 118-168 mL · L-1 from a V˙O2max of 2-4.5 L · min-1 followed by a reduction (second-order polynomial: R2  = .27). After accounting for a hypoxemia-induced decrease in arterial O2 content with increasing V˙O2max (R2  = .17; P < .001), systemic O2 extraction fraction increased up to ~90% ( V˙O2max : 4.5 L · min-1 ) with no further change (exponential decay model: R2  = .42). Likewise, leg O2 extraction fraction increased with V˙O2max to approach a maximal value of ~90-95% (R2  = .83). Muscle O2 diffusing capacity and the equilibration index Y increased linearly with V˙O2max (R2  = .77 and R2  = .31, respectively; both P < .01), reflecting decreasing O2 diffusional limitations and accentuating O2 delivery limitations. In conclusion, although O2 delivery is the main limiting factor to V˙O2max , enhanced O2 extraction fraction (≥90%) contributes to the remarkably high V˙O2max in endurance-trained individuals.

Increased oxygen extraction and mitochondrial protein expression after small muscle mass endurance training.

When exercising with a small muscle mass, the mass-specific O2 delivery exceeds the muscle oxidative capacity resulting in a lower O2 extraction compared with whole-body exercise. We elevated the muscle oxidative capacity and tested its impact on O2 extraction during small muscle mass exercise. Nine individuals conducted six weeks of one-legged knee extension (1L-KE) endurance training. After training, the trained leg (TL) displayed 45% higher citrate synthase and COX-IV protein content in vastus lateralis and 15%-22% higher pulmonary oxygen uptake ( V ˙ O 2 peak ) and peak power output ( W ˙ peak ) during 1L-KE than the control leg (CON; all P < .05). Leg O2 extraction (catheters) and blood flow (ultrasound Doppler) were measured while both legs exercised simultaneously during 2L-KE at the same submaximal power outputs (real-time feedback-controlled). TL displayed higher O2 extraction than CON (main effect: 1.7 ± 1.6% points; P = .010; 40%-83% of W ˙ peak ) with the largest between-leg difference at 83% of W ˙ peak (O2 extraction: 3.2 ± 2.2% points; arteriovenous O2 difference: 7.1 ± 4.8 mL· L-1 ; P < .001). At 83% of W ˙ peak , muscle O2 conductance (DM O2 ; Fick law of diffusion) and the equilibration index Y were higher in TL (P < .01), indicating reduced diffusion limitations. The between-leg difference in O2 extraction correlated with the between-leg ratio of citrate synthase and COX-IV (r = .72-.73; P = .03), but not with the difference in the capillary-to-fiber ratio (P = .965). In conclusion, endurance training improves O2 extraction during small muscle mass exercise by elevating the muscle oxidative capacity and the recruitment of DM O2, especially evident during high-intensity exercise exploiting a larger fraction of the muscle oxidative capacity.

Supplementation with a Mango Leaf Extract (Zynamite®) in Combination with Quercetin Attenuates Muscle Damage and Pain and Accelerates Recovery after Strenuous Damaging Exercise.

Prolonged or unusual exercise may cause exercise-induced muscle damage (EIMD). To test whether Zynamite®, a mango leaf extract rich in the natural polyphenol mangiferin, administered in combination with quercetin facilitates recovery after EIMD, 24 women and 33 men were randomly assigned to two treatment groups matched by sex and 5 km running performance, and ran a 10 km race followed by 100 drop jumps to elicit EIMD. One hour before the competition, and every 8 hours thereafter for 24 hours, they ingested placebo (728 mg of maltodextrin) or 140 mg of Zynamite® combined with 140 mg of quercetin (double-blind). Although competition times were similar, polyphenol supplementation attenuated the muscle pain felt after the competition (6.8 ± 1.5 and 5.7 ± 2.2 a.u., p = 0.035) and the loss of jumping performance (9.4 ± 11.5 and 3.9 ± 5.2%, p = 0.036; p = 0.034) and mechanical impulse (p = 0.038) 24 hours later. The polyphenols attenuated the increase of serum myoglobin and alanine aminotransferase in men, but not in women (interaction p < 0.05). In conclusion, a single dose of 140 mg Zynamite® combined with 140 mg of quercetin, administered one hour before competition, followed by three additional doses every eight hours, attenuates muscle pain and damage, and accelerates the recovery of muscle performance.

A Single Dose of The Mango Leaf Extract Zynamite® in Combination with Quercetin Enhances Peak Power Output During Repeated Sprint Exercise in Men and Women.

The mango leaf extract rich in mangiferin Zynamite® improves exercise performance when combined with luteolin or quercetin ingested at least 48 h prior to exercise. To determine whether a single dose of Zynamite® administered 1 h before exercise increases repeated-sprint performance, 20 men and 20 women who were physically active were randomly assigned to three treatments following a double-blind cross-over counterbalanced design. Treatment A, 140 mg of Zynamite®, 140 mg of quercetin, 147.7 mg of maltodextrin, and 420 mg of sunflower lecithin; Treatment B, 140 mg of Zynamite®, 140 mg of quercetin, and 2126 mg of maltodextrin and Treatment C, 2548 mg of maltodextrin (placebo). Subjects performed three Wingate tests interspaced by 4 min and a final 15 s sprint after ischemia. Treatments A and B improved peak power output during the first three Wingates by 2.8% and 3.8%, respectively (treatment x sprint interaction, p = 0.01). Vastus Lateralis oxygenation (NIRS) was reduced, indicating higher O2 extraction (treatment × sprint interaction, p = 0.01). Improved O2 extraction was observed in the sprints after ischemia (p = 0.008; placebo vs. mean of treatments A and B). Blood lactate concentration was 5.9% lower after the ingestion of Zynamite® with quercetin in men (treatment by sex interaction, p = 0.049). There was a higher Vastus Lateralis O2 extraction during 60 s ischemia with polyphenols (treatment effect, p = 0.03), due to the greater muscle VO2 in men (p = 0.001). In conclusion, a single dose of Zynamite® combined with quercetin one hour before exercise improves repeated-sprint performance and muscle O2 extraction and mitochondrial O2. consumption during ischemia. No advantage was obtained from the addition of phospholipids.

Enhancement of Exercise Performance by 48 Hours, and 15-Day Supplementation with Mangiferin and Luteolin in Men.

The natural polyphenols mangiferin and luteolin have free radical-scavenging properties, induce the antioxidant gene program and down-regulate the expression of superoxide-producing enzymes. However, the effects of these two polyphenols on exercise capacity remains mostly unknown. To determine whether a combination of luteolin (peanut husk extract containing 95% luteolin, PHE) and mangiferin (mango leave extract (MLE), Zynamite®) at low (PHE: 50 mg/day; and 140 mg/day of MLE containing 100 mg of mangiferin; L) and high doses (PHE: 100 mg/day; MLE: 420 mg/day; H) may enhance exercise performance, twelve physically active men performed incremental exercise to exhaustion, followed by sprint and endurance exercise after 48 h (acute effects) and 15 days of supplementation (prolonged effects) with polyphenols or placebo, following a double-blind crossover design. During sprint exercise, mangiferin + luteolin supplementation enhanced exercise performance, facilitated muscle oxygen extraction, and improved brain oxygenation, without increasing the VO2. Compared to placebo, mangiferin + luteolin increased muscle O2 extraction during post-exercise ischemia, and improved sprint performance after ischemia-reperfusion likely by increasing glycolytic energy production, as reflected by higher blood lactate concentrations after the sprints. Similar responses were elicited by the two doses tested. In conclusion, acute and prolonged supplementation with mangiferin combined with luteolin enhances performance, muscle O2 extraction, and brain oxygenation during sprint exercise, at high and low doses.

Superior Intrinsic Mitochondrial Respiration in Women Than in Men.

Sexual dimorphism is apparent in humans, however, to date no studies have investigated mitochondrial function focusing on intrinsic mitochondrial respiration (i.e., mitochondrial respiration for a given amount of mitochondrial protein) and mitochondrial oxygen affinity (p50mito) in relation to biological sex in human. A skeletal muscle biopsy was donated by nine active women, and ten men matched for maximal oxygen consumption (VO2max) and by nine endurance trained men. Intrinsic mitochondrial respiration, assessed in isolated mitochondria, was higher in women compared to men when activating complex I (CIP) and complex I+II (CI+IIP) (p < 0.05), and was similar to trained men (CIP, p = 0.053; CI+IIP, p = 0.066). Proton leak and p50mito were higher in women compared to men independent of VO2max. In conclusion, significant novel differences in mitochondrial oxidative function, intrinsic mitochondrial respiration and p50mito exist between women and men. These findings may represent an adaptation in the oxygen cascade in women to optimize muscle oxygen uptake to compensate for a lower oxygen delivery during exercise.

Mangifera indica L. Leaf Extract in Combination With Luteolin or Quercetin Enhances VO2peak and Peak Power Output, and Preserves Skeletal Muscle Function During Ischemia-Reperfusion in Humans.

It remains unknown whether polyphenols such as luteolin (Lut), mangiferin and quercetin (Q) have ergogenic effects during repeated all-out prolonged sprints. Here we tested the effect of Mangifera indica L. leaf extract (MLE) rich in mangiferin (Zynamite®) administered with either quercetin (Q) and tiger nut extract (TNE), or with luteolin (Lut) on sprint performance and recovery from ischemia-reperfusion. Thirty young volunteers were randomly assigned to three treatments 48 h before exercise. Treatment A: placebo (500 mg of maltodextrin/day); B: 140 mg of MLE (60% mangiferin) and 50 mg of Lut/day; and C: 140 mg of MLE, 600 mg of Q and 350 mg of TNE/day. After warm-up, subjects performed two 30 s Wingate tests and a 60 s all-out sprint interspaced by 4 min recovery periods. At the end of the 60 s sprint the circulation of both legs was instantaneously occluded for 20 s. Then, the circulation was re-opened and a 15 s sprint performed, followed by 10 s recovery with open circulation, and another 15 s final sprint. MLE supplements enhanced peak (Wpeak) and mean (Wmean) power output by 5.0-7.0% (P < 0.01). After ischemia, MLE+Q+TNE increased Wpeak by 19.4 and 10.2% compared with the placebo (P < 0.001) and MLE+Lut (P < 0.05), respectively. MLE+Q+TNE increased Wmean post-ischemia by 11.2 and 6.7% compared with the placebo (P < 0.001) and MLE+Lut (P = 0.012). Mean VO2 during the sprints was unchanged, suggesting increased efficiency or recruitment of the anaerobic capacity after MLE ingestion. In women, peak VO2 during the repeated sprints was 5.8% greater after the administration of MLE, coinciding with better brain oxygenation. MLE attenuated the metaboreflex hyperpneic response post-ischemia, may have improved O2 extraction by the Vastus Lateralis (MLE+Q+TNE vs. placebo, P = 0.056), and reduced pain during ischemia (P = 0.068). Blood lactate, acid-base balance, and plasma electrolytes responses were not altered by the supplements. In conclusion, a MLE extract rich in mangiferin combined with either quercetin and tiger nut extract or luteolin exerts a remarkable ergogenic effect, increasing muscle power in fatigued subjects and enhancing peak VO2 and brain oxygenation in women during prolonged sprinting. Importantly, the combination of MLE+Q+TNE improves skeletal muscle contractile function during ischemia/reperfusion.

N1-methylnicotinamide is a signalling molecule produced in skeletal muscle coordinating energy metabolism.

Obesity is a major health problem, and although caloric restriction and exercise are successful strategies to lose adipose tissue in obese individuals, a simultaneous decrease in skeletal muscle mass, negatively effects metabolism and muscle function. To deeper understand molecular events occurring in muscle during weight-loss, we measured the expressional change in human skeletal muscle following a combination of severe caloric restriction and exercise over 4 days in 15 Swedish men. Key metabolic genes were regulated after the intervention, indicating a shift from carbohydrate to fat metabolism. Nicotinamide N-methyltransferase (NNMT) was the most consistently upregulated gene following the energy-deficit exercise. Circulating levels of N1-methylnicotinamide (MNA), the product of NNMT activity, were doubled after the intervention. The fasting-fed state was an important determinant of plasma MNA levels, peaking at ~18 h of fasting and being lowest ~3 h after a meal. In culture, MNA was secreted by isolated human myotubes and stimulated lipolysis directly, with no effect on glucagon or insulin secretion. We propose that MNA is a novel myokine that enhances the utilization of energy stores in response to low muscle energy availability. Future research should focus on applying MNA as a biomarker to identify individuals with metabolic disturbances at an early stage.

Skeletal muscle signaling, metabolism, and performance during sprint exercise in severe acute hypoxia after the ingestion of antioxidants.

The aim of this study was to determine if reactive oxygen species (ROS) could play a role in blunting Thr172-AMP-activated protein kinase (AMPK)-α phosphorylation in human skeletal muscle after sprint exercise in hypoxia and to elucidate the potential signaling mechanisms responsible for this response. Nine volunteers performed a single 30-s sprint (Wingate test) in two occasions while breathing hypoxic gas ([Formula: see text] = 75 mmHg): one after the ingestion of placebo and another following the intake of antioxidants (α-lipoic acid, vitamin C, and vitamin E), with a randomized double-blind design. Vastus lateralis muscle biopsies were obtained before, immediately after, and 30- and 120-min postsprint. Compared with the control condition, the ingestion of antioxidants resulted in lower plasma carbonylated proteins, attenuated elevation of the AMP-to-ATP molar ratio, and reduced glycolytic rate (P < 0.05) without significant effects on performance or V̇o2 The ingestion of antioxidants did not alter the basal muscle signaling. Thr172-AMPKα and Thr184/187-transforming growth factor-ß-activated kinase 1 (TAK1) phosphorylation were not increased after the sprint regardless of the ingestion of antioxidants. Thr286-CaMKII phosphorylation was increased after the sprint, but this response was blunted by the antioxidants. Ser485-AMPKα1/Ser491-AMPKα2 phosphorylation increased immediately after the sprints coincident with increased Akt phosphorylation. In summary, antioxidants attenuate the glycolytic response to sprint exercise in severe acute hypoxia and modify the muscle signaling response to exercise. Ser485-AMPKα1/Ser491-AMPKα2 phosphorylation, a known mechanism of Thr172-AMPKα phosphorylation inhibition, is increased immediately after sprint exercise in hypoxia, probably by a mechanism independent of ROS.NEW & NOTEWORTHY The glycolytic rate is increased during sprint exercise in severe acute hypoxia. This study showed that the ingestion of antioxidants before sprint exercise in severe acute hypoxia reduced the glycolytic rate and attenuated the increases of the AMP-to-ATP and the reduction of the NAD+-to-NADH.H+ ratios. This resulted in a modified muscle signaling response with a blunted Thr286-CaMKII but similar AMP-activated protein kinase phosphorylation responses in the sprints preceded by the ingestion of antioxidants.

Endurance Exercise Enhances the Effect of Strength Training on Muscle Fiber Size and Protein Expression of Akt and mTOR.

Reports concerning the effect of endurance exercise on the anabolic response to strength training have been contradictory. This study re-investigated this issue, focusing on training effects on indicators of protein synthesis and degradation. Two groups of male subjects performed 7 weeks of resistance exercise alone (R; n = 7) or in combination with preceding endurance exercise, including both continuous and interval cycling (ER; n = 9). Muscle biopsies were taken before and after the training period. Similar increases in leg-press 1 repetition maximum (30%; P<0.05) were observed in both groups, whereas maximal oxygen uptake was elevated (8%; P<0.05) only in the ER group. The ER training enlarged the areas of both type I and type II fibers, whereas the R protocol increased only the type II fibers. The mean fiber area increased by 28% (P<0.05) in the ER group, whereas no significant increase was observed in the R group. Moreover, expression of Akt and mTOR protein was enhanced in the ER group, whereas only the level of mTOR was elevated following R training. Training-induced alterations in the levels of both Akt and mTOR protein were correlated to changes in type I fiber area (r = 0.55-0.61, P<0.05), as well as mean fiber area (r = 0.55-0.61, P<0.05), reflecting the important role played by these proteins in connection with muscle hypertrophy. Both training regimes reduced the level of MAFbx protein (P<0.05) and tended to elevate that of MuRF-1. The present findings indicate that the larger hypertrophy observed in the ER group is due more to pronounced stimulation of anabolic rather than inhibition of catabolic processes.

AMPK signaling in skeletal muscle during exercise: Role of reactive oxygen and nitrogen species.

Reactive oxygen and nitrogen species (RONS) are generated during exercise depending on intensity, duration and training status. A greater amount of RONS is released during repeated high-intensity sprint exercise and when the exercise is performed in hypoxia. By activating adenosine monophosphate-activated kinase (AMPK), RONS play a critical role in the regulation of muscle metabolism but also in the adaptive responses to exercise training. RONS may activate AMPK by direct an indirect mechanisms. Directly, RONS may activate or deactivate AMPK by modifying RONS-sensitive residues of the AMPK-α subunit. Indirectly, RONS may activate AMPK by reducing mitochondrial ATP synthesis, leading to an increased AMP:ATP ratio and subsequent Thr(172)-AMPK phosphorylation by the two main AMPK kinases: LKB1 and CaMKKß. In presence of RONS the rate of Thr(172)-AMPK dephosphorylation is reduced. RONS may activate LKB1 through Sestrin2 and SIRT1 (NAD(+)/NADH.H(+)-dependent deacetylase). RONS may also activate CaMKKß by direct modification of RONS sensitive motifs and, indirectly, by activating the ryanodine receptor (Ryr) to release Ca(2+). Both too high (hypoxia) and too low (ingestion of antioxidants) RONS levels may lead to Ser(485)-AMPKα1/Ser(491)-AMPKα2 phosphorylation causing inhibition of Thr(172)-AMPKα phosphorylation. Exercise training increases muscle antioxidant capacity. When the same high-intensity training is applied to arm and leg muscles, arm muscles show signs of increased oxidative stress and reduced mitochondrial biogenesis, which may be explained by differences in RONS-sensing mechanisms and basal antioxidant capacities between arm and leg muscles. Efficient adaptation to exercise training requires optimal exposure to pulses of RONS. Inappropriate training stimulus may lead to excessive RONS formation, oxidative inactivation of AMPK and reduced adaptation or even maladaptation. Theoretically, exercise programs should be designed taking into account the intrinsic properties of different skeletal muscles, the specific RONS induction and the subsequent signaling responses.

What limits performance during whole-body incremental exercise to exhaustion in humans?

To determine the mechanisms causing task failure during incremental exercise to exhaustion (IE), sprint performance (10 s all-out isokinetic) and muscle metabolites were measured before (control) and immediately after IE in normoxia (P(IO2) 143 mmHg) and hypoxia (P(IO2): 73 mmHg) in 22 men (22 ± 3 years). After IE, subjects recovered for either 10 or 60 s, with open circulation or bilateral leg occlusion (300 mmHg) in random order. This was followed by a 10 s sprint with open circulation. Post-IE peak power output (W(peak)) was higher than the power output reached at exhaustion during IE (P < 0.05). After 10 and 60 s recovery in normoxia, W(peak) was reduced by 38 ± 9 and 22 ± 10% without occlusion, and 61 ± 8 and 47 ± 10% with occlusion (P < 0.05). Following 10 s occlusion, W(peak) was 20% higher in hypoxia than normoxia (P < 0.05), despite similar muscle lactate accumulation ([La]) and phosphocreatine and ATP reduction. Sprint performance and anaerobic ATP resynthesis were greater after 60 s compared with 10 s occlusions, despite the higher [La] and [H(+)] after 60 s compared with 10 s occlusion recovery (P < 0.05). The mean rate of ATP turnover during the 60 s occlusion was 0.180 ± 0.133 mmol (kg wet wt)(-1) s(-1), i.e. equivalent to 32% of leg peak O2 uptake (the energy expended by the ion pumps). A greater degree of recovery is achieved, however, without occlusion. In conclusion, during incremental exercise task failure is not due to metabolite accumulation or lack of energy resources. Anaerobic metabolism, despite the accumulation of lactate and H(+), facilitates early recovery even in anoxia. This points to central mechanisms as the principal determinants of task failure both in normoxia and hypoxia, with lower peripheral contribution in hypoxia.

Blood temperature and perfusion to exercising and non-exercising human limbs.

NEW FINDINGS: What is the central question of this study? Temperature-sensitive mechanisms are thought to contribute to blood-flow regulation, but the relationship between exercising and non-exercising limb perfusion and blood temperature is not established. What is the main finding and its importance? The close coupling among perfusion, blood temperature and aerobic metabolism in exercising and non-exercising extremities across different exercise modalities and activity levels and the tight association between limb vasodilatation and increases in plasma ATP suggest that both temperature- and metabolism-sensitive mechanisms are important for the control of human limb perfusion, possibly by activating ATP release from the erythrocytes. Temperature-sensitive mechanisms may contribute to blood-flow regulation, but the influence of temperature on perfusion to exercising and non-exercising human limbs is not established. Blood temperature (TB ), blood flow and oxygen uptake (V̇O2) in the legs and arms were measured in 16 healthy humans during 90 min of leg and arm exercise and during exhaustive incremental leg or arm exercise. During prolonged exercise, leg blood flow (LBF) was fourfold higher than arm blood flow (ABF) in association with higher TB and limb V̇O2. Leg and arm vascular conductance during exercise compared with rest was related closely to TB (r(2) = 0.91; P < 0.05), plasma ATP (r(2) = 0.94; P < 0.05) and limb V̇O2 (r(2) = 0.99; P < 0.05). During incremental leg exercise, LBF increased in association with elevations in TB and limb V̇O2, whereas ABF, arm TB and V̇O2 remained largely unchanged. During incremental arm exercise, both ABF and LBF increased in relationship to similar increases in V̇O2. In 12 trained males, increases in femoral TB and LBF during incremental leg exercise were mirrored by similar pulmonary artery TB and cardiac output dynamics, suggesting that processes in active limbs dominate central temperature and perfusion responses. The present data reveal a close coupling among perfusion, TB and aerobic metabolism in exercising and non-exercising extremities and a tight association between limb vasodilatation and increases in plasma ATP. These findings suggest that temperature and V̇O2 contribute to the regulation of limb perfusion through control of intravascular ATP.

Blood ammonia and lactate as markers of muscle metabolites during leg press exercise.

To examine whether blood lactate and ammonia concentrations can be used to estimate the functional state of the muscle contractile machinery with regard to muscle lactate and adenosine triphosphate (ATP) levels during leg press exercise. Thirteen men (age, 34 ± 5 years; 1 repetition maximum leg press strength 199 ± 33 kg) performed either 5 sets of 10 repetitions to failure (5×10RF), or 10 sets of 5 repetitions not to failure (10×5RNF) with the same initial load (10RM) and interset rests (2 minutes) on 2 separate sessions in random order. Capillary blood samples were obtained before and during exercise and recovery. Six subjects underwent vastus lateralis muscle biopsies at rest, before the first set and after the final exercise set. The 5×10RF resulted in a significant and marked decrease in power output (37%), muscle ATP content (24%), and high levels of muscle lactate (25.0 ± 8.1 mmol·kg wet weight), blood lactate (10.3 ± 2.6 mmol·L), and blood ammonia (91.6 ± 40.5 µmol·L). During 10×5RNF no or minimal changes were observed. Significant correlations were found between: (a) blood ammonia and muscle ATP (r = -0.75), (b) changes in peak power output and blood ammonia (r = -0.87) and blood lactate (r = -0.84), and (c) blood and muscle lactate (r = 0.90). Blood lactate and ammonia concentrations can be used as extracellular markers for muscle lactate and ATP contents, respectively. The decline in mechanical power output can be used to indirectly estimate blood ammonia and lactate during leg press exercise.