ABSTRACT
Diminished insulin and insulin-like growth factor-1 signaling extends the lifespan of invertebrates1-4; however, whether it is a feasible longevity target in mammals is less clear5-12. Clinically utilized therapeutics that target this pathway, such as small-molecule inhibitors of phosphoinositide 3-kinase p110α (PI3Ki), provide a translatable approach to studying the impact of these pathways on aging. Here, we provide evidence that dietary supplementation with the PI3Ki alpelisib from middle age extends the median and maximal lifespan of mice, an effect that was more pronounced in females. While long-term PI3Ki treatment was well tolerated and led to greater strength and balance, negative impacts on common human aging markers, including reductions in bone mass and mild hyperglycemia, were also evident. These results suggest that while pharmacological suppression of insulin receptor (IR)/insulin-like growth factor receptor (IGFR) targets could represent a promising approach to delaying some aspects of aging, caution should be taken in translation to humans.
Subject(s)
Longevity , Phosphatidylinositol 3-Kinases , Mice , Animals , Male , Humans , Female , Aging , Phosphoinositide-3 Kinase Inhibitors/pharmacology , Mammals/metabolism , Dietary SupplementsABSTRACT
The role of mitochondrial ROS in signalling muscle adaptations to exercise training has not been explored in detail. We investigated the effect of supplementation with the mitochondria-targeted antioxidant MitoQ on a) the skeletal muscle mitochondrial and antioxidant gene transcriptional response to acute high-intensity exercise and b) skeletal muscle mitochondrial content and function following exercise training. In a randomised, double-blind, placebo-controlled, parallel design study, 23 untrained men (age: 44 ± 7 years, VO2peak: 39.6 ± 7.9 ml/kg/min) were randomised to receive either MitoQ (20 mg/d) or a placebo for 10 days before completing a bout of high-intensity interval exercise (cycle ergometer, 10 × 60 s at VO2peak workload with 75 s rest). Blood samples and vastus lateralis muscle biopsies were collected before exercise and immediately and 3 h after exercise. Participants then completed high-intensity interval training (HIIT; 3 sessions per week for 3 weeks) and another blood sample and muscle biopsy were collected. There was no effect of acute exercise or MitoQ on systemic (plasma protein carbonyls and reduced glutathione) or skeletal muscle (mtDNA damage and 4-HNE) oxidative stress biomarkers. Acute exercise-induced increases in skeletal muscle peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1-α) mRNA expression were augmented in the MitoQ group. Despite this, training-induced increases in skeletal muscle mitochondrial content were similar between groups. HIIT-induced increases in VO2peak and 20 km time trial performance were also similar between groups while training-induced increases in peak power achieved during the VO2peak test were augmented in the MitoQ group. These data suggest that training-induced increases in peak power are enhanced following MitoQ supplementation, which may be related to the augmentation of skeletal muscle PGC1α expression following acute exercise. However, these effects do not appear to be related to an effect of MitoQ supplementation on exercise-induced oxidative stress or training-induced mitochondrial biogenesis in skeletal muscle.
Subject(s)
Antioxidants , Exercise , Organophosphorus Compounds/pharmacology , Ubiquinone/analogs & derivatives , Adult , Antioxidants/metabolism , Dietary Supplements , Exercise/physiology , Humans , Male , Middle Aged , Mitochondria/metabolism , Muscle, Skeletal/metabolism , Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha/genetics , Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Ubiquinone/pharmacologyABSTRACT
Unaccustomed exercise causes muscle damage resulting in loss of muscle function, which may be attributable to exercise-induced increases in skeletal muscle reactive oxygen species. This study examined the effect of mitochondria-targeted antioxidant supplementation on recovery of muscle function following exercise. Thirty-two untrained men received MitoQ (20 mg/day) or a placebo for 14 days before performing 300 maximal eccentric contractions of the knee extensor muscles of 1 leg. Muscle function was assessed using isokinetic dynamometry before, immediately after, and 24, 48, 72, and 168 hours after exercise. Muscle soreness was assessed using a visual analogue scale 24, 48, 72, and 168 hours after exercise. Blood samples were collected before, immediately after, and 2, 24, 48, 72, and 168 hours after exercise and urine samples were collected before and during the 48 hours after exercise. The reduction in maximal voluntary isometric contraction force and peak concentric torque following exercise was unaffected by MitoQ while recovery of peak eccentric torque was delayed in the MitoQ group. Exercise-induced increases in urine F2-isoprostanes were unaffected by MitoQ. MitoQ augmented exercise-induced increases in plasma creatine kinase levels, while plasma IL-6 was similar between groups. Muscle soreness was not affected by MitoQ. These results indicate that MitoQ does not attenuate post-exercise muscle soreness and may delay recovery of muscle function following eccentric exercise. Trial registration number: ACTRN12620001089921. Novelty: Post-exercise recovery of maximal voluntary isometric contraction force and peak concentric torque were unaffected by MitoQ. MitoQ delayed post-exercise recovery of peak eccentric torque. Post-exercise muscle soreness was unaffected by MitoQ.
Subject(s)
Isometric Contraction , Muscular Diseases , Antioxidants/pharmacology , Creatine Kinase , Dietary Supplements , F2-Isoprostanes , Humans , Male , Mitochondria , Muscle Contraction/physiology , Muscle, Skeletal/physiology , Myalgia/prevention & control , TorqueABSTRACT
BACKGROUND: Exercise increases skeletal muscle reactive oxygen species (ROS) production, which may contribute to the onset of muscular fatigue and impair athletic performance. Mitochondria-targeted antioxidants such as MitoQ, which contains a ubiquinone moiety and is targeted to mitochondria through the addition of a lipophilic triphenylphosphonium cation, are becoming popular amongst active individuals as they are designed to accumulate within mitochondria and may provide targeted protection against exercise-induced oxidative stress. However, the effect of MitoQ supplementation on cycling performance is currently unknown. Here, we investigate whether MitoQ supplementation can improve cycling performance measured as time to complete an 8 km time trial. METHOD: In a randomized, double-blind, placebo-controlled crossover study, 19 middle-aged (age: 44 ± 4 years) recreationally trained (VO2peak: 58.5 ± 6.2 ml·kg- 1·min- 1, distance cycled per week during 6 months prior to study enrollment: 158.3 ± 58.4 km) male cyclists completed 45 min cycling at 70% VO2peak followed by an 8 km time trial after 28 days of supplementation with MitoQ (20 mg·day- 1) and a placebo. Free F2-isoprostanes were measured in plasma samples collected at rest, after 45 min cycling at 70% VO2peak and after completion of the time trial. Respiratory gases and measures of rating of perceived exertion (RPE) were also collected. RESULTS: Mean completion time for the time trial was 1.3% faster with MitoQ (12.91 ± 0.94 min) compared to placebo (13.09 ± 0.95 min, p = 0.04, 95% CI [0.05, 2.64], d = 0.2). There was no difference in RPE during the time trial between conditions (p = 0.82) despite there being a 4.4% increase in average power output during the time trial following MitoQ supplementation compared to placebo (placebo; 270 ± 51 W, MitoQ; 280 ± 53 W, p = 0.04, 95% CI [0.49, 8.22], d = 0.2). Plasma F2-isoprostanes were lower on completion of the time trial following MitoQ supplementation (35.89 ± 13.6 pg·ml- 1) compared to placebo (44.7 ± 16.9 pg·ml- 1 p = 0.03). CONCLUSION: These data suggest that MitoQ supplementation may be an effective nutritional strategy to attenuate exercise-induced increases in oxidative damage to lipids and improve cycling performance.
Subject(s)
Antioxidants/pharmacology , Athletic Performance/physiology , Bicycling/physiology , Mitochondria, Muscle/drug effects , Organophosphorus Compounds/pharmacology , Performance-Enhancing Substances/pharmacology , Ubiquinone/analogs & derivatives , Adult , Antioxidants/metabolism , Cross-Over Studies , Double-Blind Method , F2-Isoprostanes/blood , Humans , Lipid Peroxidation , Male , Middle Aged , Mitochondria, Muscle/metabolism , Muscle, Skeletal/metabolism , Organophosphorus Compounds/metabolism , Oxidative Stress/drug effects , Oxygen Consumption , Performance-Enhancing Substances/metabolism , Physical Exertion/drug effects , Physical Exertion/physiology , Placebos/metabolism , Placebos/pharmacology , Reactive Oxygen Species/metabolism , Sports Nutritional Physiological Phenomena/drug effects , Sports Nutritional Physiological Phenomena/physiology , Time Factors , Ubiquinone/metabolism , Ubiquinone/pharmacologyABSTRACT
Measurement of skeletal muscle mitochondrial respiration requires invasive biopsy to obtain a muscle sample. Peripheral blood mononuclear cell (PBMC) mitochondrial protein content appears to reflect training status in young men; however, no studies have investigated whether there are training-induced changes in PBMC mitochondrial respiration. Therefore, we determined whether PBMC mitochondrial respiration could be used as a marker of skeletal muscle mitochondrial respiration in young healthy men and whether PBMC mitochondrial respiration responds to short-term training. Skeletal muscle and PBMC samples from 10 healthy young (18-35 yr) male participants were taken before and after a 2-wk high-intensity interval training protocol. High-resolution respirometry was used to determine mitochondrial respiration from muscle and PBMCs, and Western blotting and quantitative PCR were used to assess mitochondrial biogenesis in PBMCs. PBMC mitochondrial respiration was not correlated with muscle mitochondrial respiration at baseline ( R2 = 0.012-0.364, P > 0.05). While muscle mitochondrial respiration increased in response to training (32.1-61.5%, P < 0.05), PBMC respiration was not affected by training. Consequently, PBMCs did not predict training effect on muscle mitochondrial respiration ( R2 = 0.024-0.283, P > 0.05). Similarly, gene and protein markers of mitochondrial biogenesis did not increase in PBMCs following training. This suggests PBMC mitochondrial function does not reflect that of skeletal muscle and does not increase following short-term high-intensity training. PBMCs are therefore not a suitable biomarker for muscle mitochondrial function in young healthy men. It may be useful to study PBMC mitochondrial function as a biomarker of muscle mitochondrial function in pathological populations with different respiration capacities. NEW & NOTEWORTHY Research in primates has suggested that peripheral blood mononuclear cells (PBMCs) may provide a less-invasive alternative to a muscle biopsy for measuring muscle mitochondrial function. Furthermore, trained individuals appear to have greater mitochondrial content in PBMCs. Here we show that in healthy young men, PBMCs do not reflect skeletal muscle mitochondrial function and do not adapt in response to a training intervention that increases muscle mitochondrial function, suggesting PBMCs are a poor marker of muscle mitochondrial function in humans.
Subject(s)
Energy Metabolism , High-Intensity Interval Training , Leukocytes, Mononuclear/metabolism , Mitochondria, Muscle/metabolism , Muscle, Skeletal/metabolism , Adaptation, Physiological , Adolescent , Adult , Age Factors , Biomarkers/metabolism , Cell Respiration , Healthy Volunteers , Humans , Male , Mitochondrial Proteins/genetics , Mitochondrial Proteins/metabolism , Oxidative Phosphorylation , Sex Factors , Time Factors , Young AdultABSTRACT
Oxidation can decrease or increase the Ca2+ sensitivity of the contractile apparatus in rodent fast-twitch (type II) skeletal muscle fibres, but the reactions and molecular targets involved are unknown. This study examined whether increased Ca2+ sensitivity is due to S-glutathionylation of particular cysteine residues. Skinned muscle fibres were directly activated in heavily buffered Ca2+ solutions to assess contractile apparatus Ca2+ sensitivity. Rat type II fibres were subjected to S-glutathionylation by successive treatments with 2,2'-dithiodipyridine (DTDP) and glutathione (GSH), and displayed a maximal increase in pCa50 (−log10 [Ca2+] at half-maximal force) of â¼0.24 pCa units, with little or no effect on maximum force or Hill coefficient. Partial similar effect was produced by exposure to oxidized gluthathione (GSSG, 10 mM) for 10 min at pH 7.1, and near-maximal effect by GSSG treatment at pH 8.5. None of these treatments significantly altered Ca2+ sensitivity in rat type I fibres. Western blotting showed that both the DTDPGSH and GSSGpH 8.5 treatments caused marked S-glutathionylation of the fast troponin I isoform (TnI(f)) present in type II fibres, but not of troponin C (TnC) or myosin light chain 2. Both the increased Ca2+ sensitivity and glutathionylation of TnI(f) were blocked by N-ethylmaleimide (NEM). S-nitrosoglutathione (GSNO) also increased Ca2+ sensitivity, but only in conditions where it caused S-glutathionylation of TnI(f). In human type II fibres from vastus lateralis muscle, DTDPGSH treatment also caused similar increased Ca2+ sensitivity and S-glutathionylation of TnI(f). When the slow isoform of TnI in type I fibres of rat was partially substituted (â¼30%) with TnI(f), DTDPGSH treatment caused a significant increase in Ca2+ sensitivity (â¼0.08 pCa units). TnIf in type II fibres from toad and chicken muscle lack Cys133 present in mammalian TnIf, and such fibres showed no change in Ca2+ sensitivity with DTDPGSH nor any S-glutathionylation of TnI(f) (latter examined only in toad). Following 40 min of cycling exercise in human subjects (at â¼60% peak oxygen consumption), TnI(f) in vastus lateralis muscle displayed a marked increase in S-glutathionylation (â¼4-fold). These findings show that S-glutathionylation of TnI(f), most probably at Cys133, increases the Ca2+ sensitivity of the contractile apparatus, and that this occurs in exercising humans, with likely beneficial effects on performance.
Subject(s)
Calcium/physiology , Muscle Fibers, Fast-Twitch/physiology , Troponin I/physiology , 2,2'-Dipyridyl/analogs & derivatives , 2,2'-Dipyridyl/pharmacology , Adult , Animals , Bufo marinus , Chickens , Cysteine/physiology , Disulfides/pharmacology , Exercise/physiology , Female , Glutathione/pharmacology , Glutathione Disulfide/pharmacology , Humans , Male , Muscle Contraction/drug effects , Muscle Contraction/physiology , Muscle Fibers, Fast-Twitch/drug effects , Muscle Fibers, Slow-Twitch/drug effects , Muscle Fibers, Slow-Twitch/physiology , Rabbits , Rats , Rats, Long-Evans , Swine , Young AdultABSTRACT
AIM: To determine whether voluntary free wheel (FW) or resistance wheel (RW) exercise or reduced muscle activity would influence maturational increases in muscle mass and the number of satellite cells (SCs) and myonuclei (MN) accrued by adulthood. METHODS: Hind limb muscles of male rats housed with, or without, FWs from 4 to 5, 7 or 10 weeks of age, and rats housed with RWs from 4 to 10 week of age, were evaluated. To assess the effect of reduced muscle activity, gastrocnemius muscles of 4-week-old rats were injected with botulinum toxin (Btx) and collected at 7 weeks of age. Muscle fibre size and the frequency of Pax7-positive SCs and MN were determined in 7- and 10-week-old muscles via immunohistochemical methods. RESULTS: Free wheel exercise enhanced muscle growth and the frequency of SCs in the medial gastrocnemius (MG) (threefold) and vastus lateralis (VL) (twofold) of rats at 10 week of age. Resistance wheel exercise increased the number of SCs and MN (22-30%), with more muscle fibre nuclei being associated with larger fibre size, in the soleus, MG and VL muscles. Btx impaired the normal increases in muscle fibre size and the accrual of MN but not SCs. CONCLUSION: A greater volume of exercise during maturational growth was important for enhancing SC numbers, whereas their conversion to MN required higher-intensity exercise. The enhanced muscle fibre nuclear populations may influence the capacity of the muscle to adapt to exercise, injury or disuse in later adulthood.
Subject(s)
Aging , Cell Proliferation , Muscle Development , Muscle Fibers, Skeletal/physiology , Quadriceps Muscle/growth & development , Resistance Training , Satellite Cells, Skeletal Muscle/physiology , Age Factors , Animals , Cell Size , Hindlimb , Immunohistochemistry , Male , Muscle Fibers, Skeletal/metabolism , Organ Size , Paired Box Transcription Factors/metabolism , Quadriceps Muscle/cytology , Quadriceps Muscle/metabolism , Rats , Rats, Sprague-Dawley , Satellite Cells, Skeletal Muscle/metabolismABSTRACT
AIMS/HYPOTHESIS: Insulin activates insulin receptor protein tyrosine kinase and downstream phosphatidylinositol-3-kinase (PI3K)/Akt signalling in muscle to promote glucose uptake. The insulin receptor can serve as a substrate for the protein tyrosine phosphatase (PTP) 1B and T cell protein tyrosine phosphatase (TCPTP), which share a striking 74% sequence identity in their catalytic domains. PTP1B is a validated therapeutic target for the alleviation of insulin resistance in type 2 diabetes. PTP1B dephosphorylates the insulin receptor in liver and muscle to regulate glucose homeostasis, whereas TCPTP regulates insulin receptor signalling and gluconeogenesis in the liver. In this study we assessed for the first time the role of TCPTP in the regulation of insulin receptor signalling in muscle. METHODS: We generated muscle-specific TCPTP-deficient (Mck-Cre;Ptpn2(lox/lox)) mice (Mck, also known as Ckm) and assessed the impact on glucose homeostasis and muscle insulin receptor signalling in chow-fed versus high-fat-fed mice. RESULTS: Blood glucose and insulin levels, insulin and glucose tolerance, and insulin-induced muscle insulin receptor activation and downstream PI3K/Akt signalling remained unaltered in chow-fed Mck-Cre;Ptpn2(lox/lox) versus Ptpn2(lox/lox) mice. In addition, body weight, adiposity, energy expenditure, insulin sensitivity and glucose homeostasis were not altered in high-fat-fed Mck-Cre;Ptpn2(lox/lox) versus Ptpn2(lox/lox) mice. CONCLUSIONS/INTERPRETATION: These results indicate that TCPTP deficiency in muscle has no effect on insulin signalling and glucose homeostasis, and does not prevent high-fat diet-induced insulin resistance. Thus, despite their high degree of sequence identity, PTP1B and TCPTP contribute differentially to insulin receptor regulation in muscle. Our results are consistent with the notion that these two highly related PTPs make distinct contributions to insulin receptor regulation in different tissues.
Subject(s)
Glucose/metabolism , Muscles/metabolism , Protein Tyrosine Phosphatase, Non-Receptor Type 2/deficiency , Protein Tyrosine Phosphatase, Non-Receptor Type 2/physiology , Animals , Diabetes Mellitus, Type 2/blood , Glucose Tolerance Test , Homeostasis , Insulin/metabolism , Insulin Resistance , Mice , Mice, Inbred C57BL , Mice, Transgenic , Phosphatidylinositol 3-Kinases/metabolism , Phosphorylation , Protein Tyrosine Phosphatase, Non-Receptor Type 1/metabolism , Protein Tyrosine Phosphatase, Non-Receptor Type 2/genetics , Receptor, Insulin/metabolism , Signal Transduction , Time Factors , Tissue DistributionABSTRACT
There is evidence that reactive oxygen species (ROS) contribute to the regulation of skeletal muscle glucose uptake during highly fatiguing ex vivo contraction conditions via AMP-activated protein kinase (AMPK). In this study we investigated the role of ROS in the regulation of glucose uptake and AMPK signaling during low-moderate intensity in situ hindlimb muscle contractions in rats, which is a more physiological protocol and preparation. Male hooded Wistar rats were anesthetized, and then N-acetylcysteine (NAC) was infused into the epigastric artery (125 mg.kg(-1).h(-1)) of one hindlimb (contracted leg) for 15 min before this leg was electrically stimulated (0.1-ms impulse at 2 Hz and 35 V) to contract at a low-moderate intensity for 15 min. The contralateral leg did not receive stimulation or local NAC infusion (rest leg). NAC infusion increased (P<0.05) plasma cysteine and cystine (by approximately 360- and 1.4-fold, respectively) and muscle cysteine (by 1.5-fold, P=0.001). Although contraction did not significantly alter muscle tyrosine nitration, reduced (GSH) or oxidized glutathione (GSSG) content, S-glutathionylation of protein bands at approximately 250 and 150 kDa was increased (P<0.05) approximately 1.7-fold by contraction, and this increase was prevented by NAC. Contraction increased (P<0.05) skeletal muscle glucose uptake 20-fold, AMPK phosphorylation 6-fold, ACCbeta phosphorylation 10-fold, and p38 MAPK phosphorylation 60-fold, and the muscle fatigued by approximately 30% during contraction and NAC infusion had no significant effect on any of these responses. This was despite NAC preventing increases in S-glutathionylation with contraction. In conclusion, unlike during highly fatiguing ex vivo contractions, local NAC infusion during in situ low-moderate intensity hindlimb contractions in rats, a more physiological preparation, does not attenuate increases in skeletal muscle glucose uptake or AMPK signaling.
Subject(s)
Acetylcysteine/administration & dosage , Antioxidants/administration & dosage , Glucose/metabolism , Muscle Contraction , Muscle, Skeletal/drug effects , AMP-Activated Protein Kinases/metabolism , Acetylcysteine/metabolism , Animals , Antioxidants/metabolism , Biological Transport , Blood Pressure , Cysteine/blood , Cystine/blood , Electric Stimulation , Glutathione/metabolism , Heart Rate , Hindlimb , Infusions, Intra-Arterial , Male , Muscle Fatigue , Muscle Strength , Muscle, Skeletal/blood supply , Muscle, Skeletal/innervation , Muscle, Skeletal/metabolism , Phosphorylation , Rats , Rats, Wistar , Reactive Oxygen Species/metabolism , Regional Blood Flow , Time Factors , Tyrosine/metabolism , Vascular Resistance , p38 Mitogen-Activated Protein Kinases/metabolismABSTRACT
AIM: Hypohydration exacerbates cardiovascular and thermal strain and can impair exercise capacity in temperate and warm conditions. Yet, athletes often dehydrate in exercise, are hypervolaemic and have less cardiovascular sensitivity to acute hypervolaemia. We tested the hypothesis that trained individuals have less cardiovascular, thermoregulatory and performance affect of hypohydration during exercise. METHODS: After familiarization, six trained [VO(2 peak) = 64 (SD 8) mL kg(-1) min(-1)] and six untrained [O(2 peak) = 45 (4) mL kg(-1) min(-1)] males cycled 40 min at 70%O(2 peak) while euhydrated or hypohydrated by 1.5-2.0% body mass (crossover design), before a 40-min work trial with euhydration or ad libitum drinking (in Hypohydration trial), in temperate conditions (24.3 degrees C, RH 50%, v(a) = 4.5 m s(-1)). Baseline hydration was by complete or partial rehydration from exercise+heat stress the previous evening. RESULTS: During constant workload, heart rate and its drift were increased in Hypohydration compared with Euhydration for Untrained [drift: 33 (11) vs. 24 beats min(-1) h(-1) (10), 95% CI 5-11] but not Trained [14 (3) vs. 13 beats min(-1) h(-1) (3), CI -2 to 3; P = 0.01 vs. Untrained]. Similarly, rectal temperature drift was faster in Hypohydration for Untrained only [by 0.57 degrees C h(-1) (0.25); P = 0.03 vs. Trained], concomitant with their reduced sweat rate (P = 0.05) and its relation to plasma osmolality (P = 0.03). Performance power tended to be reduced for Untrained (-13%, CI -35 to 2) and Trained (-7%, CI: -16 to 1), without an effect of fitness (P = 0.38). CONCLUSION: Mild hypohydration exacerbated cardiovascular and thermoregulatory strain and tended to impair endurance performance, but aerobic fitness attenuated the physiological effects.