Skeletal Muscle Metabolism Research Articles

ACE-I/D Allele Modulates Improvements of Cardiorespiratory Function and Muscle Performance with Interval-Type Exercise.

Background: The prominent insertion/deletion polymorphism in the gene for the major modulator of tissue perfusion, angiotensin-converting enzyme (ACE-I/D) is associated with variability in adjustments in cardiac and skeletal muscle performance with standard forms of endurance and strength type training. Here, we tested whether the ACE-I/D genotype would be associated with variability in the effects of interval-type training on peak and aerobic performance of peripheral muscle and cardio-vasculature and post-exercise recovery. Methods: Nine healthy subjects (39.0 ± 14.7 years of age; 64.6 ± 16.1 kg, 173.6 ± 9.9) completed eight weeks of interval training on a soft robotic device based on repeated sets of a pedaling exercise at a matched intensity relative to their peak aerobic power output. Prior to and post-training, peak anaerobic and aerobic power output was assessed, mechanical work and metabolic stress (oxygen saturation and hemoglobin concentrations of Musculus vastus lateralis (VAS) and Musculus gastrocnemius (GAS), blood lactate and factors setting cardiac output such as heart rate, systolic and diastolic blood pressure were monitored during ramp-incremental exercise and interval exercise with the calculation of areas under the curve (AUC), which were put in relation to the produced muscle work. Genotyping was performed based on I- and D-allele-specific polymerase chain reactions on genomic DNA from mucosal swaps. The significance of interaction effects between training and ACE I-allele on absolute and work-related values was assessed with repeated measures ANOVA. Results: Subjects delivered 87% more muscle work/power, 106% more cardiac output, and muscles experienced ~72% more of a deficit in oxygen saturation and a ~35% higher passage of total hemoglobin during single interval exercise after the eight weeks of training. Interval training affected aspects of skeletal muscle metabolism and performance, whose variability was associated with the ACE I-allele. This concerned the economically favorable alterations in the work-related AUC for the deficit of SmO2 in the VAS and GAS muscles during the ramp exercise for the I-allele carriers and opposing deteriorations in non-carriers. Conversely, oxygen saturation in the VAS and GAS at rest and during interval exercise was selectively improved after training for the non-carriers of the I-allele when the AUC of tHb per work during interval exercise deteriorated in the carriers. Training also improved aerobic peak power output by 4% in the carriers but not the non-carriers (p = 0.772) of the ACE I-allele while reducing negative peak power (-27.0%) to a lesser extent in the ACE I-allele carriers than the non-carriers. Variability in cardiac parameters (i.e., the AUC of heart rate and glucose during ramp exercise, was similar to the time to recovery of maximal tHb in both muscles after cessation of ramp exercise, only associated with the ACE I-allele but not training per se. Diastolic blood pressure and cardiac output during recovery from exhaustive ramp exercise demonstrated a trend for training-associated differences in association with the ACE I-allele. Discussion: The exercise-type dependent manifestation of antidromic adjustments in leg muscle perfusion and associated local aerobic metabolism between carriers and non-carriers of the ACE I-allele with the interval-training highlight that non-carriers of the I-allele do not present an essential handicap to improve perfusion-related aerobic muscle metabolism but that the manifestation of responsiveness depends on the produced work. Conclusions: The deployed interval-type of exercise produced ACE I-allele-related differences in the alterations of negative anaerobic performance and perfusion-related aerobic muscle metabolism, which manifestation is exercise specific. The training-invariant ACE I-allele-associated differences in heart rate and blood glucose concentration emphasize that the repeated impact of the interval stimulus, despite a near doubling of the initial metabolic load, was insufficient to overturn ACE-related genetic influences on cardiovascular function.

Skeletal Muscle Mitochondrial Respiration and Exercise Intolerance in Patients With Heart Failure With Preserved Ejection Fraction

The pathophysiology of exercise intolerance in patients with heart failure with preserved ejection fraction (HFpEF) remains incompletely understood. Multiple lines of evidence suggest that abnormal skeletal muscle metabolism is a key contributor, but the mechanisms underlying metabolic dysfunction remain unresolved. To evaluate the associations of skeletal muscle mitochondrial function using respirometric analysis of biopsied muscle fiber bundles from patients with HFpEF with exercise performance. In this cross-sectional study, muscle fiber bundles prepared from fresh vastus lateralis biopsies were analyzed by high-resolution respirometry to provide detailed analyses of mitochondrial oxidative phosphorylation, including maximal capacity and the individual contributions of complex I-linked and complex II-linked respiration. These bioenergetic data were compared between patients with stable chronic HFpEF older than 60 years and age-matched healthy control (HC) participants and analyzed for intergroup differences and associations with exercise performance. All participants were treated at a university referral center, were clinically stable, and were not undergoing regular exercise or diet programs. Data were collected from March 2016 to December 2017, and data were analyzed from November 2020 to May 2021. Skeletal muscle mitochondrial function, including maximal capacity and respiration linked to complex I and complex II. Exercise performance was assessed by peak exercise oxygen consumption, 6-minute walk distance, and the Short Physical Performance Battery. Of 72 included patients, 50 (69%) were women, and the mean (SD) age was 69.6 (6.1) years. Skeletal muscle mitochondrial function measures were all markedly lower in skeletal muscle fibers obtained from patients with HFpEF compared with HCs, even when adjusting for age, sex, and body mass index. Maximal capacity was strongly and significantly correlated with peak exercise oxygen consumption (R = 0.69; P < .001), 6-minute walk distance (R = 0.70; P < .001), and Short Physical Performance Battery score (R = 0.46; P < .001). In this study, patients with HFpEF had marked abnormalities in skeletal muscle mitochondrial function. Severely reduced maximal capacity and complex I-linked and complex II-linked respiration were associated with exercise intolerance and represent promising therapeutic targets.

Filbertone, (2E)-5-methyl-2-hepten-4-one, regulates thermogenesis and lipid metabolism in skeletal muscle of a high-fat diet fed mice

Filbertone, the principal flavor compound of hazelnuts, is known to have preventive effects against hypothalamic inflammation, obesity and adipocity in vitro and in vivo. However, the effect of filbertone in skeletal muscle remains unknown. In the present study, we determined the effect of filbertone in skeletal muscle of mice fed a high-fat diet (HFD). To identify the underlying molecular and cellular processes of filbertone, we performed whole transcriptome sequencing in skeletal muscle. The muscle transcriptome analysis revealed that the upregulated differentially expressed genes (DEGs) in filbertone-fed mice were substantially associated with several pathways including thermogenesis, fatty acid degradation, oxidative phosphorylation, and branched chain amino acids (BCAAs) degradation. Furthermore, the expression level of thermogenic genes such as uncoupling protein 1 (Ucp1; p < 0.05), cell death-inducing DNA fragmentation factor alpha-like effector A (Cidea; p < 0.05), peroxisome proliferator-activated receptor alpha (Ppara; p < 0.05) and lipid droplet-associated protein genes such as Plin3 (p < 0.05), Plin4 (p < 0.05), and Plin5 (p < 0.05) were significantly upregulated in muscle tissue of HFD with filbertone fed mice compared to HFD only fed mice. Filbertone also elevated the protein level of UCP1 (p < 0.05) and PPARα (p < 0.05). In addition, filbertone reduced the accumulation of intracellular lipids in C2C12 myotubes (p < 0.05). On the basis of these results, we suggest that filbertone has a crucial effect in the regulation of muscle lipid metabolism and energy balance.

Sex differences in cachexia outcomes, anabolic signalling and BCAA metabolism following chemotherapy

Sex is a risk factor for cancer. Affecting nearly 80% of cancer patients is cachexia, a body and skeletal muscle-wasting syndrome. Poor nutritional status, tumour related factors and chemotherapy contribute to cachexia. Although negative effects of chemotherapy on skeletal muscle are documented, the majority of studies are completed in male animals. Although the branched-chain amino acids (BCAA: leucine, isoleucine and valine) activate anabolic signalling in skeletal muscle and have been shown to reduce some of the effects of cachexia, BCAA nutritional support does not reverse cachexia and few studies have investigated the effects of chemotherapy on BCAA metabolism in skeletal muscle. Therefore, the objective of this study is to compare the effects of chemotherapy on cachexia outcomes and BCAA metabolism between male and female mice. Three-month-old CD2F1 male and female mice were treated with either the chemotherapy drug cocktail folfiri (50mg/kg 5-fluorouracil (5FU), 90mg/kg Leucovorin and 24mg/kg CPT11) (drug) or vehicle (10% DMSO in saline) for 6-weeks. Within sex, drug treatment led to reductions in body and skeletal muscle weights. Between sex, drug-treated female mice lost more body and gastrocnemius muscle weight compared to drug-treated males. Within sex, drug treated animals exhibited reductions in anabolic signalling, specifically AKT, S6K1, S6 and 4E-BP1. Between sex, drug-treated females mice showed greater loss of anabolic signalling compared to drug-treated males. Within sex, the drug cocktail reduced amino acid transporters SNAT1 and LAT1, while reducing BCAA concentrations in the skeletal muscle. Drug-treated female mice exhibited greater loss of LAT1 compared to male-drug treated mice. However, male-drug treated mice had greater decreases in BCAA concentrations compared to female drug-treated mice. In addition, minimal differences were found within sex for key enzymes involved in BCAA metabolism. However, the inhibitory phosphorylation of branched-chain alpha-ketoacid dehydrogenase complex (BCKD) was elevated only in drug-treated male animals. Lastly, within sex, drug treatment showed elevated levels of BCAA concentrations in the liver, but only male drug-treated mice showed elevated plasma BCAAs compared to male vehicle-treated mice. Our data suggests a link between muscle atrophy and decreased BCAA metabolism during chemotherapy. We found that drug-treated female mice had worsened outcomes for cachexia measures such as body, skeletal muscle weight and anabolic signalling compared to drug-treated male mice. However, drug-treated males exhibited greater decreases in BCAA concentrations and their metabolism compared to drug-treated females. Findings from this study suggest that altered availability and metabolism of the BCAAs may contribute to muscle wasting differently in males and females during chemotherapy and interventions that correct such alterations may need to consider sex in order to better manage cachexia. Nserc, York University This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Deletion of myostatin alters obesity-induced estrogen metabolism in skeletal muscle

Objective: Obesity alters sex hormone balance in males, resulting in low testosterone and altered metabolism of estrogen, independent risk factors for cardiometabolic dysfunction. The basis of these alterations and how they are impacted by factors that improve cardiometabolic outcomes in obesity are unknown. Methods: Control and obese mice ( db/db) with or without deletion of myostatin to induce hypermuscularity were used. Mice were analyzed for plasma sex hormones via mass-spectrometry with or without 10mg/kg/day fadrazole to inhibit aromatase. Quantitative PCR was used to measure aromatase along with cytochrome P450 enzymes involved in estrogen metabolism in the liver, skeletal muscle, and visceral adipose tissue. HEK293a cells were treated with myostatin in vitro and enzyme levels assessed. Results: Obesity decreased plasma testosterone 30% and increased estrogen 20% with no change in gonadal weight. Deletion of myostatin increased muscle mass &gt;30% and resulted in a reduction in estrogen levels without a parallel increase in androgens in obese mice. Inhibition of aromatase restores estrogen levels indicating the excess estrogen in obesity is derived from testosterone conversion. Since estrogen was lowered without increasing testosterone and estrogen metabolism must be increased. Aromatase was increased in obesity but unaffected by changes in muscle mass. CYP1B1, an enzyme that converts estradiol to hydroxy estrogens, showed 2.5 fold expression in obese skeletal muscle, which is improved in the absence of myostatin, with no change in other P450s involved in estrogen metabolism. Moreover, changes in P450 enzymes in liver or fat were either absent or changed directionally opposite to observed hormone changes. In HEK cells, treatment with myostatin recapitulated changes in CYP1B1 expression seen in vitro, suggesting a direct link between the myokine and sex hormone metabolism by skeletal muscle. Conclusion: In summary, obesity causes a shift in sex hormone levels to estrogen production, which is improved by myostatin deletion, without restoration of androgens. A potential mechanism responsible could be altered estrogen catabolism, leading to decreased estrogen levels without the restoration of androgen levels. Together, these data suggest CYP1B1 drives the conversion of estrogen to hydroxestrogens, which are known pro-oxidant and pro-lipidemic influences that may contribute to cardiometabolic disease. Whether targeting CYP1B1 improves cardiometabolic function in obesity remains to be determined. D.W. Stepp and D.J. Fulton are supported by NIH 1R01HL147159 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

RIP140 deficiency enhances cardiac fuel metabolism and protects mice from heart failure.

During the development of heart failure (HF), the capacity for cardiomyocyte (CM) fatty acid oxidation (FAO) and ATP production is progressively diminished, contributing to pathologic cardiac hypertrophy and contractile dysfunction. Receptor-interacting protein 140 (RIP140, encoded by Nrip1) has been shown to function as a transcriptional corepressor of oxidative metabolism. We found that mice with striated muscle deficiency of RIP140 (strNrip1-/-) exhibited increased expression of a broad array of genes involved in mitochondrial energy metabolism and contractile function in heart and skeletal muscle. strNrip1-/- mice were resistant to the development of pressure overload-induced cardiac hypertrophy, and CM-specific RIP140-deficient (csNrip1-/-) mice were protected against the development of HF caused by pressure overload combined with myocardial infarction. Genomic enhancers activated by RIP140 deficiency in CMs were enriched in binding motifs for transcriptional regulators of mitochondrial function (estrogen-related receptor) and cardiac contractile proteins (myocyte enhancer factor 2). Consistent with a role in the control of cardiac fatty acid oxidation, loss of RIP140 in heart resulted in augmented triacylglyceride turnover and fatty acid utilization. We conclude that RIP140 functions as a suppressor of a transcriptional regulatory network that controls cardiac fuel metabolism and contractile function, representing a potential therapeutic target for the treatment of HF.

Healthy skeletal muscle atrophy and/or metabolism via FoxOs signaling pathway protects against starvation-induced fatty liver

Objective: Non-alcoholic fatty liver disease (NAFLD) with excessive triglyceride accumulation in the liver prevails in approximately 24% of adults worldwide. Prolonged fasting rapidly causes hepatic steatosis. However, the etiology of hepatic steatosis during starvation is unclear. During starvation, skeletal muscle, the central tissue for metabolism, physical exercise, and storage for protein/amino acids, adapts to energy deficit by promoting muscle proteolysis, which leads to muscle atrophy. The critical factors that cause muscle atrophy are the Forkhead Box O family transcription factors (FoxO1, 3 and 4). Although skeletal muscle FoxOs are necessary to adapt to starvation, the specific function of skeletal muscle FoxOs at the whole-body level is unclear. We hypothesized that skeletal muscle atrophy and/or metabolism would be associated with the etiology of starvation-induced fatty liver and aimed to elucidate the inter-organ crosstalk from skeletal muscle to liver via FoxOs signaling pathway. Methods: We previously generated skeletal muscle-specific FoxO1,3,4-triple knockout mice (mFoxO1,3,4-/-) using human skeletal α-actin promoter-driven Cre recombinase expression (Oyabu et al. 2022. FASEB J 36: e22152). The control FoxO1,3,4flox/flox mice (wild-type mice) and mFoxO1,3,4-/- mice were fasted for 48 h to mimic prolonged fasting. Results: Skeletal muscle mass was significantly reduced in 48 h-fasted wild-type mice. This reduction was prevented by the skeletal muscle FoxO-triple deletion. Fasted mFoxO1,3,4-/- mice showed reduced blood glucose levels, suggesting the lack of energy in 48 h-fasted mFoxO1,3,4-/- mice. Interestingly, starvation-induced lipid-droplet formation in the liver, accumulation of hepatic triglyceride, and the expression of hepatic lipid-droplet-marker protein (Plin2) were significantly increased in 48 h-fasted mFoxO1,3,4-/- mice compared to fasted wild-type mice, indicating that hepatic steatosis was accelerated by skeletal muscle FoxO-triple deletion, despite of the retention from skeletal muscle loss. This suggests that skeletal muscle FoxOs are essential for protection against starvation-induced hepatic steatosis for adaptation to starvation. Starvation induced the upregulation of genes involved in not only muscle proteolysis, but also triglyceride uptake and metabolism in skeletal muscle, which were abrogated by the skeletal muscle-specific FoxO-triple deletion. In addition, the muscle triglyceride level was lower in 48 h-fasted mFoxO1,3,4-/- mice than fasted wild-type mice. Conclusion: Here, we demonstrated a critical connection between liver and skeletal muscle via FoxOs transcription factors. Our data indicate that healthy skeletal muscle atrophy and/or metabolism via FoxOs signaling pathway is critical for prevention of starvation-induced hepatic steatosis. This study sheds light on the skeletal muscle-liver inter-organ crosstalk as the potential therapeutic targets of malnutrition-induced NAFLD. This study was supported by Grants-in-Aids for Scientific Research KAKENHI from the Japan Society for the Promotion of Science. This study was also supported by Mishima Kaiun Memorial Foundation and Grants-in-Aids for the Second Dream Challenge Planning Awards from JSBBA. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

NIRS-based in vivo assessment of skeletal muscle metabolism in young adults with cerebral palsy: an exploratory pilot study

Individuals with mild cerebral palsy (CP) have ~32% higher energy cost of walking compared to typically developing (TD) peers ( Nardon et al., 2021). In CP, the ability to walk is limited by poor body movement coordination and an early onset of neuromuscular fatigue. Analyzes of gracilis muscle biopsy in children with CP demonstrated a preserved mitochondria content but a 50-80% reduced maximal muscle mitochondrial complexes activity compared to TD ( Dayanidhi et al., 2021), suggesting the latter being an additional factor explaining the early onset of fatigue during ambulation. A novel near-infrared spectroscopy (NIRS) based approach, validated against muscle biopsy (r2=0.46; Ryan et al., 2014), allows non-invasive, in vivo determination of the skeletal muscle mitochondrial oxidative capacity ( k). We therefore aimed to evaluate the use of the NIRS-based protocol in young adults with CP; and to compare CP with TD peers. Methods: Four young men with CP (age: 24.4 ±2.6 yrs, BMI: 24.1 ±2.8 kg/m2) and four age- and sex-matched TD (age: 24.3 ±3.2 yrs, BMI: 23.8 ±1.5 kg/m2) took part in this ongoing study. Moderately active (IPAQ questionnaire) CP participants with stable motor function impairment level II-III of the Gross Motor Function Classification System were included. Exclusion criteria: epilepsy episode and/or botulinum toxin injection in the past 6mo; surgery in the past 36mo. Medial gastrocnemius muscle (chosen as representative locomotor muscle) k was measured twice by NIRS combined with intermittent arterial occlusions, after 10-15s of muscle activation. Reproducibility was determined by coefficient of variation (CV) and interclass correlation coefficient (ICC). Paired t test was used to compare groups (CP vs TD). Results: Test-retest CV (and ICC) was 9.5±7.5% (ICC=0.66) in CP and 6.2±3.5% (ICC=0.94) in TD. In the CP group, the within-subject reproducibility was large (CV range 0.4-17.0%). Resting muscle tissue saturation was within normal ranges in both groups (CP: 70.8±2.2%, CON: 66.1±6.9; p=0.35). Young adults with CP had ~37% lower k compared to TD (CP: 1.03±0.06 min-1, CON: 1.64±0.54 min-1) but this study is yet underpowered to draw solid conclusions (p=0.09). Discussion: Test-retest reliability of k in people with CP was lower than previously reported in other populations (CV ~10%; Adami &amp; Rossiter, 2018). Adding a third repeat for measuring k is recommended to improve sensitivity. Our pilot data showed ~37% lower k in the calf muscle in young adults with CP compared to TD peers, a similar loss seen in middle-age COPD (GOLD class 3-4) and multiple sclerosis patients, who also report easy fatigability during walking. Further studies in a larger group of individuals are needed to confirm NIRS-based muscle oxidative capacity as a potential clinical marker in CP, and to assess its feasibility to correlate with different severity levels. Supported by Rhode Island Foundation 2801_20190604 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Blunted reactive hyperemia after prolonged sitting is independent of altered microvascular function: contribution of skeletal muscle metabolism

Introduction: Although reduced post-occlusive reactive hyperemia (PORH) after prolonged sitting (PS) has been reported as impaired microvascular function, no specific mechanism(s) have been elucidated. One potential mechanism, independent of microvascular function, is that an altered muscle metabolic rate (MMR) may change the magnitude of PORH by modifying the oxygen deficit achieved during cuff-induced arterial occlusions. We speculated that if MMR changes during PS, this may invalidate current inferences about microvascular function during PS. Objective: Therefore, the objective of this study was to examine if peripheral leg MMR changes during PS and to ascertain whether the change in the oxygen deficit explains the change in PORH after PS. Hypothesis: We hypothesized that peripheral leg MMR would decrease during sitting and that the reduced oxygen deficit would explain the attenuation in PORH after PS. Methods: 31 subjects sat for 2.5 hours. A near-infrared spectroscopy (NIRS) sensor was placed above the medial gastrocnemius. PORH was quantified with the reperfusion slope of the NIRS tissue oxygenation index (TOI) after 5-min of a cuff-induced arterial occlusion. MMR was quantified with the TOI slope during a cuff-induced arterial occlusion. Measurements were made before and after sitting. To determine if changes in MMR and the oxygen deficit explains the change in the reperfusion slope, an artificial neural network (ANN) was trained to define the stimulus-response relationship between the oxygen deficit (i.e., difference in TOI during arterial occlusion (ΔTOI) and the area under baseline TOI during arterial occlusion (AUC)), MMR, and the reperfusion slope with 65 retrospectively collected PORH recordings. The ANN was then used to predict the experimentally collected reperfusion slopes at baseline and post-sitting. Therefore, if the ANN can replicate the experimental results, this provides evidence that the reperfusion slope after sitting is dependent on MMR and the oxygen deficit during an arterial occlusion. Results: Consistent with other studies, the reperfusion slope was reduced after PS (Δ -0.83±1.48 TOI%·s-1, p&lt;0.01). Unique to the current study, we discovered that MMR was reduced (Δ -0.05±0.05 TOI%·s-1, p&lt;0.01), ΔTOI was reduced (Δ -11.14±12.89 TOI%, p&lt;0.01), and AUC was reduced (Δ -2173.61±2502.41 TOI%·s, p&lt;0.01) after PS. Furthermore, there were no differences between the ANN-predicted and experimental reperfusion slopes at baseline (p=0.37) or post-sitting (p=0.11). Conclusions: Since attenuated MMR and the oxygen deficit play significant roles in the reduced reperfusion slope after PS, current inferences about microvascular function with PORH after PS are not valid. Therefore, these data provide evidence that microvascular function may be preserved during PS. Interestingly, the blunted MMR after PS may not be trivial and could be an important mechanism underlying the relationship between PS and disease development. This work was supported by the National Institutes of Health (R01 HD106911-01A1 and R01 AG077803), the University of Nebraska Collaboration Initiative Grant (32105), and the NASA Nebraska Space Grant Fellowship. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Skeletal Muscle Consequences of Phosphatidylethanolamine Synthesis Deficiency.

The maintenance of phospholipid homeostasis is increasingly being implicated in metabolic health. Phosphatidylethanolamine (PE) is the most abundant phospholipid on the inner leaflet of cellular membranes, and we have previously shown that mice with a heterozygous ablation of the PE synthesizing enzyme, Pcyt2 (Pcyt2+/-), develop obesity, insulin resistance, and NASH. Skeletal muscle is a major determinant of systemic energy metabolism, making it a key player in metabolic disease development. Both the total PE levels and the ratio of PE to other membrane lipids in skeletal muscle are implicated in insulin resistance; however, the underlying mechanisms and the role of Pcyt2 regulation in this association remain unclear. Here, we show how reduced phospholipid synthesis due to Pcyt2 deficiency causes Pcyt2+/- skeletal muscle dysfunction and metabolic abnormalities. Pcyt2+/- skeletal muscle exhibits damage and degeneration, with skeletal muscle cell vacuolization, disordered sarcomeres, mitochondria ultrastructure irregularities and paucity, inflammation, and fibrosis. There is intramuscular adipose tissue accumulation, and major disturbances in lipid metabolism with impaired FA mobilization and oxidation, elevated lipogenesis, and long-chain fatty acyl-CoA, diacylglycerol, and triacylglycerol accumulation. Pcyt2+/- skeletal muscle exhibits perturbed glucose metabolism with elevated glycogen content, impaired insulin signaling, and reduced glucose uptake. Together, this study lends insight into the critical role of PE homeostasis in skeletal muscle metabolism and health with broad implications on metabolic disease development.

Effect of RONS-Induced Intracellular Redox Homeostasis in 6-NBDG/Glucose Uptake in C2C12 Myotubes and Single Isolated Skeletal Muscle Fibres.

The glucose uptake in skeletal muscle is essential to produce energy through ATP, which is needed by this organ to maintain vital functions. The impairment of glucose uptake compromises the metabolism and function of skeletal muscle and other organs and is a feature of diabetes, obesity, and ageing. There is a need for research to uncover the mechanisms involved in the impairment of glucose uptake in skeletal muscle. In this study, we adapted, developed, optimised, and validated a methodology based on the fluorescence glucose analogue 6-NBDG, combined with a quantitative fluorescence microscopy image analysis, to determine the glucose uptake in two models of skeletal muscle cells: C2C12 myotubes and single fibres isolated from muscle. It was proposed that reactive oxygen and nitrogen species (RONS) and redox homeostasis play an important role in the modulation of intracellular redox signalling pathways associated with glucose uptake. In this study, we prove that the prooxidative intracellular redox environment under oxidative eustress produced by RONS such as hydrogen peroxide and nitric oxide improves glucose uptake in skeletal muscle cells. However, when oxidation is excessive, oxidative distress occurs, and cellular viability is compromised, although there might be an increase in the glucose uptake. Based on the results of this study, the determination of 6-NBDG/glucose uptake in myotubes and skeletal muscle cells is feasible, validated, and will contribute to improve future research.

A Systematic Approach for the Interpretation of Cardiopulmonary Exercise Testing in Children with Focus on Cardiovascular Diseases

Cardiopulmonary exercise testing (CPET) is the clinical standard for children with congenital heart disease (CHD), heart failure (HF) being assessed for transplantation candidacy, and subjects with unexplained dyspnea on exertion. Heart, lung, skeletal muscle, peripheral vasculature, and cellular metabolism impairment frequently lead to circulatory, ventilatory, and gas exchange abnormalities during exercise. An integrated analysis of the multi-system response to exercise can be beneficial for differential diagnosis of exercise intolerance. The CPET combines standard graded cardiovascular stress testing with simultaneous ventilatory respired gas analysis. This review addresses the interpretation and clinical significance of CPET results with specific reference to cardiovascular diseases. The diagnostic values of commonly obtained CPET variables are discussed using an easy-to-use algorithm for physicians and trained nonphysician personnel in clinical practice.

Neutral Effect of Skeletal Muscle Mineralocorticoid Receptor on Glucose Metabolism in Mice.

The mineralocorticoid receptor (MR) is able to regulate the transcription of a number of genes in the myotube, although its roles in skeletal muscle (SM) metabolism still await demonstration. SM represents a major site for glucose uptake, and its metabolic derangements play a pivotal role in the development of insulin resistance (IR). The aim of this study was to investigate the contribution of SM MR in mediating derangements of glucose metabolism in a mouse model of diet-induced obesity. We observed that mice fed a high-fat diet (HFD mice) showed impaired glucose tolerance compared to mice fed a normal diet (ND mice). Mice fed a 60% HFD treated with the MR antagonist Spironolactone (HFD + Spiro) for 12 weeks revealed an improvement in glucose tolerance, as measured with an intraperitoneal glucose tolerance test, compared with HFD mice. To investigate if blockade of SM MR could contribute to the favorable metabolic effects observed with pharmacological MR antagonism, we analyzed MR expression in the gastrocnemius, showing that SM MR protein abundance is downregulated by HFD compared to ND mice and that pharmacological treatment with Spiro was able to partially revert this effect in HFD + Spiro mice. Differently from what we have observed in adipose tissue, where HDF increased adipocyte MR expression, SM MR protein was down-regulated in our experimental model, suggesting a completely different role of SM MR in the regulation of glucose metabolism. To confirm this hypothesis, we investigated the effects of MR blockade on insulin signaling in a cellular model of IRin C2C12 myocytes, which were treated with or without Spiro. We confirmed MR protein downregulation in insulin-resistant myotubes. We also analyzed Akt phosphorylation upon insulin stimulation, and we did not observe any difference between palmitate- and palmitate + Spiro-treated cells. These results were confirmed by in vitro glucose uptake analysis. Taken together, our data indicate that reduced activity of SM MR does not improve insulin signaling in mouse skeletal myocytes and does not contribute to the favorable metabolic effects on glucose tolerance and IR induced by systemic pharmacological MR blockade.

Aerobic exercise reduced the amount of CHRONO bound to BMAL1 and ameliorated glucose metabolic dysfunction in skeletal muscle of high-fat diet-fed mice

AimsThe purpose of this study was to investigate the effects of aerobic exercise on the CHRONO-BMAL1 pathway and glucose metabolism in skeletal muscle of high-fat diet (HFD)-fed mice. Main methodsMale C57BL/6J mice were randomly allocated into four groups: normal chow diet with control (NCD + CON), NCD with exercise (NCD + EXE), HFD with control (HFD + CON) and HFD with exercise (HFD + EXE). The NCD and HFD groups were respectively fed a diet of 10 % and 60 % kilocalories from fat for 12 weeks. During the dietary intervention, EXE groups were subjected to 70 % VO2max intensity of treadmill exercise six times per week for 12 weeks. Body weight, energy intake, fat weight, serum lipid profiles, systemic glucose homeostasis, the amount of CHRONO bound to BMAL1, the enzymatic activity, mRNA and protein expression involved in glucose metabolism of skeletal muscle were measured. Key findingsThe results showed that the 12-week HFD feeding without exercise induced weight gain, serum dyslipidemia and insulin resistance. Furthermore, HFD increased the amount of CHRONO bound to BMAL1 and repressed the glucose metabolism in skeletal muscle. However, aerobic exercise prevented weight gain, serum dyslipidemia and systemic insulin resistance in the HFD-fed mice. Meanwhile, aerobic exercise also decreased the amount of CHRONO bound to BMAL1 and increased the glucose uptake, glucose oxidation and glycogenesis in skeletal muscle of the HFD-fed mice. SignificanceThese data suggested that aerobic exercise could counterbalance CHRONO interacted with BMAL1 and prevent glucose metabolism dysfunction of skeletal muscle, and finally maintain whole-body insulin sensitivity in the HFD-fed mice.

The transcription regulator ATF4 is a mediator of skeletal muscle aging

Aging slowly erodes skeletal muscle strength and mass, eventually leading to profound functional deficits and muscle atrophy. The molecular mechanisms of skeletal muscle aging are not well understood. To better understand mechanisms of muscle aging, we investigated the potential role of ATF4, a transcription regulatory protein that can rapidly promote skeletal muscle atrophy in young animals deprived of adequate nutrition or activity. To test the hypothesis that ATF4 may be involved in skeletal muscle aging, we studied fed and active muscle-specific ATF4 knockout mice (ATF4 mKO mice) at 6 months of age, when wild-type mice have achieved peak muscle mass and function, and at 22 months of age, when wild-type mice have begun to manifest age-related muscle atrophy and weakness. We found that 6-month-old ATF4 mKO mice develop normally and are phenotypically indistinguishable from 6-month-old littermate control mice. However, as ATF4 mKO mice become older, they exhibit significant protection from age-related declines in strength, muscle quality, exercise capacity, and muscle mass. Furthermore, ATF4 mKO muscles are protected from some of the transcriptional changes characteristic of normal muscle aging (repression of certain anabolic mRNAs and induction of certain senescence-associated mRNAs), and ATF4 mKO muscles exhibit altered turnover of several proteins with important roles in skeletal muscle structure and metabolism. Collectively, these data suggest ATF4 as an essential mediator of skeletal muscle aging and provide new insight into a degenerative process that impairs the health and quality of life of many older adults.

Metabolome-Based Genome-Wide Association Study of Duck Meat Leads to Novel Genetic and Biochemical Insights.

Meat is among the most consumed foods worldwide and has a unique flavor and high nutrient density in the human diet. However, the genetic and biochemical bases of meat nutrition and flavor are poorly understood. Here, 3431 metabolites and 702 volatiles in 423 skeletal muscle samples are profiled from a gradient consanguinity segregating population generated by Pekin duck × Liancheng duck crosses using metabolomic approaches. The authors identified 2862 metabolome-based genome-wide association studies (mGWAS) signals and 48 candidate genes potentially modulating metabolite and volatile levels, 79.2% of which are regulated by cis-regulatory elements. The level of plasmalogen is significantly associated with TMEM189 encoding plasmanylethanolamine desaturase 1. The levels of 2-pyrrolidone and glycerophospholipids are regulated by the gene expression of AOX1 and ACBD5, which further affects the levels of volatiles, 2-pyrrolidone and decanal, respectively. Genetic variations in GADL1 and CARNMT2 determine the levels of 49 metabolites including L-carnosine and anserine. This study provides novel insights into the genetic and biochemical basis of skeletal muscle metabolism and constitutes a valuable resource for the precise improvement of meat nutrition and flavor.

Red blood cell extracellular vesicles deliver therapeutic siRNAs to skeletal muscles for treatment of cancer cachexia

Cancer cachexia is a multifactorial syndrome characterized by a significant loss of skeletal muscle, that negatively affects the quality of life. Inhibition of myostatin (Mstn), a negative regulator of skeletal muscle growth and differentiation, has been proven to preserve muscle mass in muscle atrophy diseases, including cachexia. However, myostatin inhibitors have repeatedly failed clinical trials because of modest therapeutic effects and side effects due to the poor efficiency and toxicity of existing delivery methods. Here, we describe a novel method for delivering Mstn siRNA to skeletal muscles using red blood cell extracellular vesicles (RBCEVs) in a cancer cachectic mouse model. Our data show that RBCEVs are taken up by myofibers via intramuscular administration. Repeated intramuscular administrations with RBCEVs allowed the delivery of siRNAs, thereby inhibiting Mstn, increasing muscle growth and preventing cachexia in cancer-bearing mice. We observed the same therapeutic effects when delivering siRNAs against malonyl-CoA decarboxylase, an enzyme driving dysfunctional fatty acid metabolism in skeletal muscles during cancer cachexia. We demonstrate that intramuscular siRNA delivery by RBCEVs is safe and non-inflammatory. Hence, this method is useful to reduce the therapeutic dose of siRNAs, to avoid toxicity and off-target effects caused by systemic administration of naked siRNAs at high doses.

Stable isotope-labeled carnitine reveals its rapid transport into muscle cells and acetylation during contraction

Carnitine plays multiple roles in skeletal muscle metabolism, including fatty acid transport and buffering of excess acetyl-CoA in the mitochondria. The skeletal muscle cannot synthesize carnitine; therefore, carnitine must be taken up from the blood into the cytoplasm. Carnitine metabolism, its uptake into cells, and the subsequent reactions of carnitine are accelerated by muscle contraction. Isotope tracing enables the marking of target molecules and monitoring of tissue distribution. In this study, stable isotope-labeled carnitine tracing was combined with matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) imaging to determine carnitine distribution in mouse skeletal muscle tissues. Deuterium-labeled carnitine (d3-carnitine) was intravenously injected into the mice and diffused to the skeletal muscles for 30 and 60 min. To examine whether muscle contraction changes the distribution of carnitine and its derivatives, unilateral in situ muscle contraction was performed; 60 min muscle contraction showed increased d3-carnitine and its derivative d3-acetylcarnitine in the muscle, indicating that carnitine uptake in cells is promptly converted to acetylcarnitine, consequently, buffering accumulated acetyl-CoA. While the endogenous carnitine was localized in the slow type fibers rather than fast type, the contraction-induced distributions of d3-carnitine and acetylcarnitine were not necessarily associated with muscle fiber type. In conclusion, the combination of isotope tracing and MALDI-MS imaging can reveal carnitine flux during muscle contraction and show the significance of carnitine in skeletal muscles.

Dietary Methionine Restriction Improves Gastrocnemius Muscle Glucose Metabolism through Improved Insulin Secretion and H19/IRS-1/Akt Pathway in Middle-Aged Mice

Methionine restriction (MR) improves glucose metabolism. In skeletal muscle, H19 is a key regulator of insulin sensitivity and glucose metabolism. Therefore, this study aims to reveal the underlying mechanism of H19 upon MR on glucose metabolism in skeletal muscle. Middle-aged mice were fed MR diet for 25 weeks. Mouse islets β cell line β-TC6 cells and mouse myoblast cell line C2C12 cells were used to establish the apoptosis or insulin resistance model. Our findings showed that MR increased B-cell lymphoma-2 (Bcl-2) expression, deceased Bcl-2 associated X protein (Bax), cleaved cysteinyl aspartate-specific proteinase-3 (Caspase-3) expression in pancreas, and promoted insulin secretion of β-TC6 cells. Meanwhile, MR increased H19 expression, insulin Receptor Substrate-1/insulin Receptor Substrate-2 (IRS-1/IRS-2) value, protein Kinase B (Akt) phosphorylation, glycogen synthase kinase-3β (GSK3β) phosphorylation, and hexokinase 2 (HK2) expression in gastrocnemius muscle and promoted glucose uptake in C2C12 cells. But these results were reversed after H19 knockdown in C2C12 cells. In conclusion, MR alleviates pancreatic apoptosis and promotes insulin secretion. And MR enhances gastrocnemius muscle insulin-dependent glucose uptake and utilization via the H19/IRS-1/Akt pathway, thereby ameliorating blood glucose disorders and insulin resistance in high-fat-diet (HFD) middle-aged mice.

Sepsis-like Energy Deficit Is Not Sufficient to Induce Early Muscle Fiber Atrophy and Mitochondrial Dysfunction in a Murine Sepsis Model.

Sepsis-induced myopathy is characterized by muscle fiber atrophy, mitochondrial dysfunction, and worsened outcomes. Whether whole-body energy deficit participates in the early alteration of skeletal muscle metabolism has never been investigated. Three groups were studied: "Sepsis" mice, fed ad libitum with a spontaneous decrease in caloric intake (n = 17), and "Sham" mice fed ad libitum (Sham fed (SF), n = 13) or subjected to pair-feeding (Sham pair fed (SPF), n = 12). Sepsis was induced by the intraperitoneal injection of cecal slurry in resuscitated C57BL6/J mice. The feeding of the SPF mice was restricted according to the food intake of the Sepsis mice. Energy balance was evaluated by indirect calorimetry over 24 h. The tibialis anterior cross-sectional area (TA CSA), mitochondrial function (high-resolution respirometry), and mitochondrial quality control pathways (RTqPCR and Western blot) were assessed 24 h after sepsis induction. The energy balance was positive in the SF group and negative in both the SPF and Sepsis groups. The TA CSA did not differ between the SF and SPF groups, but was reduced by 17% in the Sepsis group compared with the SPF group (p < 0.05). The complex-I-linked respiration in permeabilized soleus fibers was higher in the SPF group than the SF group (p < 0.05) and lower in the Sepsis group than the SPF group (p < 0.01). Pgc1α protein expression increased 3.9-fold in the SPF mice compared with the SF mice (p < 0.05) and remained unchanged in the Sepsis mice compared with the SPF mice; the Pgc1α mRNA expression decreased in the Sepsis compared with the SPF mice (p < 0.05). Thus, the sepsis-like energy deficit did not explain the early sepsis-induced muscle fiber atrophy and mitochondrial dysfunction, but led to specific metabolic adaptations not observed in sepsis.