Robinson, Matthew M.

We investigated the link between enhancement of SI (by hyperinsulinemic–euglycemic clamp) and muscle metabolites after 12 weeks of aerobic (high-intensity interval training [HIIT]), resistance training (RT), or combined training (CT) exercise in 52 lean healthy individuals. Muscle RNA sequencing revealed a significant association between SI after both HIIT and RT and the branched-chain amino acid (BCAA) metabolic pathway. Concurrently with increased expression and activity of branched-chain ketoacid dehydrogenase enzyme, many muscle amino metabolites, including BCAAs, glutamate, phenylalanine, aspartate, asparagine, methionine, and γ-aminobutyric acid, increased with HIIT, supporting the substantial impact of HIIT on amino acid metabolism. Short-chain C3 and C5 acylcarnitines were reduced in muscle with all three training modes, but unlike RT, both HIIT and CT increased tricarboxylic acid metabolites and cardiolipins, supporting greater mitochondrial activity with aerobic training. Conversely, RT and CT increased more plasma membrane phospholipids than HIIT, suggesting a resistance exercise effect on cellular membrane protection against environmental damage. Sex and age contributed modestly to the exercise-induced changes in metabolites and their association with cardiometabolic parameters. Integrated transcriptomic and metabolomic analyses suggest various clusters of genes and metabolites are involved in distinct effects of HIIT, RT, and CT. These distinct metabolic signatures of different exercise modes independently link each type of exercise training to improved SI and cardiometabolic risk. Article Highlights We aimed to understand the link between skeletal muscle metabolites and cardiometabolic health after exercise training. Although aerobic, resistance, and combined exercise training each enhance muscle insulin sensitivity as well as other cardiometabolic parameters, they disparately alter amino and citric acid metabolites as well as the lipidome, linking these metabolomic changes independently to the improvement of cardiometabolic risks with each exercise training mode. These findings reveal an important layer of the unique exercise mode–dependent changes in muscle metabolism, which may eventually lead to more informed exercise prescription for improving SI.

We assessed the effects of 12 wk of supervised high-intensity interval training (HIIT), resistance training, and combined training (CT) on skeletal muscle mitochondrial abundance and markers of fission and fusion. HIIT increased mitochondrial area and size and promoted protein changes indicative of increased mitochondrial fusion, whereas lessor effects were observed after CT and no changes were observed after RT. Furthermore, increased mitochondrial area and size after HIIT associated with improved mitochondrial respiration, cardiorespiratory fitness, and insulin sensitivity.

Maximal aerobic exercise capacity [maximal oxygen consumption (V̇o 2max )] is one of the strongest predictors of morbidity and mortality. Aerobic exercise training can increase V̇o 2max , but inter-individual variability is marked and unexplained physiologically. The mechanisms underlying this variability have major clinical implications for extending human healthspan. Here, we report a novel transcriptome signature related to ΔV̇o 2max with exercise training detected in whole blood RNA. We used RNA-Seq to characterize transcriptomic signatures of ΔV̇o 2max in healthy women who completed a 16-wk randomized controlled trial comparing supervised, higher versus lower aerobic exercise training volume and intensity (4 training groups, fully crossed). We found significant baseline gene expression differences in subjects who responded to aerobic exercise training with robust versus little/no ΔV̇o 2max , and differentially expressed genes/transcripts were mostly related to inflammatory signaling and mitochondrial function/protein translation. Baseline gene expression signatures associated with robust versus little/no ΔV̇o 2max were also modulated by exercise training in a dose-dependent manner, and they predicted ΔV̇o 2max in this and a separate dataset. Collectively, our data demonstrate the potential utility of using whole blood transcriptomics to study the biology of inter-individual variability in responsiveness to the same exercise training stimulus.

Excess fat intake can increase lipid oxidation and expression of mitochondrial proteins, indicating remodeling of the mitochondrial proteome. Yet intermediates of lipid oxidation also accumulate, indicating a relative insufficiency to completely oxidize lipids. We investigated remodeling of the mitochondrial proteome to determine mechanisms of changes in lipid oxidation following high-fat feeding. C57BL/6J mice consumed a high-fat diet (HFD, 60% fat from lard) or a low-fat diet (LFD, 10% fat) for 12 wk. Mice were fasted for 4 h and then anesthetized by pentobarbital sodium overdose for tissue collection. A mitochondrial-enriched fraction was prepared from gastrocnemius muscles and underwent proteomic analysis by high-resolution mass spectrometry. Mitochondrial respiratory efficiency was measured as the ratio of ATP production to O2 consumption. Intramuscular acylcarnitines were measured by liquid chromatography-mass spectrometry. A total of 658 mitochondrial proteins were identified: 40 had higher abundance and 14 had lower abundance in mice consuming the HFD than in mice consuming the LFD. Individual proteins that changed with the HFD were primarily related to β-oxidation; there were fewer changes to the electron transfer system. Gene set enrichment analysis indicated that the HFD increased pathways of lipid metabolism and β-oxidation. Intramuscular concentrations of select acylcarnitines (C18:0) were greater in the HFD mice and reflected dietary lipid composition. Mitochondrial respiratory ATP production-to-O2 consumption ratio for lipids was not different between LFD and HFD mice. After the 60% fat diet, remodeling of the mitochondrial proteome revealed upregulation of proteins regulating lipid oxidation that was not evident for all mitochondrial pathways. The accumulation of lipid metabolites with obesity may occur without intrinsic dysfunction to mitochondrial lipid oxidation.

Abstract Context Aerobic exercise training can increase brain volume and blood flow, but the impact on brain metabolism is less known. Objective We determined whether high-intensity interval training (HIIT) increases brain metabolism by measuring brain glucose uptake in younger and older adults. Design Brain glucose uptake was measured before and after HIIT or a sedentary (SED) control period within a larger exercise study. Setting Study procedures were performed at the Mayo Clinic in Rochester, MN. Participants Participants were younger (18 to 30 years) or older (65 to 80 years) SED adults who were free of major medical conditions. Group sizes were 15 for HIIT (nine younger and six older) and 12 for SED (six younger and six older). Intervention Participants completed 12 weeks of HIIT or SED. HIIT was 3 days per week of 4 × 4 minute intervals at over 90% of peak aerobic capacity (VO2peak) with 2 days per week of treadmill walking at 70% VO2peak. Main Outcome Measures Resting brain glucose uptake was measured using 18F-fluorodeoxyglucose positron emission tomography scans at baseline and at week 12. Scans were performed at 96 hours after exercise. VO2peak was measured by indirect calorimetry. Results Glucose uptake increased significantly in the parietal-temporal and caudate regions after HIIT compared with SED. The gains with HIIT were not observed in all brain regions. VO2peak was increased for all participants after HIIT and did not change with SED. Conclusion We demonstrate that brain glucose metabolism increased after 12 weeks of HIIT in adults in regions where it is reduced in Alzheimer’s disease.

Skeletal muscle mitochondrial protein synthesis is regulated in part by insulin. The development of insulin resistance with diet-induced obesity may therefore contribute to impairments to protein synthesis and decreased mitochondrial respiration. Yet the impact of diet-induced obesity and insulin resistance on mitochondrial energetics is controversial, with reports varying from decreases to increases in mitochondrial respiration. We investigated the impact of changes in insulin sensitivity on long-term rates of mitochondrial protein synthesis as a mechanism for changes to mitochondrial respiration in skeletal muscle. Insulin resistance was induced in C57BL/6J mice using 4 wk of a high-fat compared with a low-fat diet. For 8 additional weeks, diets were enriched with pioglitazone to restore insulin sensitivity compared with nonenriched control low-fat or high-fat diets. Skeletal muscle mitochondrial protein synthesis was measured using deuterium oxide labeling during weeks 10–12. High-resolution respirometry was performed using palmitoyl-l-carnitine, glutamate+malate, and glutamate+malate+succinate as substrates for mitochondria isolated from quadriceps. Mitochondrial protein synthesis and palmitoyl- l-carnitine oxidation were increased in mice consuming a high-fat diet, regardless of differences in insulin sensitivity with pioglitazone treatment. There was no effect of diet or pioglitazone treatment on ADP-stimulated respiration or H2O2 emission using glutamate+malate or glutamate+malate+succinate. The results demonstrate no impairments to mitochondrial protein synthesis or respiration following induction of insulin resistance. Instead, mitochondrial protein synthesis was increased with a high-fat diet and may contribute to remodeling of the mitochondria to increase lipid oxidation capacity. Mitochondrial adaptations with a high-fat diet appear driven by nutrient availability, not intrinsic defects that contribute to insulin resistance.

Insulin regulates skeletal muscle protein degradation, but the types of proteins being degraded in vivo remain to be determined due to methodological limitations. We present a method to assess the types of skeletal muscle proteins that are degraded by extracting their degradation products as low-molecular weight (LMW) peptides from muscle samples. High-resolution mass spectrometry was used to identify the original intact proteins that generated the LMW peptides, which we validated in rodents and then applied to humans. We deprived insulin from insulin-treated streptozotocin (STZ) diabetic mice for 6 and 96 h and for 8 h in type 1 diabetic humans (T1D) for comparison with insulin-treated conditions. Protein degradation was measured using activation of autophagy and proteasome pathways, stable isotope tracers, and LMW approaches. In mice, insulin deprivation activated proteasome pathways and autophagy in muscle homogenates and isolated mitochondria. Reproducibility analysis of LMW extracts revealed that ∼80% of proteins were detected consistently. As expected, insulin deprivation increased whole body protein turnover in T1D. Individual protein degradation increased with insulin deprivation, including those involved in mitochondrial function, proteome homeostasis, nDNA support, and contractile/cytoskeleton. Individual mitochondrial proteins that generated more LMW fragment with insulin deprivation included ATP synthase subunit-γ (+0.5-fold, P = 0.007) and cytochrome c oxidase subunit 6 (+0.305-fold, P = 0.03). In conclusion, identifying LMW peptide fragments offers an approach to determine the degradation of individual proteins. Insulin deprivation increases degradation of select proteins and provides insight into the regulatory role of insulin in maintaining proteome homeostasis, especially of mitochondria.

Context:Numerous factors are purported to influence insulin sensitivity including age, adiposity, mitochondrial function, and physical fitness. Univariate associations cannot address the complexity of insulin resistance or the interrelationship among potential determinants.Objective:The objective of the study was to identify significant independent predictors of insulin sensitivity across a range of age and adiposity in humans.Design, Setting, and Participants:Peripheral and hepatic insulin sensitivity were measured by two stage hyperinsulinemic-euglycemic clamps in 116 men and women (aged 19–78 y). Insulin-stimulated glucose disposal, the suppression of endogenous glucose production during hyperinsulinemia, and homeostatic model assessment of insulin resistance were tested for associations with 11 potential predictors. Abdominal subcutaneous fat, visceral fat (AFVISC), intrahepatic lipid, and intramyocellular lipid (IMCL) were quantified by magnetic resonance imaging and spectroscopy. Skeletal muscle mitochondrial respiratory capacity (state 3), coupling efficiency, and reactive oxygen species production were evaluated from muscle biopsies. Aerobic fitness was measured from whole-body maximum oxygen uptake (VO2 peak), and metabolic flexibility was determined using indirect calorimetry.Results:Multiple regression analysis revealed that AFVISC (P < .0001) and intrahepatic lipid (P = .002) were independent negative predictors of peripheral insulin sensitivity, whereas VO2 peak (P = .0007) and IMCL (P = .023) were positive predictors. Mitochondrial capacity and efficiency were not independent determinants of peripheral insulin sensitivity. The suppression of endogenous glucose production during hyperinsulinemia model of hepatic insulin sensitivity revealed percentage fat (P < .0001) and AFVISC (P = .001) as significant negative predictors. Modeling homeostatic model assessment of insulin resistance identified AFVISC (P < .0001), VO2 peak (P = .001), and IMCL (P = .01) as independent predictors.Conclusion:The reduction in insulin sensitivity observed with aging is driven primarily by age-related changes in the content and distribution of adipose tissue and is independent of muscle mitochondrial function or chronological age.

Caloric restriction (CR) improves insulin sensitivity and reduces the incidence of diabetes in obese individuals. The underlying mechanisms whereby CR improves insulin sensitivity are not clear. We evaluated the effect of 16 weeks of CR on whole-body insulin sensitivity by pancreatic clamp before and after CR in 11 obese participants (BMI = 35 kg/m2) compared with 9 matched control subjects (BMI = 34 kg/m2). Compared with the control subjects, CR increased the glucose infusion rate needed to maintain euglycemia during hyperinsulinemia, indicating enhancement of peripheral insulin sensitivity. This improvement in insulin sensitivity was not accompanied by changes in skeletal muscle mitochondrial oxidative capacity or oxidant emissions, nor were there changes in skeletal muscle ceramide, diacylglycerol, or amino acid metabolite levels. However, CR lowered insulin-stimulated thioredoxin-interacting protein (TXNIP) levels and enhanced nonoxidative glucose disposal. These results support a role for TXNIP in mediating the improvement in peripheral insulin sensitivity after CR.