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Am J Physiol Endocrinol Metab 298: E49-E58, 2010. First published November 3, 2009; doi:10.1152/ajpendo.00317.2009.--Insulin resistance in skeletal muscle in obesity and T2DM is associated with reduced muscle oxidative capacity, reduced expression in nuclear genes responsible for oxidative metabolism, and reduced activity of mitochondrial electron transport chain. The presented study was undertaken to analyze mitochondrial content and mitochondrial enzyme profile in skeletal muscle of sedentary lean individuals and to compare that with our previous data on obese or obese T2DM group. Frozen skeletal muscle biopsies obtained from lean volunteers were used to estimate cardiolipin content, mtDNA (markers of mitochondrial mass), NADH oxidase activity of mitochondrial electron transport chain (ETC), and activity of citrate synthase and [beta]-hydroxyacyl-CoA dehydrogenase ([beta]-HAD), key enzymes of TCA cycle and [beta]-oxidation pathway, respectively. Frozen biopsies collected from obese or T2DM individuals in our previous studies were used to estimate activity of [beta]-HAD. The obtained data were complemented by data from our previous studies and statistically analyzed to compare mitochondrial content and mitochondrial enzyme profile in lean, obese, or T2DM cohort. The total activity of NADH oxidase was reduced significantly in obese or T2DM subjects. The cardiolipin content for lean or obese group was similar, and although for T2DM group cardiolipin showed a tendency to decline, it was statistically insignificant. The total activity of citrate synthase for lean and T2DM group was similar; however, it was increased significantly in the obese group. Activity of [beta]-HAD and mtDNA content was similar for all three groups. We conclude that the total activity of NADH oxidase in biopsy for lean group is significantly higher than corresponding activity for obese or T2DM cohort. The specific activity of NADH oxidase (per mg cardiolipin) and NADH oxidase/citrate synthase and NADH oxidase/ [beta]-HAD ratios are reduced two- to threefold in both T2DM and obesity. insulin resistance; [beta]-oxidation; cardiolipin; reduced nicotinamide adenine dinucleotide/nicotinamide adenine dinucleotide ratio; trichloroacetic acid cycle

Type 2 diabetes is characterized by fasting hyperglycemia, secondary to hepatic insulin resistance and increased glucose production. Peroxisome proliferator-activated receptor-[gamma] coactivator-1[alpha] (PGC-1[alpha]) is a transcriptional coactivator that is thought to control adaptive responses to physiological stimuli. In liver, PGC-1[alpha] expression is induced by fasting, and this effect promotes gluconeogenesis. To examine whether PGC-1[alpha] is involved in the pathogenesis of hepatic insulin resistance, we generated transgenic (TG) mice with whole body overexpression of human PGC-1[alpha] and evaluated glucose homeostasis with a euglycemic-hyperinsulinemic clamp. PGC-1[alpha] was moderately (~2-fold) overexpressed in liver, skeletal muscle, brain, and heart of TG mice. In liver, PGC-l[alpha] overexpression resulted in increased expression of hepatocyte nuclear factor-4[alpha] and the gluconeogenic enzymes phosphoenolpyruvate carboxykinase and glucose-6-phosphatase. PGC-1[alpha] overexpression caused hepatic insulin resistance, manifested by higher glucose production and diminished insulin suppression of gluconeogenesis. Paradoxically, PGC-1[alpha] overexpression improved muscle insulin sensitivity, as evidenced by elevated insulin-stimulated Akt phosphorylation and peripheral glucose disposal. Content of myoglobin and troponin I slow protein was increased in muscle of TG mice, indicating fiber-type switching. PGC-1[alpha] overexpression also led to lower reactive oxygen species production by mitochondria and reduced IKK/I[kappa]B signaling in muscle. Feeding a high-fat diet to TG mice eliminated the increased muscle insulin sensitivity. The dichotomous effect of PGC-1[alpha] overexpression in liver and muscle suggests that PGC-1[alpha] is a fuel gauge that couples energy demands (muscle) with the corresponding fuel supply (liver). Thus, under conditions of physiological stress (i.e., prolonged fast and exercise training), increased hepatic glucose production may help sustain glucose utilization in peripheral tissues. diabetes; phosphoenolpyruvate carboxykinase; glucose-6-phosphatase; gluconeogenesis

We previously reported an 'athlete's paradox' in which endurance-trained athletes, who possess a high oxidative capacity and enhanced insulin sensitivity, also have higher intramyocellular lipid (IMCL) content. The purpose of this study was to determine whether moderate exercise training would increase IMCL, oxidative capacity of muscle, and insulin sensitivity in previously sedentary overweight to obese, insulin-resistant, older subjects. Twenty-five older (66.4 [+ or -] 0.8 yr) obese (BMI = 30.3 [+ or -] 0.7 kg/[m.sup.2]) men (n = 9) and women (n = 16) completed a 16-wk moderate but progressive exercise training program. Body weight and fat mass modestly but significantly (P < 0.01) decreased. Insulin sensitivity, measured using the euglycemic hyperinsulinemic clamp, was increased (21%, P = 0.02), with modest improvements (7%, P = 0.04) in aerobic fitness ([Vo.sub.2peak]). Histochemical analyses of IMCL (Oil Red O staining), oxidative capacity [succinate dehydrogenase activity (SDH)], glycogen content, capillary density, and fiber type were performed on skeletal muscle biopsies. Exercise training increased IMCL by 21%. In contrast, diacylglycerol and ceramide, measured by mass spectroscopy, were decreased (n = 13; -29% and -24%, respectively, P < 0.05) with exercise training. SDH (19%), glycogen content (15%), capillary density (7%), and the percentage of type I slow oxidative fibers (from 50.8 to 55.7%), all P [less than or equal to] 0.05, were increased after exercise. In summary, these results extend the athlete's paradox by demonstrating that chronic exercise in overweight to obese older adults improves insulin sensitivity in conjunction with favorable alterations in lipid partitioning and an enhanced oxidative capacity within muscle. Therefore, several key deleterious effects of aging and/or obesity on the metabolic profile of skeletal muscle can be reversed with only moderate increases in physical activity. insulin sensitivity; aging; diacylglycerol; ceramide

The current study was undertaken to address responsiveness of skeletal muscle mitochondrial electron transport chain (ETC) activity to weight loss (WL) and exercise in overweight or obese, sedentary volunteers. Fourteen middle-aged participants (7 male/7 female) had assessments of mitochondrial ETC activity and mitochondrial (mt)DNA in vastus lateralis muscle, obtained by percutaneous biopsy, before and after a 16-wk intervention. Mean WL was 9.7 (1.5%) and the mean increase in [VO.sub.2 max] was [means (SD)] 21.7 (3.7)%. Total ETC activity increased significantly, from 0.13 (0.02) to 0.19 (0.03) U/mU creatine kinase (CK; P < 0.001). ETC activity was also assessed in mitochondria isolated into subsarcolemmal (SSM) and intermyofibrillar (IMF-M) fractions. In response to intervention, there was a robust increase of ETC activity in SSM (0.028 (0.007) to 0.046 (0.011) U/mU CK, P < 0.001), and in IMF-M [0.101 (0.015) to 0.148 (0.018) U/mU CK, P < 0.005]. At baseline, the percentage of ETC activity contained in the SSM fraction was low and remained unchanged following intervention [19 (3) vs. 22 (2)%], despite the increase in ETC activity. Also, muscle mtDNA content did not change significantly [1665 (213) vs. 1874 (214) mtDNA/nuclear DNA], denoting functional improvement rather than proliferation of mitochondria as the principal mechanism of enhanced ETC activity. Increases in ETC activity were correlated with energy expenditure during exercise sessions, and ETC activity in SSM correlated with insulin sensitivity after adjustment for [VO.sub.2 max]. In summary, skeletal muscle ETC activity is increased by WL and exercise in previously sedentary obese men and women. We conclude that improved skeletal muscle ETC activity following moderate WL and improved aerobic capacity contributes to associated alleviation of insulin resistance. insulin resistance; exercise; mitochondria; mitochondrial DNA; succinate oxidase

Exercise training increases intramyocellular lipid and oxidative capacity in older adults. Am J Physiol Endocrinol Metab 287: E857-E862, 2004. First published June 29, 2004; doi:10.1152/ajpendo.00459.2003.--Intramyocellular lipid (IMCL) has been associated with insulin resistance. However, an association between IMCL and insulin resistance might be modulated by oxidative capacity in skeletal muscle. We examined the hypothesis that 12 wk of exercise training would increase both IMCL and the oxidative capacity of skeletal muscle in older (67.3 [+ or -] 0.7 yr), previously sedentary subjects (n = 13; 5 men and 8 women). Maximal aerobic capacity ([Vo.sub.2 max]) increased from 1.65 [+ or -] 0.20 to 1.85 [+ or -] 0.14 l/min (P < 0.05), and systemic fat oxidation induced by 1 h of cycle exercise at 45% of [Vo.sub.2 max] increased (P < 0.05) from 15.03 [+ or -] 40 to 19.29 [+ or -] 0.80 ([micro]mol*[min.sup.-1]*kg fat-free [mass.sup.-1]). IMCL, determined by quantitative histological staining in vastus lateralis biopsies, increased (P < 0.05) from 22.9 [+ or -] 1.9 to 25.9 [+ or -] 2.6 arbitrary units (AU). The oxidative capacity of muscle, determined by succinate dehydrogenase staining intensity, significantly increased (P < 0.05) from 75.2 [+ or -] 5.2 to 83.9 [+ or -] 3.6 AU. The percentage of type I fibers significantly increased (P < 0.05) from 35.4 [+ or -] 2.1 to 40.1 [+ or -] 2.3%. In conclusion, exercise training increases IMCL in older persons in parallel with an enhanced capacity for fat oxidation. skeletal muscle; physical activity; fiber type; triacylglycerol

Subdivisions of subcutaneous abdominal adipose tissue and insulin resistance. Am J Physiol Endocrinol Metab 278: E941-E948, 2000.--Whereas truncal (central) adiposity is strongly associated with the insulin resistant metabolic syndrome, it is uncertain whether this is accounted for principally by visceral adiposity (VAT). Several recent studies find as strong or stronger association between subcutaneous abdominal adiposity (SAT) and insulin resistance. To reexamine the issue of truncal adipose tissue depots, we performed cross-sectional abdominal computed tomography, and we undertook the novel approach of partitioning SAT into the plane superficial to the fascia within subcutaneous adipose tissue (superficial SAT) and that below this fascia (deep SAT), as well as measurement of VAT. Among 47 lean and obese glucose-tolerant men and women, insulin-stimulated glucose utilization, measured by euglycemic clamp, was strongly correlated with both VAT and deep SAT (r = -0.61 and -0.64, respectively; both P [is less than] 0.001), but not with superficial SAT (r = -0.29, not significant). Also, VAT and deep SAT followed a highly congruent pattern of associations with glucose and insulin area under the curve (75-g oral glucose tolerance test), mean arterial blood pressure, apoprotein-B, high-density lipoprotein cholesterol, and triglyceride. Superficial SAT had markedly weaker association with all these parameters and instead followed the pattern observed for thigh subcutaneous adiposity. We conclude that there are two functionally distinct compartments of adipose tissue within abdominal subcutaneous fat and that the deep SAT has a strong relation to insulin resistance. obesity; insulin resistance; visceral adiposity

Kelley, David E., Bret Goodpaster, Rena R. Wing, and Jean-Aime Simoneau. Skeletal muscle fatty acid metabolism in association with insulin resistance, obesity, and weight loss. Am. J. Physiol. 277 (Endocrinol. Metab. 40): E1130-E1141, 1999.--The current study was undertaken to investigate fatty acid metabolism by skeletal muscle to examine potential mechanisms that could lead to increased muscle triglyceride in obesity. Sixteen lean and 40 obese research volunteers had leg balance measurement of glucose and free fatty acid (FFA) uptake (fractional extraction of [9,10 [sup.3]H]oleate) and indirect calorimetry across the leg to determine substrate oxidation during fasting and insulin-stimulated conditions. Muscle obtained by percutaneous biopsy had lower carnitine palmitoyl transferase (CPT) activity and oxidative enzyme activity in obesity (P [is less than] 0.05). During fasting conditions, obese subjects had an elevated leg respiratory quotient (RQ, 0.83 [+ or -] 0.02 vs. 0.90 [+ or -] 0.01; P [is less than] 0.01) and reduced fat oxidation but similar FFA uptake across the leg. During insulin infusions, fat oxidation by leg tissues was suppressed in lean but not obese subjects; rates of FFA uptake were similar. Fasting values for leg RQ correlated with insulin sensitivity (r = -0.57, P [is less than] 0.001). Thirty-two of the obese subjects were restudied after weight loss (WL, - 14.0 [+ or -] 0.9 kg); insulin sensitivity and insulin suppression of fat oxidation improved (P [is less than] 0.01), but fasting leg RQ (0.90 [+ or -] 0.02 vs. 0.90 [+ or -] 0.02, pre-WL vs. post-WL) and muscle CPT activity did not change. The findings suggest that triglyceride accumulation in skeletal muscle in obesity derives from reduced capacity for fat oxidation and that inflexibility in regulating fat oxidation, more than fatty acid uptake, is related to insulin resistance. arteriovenous balance; indirect calorimetry; fatty acid metabolism

We examined single muscle fiber contractile function of the oldest-old (3F/2M, 89 ± 1 yr old) enrolled in The Health, Aging, and Body Composition Study (The Health ABC Study). Vastus lateralis muscle biopsies were obtained and single muscle fiber function was determined (n = 105) prior to myosin heavy chain (MHC) isoform identification with sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Cross-sectional area of MHC I muscle fibers (5,576 ± 333 μm2; n = 58) was 21% larger (P < 0.05) than MHC IIa fibers (4,518 ± 386 μm2; n = 47). Normalized power (an indicator of muscle fiber quality incorporating size, strength, and speed) of MHC I and IIa muscle fibers was 2.3 ± 0.1 and 17.4 ± 0.8 W/l, respectively. Compared with previous research from our lab using identical procedures, MHC I normalized power was 28% higher than healthy 20 yr olds and similar to younger octogenarians (∼80 yr old). Normalized power of MHC IIa fibers was 63% greater than 20 yr olds and 39% greater than younger octogenarians. These comparative data suggest that power output per unit size (i.e., muscle quality) of remaining muscle fibers improves with age, a phenomenon more pronounced in MHC IIa fibers. Age-related single muscle fiber quality improvements may be a compensatory mechanism to help offset decrements in whole muscle function. [ABSTRACT FROM AUTHOR]

Type 2 diabetes is characterized by fasting hyperglycemia, secondary to hepatic insulin resistance and increased glucose production. Peroxisome proliferator-activated re- ceptor-γ coactivator-1α (PGC-1α) is a transcriptional coactivator that is thought to control adaptive responses to physiological stimuli. In liver, PGC-1α expression is induced by fasting, and this effect promotes gluconeogenesis. To examine whether PGC-1α is involved in the pathogenesis of hepatic insulin resistance, we generated transgenic (TO) mice with whole body overexpression of human PGC-1α and evaluated glucose homeostasis with a euglycemic-hyperinsulinemic clamp. PGC-1α was moderately (~P2-fold) overexpressed in liver, skeletal muscle, brain, and heart of TO mice. In liver, PGC-1α overexpression resulted in increased expression of hepatocyte nuclear factor-4α and the gluconeogenic enzymes phosphoeno/pyruvate carboxykinase and glucose-6-phosphatase. PGC-1α overexpression caused hepatic insulin resistance, manifested by higher glucose production and diminished insulin suppression of gluconeogenesis. Paradoxically, PGC-1α overexpression improved muscle insulin sensitivity, as evidenced by elevated insulin-stimulated Akt phosphorylation and peripheral glucose disposal. Content of myoglobin and troponin I slow protein was increased in muscle of TO mice, indicating fiber-type switching. PGC-1α overexpression also led to lower reactive oxygen species production by mitochondria and reduced 1KK/1κB signaling in muscle. Feeding a high-fat diet to TO mice eliminated the increased muscle insulin sensitivity. The dichotomous effect of POC-1α overexpression in liver and muscle suggests that PGC-1α is a fuel gauge that couples energy demands (muscle) with the corresponding fuel supply (liver). Thus, under conditions of physiological stress (i.e., prolonged fast and exercise training), increased hepatic glucose production may help sustain glucose utilization in peripheral tissues. [ABSTRACT FROM AUTHOR]

Physical activity, including structured exercise, is associated with favorable health-related chronic disease outcomes. Although there is evidence of various molecular pathways that affect these responses, a comprehensive molecular map of these molecular responses to exercise has not been developed. The Molecular Transducers of Physical Activity Consortium (MoTrPAC) is a multicenter study designed to isolate the effects of structured exercise training on the molecular mechanisms underlying the health benefits of exercise and physical activity. MoTrPAC contains both a preclinical and human component. The details of the human studies component of MoTrPAC that include the design and methods are presented here. The human studies contain both an adult and pediatric component. In the adult component, sedentary participants are randomized to 12 wk of Control, Endurance Exercise Training, or Resistance Exercise Training with outcomes measures completed before and following the 12 wk. The adult component also includes recruitment of highly active endurance-trained or resistance-trained participants who only complete measures once. A similar design is used for the pediatric component; however, only endurance exercise is examined. Phenotyping measures include weight, body composition, vital signs, cardiorespiratory fitness, muscular strength, physical activity and diet, and other questionnaires. Participants also complete an acute rest period (adults only) or exercise session (adults, pediatrics) with collection of biospecimens (blood only for pediatrics) to allow for examination of the molecular responses. The design and methods of MoTrPAC may inform other studies. Moreover, MoTrPAC will provide a repository of data that can be used broadly across the scientific community. NEW & NOTEWORTHY The Molecular Transducers of Physical Activity Consortium (MoTrPAC) will be the first large trial to isolate the effects of structured exercise training on the molecular mechanisms underlying the health benefits of exercise and physical activity. By generating a compendium of the molecular responses to exercise, MoTrPAC will lay the foundation for a new era of biomedical research on Precision Exercise Medicine. Presented here is the design, protocols, and procedures for the MoTrPAC human studies.

Age-related declines in cardiorespiratory fitness and physical function are mitigated by regular endurance exercise in older adults. This may be due, in part, to changes in the transcriptional program of skeletal muscle following repeated bouts of exercise. However, the impact of chronic exercise training on the transcriptional response to an acute bout of endurance exercise has not been clearly determined. Here, we characterized baseline differences in muscle transcriptome and exercise-induced response in older adults who were active/endurance trained or sedentary. RNA-sequencing was performed on vastus lateralis biopsy specimens obtained before, immediately after, and 3 h following a bout of endurance exercise (40 min of cycling at 60%-70% of heart rate reserve). Using a recently developed bioinformatics approach, we found that transcript signatures related to type I myofibers, mitochondria, and endothelial cells were higher in active/endurance-trained adults and were associated with key phenotypic features including V̇o 2peak , ATP max , and muscle fiber proportion. Immune cell signatures were elevated in the sedentary group and linked to visceral and intermuscular adipose tissue mass. Following acute exercise, we observed distinct temporal transcriptional signatures that were largely similar among groups. Enrichment analysis revealed catabolic processes were uniquely enriched in the sedentary group at the 3-h postexercise timepoint. In summary, this study revealed key transcriptional signatures that distinguished active and sedentary adults, which were associated with difference in oxidative capacity and depot-specific adiposity. The acute response signatures were consistent with beneficial effects of endurance exercise to improve muscle health in older adults irrespective of exercise history and adiposity. NEW & NOTEWORTHY Muscle transcript signatures associated with oxidative capacity and immune cells underlie important phenotypic and clinical characteristics of older adults who are endurance trained or sedentary. Despite divergent phenotypes, the temporal transcriptional signatures in response to an acute bout of endurance exercise were largely similar among groups. These data provide new insight into the transcriptional programs of aging muscle and the beneficial effects of endurance exercise to promote healthy aging in older adults.

Skeletal muscle atrophy is a clinically important outcome of disuse because of injury, immobilization, or bed rest. Disuse atrophy is accompanied by mitochondrial dysfunction, which likely contributes to activation of the muscle atrophy program. However, the linkage of muscle mass and mitochondrial energetics during disuse atrophy and its recovery is incompletely understood. Transcriptomic analysis of muscle biopsies from healthy older adults subject to complete bed rest revealed marked inhibition of mitochondrial energy metabolic pathways. To determine the temporal sequence of muscle atrophy and changes in intramyocellular lipid and mitochondrial energetics, we conducted a time course of hind limb unloading-induced atrophy in adult mice. Mitochondrial respiration and calcium retention capacity were diminished, whereas H 2 O 2 emission was increased within 3 days of unloading before significant muscle atrophy. These changes were associated with a decrease in total cardiolipin and profound changes in remodeled cardiolipin species. Hind limb unloading performed in muscle-specific peroxisome proliferator-activated receptor-γ coactivator-1α/β knockout mice, a model of mitochondrial dysfunction, did not affect muscle atrophy but impacted muscle function. These data suggest early mitochondrial remodeling affects muscle function but not mass during disuse atrophy. Early alterations in mitochondrial energetics and lipid remodeling may represent novel targets to prevent muscle functional impairment caused by disuse and to enhance recovery from periods of muscle atrophy.

We hypothesized that acute lipid-induced insulin resistance would be attenuated in high-oxidative muscle of lean trained (LT) endurance athletes due to their enhanced metabolic flexibility and mitochondrial capacity. Lean sedentary (LS), obese sedentary (OS), and LT participants completed two hyperinsulinemic euglycemic clamp studies with and without (glycerol control) the coinfusion of Intralipid. Metabolic flexibility was measured by indirect calorimetry as the oxidation of fatty acids and glucose during fasted and insulin-stimulated conditions, the latter with and without lipid oversupply. Muscle biopsies were obtained for mitochondrial and insulin-signaling studies. During hyperinsulinemia without lipid, glucose infusion rate (GIR) was lowest in OS due to lower rates of nonoxidative glucose disposal (NOGD), whereas state 4 respiration was increased in all groups. Lipid infusion reduced GIR similarly in all subjects and reduced state 4 respiration. However, in LT subjects, fat oxidation was higher with lipid oversupply, and although glucose oxidation was reduced, NOGD was better preserved compared with LS and OS subjects. Mitochondrial performance was positively associated with better NOGD and insulin sensitivity in both conditions. We conclude that enhanced mitochondrial performance with exercise is related to better metabolic flexibility and insulin sensitivity in response to lipid overload., (Copyright © 2014 the American Physiological Society.)

Excess amounts of abdominal subcutaneous (SAT) and visceral (VAT) adipose tissue (AT) are associated with insulin resistance, even in normal-weight subjects. In contrast, gluteal-femoral AT (GFAT) is hypothesized to offer protection against insulin resistance. Dynamic PET imaging studies were undertaken to examine the contributions of both metabolic activity and size (volume) of these depots in systemic glucose metabolism. Nonobese, healthy volunteers (n = 15) underwent dynamic PET imaging uptake of [¹⁸F]FDG at a steady-state (20 mU·m⁻²·min⁻¹) insulin infusion. PET images of tissue [¹⁸F]FDG activity were coregistered with MRI to derive K values for insulin-stimulated rates of fractional glucose uptake within tissue. Adipose tissue volume was calculated from DEXA and MRI. VAT had significantly higher rates of fractional glucose uptake per volume than SAT (P < 0.05) or GFAT (P < 0.01). K(GFAT) correlated positively (r = 0.67, P < 0.01) with systemic insulin sensitivity [glucose disappearance rate (R(d))] and negatively with insulin-suppressed FFA (r = -0.71, P < 0.01). SAT (r = -0.70, P < 0.01) and VAT mass (r = -0.55, P < 0.05) correlated negatively with R(d), but GFAT mass did not. We conclude that rates of fractional glucose uptake within GFAT and VAT are significantly and positively associated with systemic insulin sensitivity in nonobese subjects. Furthermore, whereas SAT and VAT amounts are confirmed to relate to systemic insulin resistance, GFAT amount is not associated with insulin resistance. These dynamic PET imaging studies indicate that both quantity and quality of specific AT depots have distinct roles in systemic insulin resistance and may help explain the metabolically obese but normal-weight phenotype.