Young, healthy males and females present cardiometabolic protection against the detrimental effects of a 7-day high-fat high-calorie diet.

PURPOSE: High-fat, high-calorie (HFHC) diets have been used as a model to investigate lipid-induced insulin resistance. Short-term HFHC diets reduce insulin sensitivity in young healthy males, but to date, no study has directly compared males and females to elucidate sex-specific differences in the effects of a HFHC diet on functional metabolic and cardiovascular outcomes. METHODS: Eleven males (24 ± 4 years; BMI 23 ± 2 kg.m-2; V̇O2 peak 62.3 ± 8.7 ml.min-1.kg-1FFM) were matched to 10 females (25 ± 4 years; BMI 23 ± 2 kg.m-2; V̇O2 peak 58.2 ± 8.2 ml.min-1.kg-1FFM). Insulin sensitivity, measured via oral glucose tolerance test, metabolic flexibility, arterial stiffness, body composition and blood lipids and liver enzymes were measured before and after 7 days of a high-fat (65% energy) high-calorie (+ 50% kcal) diet. RESULTS: The HFHC diet did not change measures of insulin sensitivity, metabolic flexibility or arterial stiffness in either sex. There was a trend towards increased total body fat mass (kg) after the HFHC diet (+ 1.8% and + 2.3% for males and females, respectively; P = 0.056). In contrast to females, males had a significant increase in trunk to leg fat mass ratio (+ 5.1%; P = 0.005). CONCLUSION: Lean, healthy young males and females appear to be protected from the negative cardio-metabolic effects of a 7-day HFHC diet. Future research should use a prolonged positive energy balance achieved via increased energy intake and reduced energy expenditure to exacerbate negative metabolic and cardiovascular functional outcomes to determine whether sex-specific differences exist under more metabolically challenging conditions.

Decreased Aerobic Exercise Capacity After Long-Term Remission From Cushing Syndrome: Exploration of Mechanisms.

BACKGROUND: Although major improvements are achieved after cure of Cushing syndrome (CS), fatigue and decreased quality of life persist. This is the first study to measure aerobic exercise capacity in patients in remission of CS for more than 4 years in comparison with matched controls, and to investigate whether the reduction in exercise capacity is related to alterations in muscle tissue. METHODS: Seventeen patients were included. A control individual, matched for sex, estrogen status, age, body mass index, smoking, ethnicity, and physical activity level was recruited for each patient. Maximal aerobic capacity (VO2peak) was assessed during incremental bicycle exercise to exhaustion. In 8 individually matched patients and controls, a percutaneous muscle biopsy was obtained and measures were made of cross-sectional areas, capillarization, and oxphos complex IV (COXIV) protein content as an indicator of mitochondrial content. Furthermore, protein content of endothelial nitric oxide synthase (eNOS) and eNOS phosphorylated on serine1177 and of the NAD(P)H-oxidase subunits NOX2, p47phox, and p67phox were measured in the microvascular endothelial layer. FINDINGS: Patients showed a lower mean VO2peak (SD) (28.0 [7.0] vs 34.8 [7.9] ml O2/kg bw/min, P < .01), maximal workload (SD) (176 [49] vs 212 [67] watt, P = .01), and oxygen pulse (SD) (12.0 [3.7] vs 14.8 [4.2] ml/beat, P < .01) at VO2peak. No differences were seen in muscle fiber type-specific cross-sectional area, capillarization measures, mitochondrial content, and protein content of eNOS, eNOS-P-ser1177, NOX2, p47phox, and p67phox. INTERPRETATION: Because differences in muscle fiber and microvascular outcome measures are not statistically significant, we hypothesize that cardiac dysfunction, seen in active CS, persists during remission and limits blood supply to muscles.

Visualization and quantitation of GLUT4 translocation in human skeletal muscle following glucose ingestion and exercise.

Insulin- and contraction-stimulated increases in glucose uptake into skeletal muscle occur in part as a result of the translocation of glucose transporter 4 (GLUT4) from intracellular stores to the plasma membrane (PM). This study aimed to use immunofluorescence microscopy in human skeletal muscle to quantify GLUT4 redistribution from intracellular stores to the PM in response to glucose feeding and exercise. Percutaneous muscle biopsy samples were taken from the m. vastus lateralis of ten insulin-sensitive men in the basal state and following 30 min of cycling exercise (65% VO2 max). Muscle biopsy samples were also taken from a second cohort of ten age-, BMI- and VO2 max-matched insulin-sensitive men in the basal state and 30 and 60 min following glucose feeding (75 g glucose). GLUT4 and dystrophin colocalization, measured using the Pearson's correlation coefficient, was increased following 30 min of cycling exercise (baseline r = 0.47 ± 0.01; post exercise r = 0.58 ± 0.02; P < 0.001) and 30 min after glucose ingestion (baseline r = 0.42 ± 0.02; 30 min r = 0.46 ± 0.02; P < 0.05). Large and small GLUT4 clusters were partially depleted following 30 min cycling exercise, but not 30 min after glucose feeding. This study has, for the first time, used immunofluorescence microscopy in human skeletal muscle to quantify increases in GLUT4 and dystrophin colocalization and depletion of GLUT4 from large and smaller clusters as evidence of net GLUT4 translocation to the PM.

Improvement in cardiac energetics by perhexiline in heart failure due to dilated cardiomyopathy.

OBJECTIVES: The aim of this study was to determine whether short-term treatment with perhexiline improves cardiac energetics, left ventricular function, and symptoms of heart failure by altering cardiac substrate utilization. BACKGROUND: Perhexiline improves exercise capacity and left ventricular ejection fraction (LVEF) in patients with heart failure (HF). (31)P cardiac magnetic resonance spectroscopy can be used to quantify the myocardial phosphocreatine/adenosine triphosphate ratio. Because improvement of HF syndrome can improve cardiac energetics secondarily, we investigated the effects of short-term perhexiline therapy. METHODS: Patients with systolic HF of nonischemic etiology (n = 50, 62 ± 1.8 years of age, New York Heart Association functional class II to IV, LVEF: 27.0 ± 1.44%) were randomized to receive perhexiline 200 mg or placebo for 1 month in a double-blind fashion. Clinical assessment, echocardiography, and (31)P cardiac magnetic resonance spectroscopy were performed at baseline and after 1 month. A substudy of 22 patients also underwent cross-heart blood sampling at completion of the study to quantify metabolite utilization. RESULTS: Perhexiline therapy was associated with a 30% increase in the phosphocreatine/adenosine triphosphate ratio (from 1.16 ± 0.39 to 1.51 ± 0.51; p < 0.001) versus a 3% decrease with placebo (from 1.36 ± 0.31 to 1.34 ± 0.31; p = 0.37). Perhexiline therapy also led to an improvement in New York Heart Association functional class compared with placebo (p = 0.036). Short-term perhexiline therapy did not change LVEF. Cross-heart measures of cardiac substrate uptake and respiratory exchange ratio (which reflects the ratio of substrates used) did not differ between patients who received perhexiline versus placebo. CONCLUSIONS: Perhexiline improves cardiac energetics and symptom status with no evidence of altered cardiac substrate utilization. No change in LVEF is seen at this early stage. (Metabolic Manipulation in Chronic Heart Failure; NCT00841139).

Quantitative immunofluorescence microscopy of subcellular GLUT4 distribution in human skeletal muscle: effects of endurance and sprint interval training.

Increases in insulin-mediated glucose uptake following endurance training (ET) and sprint interval training (SIT) have in part been attributed to concomitant increases in glucose transporter 4 (GLUT4) protein content in skeletal muscle. This study used an immunofluorescence microscopy method to investigate changes in subcellular GLUT4 distribution and content following ET and SIT. Percutaneous muscle biopsy samples were taken from the m. vastus lateralis of 16 sedentary males in the overnight fasted state before and after 6 weeks of ET and SIT. An antibody was fully validated and used to show large (> 1 µm) and smaller (<1 µm) GLUT4-containing clusters. The large clusters likely represent trans-Golgi network stores and the smaller clusters endosomal stores and GLUT4 storage vesicles (GSVs). Density of GLUT4 clusters was higher at the fibre periphery especially in perinuclear regions. A less dense punctate distribution was seen in the rest of the muscle fibre. Total GLUT4 fluorescence intensity increased in type I and type II fibres following both ET and SIT. Large GLUT4 clusters increased in number and size in both type I and type II fibres, while the smaller clusters increased in size. The greatest increases in GLUT4 fluorescence intensity occurred within the 1 µm layer immediately adjacent to the PM. The increase in peripheral localisation and protein content of GLUT4 following ET and SIT is likely to contribute to the improvements in glucose homeostasis observed after both training modes.

Sprint interval and endurance training are equally effective in increasing muscle microvascular density and eNOS content in sedentary males.

Sprint interval training (SIT) has been proposed as a time efficient alternative to endurance training (ET) for increasing skeletal muscle oxidative capacity and improving certain cardiovascular functions. In this study we sought to make the first comparisons of the structural and endothelial enzymatic changes in skeletal muscle microvessels in response to ET and SIT. Sixteen young sedentary males (age 21 ± SEM 0.7 years, BMI 23.8 ± SEM 0.7 kg m(-2)) were randomly assigned to 6 weeks of ET (40-60 min cycling at ∼65% , 5 times per week) or SIT (4-6 Wingate tests, 3 times per week). Muscle biopsies were taken from the m. vastus lateralis before and following 60 min cycling at 65% to measure muscle microvascular endothelial eNOS content, eNOS serine(1177) phosphorylation, NOX2 content and capillarisation using quantitative immunofluorescence microscopy. Whole body insulin sensitivity, arterial stiffness and blood pressure were also assessed. ET and SIT increased skeletal muscle microvascular eNOS content (ET 14%; P < 0.05, SIT 36%; P < 0.05), with a significantly greater increase observed following SIT (P < 0.05). Sixty minutes of moderate intensity exercise increased eNOS ser(1177) phosphorylation in all instances (P < 0.05), but basal and post-exercise eNOS ser(1177) phosphorylation was lower following both training modes. All microscopy measures of skeletal muscle capillarisation (P < 0.05) were increased with SIT or ET, while neither endothelial nor sarcolemmal NOX2 was changed. Both training modes reduced aortic stiffness and increased whole body insulin sensitivity (P < 0.05). In conclusion, in sedentary males SIT and ET are effective in improving muscle microvascular density and eNOS protein content.

Adipophilin distribution and colocalization with lipid droplets in skeletal muscle.

Intramyocellular lipids (IMCL) are stored as discrete lipid droplets which are associated with a number of proteins. The lipid droplet-associated protein adipophilin (the human orthologue of adipose differentiation-related protein) is ubiquitously expressed and is one of the predominant lipid droplet-proteins in skeletal muscle. The aim of this study was to investigate the subcellular distribution of adipophilin in human muscle fibres and to measure the colocalization of adipophilin with IMCL. Muscle biopsies from six lean male cyclists (BMI 23.4 +/- 0.4, aged 31 +/- 2 years, W (max) 346 +/- 8) were stained for myosin heavy chain type 1, IMCL, adipophilin and mitochondria using immunofluorescence and viewed with widefield and confocal fluorescence microscopy. The present study shows that like IMCL, the adipophilin content is ~twofold greater in type I skeletal muscle fibres and is situated in the areas between the mitochondrial network. Colocalization analysis demonstrated that 61 +/- 2% of IMCL contain adipophilin. Although the majority of adipophilin is contained within IMCL, 36 +/- 4% of adipophilin is not associated with IMCL. In conclusion, this study indicates that the IMCL pool is heterogeneous, as the majority but not all IMCL contain adipophilin.

Substrate source use in older, trained males after decades of endurance training.

PURPOSE: The purpose of this study was to compare substrate source use in older, long-term exercising, endurance-trained males with sedentary controls. METHODS: [U-C]palmitate and [6,6-H2]glucose tracers were applied to assess plasma free fatty acid (FFA) and glucose oxidation rates, and to estimate muscle- and/or lipoprotein-derived triacylglycerol (TG) and muscle glycogen use. Subjects were 10 long-term exercising, endurance-trained males and 10 sedentary controls (age 57 +/- 1 and 60 +/- 2 yr, respectively). Muscle biopsy samples were collected before and after exercise to assess muscle fiber type-specific intramyocellular lipid and glycogen content. RESULTS: During exercise, plasma palmitate Ra, Rd, and Rox were significantly greater in the trained subjects compared with the controls (Ra: 0.36 +/- 0.02 and 0.25 +/- 0.02; Rd: 0.36 +/- 0.03 and 0.24 +/- 0.02; Rox: 0.31 +/- 0.02 and 0.20 +/- 0.02 mmol.min, respectively, P < 0.01). This resulted in greater plasma FFA and total fat oxidation rates in the trained versus sedentary subjects (P < 0.001). Muscle- and/or lipoprotein-derived TG use contributed 10 +/- 2 and 11 +/- 3% in the trained and control groups, respectively (NS). No significant net changes in muscle fiber lipid content were observed. CONCLUSIONS: Older, endurance-trained males oxidize more fat during moderate-intensity exercise than do sedentary controls. This greater total fat oxidation rate is attributed to a higher plasma FFA release, uptake, and oxidation rate. In contrast, intramyocellular triacylglycerol does not seem to represent a major substrate source during 1 h of moderate-intensity exercise in older trained or sedentary men.

Creatine supplementation increases glycogen storage but not GLUT-4 expression in human skeletal muscle.

It has been speculated that creatine supplementation affects muscle glucose metabolism in humans by increasing muscle glycogen storage and up-regulating GLUT-4 protein expression. In the present study, we assessed the effects of creatine loading and prolonged supplementation on muscle glycogen storage and GLUT-4 mRNA and protein content in humans. A total of 20 subjects participated in a 6-week supplementation period during which creatine or a placebo was ingested. Muscle biopsies were taken before and after 5 days of creatine loading (20 g.day(-1)) and after 6 weeks of continued supplementation (2 g.day(-1)). Fasting plasma insulin concentrations, muscle creatine, glycogen and GLUT-4 protein content as well as GLUT-4, glycogen synthase-1 (GS-1) and glycogenin-1 (Gln-1) mRNA expression were determined. Creatine loading significantly increased total creatine, free creatine and creatine phosphate content with a concomitant 18 +/- 5% increase in muscle glycogen content (P<0.05). The subsequent use of a 2 g.day(-1) maintenance dose for 37 days did not maintain total creatine, creatine phosphate and glycogen content at the elevated levels. The initial increase in muscle glycogen accumulation could not be explained by an increase in fasting plasma insulin concentration, muscle GLUT-4 mRNA and/or protein content. In addition, neither muscle GS-1 nor Gln-1 mRNA expression was affected. We conclude that creatine ingestion itself stimulates muscle glycogen storage, but does not affect muscle GLUT-4 expression.

Coronary sinus catheter placement: assessment of placement criteria and cardiac complications.

STUDY OBJECTIVES: To evaluate the placement and complications of a coronary sinus (CS) catheter in human subjects. DESIGN: Sixty-two CS catheters inserted in patients scheduled for coronary artery bypass graft surgery (CABG). SETTING: University hospital, anesthesia and cardiothoracic surgery departments. PATIENTS: Sixty-two patients without valvular or concomitant diseases undergoing CABG. INTERVENTIONS: CS fluoroscopy, measurements of CS flow, CS oxygen saturation, and CS distal tip pressure before incision, after incision, 20 min after aortic cross-clamp release (X-off), 50 min after X-off, 2 h after X-off, 4 h after X-off, and 6 h after X-off. RESULTS: In 57 patients (92%), we achieved successful CS catheter placement. In five patients (8%), CS catheter positioning was not possible. Of the 57 CS catheters placed, dislocation occurred during the operation in six patients (11%) and postoperatively in three patients (6%). Cardiac complications of CS catheter placement occurred in nine patients (15%). Four patients (6%) acquired hemopericardium. Three of these patients had a small hematoma in the right ventricle. In two other patients, contrast medium appeared in the right ventricular wall during catheterization. No hemodynamic signs of these complications were detected clinically. Irregular heart rhythm was observed in only three patients. CS blood oxygen saturation ranged from 40 to 60%. CS flow amounted to 3% of cardiac output. Variations in CS flow paralleled changes in cardiac output. CONCLUSIONS: A CS catheter is a useful tool for clinical human cardiac research; however, the placement of a CS catheter can cause minor myocardial damage in > 10% of patients. Importantly, this damage may not be clinically evident, but only observed after thoracotomy. CS oxygen saturation, CS flow, distal tip pressure, and fluoroscopy are reliable tools to assess a safe and correct positioning of the CS catheter.

Intramyocellular lipids form an important substrate source during moderate intensity exercise in endurance-trained males in a fasted state.

Both stable isotope methodology and fluorescence microscopy were applied to define the use of intramuscular triglyceride (IMTG) stores as a substrate source during exercise on a whole-body as well as on a fibre type-specific intramyocellular level in trained male cyclists. Following an overnight fast, eight subjects were studied at rest, during 120 min of moderate intensity exercise (60 % maximal oxygen uptake capacity (VO2,max)) and 120 min of post-exercise recovery. Continuous infusions of [U-13C]palmitate and [6,6-2H2]glucose were administered at rest and during subsequent exercise to quantify whole-body plasma free fatty acid (FFA) and glucose oxidation rates and the contribution of other fat sources (sum of muscle- plus lipoprotein-derived TG) and muscle glycogen to total energy expenditure. Fibre type-specific intramyocellular lipid content was determined in muscle biopsy samples collected before, immediately after and 2 h after exercise. At rest, fat oxidation provided 66 +/- 5 % of total energy expenditure, with FFA and other fat sources contributing 48 +/- 6 and 17 +/- 3 %, respectively. FFA oxidation rates increased during exercise, and correlated well with the change in plasma FFA concentrations. Both the use of other fat sources and muscle glycogen declined with the duration of exercise, whereas plasma glucose production and utilisation increased (P < 0.001). On average, FFA, other fat sources, plasma glucose and muscle glycogen contributed 28 +/- 3, 15 +/- 2, 12 +/- 1 and 45 +/- 4 % to total energy expenditure during exercise, respectively. Fluorescence microscopy revealed a 62 +/- 7 % net decline in muscle lipid content following exercise in the type I fibres only, with no subsequent change during recovery. We conclude that IMTG stores form an important substrate source during moderate intensity exercise in endurance-trained male athletes following an overnight fast, with the oxidation rate of muscle- plus lipoprotein-derived TG being decreased with the duration of exercise.

Effects of creatine loading and prolonged creatine supplementation on body composition, fuel selection, sprint and endurance performance in humans.

Most research on creatine has focused on short-term creatine loading and its effect on high-intensity performance capacity. Some studies have investigated the effect of prolonged creatine use during strength training. However, studies on the effects of prolonged creatine supplementation are lacking. In the present study, we have assessed the effects of both creatine loading and prolonged supplementation on muscle creatine content, body composition, muscle and whole-body oxidative capacity, substrate utilization during submaximal exercise, and on repeated supramaximal sprint, as well as endurance-type time-trial performance on a cycle ergometer. Twenty subjects ingested creatine or a placebo during a 5-day loading period (20 g.day(-1)) after which supplementation was continued for up to 6 weeks (2 g.day(-1)). Creatine loading increased muscle free creatine, creatine phosphate (CrP) and total creatine content ( P <0.05). The subsequent use of a 2 g.day(-1) maintenance dose, as suggested by an American College of Sports Medicine Roundtable, resulted in a decline in both the elevated CrP and total creatine content and maintenance of the free creatine concentration. Both short- and long-term creatine supplementation improved performance during repeated supramaximal sprints on a cycle ergometer. However, whole-body and muscle oxidative capacity, substrate utilization and time-trial performance were not affected. The increase in body mass following creatine loading was maintained after 6 weeks of continued supplementation and accounted for by a corresponding increase in fat-free mass. This study provides definite evidence that prolonged creatine supplementation in humans does not increase muscle or whole-body oxidative capacity and, as such, does not influence substrate utilization or performance during endurance cycling exercise. In addition, our findings suggest that prolonged creatine ingestion induces an increase in fat-free mass.

Impaired performance of skeletal muscle in alpha-glucosidase knockout mice.

Glycogen storage disease type II (GSD II) is an inherited progressive muscle disease in which lack of functional acid alpha-glucosidase (AGLU) results in lysosomal accumulation of glycogen. We report on the impact of a null mutation of the acid alpha-glucosidase gene (AGLU(-/-)) in mice on the force production capabilities, contractile mass, oxidative capacity, energy status, morphology, and desmin content of skeletal muscle. Muscle function was assessed in halothane-anesthetized animals, using a recently designed murine isometric dynamometer. Maximal torque production during single tetanic contraction was 50% lower in the knockout mice than in wild type. Loss of developed torque was found to be disproportionate to the 20% loss in muscle mass. During a series of supramaximal contraction, fatigue, expressed as percentile decline of developed torque, did not differ between AGLU(-/-) mice and age-matched controls. Muscle oxidative capacity, energy status, and protein content (normalized to either dry or wet weight) were not changed in knockout mice compared to control. Alterations in muscle cell morphology were clearly visible. Desmin content was increased, whereas alpha-actinin was not. As the decline in muscle mass is insufficient to explain the degree in decline of mechanical performance, we hypothesize that the large clusters of noncontractile material present in the cytoplasm hamper longitudinal force transmission, and hence muscle contractile function. The increase in muscular desmin content is most likely reflecting adaptations to altered intracellular force transmission.