Apo AI Nanoparticles Delivered Post Myocardial Infarction Moderate Inflammation.

RATIONALE: Decades of research have examined immune-modulatory strategies to protect the heart after an acute myocardial infarction and prevent progression to heart failure but have failed to translate to clinical benefit. OBJECTIVE: To determine anti-inflammatory actions of n-apo AI (Apo AI nanoparticles) that contribute to cardiac tissue recovery after myocardial infarction. METHODS AND RESULTS: Using a preclinical mouse model of myocardial infarction, we demonstrate that a single intravenous bolus of n-apo AI (CSL111, 80 mg/kg) delivered immediately after reperfusion reduced the systemic and cardiac inflammatory response. N-apo AI treatment lowered the number of circulating leukocytes by 30±7% and their recruitment into the ischemic heart by 25±10% (all P<5.0×10-2). This was associated with a reduction in plasma levels of the clinical biomarker of cardiac injury, cardiac troponin-I, by 52±17% (P=1.01×10-2). N-apo AI reduced the cardiac expression of chemokines that attract neutrophils and monocytes by 60% to 80% and lowered surface expression of integrin CD11b on monocytes by 20±5% (all P<5.0×10-2). Fluorescently labeled n-apo AI entered the infarct and peri-infarct regions and colocalized with cardiomyocytes undergoing apoptosis and with leukocytes. We further demonstrate that n-apo AI binds to neutrophils and monocytes, with preferential binding to the proinflammatory monocyte subtype and partially via SR-BI (scavenger receptor BI). In patients with type 2 diabetes, we also observed that intravenous infusion of the same n-apo AI (CSL111, 80 mg/kg) similarly reduced the level of circulating leukocytes by 12±5% (all P<5.0×10-2). CONCLUSIONS: A single intravenous bolus of n-apo AI delivered immediately post-myocardial infarction reduced the systemic and cardiac inflammatory response through direct actions on both the ischemic myocardium and leukocytes. These data highlight the anti-inflammatory effects of n-apo AI and provide preclinical support for investigation of its use for management of acute coronary syndromes in the setting of primary percutaneous coronary interventions.

Acute metabolic and cardiovascular effects of mirabegron in healthy individuals.

AIMS: To quantify acute energy expenditure, supraclavicular skin temperature and cardiovascular responses to four doses of the ß3-adrenoceptor agonist, mirabegron. MATERIALS AND METHODS: A total of 17 individuals (11 men, six women) participated in this ascending-dose study, receiving single 50-, 100-, 150- and 200-mg doses of mirabegron on four separate days with 3 to 14 days wash-out between each dose. All variables were measured each visit from baseline to 180 minutes post mirabegron treatment. To determine brown adipose tissue (BAT) thermogenic efficacy at each dose, energy expenditure and supraclavicular skin temperature were compared from baseline to 180 minutes post mirabegron treatment. To examine safety, changes in cardiovascular variables at 100, 150 and 200 mg were compared with the standard clinical dose of 50 mg. RESULTS: Energy expenditure significantly increased after the 100- (35.6 ± 5.4 kJ/h) and 200-mg (35.6 ± 13.1 kJ/h) doses (P ≤ 0.05), and trended towards an increase after 150 mg (24.1 ± 13.6 kJ/h). Supraclavicular skin temperature increased after 50- (0.22 ± 0.1°C), 100- (0.30 ± 0.1°C) and 150-mg mirabegron doses (0.29 ± 0.1°C; P ≤ 0.05). The change in systolic blood pressure was greater after 150- (7.1 ± 1.3 mm Hg) and 200-mg doses (9.3 ± 1.9 mm Hg) than after the 50-mg dose (2.2 ± 1.3 mm Hg; P ≤ 0.05). The change in heart rate was greater after 200 mg (9.0 ± 2.2 bpm) compared with 50 mg (2.9 ± 1.4 bpm; P ≤ 0.05). CONCLUSIONS: A 100-mg dose of mirabegron increases energy expenditure and supraclavicular skin temperature in a ß3-adrenoceptor-specific manner, without the off-target elevations in blood pressure or heart rate observed at higher doses.

Pioglitazone reduces cold-induced brown fat glucose uptake despite induction of browning in cultured human adipocytes: a randomised, controlled trial in humans.

AIMS/HYPOTHESIS: Increasing brown adipose tissue (BAT) activity is a possible therapeutic strategy to increase energy expenditure and glucose and lipid clearance to ameliorate obesity and associated comorbidities. The thiazolidinedione (TZD) class of glucose-lowering drugs increase BAT browning in preclinical experimental models but whether these actions extend to humans in vivo is unknown. The aim of this study was to determine the effect of pioglitazone treatment on adipocyte browning and adaptive thermogenesis in humans. METHODS: We first examined whether pioglitazone treatment of cultured human primary subacromioclavicular-derived adipocytes induced browning. Then, in a blinded, placebo-controlled, parallel trial, conducted within the Baker Institute clinical research laboratories, 14 lean male participants who were free of cardiometabolic disease were randomised to receive either placebo (lactose; n = 7, age 22 ± 1 years) or pioglitazone (45 mg/day, n = 7, age 21 ± 1 years) for 28 days. Participants were allocated to treatments by Alfred Hospital staff independent from the study via electronic generation of a random number sequence. Researchers conducting trials and analysing data were blind to treatment allocation. The change in cold-stimulated BAT activity, assessed before and after the intervention by [18F]fluorodeoxyglucose uptake via positron emission tomography/computed tomography in upper thoracic and cervical adipose tissue, was the primary outcome measure. Energy expenditure, cardiovascular responses, core temperature, blood metabolites and hormones were measured in response to acute cold exposure along with body composition before and after the intervention. RESULTS: Pioglitazone significantly increased in vitro browning and adipogenesis of adipocytes. In the clinical trial, cold-induced BAT maximum standardised uptake value was significantly reduced after pioglitazone compared with placebo (-57 ± 6% vs -12 ± 18%, respectively; p < 0.05). BAT total glucose uptake followed a similar but non-significant trend (-50 ± 10% vs -6 ± 24%, respectively; p = 0.097). Pioglitazone increased total and lean body mass compared with placebo (p < 0.05). No other changes between groups were detected. CONCLUSIONS/INTERPRETATION: The disparity in the actions of pioglitazone on BAT between preclinical experimental models and our in vivo human trial highlight the imperative to conduct human proof-of-concept studies as early as possible in BAT research programmes aimed at therapeutic development. Our clinical trial findings suggest that reduced BAT activity may contribute to weight gain associated with pioglitazone and other TZDs. TRIAL REGISTRATION: ClinicalTrials.gov NCT02236962 FUNDING: This work was supported by the Diabetes Australia Research Program and OIS scheme from the Victorian State Government.

Correction to: Pioglitazone reduces cold-induced brown fat glucose uptake despite induction of browning in cultured human adipocytes: a randomised, controlled trial in humans.

The baseline insulin data given in Table 1 for the placebo group were incorrectly reported as 51 ± 10 pmol/l instead of 48 ± 10 pmol/l. This mistake also impacts on data reported in Table 4.

Chronic ephedrine administration decreases brown adipose tissue activity in a randomised controlled human trial: implications for obesity.

AIMS/HYPOTHESIS: Brown adipose tissue (BAT) activation increases energy expenditure and may have therapeutic potential to combat obesity. The primary activating and adaptive signal for BAT is via ß-adrenergic signalling. We previously demonstrated that human BAT is acutely responsive to oral administration of the sympathomimetic, ephedrine. Here we aimed to determine whether adaptive thermogenesis can be induced via chronic treatment with ephedrine. METHODS: Twenty-three healthy young men, recruited from the general public in Melbourne, Australia, who were non-smokers, physically inactive and non-medicated with no prior history of cardiovascular disease or diabetes were recruited for this study. They were assigned to receive either 1.5 mg kg(-1) day(-1) ephedrine ('active' group; n = 12, age 23 ± 1 years, BMI 24 ± 1 kg/m(2)) or placebo (n = 11; 22 ± 2 years, 23 ± 2 kg/m(2)) for 28 days in a randomised (computer-generated random order sequence), placebo-controlled, parallel-group trial. Participants and all investigators were blinded to treatments. Body composition was measured before and after the intervention by dual energy X-ray absorptiometry. BAT activity, measured via (18)F-fluorodeoxyglucose positron emission tomography-computed tomography, in response to a single dose of 2.5 mg/kg ephedrine, was the primary outcome measure to be determined before and after the 28 day treatment period. RESULTS: Twenty-eight individuals were randomised and consented to the study. Twenty-three completed the trial and only these participants were included in the final analyses. After 28 days of treatment, the active group lost a significant amount of total body fat (placebo 1.1 ± 0.3 kg, ephedrine -0.9 ± 0.5 kg; p < 0.01) and visceral fat (placebo 6.4 ± 19.1 g, ephedrine -134 ± 43 g; p < 0.01), with no change in lean mass or bone mineral content compared with the placebo group. In response to acute ephedrine, BAT activity (change in mean standardised uptake value: placebo -3 ± 7%, ephedrine -22 ± 6%) and the increase in systolic blood pressure were significantly reduced (p < 0.05) in the active group compared with placebo. CONCLUSIONS/INTERPRETATION: Chronic ephedrine treatment reduced body fat content, but this was not associated with an increase in BAT activity. Rather, chronic ephedrine suppressed BAT glucose disposal, suggesting that chronic ephedrine treatment decreased, rather than increased, BAT activity. TRIAL REGISTRATION: ClinicalTrials.gov NCT02236962 FUNDING: This study was funded by the National Health and Medical Research Council of Australia Program Grant (1036352) and the OIS scheme from the Victorian State Government.

Effects of high-density lipoprotein elevation with cholesteryl ester transfer protein inhibition on insulin secretion.

RATIONALE: High-density lipoprotein cholesterol elevation via cholesteryl ester transfer protein (CETP) inhibition represents a novel therapy for atherosclerosis, which also may have relevance for type 2 diabetes mellitus. OBJECTIVE: The current study assessed the effects of a CETP inhibitor on postprandial insulin, ex vivo insulin secretion, and cholesterol efflux from pancreatic ß-cells. METHODS AND RESULTS: Healthy participants received a daily dose of CETP inhibitor (n=10) or placebo (n=15) for 14 days in a randomized double-blind study. Insulin secretion and cholesterol efflux from MIN6N8 ß-cells were determined after incubation with treated plasma. CETP inhibition increased plasma high-density lipoprotein cholesterol, apolipoprotein AI, and postprandial insulin. MIN6N8 ß-cells incubated with plasma from CETP inhibitor-treated individuals (compared with placebo) exhibited an increase in both glucose-stimulated insulin secretion and cholesterol efflux over the 14-day treatment period. CONCLUSIONS: CETP inhibition increased postprandial insulin and promoted ex vivo ß-cell glucose-stimulated insulin secretion, potentially via enhanced ß-cell cholesterol efflux.

Can exercise training rescue the adverse cardiometabolic effects of low birth weight and prematurity?

Being born preterm and/or small for gestational age are well-established risk factors for cardiometabolic disease in adulthood. Physical activity has the potential to mitigate against the detrimental cardiometabolic effects of low birth weight from two perspectives: (i) maternal exercise prior to and during pregnancy; and (ii) exercise during childhood or adulthood for those born small or prematurely. Evidence from epidemiological birth cohort studies suggests that the effects of moderate-intensity physical activity during pregnancy on mean birth weight are small, but reduce the risk of either high or low birth weight infants. In contrast, vigorous and/or high-intensity exercise during pregnancy has been associated with reduced birth weight. In childhood and adolescence, exercise ability is compromised in extremely low birth weight individuals (< 1000 g), but only marginally reduced in those of very low to low birth weight (1000-2500 g). Epidemiological studies show that the association between birth weight and metabolic disease is lost in physically fit individuals and, consistently, that the association between low birth weight and metabolic syndrome is accentuated in unfit individuals. Physical activity intervention studies indicate that most cardiometabolic risk factors respond to exercise in a protective manner, independent of birth weight. The mechanisms by which exercise may protect low birth weight individuals include restoration of muscle mass, reduced adiposity and enhanced ß-cell mass and function, as well as effects on both aerobic and anaerobic muscle metabolism, including substrate utilization and mitochondrial function. Vascular and cardiac adaptations are also likely important, but are less well studied.

Tumor necrosis factor alpha-induced skeletal muscle insulin resistance involves suppression of AMP-kinase signaling.

Elevated levels of tumor necrosis factor (TNFalpha) are implicated in the development of insulin resistance, but the mechanisms mediating these chronic effects are not completely understood. We demonstrate that TNFalpha signaling through TNF receptor (TNFR) 1 suppresses AMPK activity via transcriptional upregulation of protein phosphatase 2C (PP2C). This in turn reduces ACC phosphorylation, suppressing fatty-acid oxidation, increasing intramuscular diacylglycerol accumulation, and causing insulin resistance in skeletal muscle, effects observed both in vitro and in vivo. Importantly even at pathologically elevated levels of TNFalpha observed in obesity, the suppressive effects of TNFalpha on AMPK signaling are reversed in mice null for both TNFR1 and 2 or following treatment with a TNFalpha neutralizing antibody. Our data demonstrate that AMPK is an important TNFalpha signaling target and is a contributing factor to the suppression of fatty-acid oxidation and the development of lipid-induced insulin resistance in obesity.

Reconstituted high-density lipoprotein infusion modulates fatty acid metabolism in patients with type 2 diabetes mellitus.

We recently demonstrated that reconstituted high-density lipoprotein (rHDL) modulates glucose metabolism in humans via both AMP-activated protein kinase (AMPK) in muscle and by increasing plasma insulin. Given the key roles of both AMPK and insulin in fatty acid metabolism, the current study investigated the effect of rHDL infusion on fatty acid oxidation and lipolysis. Thirteen patients with type 2 diabetes received separate infusions of rHDL and placebo in a randomized, cross-over study. Fatty acid metabolism was assessed using steady-state tracer methodology, and plasma lipids were measured by mass spectrometry (lipidomics). In vitro studies were undertaken in 3T3-L1 adipocytes. rHDL infusion inhibited fasting-induced lipolysis (P = 0.03), fatty acid oxidation (P < 0.01), and circulating glycerol (P = 0.04). In vitro, HDL inhibited adipocyte lipolysis in part via activation of AMPK, providing a possible mechanistic link for the apparent reductions in lipolysis observed in vivo. In contrast, circulating NEFA increased after rHDL infusion (P < 0.01). Lipidomic analyses implicated phospholipase hydrolysis of rHDL-associated phosphatidylcholine as the cause, rather than lipolysis of endogenous fat stores. rHDL infusion inhibits fasting-induced lipolysis and oxidation in patients with type 2 diabetes, potentially through both AMPK activation in adipose tissue and elevation of plasma insulin. The phospholipid component of rHDL also has the potentially undesirable effect of increasing circulating NEFA.

Acute signalling responses to intense endurance training commenced with low or normal muscle glycogen.

We have previously demonstrated that well-trained subjects who completed a 3 week training programme in which selected high-intensity interval training (HIT) sessions were commenced with low muscle glycogen content increased the maximal activities of several oxidative enzymes that promote endurance adaptations to a greater extent than subjects who began all training sessions with normal glycogen levels. The aim of the present study was to investigate acute skeletal muscle signalling responses to a single bout of HIT commenced with low or normal muscle glycogen stores in an attempt to elucidate potential mechanism(s) that might underlie our previous observations. Six endurance-trained cyclists/triathletes performed a 100 min ride at approximately 70% peak O(2) uptake (AT) on day 1 and HIT (8 x 5 min work bouts at maximal self-selected effort with 1 min rest) 24 h later (HIGH). Another six subjects, matched for fitness and training history, performed AT on day 1 then 1-2 h later, HIT (LOW). Muscle biopsies were taken before and after HIT. Muscle glycogen concentration was higher in HIGH versus LOW before the HIT (390 +/- 28 versus 256 +/- 67 micromol (g dry wt)(1)). After HIT, glycogen levels were reduced in both groups (P < 0.05) but HIGH was elevated compared with LOW (229 +/- 29 versus 124 +/- 41 micromol (g dry wt)(1); P < 0.05). Phosphorylation of 5 AMP-activated protein kinase (AMPK) increased after HIT, but the magnitude of increase was greater in LOW (P < 0.05). Despite the augmented AMPK response in LOW after HIT, selected downstream AMPK substrates were similar between groups. Phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK) was unchanged for both groups before and after the HIT training sessions. We conclude that despite a greater activation AMPK phosphorylation when HIT was commenced with low compared with normal muscle glycogen availability, the localization and phosphorylation state of selected downstream targets of AMPK were similar in response to the two interventions.

Short-term endurance training does not alter the oxidative capacity of human subcutaneous adipose tissue.

Endurance training results in adaptations that enhance regulation of energy storage and expenditure at rest and during exercise. While processes involved in skeletal muscle oxidative remodelling are well described, it is unknown whether oxidative capacity of human subcutaneous white adipose tissue (WAT) is modified by endurance training. Since human WAT retains rudimentary characteristics required for upregulation of oxidative function, we hypothesised that 10 days of intense endurance training would promote changes in WAT that favour an increase in oxidative capacity. Eleven untrained males (age 22 +/- 1 years, body mass 81 +/- 5 kg, peak oxygen uptake (VO(2peak)) 3.7 +/- 0.2 l/min) undertook a 10-day endurance training protocol. Subcutaneous adipose tissue biopsies were taken from the abdomen prior to and 1 day after completion of training and analysed for fatty acid oxidative capacity, citrate synthase activity, and mitochondrial content via electron microscopy and gene expression analyses. There was a reduction in whole-body rates of carbohydrate oxidation, and concomitant increases in fat oxidation rate measured during 20-min of submaximal cycling (70% of pre-training VO(2peak)) and an increase in basal GLUT4 protein in skeletal muscle. Despite these training-induced adaptations, there were no changes in WAT of ex-vivo fat oxidation rate, maximal citrate synthase activity, mitochondrial volume or in selected genes involved in adipose tissue oxidative capacity. We conclude that 10 days training in previously untrained subjects results in adaptations in skeletal muscle but does not increase the oxidative capacity of WAT.

Matrix-Induced Autologous Chondrocyte Implantation (MACI) Grafting for Osteochondral Lesions of the Talus.

BACKGROUND: Matrix-induced autologous chondrocyte implantation (MACI) is an established treatment method for larger joints and has shown promising results in the ankle as well. We present a series of patients after ankle MACI with long-term follow-up of clinical and radiological outcomes. METHODS: We present the follow-up of 15 patients who underwent MACI grafting from August 2003 to February 2006. The mean follow-up was 12.9 years. Clinical evaluations were conducted using the American Orthopaedic Foot & Ankle Society (AOFAS), Foot and Ankle Activity Measurement (FAAM), and visual analog scale (VAS) scoring systems and the magnetic resonance observation of cartilage repair tissue (MOCART) scoring system for radiological evaluation. RESULTS: The mean size of the talar osteochondral defects was 204 mm2. We found a significant improvement in mean AOFAS score from 60 preoperatively to a mean of 84 at 12 years postoperatively. The 12-year FAAM score for Activities of Daily Living was 89% (range, 62%-99%). The mean 12-year MOCART score was 65 points (range, 30-100 points) with significant agreement between assessors (P < .001). However, the MOCART scores did not correlate with the FAAM scores (P = .86). CONCLUSION: Considering our long-term follow-up, we believe MACI is a reliable treatment method for talar osteochondral defects providing lasting pain relief and satisfying clinical results. However, with an equivalent outcome, but at higher costs, and the requirement for 2 operative procedures, the results do not seem to be superior to other established methods. The clinical utility of the MOCART score requires further scrutiny since we were not able to show any correlation between the score and clinical outcome. LEVEL OF EVIDENCE: Level IV, case series.

Plasma Docosahexaenoic Acid and Eicosapentaenoic Acid Concentrations Are Positively Associated with Brown Adipose Tissue Activity in Humans.

Brown adipose tissue (BAT) activation is a possible therapeutic strategy to increase energy expenditure and improve metabolic homeostasis in obesity. Recent studies have revealed novel interactions between BAT and circulating lipid species-in particular, the non-esterified fatty acid (NEFA) and oxylipin lipid classes. This study aimed to identify individual lipid species that may be associated with cold-stimulated BAT activity in humans. A panel of 44 NEFA and 41 oxylipin species were measured using mass-spectrometry-based lipidomics in the plasma of fourteen healthy male participants before and after 90 min of mild cold exposure. Lipid measures were correlated with BAT activity measured via 18F-fluorodeoxyglucose ([18F]FDG) positron emission tomography/computed tomography (PET/CT), along with norepinephrine (NE) concentration (a surrogate marker of sympathetic activity). The study identified a significant increase in total NEFA concentration following cold exposure that was positively associated with NE concentration change. Individually, 33 NEFA and 11 oxylipin species increased significantly in response to cold exposure. The concentration of the omega-3 NEFA, docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) at baseline was significantly associated with BAT activity, and the cold-induced change in 18 NEFA species was significantly associated with BAT activity. No significant associations were identified between BAT activity and oxylipins.

Acute effects of active breaks during prolonged sitting on subcutaneous adipose tissue gene expression: an ancillary analysis of a randomised controlled trial.

Active breaks in prolonged sitting has beneficial impacts on cardiometabolic risk biomarkers. The molecular mechanisms include regulation of skeletal muscle gene and protein expression controlling metabolic, inflammatory and cell development pathways. An active communication network exists between adipose and muscle tissue, but the effect of active breaks in prolonged sitting on adipose tissue have not been investigated. This study characterized the acute transcriptional events induced in adipose tissue by regular active breaks during prolonged sitting. We studied 8 overweight/obese adults participating in an acute randomized three-intervention crossover trial. Interventions were performed in the postprandial state and included: (i) prolonged uninterrupted sitting; or prolonged sitting interrupted with 2-minute bouts of (ii) light- or (iii) moderate-intensity treadmill walking every 20 minutes. Subcutaneous adipose tissue biopsies were obtained after each condition. Microarrays identified 36 differentially expressed genes between the three conditions (fold change ≥0.5 in either direction; p < 0.05). Pathway analysis indicated that breaking up of prolonged sitting led to differential regulation of adipose tissue metabolic networks and inflammatory pathways, increased insulin signaling, modulation of adipocyte cell cycle, and facilitated cross-talk between adipose tissue and other organs. This study provides preliminary insight into the adipose tissue regulatory systems that may contribute to the physiological effects of interrupting prolonged sitting.

HDL Phospholipids, but Not Cholesterol Distinguish Acute Coronary Syndrome From Stable Coronary Artery Disease.

Background Although acute coronary syndromes (ACS) are a major cause of morbidity and mortality, relationships with biologically active lipid species potentially associated with plaque disruption/erosion in the context of their lipoprotein carriers are indeterminate. The aim was to characterize lipid species within lipoprotein particles which differentiate ACS from stable coronary artery disease. Methods and Results Venous blood was obtained from 130 individuals with de novo presentation of an ACS (n=47) or stable coronary artery disease (n=83) before coronary catheterization. Lipidomic measurements (533 lipid species; liquid chromatography electrospray ionization/tandem mass spectrometry) were performed on whole plasma as well as 2 lipoprotein subfractions: apolipoprotein A1 (apolipoprotein A, high-density lipoprotein) and apolipoprotein B. Compared with stable coronary artery disease, ACS plasma was lower in phospholipids including lyso species and plasmalogens, with the majority of lipid species differing in abundance located within high-density lipoprotein (high-density lipoprotein, 113 lipids; plasma, 73 lipids). Models including plasma lipid species alone improved discrimination between the stable and ACS groups by 0.16 (C-statistic) compared with conventional risk factors. Models utilizing lipid species either in plasma or within lipoprotein fractions had a similar ability to discriminate groups, though the C-statistic was highest for plasma lipid species (0.80; 95% CI, 0.75-0.86). Conclusions Multiple lysophospholipids, but not cholesterol, featured among the lipids which were present at low concentration within high-density lipoprotein of those presenting with ACS. Lipidomics, when applied to either whole plasma or lipoprotein fractions, was superior to conventional risk factors in discriminating ACS from stable coronary artery disease. These associative mechanistic insights elucidate potential new preventive, prognostic, and therapeutic avenues for ACS which require investigation in prospective analyses.

Skeletal muscle adaptation and performance responses to once a day versus twice every second day endurance training regimens.

We determined the effects of a cycle training program in which selected sessions were performed with low muscle glycogen content on training capacity and subsequent endurance performance, whole body substrate oxidation during submaximal exercise, and several mitochondrial enzymes and signaling proteins with putative roles in promoting training adaptation. Seven endurance-trained cyclists/triathletes trained daily (High) alternating between 100-min steady-state aerobic rides (AT) one day, followed by a high-intensity interval training session (HIT; 8 x 5 min at maximum self-selected effort) the next day. Another seven subjects trained twice every second day (Low), first undertaking AT, then 1-2 h later, the HIT. These training schedules were maintained for 3 wk. Forty-eight hours before and after the first and last training sessions, all subjects completed a 60-min steady-state ride (60SS) followed by a 60-min performance trial. Muscle biopsies were taken before and after 60SS, and rates of substrate oxidation were determined throughout this ride. Resting muscle glycogen concentration (412 +/- 51 vs. 577 +/- 34 micromol/g dry wt), rates of whole body fat oxidation during 60SS (1,261 +/- 247 vs. 1,698 +/- 174 micromol.kg(-1).60 min(-1)), the maximal activities of citrate synthase (45 +/- 2 vs. 54 +/- 1 mmol.kg dry wt(-1).min(-1)), and beta-hydroxyacyl-CoA-dehydrogenase (18 +/- 2 vs. 23 +/- 2 mmol.kg dry wt(-1).min(-1)) along with the total protein content of cytochrome c oxidase subunit IV were increased only in Low (all P < 0.05). Mitochondrial DNA content and peroxisome proliferator-activated receptor-gamma coactivator-1alpha protein levels were unchanged in both groups after training. Cycling performance improved by approximately 10% in both Low and High. We conclude that compared with training daily, training twice every second day compromised high-intensity training capacity. While selected markers of training adaptation were enhanced with twice a day training, the performance of a 1-h time trial undertaken after a 60-min steady-state ride was similar after once daily or twice every second day training programs.

Oxidative stress-induced insulin resistance in skeletal muscle cells is ameliorated by gamma-tocopherol treatment.

BACKGROUND: Oxidative stress-induced reactive oxygen species are associated with the clinical manifestation of insulin resistance. Evidence suggests that antioxidant treatment may reduce this incidence. AIM OF THE STUDY: This study determined whether glucose oxidase (GO)-induced insulin resistance in cultured skeletal muscle cells could be ameliorated by pre-treatment with gamma-tocopherol (GT). METHODS: Insulin sensitivity in L6 myotubes was assessed by 2-deoxy-D: -[(3)H]-glucose uptake. The phosphorylation of distal insulin signaling proteins Akt and the Akt substrate AS160 were determined by western blot. RESULTS: One hour treatment with 100 mU/ml GO decreased insulin-stimulated glucose uptake (P < 0.001). Pre-treatment with GT either partially (100 microM) or completely (200 microM) restored insulin-stimulated glucose uptake in cells after GO-induced insulin resistance. GO-induced oxidative stress did not impair insulin stimulated phosphorylation of Akt or AS160, but 200 microM GT increased insulin-stimulated phosphorylation of these key signaling proteins (P < 0.05). CONCLUSIONS: High-dose (200 microM) GT treatment ameliorated oxidative stress-induced insulin resistance in cultured rat L6 skeletal muscle cells.

Brown adipose tissue and lipid metabolism: New strategies for identification of activators and biomarkers with clinical potential.

Development of therapeutic agents directed towards increasing brown adipose tissue (BAT) energy expenditure to combat obesity and its comorbidities is currently an area of intense research. Both preclinical and clinical studies have suggested a potentially significant role for BAT in regulating whole body energy expenditure as well as glucose and lipid metabolism. Lipids, particularly long chain fatty acids (LCFAs), are recognized as integral substrates in mediating the primary heat-producing functions of BAT, and to date thought to be principally sourced from stored intracellular lipid droplets. While this prior understanding is not disputed, recent evidence has demonstrated the importance of lipids derived from the circulation, including those from dietary sources and from tissue lipolysis, especially white adipose tissue lipolysis. Moreover, recent studies have shed further light on a potential role for BAT as an autocrine, paracrine and endocrine organ, with lipids as key signaling molecules. Advances in metabolomics have enabled high-resolution exploration of biomolecules that may be associated with various physiological processes and potentially pathological states. Such approaches have led to several novel lipid species recently being associated with BAT function and dysfunction. Further exploration of the circulating lipidome will likely reveal additional novel BAT biomarkers that can inform development of BAT-directed therapies. This review will address current progress and new strategies to identify and characterize BAT-associated lipids which may represent both novel activators and/or activity biomarkers with both research and clinical utility.

Interleukin-6 increases insulin-stimulated glucose disposal in humans and glucose uptake and fatty acid oxidation in vitro via AMP-activated protein kinase.

Although interleukin-6 (IL-6) has been associated with insulin resistance, little is known regarding the effects of IL-6 on insulin sensitivity in humans in vivo. Here, we show that IL-6 infusion increases glucose disposal without affecting the complete suppression of endogenous glucose production during a hyperinsulinemic-euglycemic clamp in healthy humans. Because skeletal muscle accounts for most of the insulin-stimulated glucose disposal in vivo, we examined the mechanism(s) by which IL-6 may affect muscle metabolism using L6 myotubes. IL-6 treatment increased fatty acid oxidation, basal and insulin-stimulated glucose uptake, and translocation of GLUT4 to the plasma membrane. Furthermore, IL-6 rapidly and markedly increased AMP-activated protein kinase (AMPK). To determine whether the activation of AMPK mediated cellular metabolic events, we conducted experiments using L6 myotubes infected with dominant-negative AMPK alpha-subunit. The effects described above were abrogated in AMPK dominant-negative-infected cells. Our results demonstrate that acute IL-6 treatment enhances insulin-stimulated glucose disposal in humans in vivo, while the effects of IL-6 on glucose and fatty acid metabolism in vitro appear to be mediated by AMPK.

High-density lipoprotein delivered after myocardial infarction increases cardiac glucose uptake and function in mice.

Protecting the heart after an acute coronary syndrome is a key therapeutic goal to support cardiac recovery and prevent progression to heart failure. A potential strategy is to target cardiac glucose metabolism at the early stages after ischemia when glycolysis is critical for myocyte survival. Building on our discovery that high-density lipoprotein (HDL) modulates skeletal muscle glucose metabolism, we now demonstrate that a single dose of reconstituted HDL (rHDL) delivered after myocardial ischemia increases cardiac glucose uptake, reduces infarct size, and improves cardiac remodeling in association with enhanced functional recovery in mice. These findings applied equally to metabolically normal and insulin-resistant mice. We further establish direct effects of HDL on cardiomyocyte glucose uptake, glycolysis, and glucose oxidation via the Akt signaling pathway within 15 min of reperfusion. These data support the use of infusible HDL preparations for management of acute coronary syndromes in the setting of primary percutaneous interventions.