Running on empty:A metabolomics approach to investigating changing energy metabolism during fasted exercise and rest

Understanding the metabolic processes in energy metabolism, particularly during fasted exercise, is a growing area of research. Previous work has focused on measuring metabolites pre and post exercise. This can provide information about the final state of energy metabolism in the participants, but it does not show how these processes vary during the exercise and any subsequent post-exercise period. To address this, the work described here took fasted participants and subjected them to an exercise and rest protocol under laboratory settings, which allowed for breath and blood sampling both pre, during and post exercise. Analysis of the data produced from both the physiological measurements and the untargeted metabolomics measurements showed clear switching between glycolytic and ketolytic metabolism, with the liquid chromatography-mass spectrometry (LC-MS) data showing the separate stages of ketolytic metabolism, notably the transport, release and breakdown of long chain fatty acids. Several signals, putatively identified as short peptides, were observed to change in a pattern similar to that of the ketolytic metabolites. This work highlights the power of untargeted metabolomic methods as an investigative tool for exercise science, both to follow known processes in a more complete way and discover possible novel biomarkers

Pharmacological and genetic approaches to modulate energy metabolism in skeletal muscle cells

The prevalence of obesity and related metabolic illnesses have rapidly increased over the last decades. One of these diseases is type 2 diabetes (T2D), characterized by chronic hyperglycaemia and insulin resistance in several organs including skeletal muscle, an organ of particular interest when it comes to further understand metabolic conditions. In this thesis, we aimed to study the effects of different pharmacological and genetic approaches on skeletal muscle energy metabolism. The results presented in this thesis shows that activation of β2-adrenergic receptors and the cold-sensing transient receptor potential ion channels (TRP), inhibiting diacylglycerol acyltransferase (DGAT) and ablation of the α2 subunit of adenosine 5’-monophosphate (AMP)-activated protein kinase (AMPK) affected energy metabolism in myotubes. The β2- agonist terbutaline increased glucose uptake, glucose and fatty acid oxidation, upregulated mitochondrial and oxidative pathways and increased protein synthesis. Activation of TRPA1 resulted in an increased glucose uptake and oxidation, whereas inhibiting of DGAT and ablation of α2 subunit of AMPK resulted in alterations in lipid metabolism

Treatment of human skeletal muscle cells with inhibitors of diacylglycerol acyltransferases 1 and 2 to explore isozyme-specific roles on lipid metabolism.

Diacylglycerol acyltransferases (DGAT) 1 and 2 catalyse the final step in triacylglycerol (TAG) synthesis, the esterification of fatty acyl-CoA to diacylglycerol. Despite catalysing the same reaction and being present in the same cell types, they exhibit different functions on lipid metabolism in various tissues. Yet, their roles in skeletal muscle remain poorly defined. In this study, we investigated how selective inhibitors of DGAT1 and DGAT2 affected lipid metabolism in human primary skeletal muscle cells. The results showed that DGAT1 was dominant in human skeletal muscle cells utilizing fatty acids (FAs) derived from various sources, both exogenously supplied FA, de novo synthesised FA, or FA derived from lipolysis, to generate TAG, as well as being involved in de novo synthesis of TAG. On the other hand, DGAT2 seemed to be specialised for de novo synthesis of TAG from glycerol-3-posphate only. Interestingly, DGAT activities were also important for regulating FA oxidation, indicating a key role in balancing FAs between storage in TAG and efficient utilization through oxidation. Finally, we observed that inhibition of DGAT enzymes could potentially alter glucose–FA interactions in skeletal muscle. In summary, treatment with DGAT1 or DGAT2 specific inhibitors resulted in different responses on lipid metabolism in human myotubes, indicating that the two enzymes play distinct roles in TAG metabolism in skeletal muscle

Chronic treatment with terbutaline increases glucose and oleic acid oxidation and protein synthesis in cultured human myotubes

Objective: In vivo studies have reported several beneficial metabolic effects of β-adrenergic receptor agonist administration in skeletal muscle, including increased glucose uptake, fatty acid metabolism, lipolysis and mitochondrial biogenesis. Although these effects have been widely studied in vivo, the in vitro data are limited to mouse and rat cell lines. Therefore, we sought to discover the effects of the β2-adrenergic receptor agonist terbutaline on metabolism and protein synthesis in human primary skeletal muscle cells. Methods: Human cultured myotubes were exposed to terbutaline in various concentrations (0.01–30 ​μM) for 4 or 96 ​h. Thereafter uptake of [14C]deoxy-D-glucose, oxydation of [14C]glucose and [14C]oleic acid were measured. Incorporation of [14C]leucine, gene expression by qPCR and proteomics analyses by mass spectrometry by the STAGE-TIP method were performed after 96 ​h exposure to 1 and 10 ​μM of terbutaline. Results: The results showed that 4 ​h treatment with terbutaline in concentrations up to 1 ​μM increased glucose uptake in human myotubes, but also decreased both glucose and oleic acid oxidation along with oleic acid uptake in concentrations of 10–30 ​μM. Moreover, administration of terbutaline for 96 ​h increased glucose uptake (in terbutaline concentrations up to 1 ​μM) and oxidation (1 ​μM), as well as oleic acid oxidation (0.1–30 ​μM), leucine incorporation into cellular protein (1–10 ​μM) and upregulated several pathways related to mitochondrial metabolism (1 ​μM). Data are available via ProteomeXchange with identifier PXD024063. Conclusion: These results suggest that β2-adrenergic receptor have direct effects in human skeletal muscle affecting fuel metabolism and net protein synthesis, effects that might be favourable for both type 2 diabetes and muscle wasting disorders

Additional file 1 of Effect of noradrenaline on propofol-induced mitochondrial dysfunction in human skeletal muscle cells

Additional file 1: Table S1. Study subject characteristics on biopsy day. Figure S1. Cell viability. Results are expressed as the percentage of cell viability relative to the control (= non-treated cells). a) Individual groups represent viability of cells exposed for 96 h to different concentrations of noradrenaline. Data are presented a s the mean ± SEM (n = 4 subjects). Values for each experimental condition were measured in triplicates in each subject. b) Individual groups represent viability of cells exposed for 96 h to ethanol (= propofol vehicle; 0.1%), 0.1 mM noradrenaline alone and c) different concentrations (μg/mL) of either propofol alone or mixture of propofol and 0.1 mM noradrenaline. Data are presented as the mean ± SEM (n = 7 subjects). Values for each experimental condition were measured in triplicates in each subject. Note: NA = noradrenaline. *** p < 0.001 vs. control group. Figure S2. Kinetic graph on XF24 Analyzer demonstrates changes after propofol at various concentrations. Real-time measurement of OCR at baseline and after sequential injection of oligomycin, FCCP and Antimycin A. Each data-point represents the mean of 7 independent samples (subjects) measured in tri- or tetraplicates normalized to protein content. Error bars indicate standard error of the mean. Different colours represent different groups exposed to propofol (0; 2.5; 10 µg/mL). Figure S3. A) Global mitochondrial parameters. Basal respiration, maximal respiratory capacity, ATP production and non-mitochondrial respiration. N = 7 replicates with 21–28 wells for each condition normalized to protein content. Error bars indicate standard error of the mean. B) Mitochondrial mass calculated as a fraction (%) of a cell surface area in 2D cross-sectional images. Figure S4. Uptake of CO2 production and lactic acid metabolism. A) Uptake of [14C]palmitic acid. B) [14C]lactic acid oxidation. C) [14C]lactic acid uptake. Note: NA = noradrenaline. Error bars in each graph indicate standard error of the mean. Note: NA = noradrenaline. Figure S5. Flow cytometry. Histogram showing MTG intensity of individual cell groups. Data are presented as the mean ± SEM (n = 2–3 experiments per each group). Note: NA = noradrenaline. Figure S6. Mitochondrial membrane potential. A) Myoblasts after staining with MitoTracker™ Green FM (left), TMRE (in the middle) and after staining of both agents (right). Experiments were performed at least at 60 cells per each group from n = 3 independent experiments (cells from 3 individual subjects). B) Determination of ∆ψm was expressed as TMRE/MTG ratio. The mitochondrial uncoupling agent FCCP was used as a positive control. Note: MTG = MitoTracker™ Green FM; TRME = tetramethylrhodamine ethyl ester; FCCP = carbonyl cyanide-4-(trifluoromethoxy)phenylhydrazone, NA = noradrenaline. Error bars indicate standard error of the mean. *p < 0.05, ***p < 0.001 vs. control group. Figure S7. Analysis of lipid droplets. A) Quantification of LD mass by cross-sectional area of BODIPY 493/503 normalized to cell area. B) LD size assessed by cross-sectional area of individual LDs. C) LD number normalized to cell area. Experiments were performed at least at 38 cells per each condition from 2 independent measurements (= 2 individual subjects). Error bars indicate standard error of the mean. ***p < 0.001 vs. control group

Three-Phase Fuel Deposition in a Long-Distance Migrant, the Red Knot (Calidris canutus piersmai), before the Flight to High Arctic Breeding Grounds

<p>Refuelling by migratory birds before take-off on long flights is generally considered a two-phase process, with protein accumulation preceding rapid fat deposition. The first phase expresses the demands for a large digestive system for nutrient storage after shrinkage during previous flights, the second phase the demands for fat stores to fuel the subsequent flight. At the last staging site in northward migration, this process may include expression of selection pressures both en route to and after arrival at the breeding grounds, which remains unascertained. Here we investigated changes in body composition during refuelling of High Arctic breeding red knots (Calidris canutus piersmai) in the northern Yellow Sea, before their flight to the tundra. These red knots followed a three-phase fuel deposition pattern, with protein being stored in the first and last phases, and fat being deposited mainly in the second phase. Thus, they did not shrink nutritional organs before take-off, and even showed hypertrophy of the nutritional organs. These suggest the build up of strategic protein stores before departure to cope with a protein shortage upon arrival on the breeding grounds. Further comparative studies are warranted to examine the degree to which the deposition of stores by migrant birds generally reflects a balance between concurrent and upcoming environmental selection pressures.</p>

Endovascular thrombectomy for acute ischaemic stroke with established large infarct: multicentre, open-label, randomised trial

International audienc

Rationale, design, and baseline characteristics in Evaluation of LIXisenatide in Acute Coronary Syndrome, a long-term cardiovascular end point trial of lixisenatide versus placebo

BACKGROUND: Cardiovascular (CV) disease is the leading cause of morbidity and mortality in patients with type 2 diabetes mellitus (T2DM). Furthermore, patients with T2DM and acute coronary syndrome (ACS) have a particularly high risk of CV events. The glucagon-like peptide 1 receptor agonist, lixisenatide, improves glycemia, but its effects on CV events have not been thoroughly evaluated. METHODS: ELIXA (www.clinicaltrials.gov no. NCT01147250) is a randomized, double-blind, placebo-controlled, parallel-group, multicenter study of lixisenatide in patients with T2DM and a recent ACS event. The primary aim is to evaluate the effects of lixisenatide on CV morbidity and mortality in a population at high CV risk. The primary efficacy end point is a composite of time to CV death, nonfatal myocardial infarction, nonfatal stroke, or hospitalization for unstable angina. Data are systematically collected for safety outcomes, including hypoglycemia, pancreatitis, and malignancy. RESULTS: Enrollment began in July 2010 and ended in August 2013; 6,068 patients from 49 countries were randomized. Of these, 69% are men and 75% are white; at baseline, the mean ± SD age was 60.3 ± 9.7 years, body mass index was 30.2 ± 5.7 kg/m(2), and duration of T2DM was 9.3 ± 8.2 years. The qualifying ACS was a myocardial infarction in 83% and unstable angina in 17%. The study will continue until the positive adjudication of the protocol-specified number of primary CV events. CONCLUSION: ELIXA will be the first trial to report the safety and efficacy of a glucagon-like peptide 1 receptor agonist in people with T2DM and high CV event risk