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Abstract book:

Workshop at cambridge universityWhy Is Exercise Good For Us? Complementing Genetics For Better Metabolic Regulation10-11 December 2015 Danish Diabetes Academy & Cambridge Metabolic Network

Danish Diabetes AcademyFunded by the Novo Nordisk Foundation

This workshop entitled “Why is exercise good for us? Complementing genetics for better metabolic regulation” is a joint meeting between the Danish Diabetes Academy and the Cambridge Metabolic Network.

With focus on exercise and metabolism, the purpose of the workshop is to present and discuss research with intentions to advance awareness of ongoing projects and nurture opportunities for research collaboration where complementary expertise will synergise progress in metabolic research.

The workshop will include leading researchers from Cambridge, affiliated with Cambridge Metabolic Network and members of the Danish Diabetes Academy with approximately 45 participants from each country.

On behalf of the organizers

Prof Anne Ferguson-Smith, Dept. of Genetics, University of CambridgeProf Ken Siddle, Dept. of Clin. Biochem., University of CambridgeDr Mike Murphy, MRC Mitochondrial Biology UnitProf Jørgen Jensen, Norwegian School of Sports SciencesAss Prof Niels Jessen, Aarhus UniversityProf Jørgen Wojtaszewski, University of Copenhagen

Preface

4 Danish Diabetes Academy

Akhilesh Reddy Redox and metabolic oscillations in the circadian (24 hour) clockwork p. 08

Bente Stallknecht Fitness to fight fat p. 09

Bjørn Richelsen Effect of intake of proteins high in ketogenic amino acids (e.G. Leucine) together with resistance training in elderly frail individuals for four months. Implications for muscle, bones, and physical function p. 10

Clara Prats Compartmentalization of skeletal muscle metabolic pathways - effect of physical activity and inactivity p. 11

Fiona Gribble Signalling in the enteroendocrine axis p. 12

Dino a. Giussani Heart disease link to fetal hypoxia and oxidative stress p. 13

Jørgen Wojtaszewski The amp activated protein kinase - a regulator of muscle insulin sensitivity p. 14

Kay-Tee Khaw Diet and metabolic health: some current controversies p. 15

Kurt Højlund Effect of exercise on insulin sensitivity and glycogen synthase activation p. 16 Margaret Ashcroft Hif and metabolismp. 17

Mike Murphy How do mitochondria respond to exercise? p. 18

Andrew Murray The genetic and metabolic basis of performance at high altitude: the secrets of sherpa success p. 19

Niels Jessen Signalling in skeletal muscles after fasting and exercise p. 20

Niels Møller Metabolic stress models in experimental clinical research p. 21

Robert Scott What can human genetics tell us about the determinants of fitness? Lessons from metabolic disease p. 22

Soren Brage Dose-response relationships between habitual physical activity and glucose metabolism: observations from a large cohort study p. 23

Torben Hansen Genetics of metabolism – lessons from greenland p. 24

Content

Speaker abstracts

5 Workshop at Cambridge University / 10-11 December 2015

01 Early life exposure to maternal obesity perturbs renal morphology in mice p. 26

02 A novel murine model of short syndrome p. 27

03 Autophagic regulation in human skeletal muscle in response to physical activity and inactivity p. 28

04 A low-cost, hand-held breathalyser that measures the human metabolism p. 29

05 Interactions between fto, physical activity and insulin sensitivity; one or both? p. 30

06 The role of the bone morphogenetic protein 9 in atherosclerosis p. 31

07 Ceramide and diacylglycerol content is not influenced by 10 weeks exercise training in first-degree relatives of type 2 diabetic patients p. 32

08 The identification of novel metabolites that track with improvements in glycaemia following a 12-week lifestyle intervention in high risk individuals p. 33

09 Improved insulin sensitivity decrease the genetic effect on triglyceride levels in blood p. 34

10 Do genes associated with obesity cause the dysfunction of hypothalamic pomc and agrp neurons? p. 35

11 Central carbon metabolism regulates circadian oscillations p. 36

12 Whole-genome sequencing reveals low frequency variants associated with body mass index and height p. 37

13 Fibro-adipogenic progenitors in response to acute and prolonged resistance exercise p. 38

14 Effects of training in vivo is preserved in cultured human myotubes p. 39

15 Effect of maternal exercise during obese pregnancy on the cardiovascular health of male offspring p. 40

16 Prolonged activation of glycogen synthase after exercise despite substantial glycogen synthesis during recovery p. 41

17 Post-weaning high-fat diet exacerbates insulin resistance programmed by maternal obesity in offspring adipose tissue p. 42

18 Reduced CD300LG mrna tissue expression, increased intramyocellular lipid content and impaired glucose metabolism in healthy male carriers of Arg82Cys in CD300LG p. 43

19 Low physical activity is associated with increased arterial stiffness in patients recently diagnosed with type 2 diabetes p. 44

20 Inorganic nitrate increases exercise efficiency, tolerance, and performance, and is growing in popularity as a nutritional sport supplement p. 45

Poster abstracts


21 Impaired lipid storage and mitochondrial network organization in morbid obese human skeletal muscle. Improvements associated to gastric bypass and link to insulin resistance remission p. 46

22 Genomic, transcriptomic, and metabolic variations shape the complex adaptation landscape of bacteria to varying environmental conditions p. 47

23 Identification of novel genetic determinants of erythrocyte membrane fatty acid composition among greenlandic inuit p. 48

24 Anabolic compounds during human catabolic and inactive conditions p. 49

25 High intensity interval training improves glycaemic control and pancreatic β cell function of type 2 diabetes patients p. 50

26 Transcriptional regulators of metabolic adaptation in muscle are robustly induced by acute exercise in type 2 diabetics. No evidence of exercise resistance p. 51

27 Are the effects of adiposity loci modified by cardiorespiratory fitness among danish adults? p. 52

28 Dissecting skeletal muscle mechanical stress x ampk signalling interactions using phosphoproteomics p. 53

29 Predicting physical activity energy expenditure from wrist accelerometry: independent contributions of acceleration magnitude and orientation p. 54


Speaker abstracts


Redox and metabolic oscillations in the circadian (24 hour) clockwork

Akhilesh Reddy, Dr.

Metabolic Research Laboratories, University of Cambridge

Much is now understood about the nature of transcriptional oscillations in multiple organisms. Until recently, however, little was known about non-transcriptional mechanisms that may contribute to circadian timekeeping. Our recent work in red blood cells and marine algae pointed towards redox oscillations playing an unanticipated role in 24 hour timekeeping.

A family of proteins called the peroxiredoxins appear to be key readouts of the non-transcriptional clockwork, and their circadian oscillation is, remarkably, conserved in all phylogenetic domains, including bacteria, archaea and eukaryotes. Thus, redox mechanisms are likely to be deeply embedded within the clockwork of multiple species, in stark contrast to the lack of evolutionary conservation of transcriptional components of the clockwork.

Our recent work highlights the importance of central carbon metabolism in modulating the transcriptional oscillator, suggesting one connection between non-transcriptional and transcriptional clock networks.


Fitness to fight fat

1. Malhotra et al., BJSM 20152. Nordby et al., Obesity 20123. Rosenkilde et al., Am.J.Physiol.Regul.Integr.

Comp.Physiol. 20124. Rosenkilde et al., J.Appl.Physiol. 20135. Nordby, Rosenkilde et al., J.Appl.Physiol. 20156. Rosenkilde et al. Scand.J.Med.Sci.Sports 20137. Auerbach et al. Am.J.Physiol.Regul.Integr.

Comp.Physiol. 20138. Reichkendler et al. Am.J.Physiol.Endocrinol.

Metab. 20139. Stallknecht et al. Am.J.Physiol.Endocrinol.

Metab. 199110. Stallknecht et al. Am.J.Physiol.Endocrinol.

Metab. 1993

Bente Stallknecht, Professor and Acting Head of Department, MD, PhD, DMSc

Department of Biomedical Sciences, Faculty of Health and Medical Sciences, University of Copenhagen

An editorial in British Journal of Sports Medicine claimed that ‘you cannot outrun a bad diet’ (1) and this has sparked both scientific and public debate.

Diet is definitely of importance for health, but so is physical exercise. In two 12-week studies of overweight men, ‘Project Training per se’ and ‘Project FINE’, we showed marked effects of daily endurance exercise on body weight and fat mass loss (2-4), peak fat oxidation during exercise (5, 6) and adipose tissue function (7, 8).

Also, in older 10-week endurance training studies of rats we found marked increases in mitochondrial enzyme activity (9) and insulin-stimulated glucose transport (10) in adipose tissue.

We conclude that physical exercise can markedly affect energy balance, fat oxidation capacity and adipose tissue function showing that ‘fitness can fight fat’.


Effect of intake of proteins high in ketogenic amino acids (e.g. Leucine) together with resistance training in elderly frail individuals for four months. Implications for muscle, bones, and physical function

Bjørn Richelsen, Jesper Mehlsen, Nikolaj Rittig, Kristina Nørregaard, Niels Møller

Endocrinology and Internal Medicine, Aarhus University Hospital

The objective was to investigate whether a high protein intake particularly of ketogenic amino acids (e.g. high leucine) together with training could enhance muscle mass and improve the physical function in elderly fragile individuals , in addition, to the effect of circulating microRNAs.

Study designIt is a 4 month randomized, blinded study with three intervention arms – 1) placebo – isocaloric without protein (+ maltrodrextrine), 2) Soy protein – high protein low leucine (HPLL – 6.7% leucine), and 3) Whey protein – high protein high leucine (HPHL – 13.4% leucine). The protein was taken twice daily giving a total amount of 45.8 g protein daily in both groups. In addition, all participants performed resistance training three times per week during the intervention period, primarily in order to possible enhance the effects of the protein supplements. Inclusion criteria was age between 60-85 years with DXA-determined osteopenia (BMD score < -1). Outcomes were DXA determined muscle and bone mass and physical function was determined by two walking tests, a balance test, and an ergometric test etc. Moreover, circulating levels of microRNAs were measured by RT-qPCR.

Results23 individuals were included in each of the protein groups and 10 individuals in the placebo. At baseline a positive correlation was found between intake of protein and LBM (r=52, P<0.001). The results from the intervention are still preliminary. In the placebo group there was a higher weight gain during the intervention ( 1.1 kg) with stable body weight in the two protein groups. There was a significant increase in LBM in the soy group with no changes in the two other groups. On the other hand, in the HPHL group the 6-minute walking test improved significantly with no changes in the placebo group. Concerning circulating miRNAs we found that resistant training increased several miRNAs such as miR-113a-3p, miR-499a-3p, and miR-399a-5p whereas no effects of the protein supplements were detected.

ConclusionsThe protein supplements seem to enhance some outcomes such as LBM and the 6 -minute walk test but without a clear picture of any specific effect of high leucine. Whereas, resistance training affected the circulating levels of miRNAs.


Compartmentalization of skeletal muscle metabolic pathways - effect of physical activity and inactivity

Clara Prats, PhD

Center for Healthy Aging, the Core Facility for In-tegrated Microscopy, Department of Biomedical Sciences, Faculty of Health and Medical Sciences, University of Copenhagen

Skeletal muscle fibers are not bags full of proteins and metabolites; their intracellular organization is complex and compact. Thus, metabolic events can only be understood as highly compartmentalized and tightly regulated processes.

This talk will review the current knowledge on physical activity/inactivity-induced adaptations of skeletal muscle glycogen metabolism and mitochondrial dynamics, with focus on the involved intracellular compartments and the dynamic structural re-organization.

Results from human skeletal muscle after several interventions: exhaustive exercise, muscle immobilization and gastric-by-pass induced weight loss indicate that intracellular compartmentalization of metabolic events and structural re-organization of intracellular compartments are key regulators skeletal muscle metabolic health.

Danish Diabetes Academy

Signalling in the enteroendocrine axis

Fiona Gribble, Professor


The gut endocrine system produces a repertoire of hormones that orchestrate post-prandial metabolism. GLP-1, for example, is secreted after food ingestion and acts to coordinate rates of insulin release with rates of nutrient absorption, and also promotes satiety.

The clinical effectiveness of GLP-1 mimetics and DPP4 inhibitors illustrate the physiological and translational importance of understanding the function of the gut-pancreatic-brain axis.

The marked ability of gastric bypass surgery to reduce appetite and resolve many cases of type 2 diabetes has leant extra weight to the idea that therapies should be developed that target or otherwise harness the gut endocrine system. Understanding the cellular signalling pathways underlying the physiological functioning of the enteroendocrine axis is crucial for the development of new therapies targeting the gut endocrine system.


Heart disease link to fetal hypoxia and oxidative stress

Dino A. Giussani, Professor

Department of Physiololgy, Development & Neuroscience, University of Cambridge

The quality of the intrauterine environment interacts with our genetic makeup to shape the risk of developing disease in later life. Fetal hypoxia is a common complication of pregnancy. It programmes cardiac and endothelial dysfunction in the offspring in adult life. However, the mechanisms via which this occurs remain elusive, precluding the identification of potential therapy.

Using an integrative approach in large and small animal species at the in vivo, isolated organ, cellular and molecular levels, my programmes of work have raised the hypothesis that oxidative stress in the fetal heart and vasculature underlies the mechanism via which prenatal hypoxia programmes cardiovascular dysfunction in later life.

We have shown that developmental hypoxia independent of changes in maternal nutrition promotes fetal growth restriction and induces changes in the cardiovascular, metabolic and endocrine systems of the adult offspring, which are normally associated with disease states during ageing (Camm et al. FASEB 25(1):420, 2011; Giussani et al. PLoS One 7(2):e31017, 2012).

Treatment with antioxidants, such as melatonin, allopurinol or statins of animal pregnancies complicated with reduced oxygen delivery to the fetus prevents the alterations in fetal growth, and the cardiovascular, metabolic and endocrine dysfunction in the fetal and adult offspring. We also show the intergenerational inheritance of cardiovascular disease risk induced by chronic fetal hypoxia via the paternal line.

Combined, the data support the hypothesis tested and the work offers both insight into mechanisms and possible therapeutic targets for clinical intervention against the early origin of cardiovascular disease in risky pregnancy.


The amp activated protein kinase - a regulator of muscle insulin sensitivity

We have data to suggest that the signaling interplay takes place at a least one important signaling node regulating glucose transport i.e. the GTPase activating protein TBC1D4. We have signaling profiling in support of this mechanism in human skeletal muscle. Thus, we hypothesize that AMPK is important in regulating muscle insulin sensitivity and therefore also for whole body insulin action. We suggest that AMPK is a key factor regulating metabolism not only during exercise but in particular in the post exercise recovery period. We propose AMPK as a potential drug target in treatment/prevention of diseases as-sociated with insulin resistance.

Jørgen Wojtaszewski, Professor, PhD Co-authors; Janne Rasmus Hingst, Rasmus Kjøbsted, Jesper Birk, Erik Richter, Bente Kiens

Dept. of Molecular Physiology, Faculty of Science, University of Copenhagen, Denmark

40 years ago it was hypothesized that muscle contraction/exercise induced a potentiation of insulin action in the prior active muscle fibers. This phenomenon was proven to exist 25 years ago and now we can present a novel mechanism.

Surprisingly, our studies show that the cellular fuel/energy sensor AMP activated protein kinase traditionally view upon as a “catabolic” signal transmitter interacts with the anabolic insulin signaling. Thus, pharmacologic treatment with the AMPK activator AICAR increases subsequently the metabolic action of insulin.

In transgenic mice ablated for AMPK catalytic activity specifically in muscle this effect of AICAR is not present. Thus, activation of AMPK is sufficient to increase muscle insulin sensitivity. Muscle contraction is a potent stimulus for AMPK. We have now data to suggest that one important physiological function of AMPK in muscle is to mediate the post contraction insulin sensitivity, i.e. in mice lacking AMPK in skeletal muscle contraction no longer mediates insulin sensitization.


Diet and metabolic health: some current controversies

Kay-Tee Khaw, Professor of Clinical Gerontology and Fellow of Gonville and Caius College

Department of Public Health and Primary Care, University of Cambridge, UK

I will discuss some current controversies about the relationship between diet and metabolic health with examples from the debate over the role of dietary sugars and fats.

Bio: Professor Khaw trained in medicine at Girton College, University of Cambridge and St. Mary’s Hospital, London and in epidemiology at the London School of Hygiene and Tropical Medicine, with subsequent clinical and academic posts in the University of London and University of California San Diego.

Her research interests are the maintenance of health in later life and the causes and prevention of chronic diseases including cardiovascular disease, cancer and osteoporosis with a focus on nutrition and hormones. She is a principal investigator in the European Prospective Investigation in Cancer in Norfolk, part of a ten country half million participant research collaboration.


Effect of exercise on insulin sensitivity and glycogen synthase activation

The exercise-induced changes in GS activity, substrate affinity and phosphorylation were sustained 3-h post-exercise and further enhanced by insulin. However, post-exercise, reduced GS activity and substrate affinity as well as increased sites 2+2a phosphorylation were found in T2D patients.

In conclusion, activation of muscle GS by acute exercise in T2D and controls involves site-specific dephosphorylation of GS and enhanced substrate affinity. Endurance-training did not rescue an impaired insulin activation of muscle GS activity in T2D, and reduced GS activity and dephosphorylation in T2D during recovery imply impaired adaptation to acute exercise.

Kurt Højlund, Professor, PhD, MD

Department of Endocrinology, Odense University Hospital,DK-5000 Odense C

Impaired insulin-stimulated glycogen synthesis is a hallmark of insulin resistance in human skeletal muscle. Recent reports indicate an impaired response to exercise in some cohorts with insulin resistance.

Insulin and exercise stimulate muscle glycogen synthase (GS) activity by dephosphorylation and changes in kinetic properties. In two recent studies, we have examined the effects acute exercise (60 min at 70% of VO2max) and 10-weeks training (4-5 sessions weekly at ~65% of VO2max) on basal and insulin-stimulated glucose metabolism and muscle GS phosphorylation, activity, and substrate affinity in patients with type 2 diabetes (T2D) and weight-matched controls.

Insulin sensitivity was lower in T2D patients vs controls with or without prior acute exercise or training. A defect in insulin activation of GS fractional velocity (FV) in T2D was not rescued by training, but training increased GS phosphorylation due to increased GS protein in both groups. Insulin and, more potently, acute exercise increased GS FV, substrate affinity and dephosphorylation in both groups.


HIF and metabolism

Margaret Ashcroft, Professor

Department of Medicine, University of Cambridge, UKEmail: [email protected]

Research over the last decade has greatly increased our understanding of how cells sense and respond to changes in cellular oxygen levels. The hypoxia inducible factor (HIF) family of bHLH-PAS transcription factors are central components of the cellular oxygen-sensing machinery in mammalian cells.

Previous studies have highlighted an important role for mitochondria in regulating HIF and the hypoxic response (1, 2). However, the molecular mechanisms that interface mitochondria with the HIF response remain to be unraveled.

We have identified and characterized a family of mitochondrial proteins that regulate cellular oxygen consumption rate and metabolism, and provide a critical role in HIF signaling in disease (1). These novel molecular links between mitochondria and the HIF pathway provide new molecular insights into metabolic adaptive responses in disease.

(1) Yang J, Staples O, Thomas LW, Briston T, Robson M, Poon E, Simões ML, El-Emir E, Buffa FM, Ahmed A, Annear NP, Shukla D, Pedley BR, Maxwell PH, Harris AL, Ashcroft M (2012). Human CHCHD4 mitochondrial proteins regulate cellular oxygen consumption rate and metabolism and provide a critical role in hypoxia signaling and tumor progression. J Clin Invest. 122(2):600-11.

(2) Briston T, Yang J, Ashcroft M (2011). HIF-1α localization with mitochondria: a new role for an old favorite? Cell Cycle. 10(23):4170-1.


How do mitochondria respond to exercise?

Mike Murphy, Dr.

MRC Mitochondrial Biology Unit, Cambridge

Mitochondria respond to exercise with both long-term and acute responses. In this talk I will survey the various ways in which mitochondria respond to exercise. I will then go to focus on the role of mitochondrial ROS production in regulating the acute response of mitochondrial function to this putative redox signal.


The genetic and metabolic basis of performance at high altitude: the secrets of Sherpa success

Andrew Murray, Dr.

Department of Physiology, Development & Neuroscience, University of Cambridge, UK

The Himalayan Sherpas, a human population of Tibetan descent, are highly adapted to life in the hypobaric hypoxia of high altitude. Physiological features and genetic signals in Tibetan groups have hinted at adaptations in oxygen supply to the tissues, however little is known about oxygen utilisation.

We therefore investigated Sherpa and Lowlander subjects before and during exposure to hypobaric hypoxia during an ascent to Mt Everest Base Camp, with particular focus on the mechanism by which a putatively advantageous allele for the PPARA gene might influence energy metabolism. Here we find that Sherpas have a metabolic adaptation to life at high altitude that is associated with enhanced efficiency of oxygen utilisation, improved muscle energetics and protection against oxidative stress compared with Lowlanders.

Our findings suggest that metabolic adaptations underpin human evolution to life at high altitude, and could impact upon our understanding of human diseases where hypoxia is a feature.


Signalling in skeletal muscles after fasting and exercise

Our data demonstrate that moderate exercise activates signaling to autophagy independent of nutritional status. Prolonged fasting induces severe insulin resistance, but insulin signaling to glucose uptake is preserved. Cellular energetics determined as AMP, ADP, ATP and phosphocreatinine levels are preserved during fasting and prolonged fasting does not deplete glycogen stores.

Our data indicate that substrate availability in skeletal muscle during prolonged fasting is not severely limiting exercise performance. Furthermore we show that exercise activates autophagy signaling, but this is nutrient independent and is therefore more likely involved in protein remodeling after exercise than supplying substrates for metabolism.

Niels Jessen, Associate Professor

Department of Clinical Medicine, Aarhus University

The energy demand in skeletal muscle is dependable on the level of physical activity, and muscles are capable of oxidizing both lipids and glucose for its energy demand. However, availability of these substrates depends are nutrient-dependent, and uptake of glucose into skeletal muscle is reduced during prolonged fasting. We have tested if glycogen metabolism and signaling to autophagy during exercise is nutrient-dependent. In addition, we have tested insulin sensitivity in skeletal muscle during prolonged fasting in healthy subjects using hyperinsulinemic euglycemic clamp with skeletal muscle biopsies, and subsequent signaling- and metabolomics analysis.


Metabolic stress models in experimental clinical research

Niels Møller, Professor, MD

Medical Research Laboratories, Clinical Institute/Medical Dep MEA., Aarhus University Hospital

Our research laboratory conducts clinical, experimental studies as outlined below. We measure circulating and interstitial levels of metabolites and hormones, use dilution techniques to assess turnover rates and exploit a/v techniques and muscle and fat biopsies to analyze regional metabolism and tissue signaling mechanisms. Acute metabolic stress with particular focus on hormonal control of protein, glucose and lipid metabolism in diabetes and normal physiology, using exercise, fasting and administration of pyrogens(endotoxin and TNF-α) as stress models.

Core methodology: Epinephrine, Insulin. Glucose, Phenylalanine, Tyrosine (GCMS), Amino acids (HPLC), FFA, Palmitate(HPLC).

Hormone & metabolite assays: Cytokines, MBL, Adiponectin, IGF’s, GH, Ghrelin, Cortisol. Dilution: 3H-glucose, 3H-palmitate, 2H-phenylalanine&tyrosine, 15N-tyrosine, 13C-leucine, 13C-urea (GCMS) – for assessment of

whole body and regional turnover.

Clamp-techniques: Glucose clamps (eu-, hypo- & hyper-), hormone clamps(somatostatin), FFA clamps(nicotinic acid derivatives – acipimox) – for assessment of hormone/insulin sensitivity.

Indirect calorimetry: Deltatrac monitors – for assessment of energy expenditure, glucose, lipid and protein oxidation.

Microdialysis: Glycerol (&lactate, glucose, urea) – for assessment of regional lipolysis Muscle & fat biopsies: Hormone, glucose, lipid and protein metabolic signals.

Arterio-venous techniques: Forearm, leg,( heart, kidney &splanchnic bed) – blood flow by plethysmography, dilution(thermo-/cardiogreen) or Doppler – for assessment of regional (muscle) metabolism. Selective leg (femoral artery) perfusion to assess direct local effects of metabolic agents.


What can human genetics tell us about the determinants of fitness? Lessons from metabolic disease

Robert Scott, Dr.

MRC Epidemiology Unit, University of Cambridge

Human genetics is a powerful tool to offer novel insights into the aetiology of complex traits. A range of components of fitness are heritable, and associated with morbidity and mortality, yet little is known about the determinants of inter-individual variation in fitness.

In this talk, I will describe understanding of the genetic basis of fitness, past and current approaches and recent results on the genetic aetiology of fitness. I will also discuss how insights from genetic of complex disease can inform our understanding of the genetic basis of fitness.


Dose-response relationships between habitual physical activity and glucose metabolism: observations from a large cohort study

Soren Brage, Dr.


This talk will cover dose-response relationships between habitual physical activity and glucose metabolism using observational epidemiological data from the Fenland cohort (n=12435), with a focus on intensity and duration aspects of activity exposures.


Genetics of metabolism – lessons from Greenland

The indigenous people of Greenland, the Inuit, are also of interest because of their adaptation to the extreme conditions in the Arctic, including low annual temperatures and a diet rich in marine ω-3 polyunsaturated fatty acids (PUFAs). A scan of Inuit genomes for signatures of adaptation reveals signals in several loci, with the strongest signal located in a cluster of fatty acid desaturases that determine PUFA levels. The selected alleles are associated with multiple metabolic and anthropometric phenotypes and have large effect sizes for weight and height, with the effect on height replicated in Europeans. Analyzing membrane lipids, we show that the selected alleles modulate fatty acid composition. Thus, the Inuit have genetic and physiological adaptations to a diet rich in PUFAs.

Our studies in Greenlanders provide proof of the value of conducting genetic association studies outside the traditional setting of large homogenous populations.

Torben Hansen, Professor

University of Copenhagen

Genetic studies of small and isolated populations provide a foundation for investigating genetic factors of importance for metabolism. We recently discovered a nonsense p.Arg684Ter variant in TBC1D4 with an allele frequency of 17%. Under a recessive model, this variant strongly increases the levels of plasma glucose (β=3.8 mmol/L, P=2.5×10-35) and serum insulin (β=165 pmol/L, P=1.5×10-20) two hours after an oral glucose load, and increases T2D risk (OR 10.3, P=1.6×10-24), while it modestly reduces fasting plasma glucose (β=-0.18 mmol/L, P=1.1×10-6) and fasting serum insulin (-8.3 pmol/L, P=0.0014). The observed effect sizes are several times larger than any previous findings of large-scale genome-wide association studies for these traits.


Poster abstracts


01 Early life exposure to maternal obesity perturbs renal morphology in mice

groups (p= 0.95). Offspring exposed to a maternal obesogenic diet had significantly larger combined renal cortex and medulla volumes than offspring exposed to a maternal chow diet (p=0.004).

However, the number of nephrons/mm3 within the cortex and medulla was significantly reduced in offspring of obese pregnancies when compared to controls (p=0.0089). These results suggest that individuals exposed to maternal obesity during gestation and suckling may be at more risk of developing renal disease and associated CVD in later life.

These findings highlight the importance of developing an effective maternal intervention for obesity during pregnancy, both to dissect the mechanisms responsible for renal perturbations and to prevent renal pathologies in future generations.

Adele Pinnock, Heather Blackmore, Tom Ashmore and Susan Ozanne


The incidence of Chronic Kidney Disease (CKD) has risen globally by 83% since 1990, concurrently with type 2 diabetes and the metabolic syndrome. Studies of maternal under-nutrition have highlighted that the kidney can be adversely “programmed” resulting in fewer filtration units, a factor shown to promote the pathogenesis of CKD, cardiovascular disease (CVD) and hypertension. Despite the dramatic increase in obesity in recent years, the effect of maternal obesity on offspring kidney structure and function remains largely unexplored.

The aim of the current study was to define the effects of maternal over-nutrition on offspring kidney structure using a mouse model of maternal diet-induced obesity. Female C57BL/6 mice were fed a high fat diet supplemented with condensed milk for six weeks prior to pregnancy and throughout gestation and lactation. Male offspring were studied at 3 weeks of age. Kidneys were harvested, sectioned and stained with Haematoxylin and Eosin.

The number of nephrons were counted in pairs of sections at even interspaces throughout the kidney to accurately estimate the total number of nephrons within the three dimensional organ. There was no difference in offspring kidney weights between the 2


02 A novel murine model of SHORT syndrome

Kwok A. Zvetkova I., Jensen J., Hart D., Virtue S., Knox R., O’Rahilly S., Voshol P., Semple R.

University of Cambridge Metabolic Research Laboratories, Level 4, Wellcome Trust-MRC Institute of Metabolic Science, Box 289, Addenbrooke’s Hospital Cambridge, CB2 0QQ, UK

Pik3r1 encodes three regulatory subunits (p85α, p55α and p50α) of the class IA phosphatidylinositide 3 kinase (PI3K), and is critical in regulating growth and metabolism. Paradoxically, knockout of either p85α or p55α and p50α resulted in an improvement of insulin sensitivity. However, recent human genetic studies have shown that mutations of PIK3R1 produce SHORT syndrome, which is characterised by Short stature, Hyperextensibility of joints and/or Hernias, Ocular depression, Rieger anomaly, and delays of Tooth eruption, and severe insulin resistance, often with lipodystrophy. We have generated a novel murine model expressing the truncated Pik3r1 to dissect their roles in the pathophysiology of the syndrome.

Homozygous mice were not identified, however heterozygous (Het) mice were viable and showed linear growth impairment. Compared to wild type (WT) mice, Het mice had a reduced fat gain when fed on a high fat diet, and were glucose intolerant and insulin resistant. In particular, het mice had an impaired suppression of glucose production by the liver, reduced lipolysis in the white adipose tissue and weakened deoxyglucose uptake in the skeletal muscle under insulin stimulation. Mechanistically, there was a modest impairment of Akt signalling in these tissues, despite severe insulin resistance. In future, we are investigating the hepatic lipid content and metabolism, and the non-canonical signalling roles of Pik3r1.


03 Autophagic regulation in human skeletal muscle in response to physical activity and inactivity

In study 2 (n=11), subjects had one lower limb immobilized for 2 weeks using a Donjoy Splint fixed in 40 degrees, followed by 6 weeks physical rehabilitation. Exercise increased ULK1 Ser555 phosphorylation, an inducer of autophagy, in both prandial and fasting conditions, and this was associated with decreased lipidation of LC3B. pULK1 Ser555 correlated positively with AMPKα phosphorylation at Thr172, suggesting signaling through AMPKα/ULK1 as a mechanism by which exercise induces autophagy. Immobilization reduced quadriceps femoris cross-sectional area by ~10% and this was completely restored during rehabilitation. This was associated with increased ULK1 and p62 protein expression and decreased FOXO3a phosphorylation at Ser318/321, which indicate transcriptional activation.

In accordance to this, mRNA expression of several autophagy-related genes controlled by FOXO3a (LC3B, p62, BECLIN1, BNIP3) increased during immobilization. FOXO3a phosphorylation increased to baseline levels during rehabilitation and the same did the expression of all autophagy-related genes and proteins.

These data demonstrate that autophagy is regulated through distinct mechanisms in response to physical activity and inactivity in human skeletal muscle. Transcriptional regulation of autophagy seems to be involved in regulation of skeletal muscle mass, while non-transcriptional activation of autophagy seems to serve as an integrate response to physical exercise that enables adaptation to a trained state.

A.B. Møller1,*, M.H. Vendelbo, B2. Christensen1, J.O.L. Jørgensen2, N. Møller2, P. Schjerling3, C. Couppe3, M. Kjær3, M. Hansen3,4, N. Jessen1

1 Research Laboratory for Biochemical Pathology, Department for Clinical Medicine, Aarhus University Hospital2 Medical Research Laboratory, Department for Clinical Medicine, Aarhus University Hospital3 Institute of Sports Medicine Copenhagen, Bispebjerg Hospital, DK4 Section for Sport Science, Department for Public Health, Aarhus University, DK* E-mail address of corresponding author: [email protected]

Autophagy is a catabolic process that enables protein degradation through the lysosomes, and in mice, initiation of autophagy is a prerequisite for adaptation to a trained state. In contrast, excessive autophagy can be detrimental in conditions of muscle loss, such as in critical illness. Autophagy is transcriptionally regulated by FOXO3a and non-transcriptionally regulated by ULK1.

The aim of the present study was to investigate autophagy signaling in response to physical activity and inactivity in human skeletal muscle. Skeletal muscle biopsies from two studies in healthy human subjects were examined. In study 1 subjects (n=7) performed 1 h cycling exercise at 50% VO2-max on two occasions: i) during 36 hours fast, and ii) during continuous glucose infusion to imitate a prandial condition.


04 A low-cost, hand-held breathalyser that measures the human metabolism

Apostolos Atsalakis

Department of Chemical Engineering and Biotechnology, University of Cambridge

The resting metabolic rate (RMR) counts to more than 75% of the the total energy expenditure of an individual (TEE - total energy expenditure). There are many statistical methods to predict the RMR (i.e. Schofield equations) but all of them fail dramatically in special cases (obesity, patients of hospitals, pregnant women etc.). Indirect calorimetry on the contrary, is a known technique for its accuracy in determining the human metabolism.

This study reports the proof of concept and the development of a low-cost, hand-held breath analysis device that measures the human metabolism by detecting certain biomarkers in the human breath.


05 Interactions between FTO, physical activity and insulin sensitivity; one or both?

assessment of insulin resistance (HOMAIR) was used to assess insulin sensitivity, and an interaction with FTO rs9939609 was observed (P=0.0025). The interaction was unaffected by adjustment for PA (P=0.0028). Dividing the population at the median HOMAIR showed a per risk-allele effect of FTO rs9939609 of 0.35 kg/m2 in the more insulin sensitive group and 0.57 kg/m2 in the less insulin sensitive group. On the contrary the FTOxPA interaction (P=0.004) was attenuated by adjustments for HOMAIR (P=0.07).

These results led us to the conclusion that the established interaction between variants in FTO and PA might be driven by a decrease in insulin sensitivity.

Camilla H. Sandholt1, Kristine H. Allin1, Allan Linneberg2,3, Charlotta Pisinger2, Oluf Pedersen1,4

and Torben Hansen5

1The Novo Nordisk Foundation Center for Basic Metabolic Research, Section of Metabolic Genetics, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, DK2Research Centre for Prevention and Health, the Capital Region of Denmark, Copenhagen, DK 3Department of Clinical Experimental Research, Rigshospitalet, Glostrup, DK4Faculty of Health Sciences, University of Aarhus, Aarhus, DK5Faculty of Health Sciences, University of Southern Denmark, Odense, DK

Gene-environment (GxE) interaction analyses can be used to pinpoint genetic variants for which the exerted effect is modulated by environmental factors. Shortly after being identified in genome-wide association studies having pronounced effect on BMI, the effect of a variant in FTO was reported to be modulated by the level of physical activity (PA) as well as insulin sensitivity. The FTOxPA has subsequently been confirmed in a large meta-analysis, whereas, no further elaborations have been made on the interaction with insulin sensitivity.

We aimed at confirming this interaction between FTO and insulin sensitivity in five Danish population-based cohorts (Ntotal=12,274). Moreover, we aimed to test whether this interaction reflects the same underlying mechanism as the interaction with PA, due to the tight correlation between these two phenotypes. The homeostatic model


06 The role of the bone morphogenetic protein 9 in atherosclerosis

Claudia-Gabriela Mitrofan, Sarah Appleby, Nick Morrell

Department of Medicine, Addenbrooke’s Hospital, University of Cambridge, Cambridge, UK

Atherosclerosis is characterised chronic inflammation in the vascular wall. Bone morphogenetic protein (BMP) signalling has been associated with atherosclerosis. These adverse effects are thought to be driven by the BMP ligands 2, 4 and 6, via the type-I Activin like kinase (ALK) receptors ALK2, ALK3 and ALK6. Type I receptors combine with type II receptors to form a signalling complex. Of these, the bone morphogenetic protein receptor II (BMPR-II) has been implicated in atherosclerosis. The endothelial BMP receptor, ALK1, in complex with BMPR-II, signals specifically in response to the circulating BMP9, which is thought to act as a vascular quiescence factor. We therefore questioned whether BMP9 confers protection against endothelial inflammation by maintaining endothelial integrity and homeostasis and regulating monocyte activation.

BMP9 concentration-dependently increased monocyte recruitment to TNF-α-stimulated HAECs. BMP9 also increased E-selectin and VCAM-1 surface expression on TNF-α-stimulated endothelial cells. This effect occurred using BMP9 concentrations greater than 1.5 ng/mL. Knockdown of ALK2, but not of ALK-1, reduced the surface expression levels of adhesion molecules.

Pre-treating the endothelium with high concentrations of BMP9 followed by TNF-α treatment enhanced the inflammatory response through increased expression of selectins and adhesions molecules and ultimately by upregulating monocyte recruitment. We propose that at high concentrations, BMP9 signals through the ALK2 receptor. These findings are relevant to the therapeutic use of BMP9 in atherosclerosis and other cardiovascular disease.


07 Ceramide and diacylglycerol content is not influenced by 10 weeks exercise training in first-degree relatives of type 2 diabetic patients

Results Insulin sensitivity were higher in Controls compared to Offsprings (Con: pre: 7.58 ± 0.8, post: 8.47 ± 0.7; Off: pre: 5.35 ± 0.4, post: 6.27 ± 0.5; p=0.008) and insulin sensitivity and VO2max increased in both groups after exercise training (p<0.05). AKTpan protein expression increased in both groups after exercise training (p=0.03). Total ceramide and DAG muscle content and subspecies measured did not differ between groups and remained unchanged after exercise training, except for a decrease in C22:0-Cer (p=0.017).

Conclusion 10 weeks exercise training improves VO2max and insulin sensitivity in Controls and Offsprings but does not alter ceramide and DAG content in muscle.

Søgaard D1, ,Østergård T, Baranowski M, Dela F, Helge JW

1Xlab. Center of Healthy Aging, Dept. of Biomedical Sciences, University of Copenhagen, DK

Aim Evaluate muscle ceramide and diacylglycerol (DAG) content and insulin sensitivity in first-degree relatives to type 2 diabetics and controls and the effect of 10 weeks exercise training.

Material and methods The study is taking advantage of a previous performed study by Østergård T. et al. The exercise protocol involved 10 weeks exercise training performed 3 times a week 45 min. at 70% of VO2max on a bicycle ergometer. Before and after 10 weeks exercise intervention a VO2max test and a hyperinsulinemic-euglycemic clamp (1.0 mU ● kg-1 ● min-1) were carried out and muscle biopsies and blood samples obtained. We selected the highest possible number of available biopsies, 16 control and 19 first degree offsprings to type 2 diabetic patients. We applied lipidomics to analyze the content of ceramide and DAG in skeletal muscle and performed Western blot to measure key enzymes in ceramide and DAG metabolism and proteins in insulin signaling.


08 The identification of novel metabolites that track with improvements in glycaemia following a 12-week lifestyle intervention in high risk individuals

Donal O’Gorman1, Aileen Kennedy1, Jeff Cobb2, Grainne O’Donoghue4, Gregers Anderson4, Noel McCaffrey1, Stephen Cleary1, Eoin Durkan1, Helena Kenny1, Klaus-Peter Adam2, Margaret Sinnot3, Bernadette Carr3, Tanja Thybo4, John Nolan4.

13U Diabetes Partnership and School of Health and Human Performance, Dublin City University, Ireland2Metabolon, Inc., 617 Davis Drive, Suite 400, Durham N.C., U.S 3Vhi Healthcare, Lower Abbey St., Dublin, Ireland 4Steno Diabetes Centre, Copenhagen, Denmark

Background and aimsA number of studies have identifed fasting metabolites associated with insulin resistance, dysglycemia and type 2 diabetes. It is not known if metabolites respond to lifestyle modifications that reduce the risk of type 2 diabetes. The purpose of this study was to identify metabolite biomarkers, linked to dysglycaemia, that track with improvements in plasma glucose following a lifestyle intervention in high risk individuals.

Materials and methodsAs part of the DEXLIFE programme, individuals with a FINDRISC score >12 or impaired fasting/2-hr glucose volunteered to participate in a 12-week diet and exercise intervention (n=104). An oral glucose tolerance test, body composition assessment (DEXA) and maximal aerobic capacity were determined before and after the intervention.

Using the stable isotope dilution technique, quantitative assays were developed for a set of 23 candidate biomarker metabolites previously linked to dysglycemia. This set included: α-hydroxybutyric acid (AHB), linoleoylglycerophosphocholine (LGPC),

oleic acid, α-ketoglutaric acid, 2-aminoadipic acid, glycine, aromatic amino acids, and the 3 branched-chain amino acids and several of their catabolites.

ResultsAfter the 12-week intervention fasting levels of 12 of the 23 metabolites were significantly different (p<0.05). In subjects where fasting glucose decreased by >10% (n=16) there was a significant decrease in plasma tyrosine, α-ketoglutarate and phenyalanine (p<0.05) as well as increased glycine and serine (p<0.05).

A >10% decrease in 2-hr glucose (n=35) was associated with significant decreases in branched-chain amino acid catabolites 3-methyl-2-oxopentanoic acid, 3-methyl-2-oxo-butyric acid, 4-methyl-2-oxopentanoic acid (p<0.05) in addition to insulin, α-ketoglutarate, tyrosine (p<0.05) and increased glycine (p<0.05).

The fold change in body weight was positively associated with the fold change in phenylalanine, tyrosine, leucine, isoleucine, 3-methyl-2-oxopentanoic acid and insulin (p<0.05) and negatively associated with glycine (p<0.05). The changes in aerobic fitness and % body fat were not associated with any of the metabolites.

ConclusionA subset of metabolites linked to dysglycaemia track with improvements in fasting and 2-hr glucose following a 12-week lifestyle intervention in high risk individuals. These metabolites are sensitive to small changes in metabolic function and may be useful for monitoring diabetes prevention programmes. Supported: DEXLIFE project (EU FP7 programme, Grant agreement no: 279228).


09 Improved insulin sensitivity decrease the genetic effect on triglyceride levels in blood

ResultsIn a fully adjusted model including changes in alcohol intake, smoking, physical activity and diet, each risk variant associated with increased triglyceride levels over 5 years (per allele effect=1.48% [1.18–1.78]; P=3.8×10-22). This triglyceride increasing effect was stronger in individuals decreasing their insulin sensitivity over the 5 year period (Pinteraction=3.29×10-5).

ConclusionIncreased genetic risk is associated with a larger increase in triglyceride levels during 5 years. The genetic risk of increased triglycerides is modifiable by insulin sensitivity, and we show that individuals carrying a high number of triglyceride-increasing genetic variants can reduce the effect of this unfavorable genetic predisposition by improving their insulin sensitivity.

Ehm A. Andersson

The Novo Nordisk Foundation Center for Basic Metabolic Research, Section of Metabolic Genetics, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, DK

High levels of circulating triglycerides are independent strong risk factor for coronary artery disease (CAD). Blood levels of triglycerides are affected by obesity and glucose homeostasis as well as lifestyle factors. In addition, triglyceride levels are highly heritable, with ~44 common genetic variants identified. The aim of the present study is to investigate the association between genetic risk of increased triglyceride on changes in triglyceride over 5 years and to test whether the triglyceride-increasing effect is modified by adiposity, insulin resistance and lifestyle factors that themselves are changing over time.

Material and methodsThe Danish Inter99 population was genotyped by metabo-chip and exome-chip and the relevant genetic variants were extracted. At baseline and after 5 years participants were enrolled for a physical examination including anthropometric and biochemical measures, as well as a 2-hour oral glucose tolerance test. Lifestyle data was self-reported based on questionnaires. A genetic risk score for triglyceride increasing variants was constructed. Linear regression was used to test for 5-year changes in triglycerides and interactions with other parameters.


10 Do genes associated with obesity cause the dysfunction of hypothalamic POMC and AGRP neurons?

Florian Merkle, Dr.


Obesity is associated with the aberrant function of specific neurone types in the hypothalamus.

I recently developed a method to generate the relevant hypothalamic neurone populations from human pluripotent stem cells. I also have developed efficient methods to engineer candidate mutations into the genomes of these cells. I am now combining these tools to generate in vitro disease models of human obesity and metabolic disease.

Specifically, I will test the hypothesis that genes associated with obesity cause the dysfunction of hypothalamic POMC and AGRP neurones.


11 Central carbon metabolism regulates circadian oscillations

Transcriptomics analyses revealed that such metabolic perturbations had a widespread impact on circadian gene expression. Moreover, the inhibition of the PPP affected the genome-wide DNA-binding activity of the circadian transcription factors BMAL1 and CLOCK, possibly through a mechanism involving histone acetylation and chromatin remodelling. Thus, manipulation of central carbon metabolism disrupts circadian rhythms, implicating NADPH production in the generation of circadian oscillations and demonstrating that redox reactions are one connection between transcriptional and metabolic circadian cycles in nucleated cells.

Guillaume Rey1, Kevin A. Feeney2, Utham K. Valekunja1, Nikolay B. Milev1, Lisa Wulund1, Alessandra Stangherlin1, Laura Bollepalli1, Vidya Velagapudi3, John S. O’Neill2, Akhilesh B. Reddy1

1 University of Cambridge Metabolic Research Laboratories, Wellcome Trust-MRC Institute of Metabolic Science, University of Cambridge, Addenbrooke’s Hospital, Cambridge CB2 0QQ, UK 2 MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge, CB2 0QH, UK3 Metabolomics Unit, Institute for Molecular Medicine Finland FIMM, Helsinki, Finland

The circadian clock is a ubiquitous timekeeping system that organizes the behaviour and physiology of organisms over the day and night. The clockwork orchestrates a multitude of metabolic processes as illustrated by previous global transcriptomics and proteomics studies, and the existence of daily rhythms of reduction and oxidation (redox) in a range of diverse species. However, the reciprocal question of whether metabolism can alter the clockwork remains unaddressed.

Here we identify the pentose phosphate pathway (PPP), a critical source of cellular reducing power in the form of NADPH, as a key modulator of the clock’s transcriptional oscillations. We systematically manipulated genes in this pathway and identified several enzymes in the oxidative branch of the PPP that affected circadian oscillations in a dose-dependent manner. Pharmacological inhibition of the PPP confirmed these effects in human cells, in mouse tissues and in Drosophila.


12 Whole-genome sequencing reveals low frequency variants associated with body mass index and height

Ioanna Tachmazidou on behalf of the UK10K anthropometric traits team

Wellcome Trust Sanger Institute, Hinxton, Cambridge, UK

Genome-wide association studies have identified many common genetic loci associated with anthropometric traits, but the role of low frequency and rare variants is still unknown. As part of the UK10K study, we investigated the genetic architecture of 12 anthropometric traits in 3,559 individuals with ~7x whole genome sequence (WGS) data from the ALSPAC and TwinsUK cohorts.

Variants discovered through WGS, along with those from the 1000 Genomes Project (1KG), were imputed into additional individuals from the ALSPAC and TwinsUK cohorts with GWAS data, as well as other existing external GWAS cohorts, increasing the total sample size to 53,500.

We investigated association between anthropometric traits and ~11.7 million variants with MAF≥0.05 and ~155 million variants with MAF 0.001-0.05. We observe a significant excess of independent not previously reported variants with p.


13 Fibro-adipogenic progenitors in response to acute and prolonged resistance exercise

Moreover, the content of CD90+ cells appears to increase following prolonged exercise, in a pattern similar to the increase in satellite cells. These results indicate that human FAPs be involved in the regulation of the muscle microenvironment following exercise.

Jean Farup1, Ulla Ramer Mikkelsen1,2, Jonas Bjerre1, Stine Klejs Rahbek1, Frank de Paoli2, Mikkel Holm Vendelbo3, Peter Schjerling2, Kristian Vissing1

1 Section of Sport Science, Dept. of Public Health, Aarhus University, DK2 Institute of Sports Medicine, Department of Orthopaedic Surgery M, Bispebjerg Hospital and Center for Healthy Aging, Faculty of Health and Medical Sciences, University of Copenhagen, DK3Dept. of Biomedicine, Aarhus University, DK3Dept. of Internal Medicine and Endocrinology, Aarhus University Hospital, DK

The mesenchymal cell population, the fibro-adipogenic progenitors (FAPs), has recently been identified and characterized in rodent and human skeletal muscle. In rodents, the FAPs appear to play a pivotal role for functional muscle regeneration as well as prevention atrophy during various pathological conditions, while also contributing to fibrosis and adipocyte accumulation when regeneration fails.

Presently, the FAP response to a physiological stimulus in human skeletal muscle, such as acute or prolonged exercise training, is unknown. In these preliminary experiments we show that in healthy young men the FAPs, identified by the expression of CD90 and PDGFR-α, proliferate in response to both acute and prolonged intensive resistance exercise.


14 Effects of training in vivo is preserved in cultured human myotubes

Jenny Lund1, Arild Rustan1, Jonathan Mudry2, Torgrim Langleite1, Hanne Gulseth1, Jørgen Jensen3, Hege Thoresen1

1 University of Oslo, Norway2 Karolinska Institutet, Sweden3 Norwegian School of Sport Sciences, Norway

Introduction and methodsPhysical activity has preventive as well as therapeutic benefits for overweight subjects. Most studies have been executed in vivo or on skeletal muscle biopsies. We studied effects of in vivo exercise on energy metabolism in cultured myotubes isolated from biopsies taken before and after 12 weeks of extensive endurance and strength training. Healthy sedentary men, aged 40-62 years, with normal weight (body mass index (BMI) <25 kg/m2) or overweight (BMI≥25 kg/m2) were included. Fatty acid and glucose metabolism were studied in myotubes using [14C]oleic acid and [14C]glucose, respectively. Gene and protein expressions, as well as DNA methylation were measured.

ResultsThe 12-week training intervention improved endurance, strength, and insulin sensitivity in vivo. Biopsy-derived cultured human myotubes from the overweight subjects after exercise showed increased oleic acid uptake (30%), oxidation (46%)

and lipid accumulation (34%). Furthermore, fractional glucose oxidation was increased by 14% as compared to cultures established prior to exercise. Myotubes established after exercise had decreased mRNA expression of insulin receptor substrate 1 (IRS1) (31%) and increased DNA methylation within first exon of IRS1 (23%).

Conclusion12 weeks of combined endurance and strength training promoted increased lipid and glucose metabolism in biopsy-derived cultured human myotubes, showing that effects of training in vivo are conserved in human myotubes in vitro.


15 Effect of maternal exercise during obese pregnancy on the cardiovascular health of male offspring

Jessica Beeson, Sarah Carr, Daniella Duque, Laura Dearden, Heather Blackmore, Denise Fernandez-Twinn and Susan Ozanne


The prevalence of obesity amongst women of childbearing age has increased dramatically. Children from obese mothers are at increased risk of premature death due to cardiovascular disease. Offspring demonstrate early indicators of metabolic syndrome with increased propensity to develop type 2 diabetes and to become obese.

There is good evidence to suggest that these effects are mediated by non-genetic “programmed” mechanisms. An evidence-based intervention is needed to prevent the propagation of obesity and metabolic dysfunction to the next generation. Previous studies have demonstrated that exercise has positive effects in pregnant rats by preventing excessive gestational weight gain and improving glycaemic control.

This study aims to investigate the effect of maternal exercise during obese pregnancy on the cardiovascular health of the offspring. Offspring from obese dams developed pathological cardiac hypertrophy, demonstrated by an increase in relative heart weight (0.58% (of body weight) ± 0.01 versus 0.69% ± 0.02, p


16 Prolonged activation of glycogen synthase after exercise despite substantial glycogen synthesis during recovery

José L Areta, Marius A Dahl, Per B Jeppesen, Thorsten Ingemann-Hansen, Henriette Pilegaard, Kristian Overgaard, Jørgen F.P. Wojtaszewski & Jørgen Jensen

Glycogen in skeletal muscle has a dual function. Skeletal muscle glycogen is the major energy substrate during high intensity exercise, and skeletal muscles make up the tissue with the largest capacity to dispose glucose into glycogen. Exercise improves metabolic regulation partly by reducing the glycogen content in skeletal muscles.

The aim of the present study was to investigate the effect of exhaustive exercise on glycogen synthase activation and whether adding protein to the carbohydrate intake afterwards influenced rate of glycogen resynthesis and glycogen synthase activation.

Participants cycled at ~75 % of maximal oxygen uptake until exhaustion (~110 min) twice and ingested (per kg) either 2.4 g carbohydrate (CHO) or 1.6 g carbohydrate and 0.8 g protein (CHO+PROT) during the first two hours recovery in a double-blinded randomised study. Muscle biopsies were taken before, immediately and 5 h after exercise.

Exercise decreased glycogen content from 454±90 mmol∙kg dw-1 to ~ 92 mmol∙kg dw-1. After 5 h recovery, glycogen content was similar in CHO (277±66 mmol∙kg dw-1) and CHO+PROT (279±79 mmol∙kg dw-1). Glucose and insulin responses were rather similar during the CHO and CHO+PROT intervention. Glycogen synthase was activated after exercise. Importantly, glycogen synthase activation and dephosphorylation persisted for 5 h after exercise despite substantial glycogen resynthesis in both interventions.

In conclusion, glycogen synthase phosphorylation remained low despite glycogen substantial resynthesis during the 5 h recovery and highlights a mechanism that uncouples the tight feedback inhibition of glycogen synthase by elevated glycogen content, which may explain the insulin sensitising effect of exercise.


17 Post-weaning high-fat diet exacerbates insulin resistance programmed by maternal obesity in offspring adipose tissue

The results revealed offspring weaned onto an obesogenic diet had reduced protein levels of IR-beta, IRS1 and p110beta in their adipose tissue. However levels of these proteins were further reduced in offspring exposed to maternal diet-induced obesity with the lowest levels observed in the programmed offspring that were weaned onto an obesogenic diet. Maternal diet-induced obesity increased expression of adipose tissue miR126 but levels of this miR were not influenced by a post weaning obesogenic diet. We conclude that maternal diet-induced obesity and a post-weaning obesognic diet both lead to reductions in key insulin signalling proteins such as IRS-1. Whilst programmed reductions in IRS-1 are associated with increased levels of miR126, the effects of an obesogenic diet on IRS-1 are mediated by miR126 independent mechanisms.

Financial support: FAPESP (2014/17012-4)

Juliana de Almeida Faria, Asha Carpenter, Elena Loche, Daniella Duque, Susan E Ozanne

IMS, University of Cambridge, UK

Strong evidence supports the idea that adverse prenatal and early postnatal environments have a significant long-term influence on risk of Type 2 Diabetes. Previously, we demonstrated that reduction in adipose tissue insulin signaling proteins, such as IRS-1, was one mechanism by which maternal diet-induced obesity leads to increased risk of type-2 diabetes in the offspring. This effect was mediated by increased expression of miR-126 in offspring adipose tissue by which IRS-1 may be silenced at the translational level. The aim of the current study was to establish if weaning onto an obesongenic diet exagerated the detrimental effects of maternal obesity on offspring adipose tissue and to explore potential underlying mechanisms. We used an established mouse model of maternal diet-induced obesity. Female C57Bl/6 mice were fed either a control diet or an obesogenic diet from six weeks prior to mating. Offspring were weaned at 21 days of age onto either standard or an obesogenic diet. Male offspring were studied at 8 weeks of age.


18 Reduced CD300LG mRNA tissue expression, increased intramyocellular lipid content and impaired glucose metabolism in healthy male carriers of Arg82Cys in CD300LG

Julie Støy1, Ulla Kampmann1, Annette Mengel2, Nils E. Magnusson2, Niels Jessen3, Niels Grarup4, Jørgen Rungby5,6, Hans Stødkilde-Jørgensen7, Ivan Brandslund8, Cramer Christensen9, Torben Hansen4, Oluf Pedersen4, Niels Møller1,2

1 Department of Internal Medicine and Endocrinol-ogy, Aarhus University Hospital, Aarhus, DK2 Medical Research Laboratories, Institute for Clinical Medicine, Aarhus University, Aarhus, DK3 Research laboratory for biochemical pathology, Institute for Clinical Medicine, Aarhus University, Aarhus, DK4 The Novo Nordisk Foundation Center for Basic Metabolic Research, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, DK5 Center for Diabetes Research and Department of Clinical Pharmacology, University Hospital of Copenhagen, DK 6 Department of Biomedicine, Aarhus University, Aarhus, DK 7 The MR research center, Aarhus University Hospital, Aarhus, DK8 Department of Clinical Biochemistry, Vejle Hospital, Vejle, DK9 Department of Medicine, Vejle Hospital, Vejle, DK

BackgroundCD300LG rs72836561 (c.313C>T, p.Arg82Cys) has in genetic-epidemiological studies been associated with the lipoprotein abnormalities of the metabolic syndrome. CD300LG belongs to the CD300-family of membrane-bound molecules which have the ability to recognize and interact with extracellular lipids. We tested whether this specific polymorphism results in

abnormal lipid accumulation in skeletal muscle and liver and other indices of metabolic dysfunction. Method40 healthy males with a mean age of 55 years were characterized metabolically including assessment of insulin sensitivity by the hyperinsulinemic euglycemic clamp, intrahepatic lipid (IHLC) and intramyocellular lipid content (IMCL) by MR-spectroscopy, and beta-cell function by an intravenous glucose tolerance test. Changes in insulin signaling and CD300LG mRNA expression were determined by western blotting and quantitative-PCR in muscle and adipose tissue.

ResultsCompared with the 20 controls (CC-carriers), the 20 CT-carriers had higher IMCL (p=0.045), a reduced fasting forearm glucose uptake (p=0.011), a trend towards lower M-values during the clamp; 6.0 mg/kg/min versus 7.1 (p=0.10), and higher IHLC (p=0.10). CT-carriers had lower CD300LG mRNA expression and CD300LG expression in muscle correlated with IMCL (β=-0.35, p=0.046), forearm glucose uptake (β=0.37, p=0.03), and tended to correlate with M-value (β=0.33, p=0.06), independently of CD300LG genotype. Beta cell function was unaffected.

ConclusionsThe CD300LG polymorphism was associated with decreased CD300LG mRNA expression in muscle and adipose tissue, increased IMCL and abnormalities of glucose metabolism. CD300LG mRNA levels correlated with IMCL and forearm glucose uptake. These findings link a specific CD300LG polymorphism with features of the metabolic syndrome suggesting a role for CD300LG in the regulation of common metabolic traits.


19 Low physical activity is associated with increased arterial stiffness in patients recently diagnosed with type 2 diabetes

(cpm), Actiheart®) worn by the participants for up to six days. High vs. low levels of physical activity was defined according to the median level of activity (cpm=31).

Results 65 patients and 65 controls were included in the final analysis (median age 59 years, 55% men, median diabetes duration 1.9 years). Participants with low physical activity had higher cfPWV compared to participants with high physical activity: A) Patients and controls combined: 9.3±1.7 m/s vs. 7.8±1.5m/s, p<0.001 B) Patients with diabetes: 9.5±1.8m/s vs. 8.3±1.6m/s, p=0.02 and C) Controls: 9.0±1.4m/s vs. 7.7±1.4m/s, p<0.01). The difference remained significant after adjustment for other determinants of cfPWV including whole body fat percentage (p<0.01). No significant interaction between diabetes and the effect of low activity was seen.

Conclusions Low physical activity is associated with increased arterial stiffness in patients recently diagnosed with type 2 diabetes and in healthy controls.

1K. L. Funck, MD; 1,2,3E. Laugesen, PhD; 1P. Høyem, PhD; J. 1Fleischer Ass. Prof.; 1S. L. Cichosz, MSc.; 1J. S. Christiansen, Prof.; 1T. K. Hansen, Prof.; 1P. L. Poulsen, MD, PhD, DmSci.

1 Department of Endocrinology and Internal Medicine and Medical Research Laboratories, Aarhus University Hospital, Nørrebrogade 44, 8000 Aarhus, DK 2 Department of Clinical Medicine, Aarhus University, Nordre Ringgade 1, 8000 Aarhus C 3 Danish Diabetes Academy, Odense University Hospital, Søndre Blvd. 29, 5000 Odense

Aims Several studies have indicated that low physical activity is associated with increased risk of cardiovascular disease (CVD) and all-cause mortality among patients with diabetes. The association between physical activity and subclinical cardiovascular changes preceding clinical events remains to be elucidated. We investigated the relationship between physical activity and arterial stiffness, an independent predictor of CVD, in patients with type 2 diabetes and controls.

Methods We included 100 patients with type 2 diabetes and 100 sex and age matched controls in a cross-sectional study. Arterial stiffness (carotid-femoral pulse wave velocity, cfPWV) was measured using the SphygmoCor® device. Physical activity was assessed by an accelerometer (count per minute


20 Inorganic nitrate increases exercise efficiency, tolerance, and performance, and is growing in popularity as a nutritional sport supplement

Lee D Roberts, Dr.

MRC Human Nutrition Research Unit, University of Cambridge

Peroxisome proliferator-activated receptor γ coactivator1α (PGC1α), a master transcriptional regulator of the muscle adaptive response to endurance exercise, coordinates the expression of typical oxidative slow twitch type I and type IIa skeletal muscle fiber genes. We find that nitrate increases the expression of PGC1α and markers of type I and IIa muscle fibers in rats in vivo and myotubes in vitro via a nitrate-nitric oxide-cGMP mediated mechanism. Nitrate-stimulated fibre-type switching is enhanced by both HIF2α stabilization and the phosphodiesterase 5 inhibitor, sildenafil. Nitrate increases the expression and secretion of exercise and PGC1α-dependent myokines FNDC5/irisin and β-aminoisobutyric acid in myotubes, rats and humans.

Both exercise and nitrate stimulated PGC1α-mediated γ-aminobutyric acid (GABA) secretion from muscle. GABA plasma concentrations were increased in exercising mice and nitrate treated rats and humans. GABA may function as an exercise/PGC-1α-mediated myokine. Both exercise training and GABA are known to increase plasma growth hormone concentrations. Nitrate treatment of humans and rodents increased circulating growth hormone levels.

Thus nitrate induces physiological responses that mimic exercise training and may underlie the beneficial effects of the molecule on exercise and cardiometabolic health.


21 Impaired lipid storage and mitochondrial network organization in morbid obese human skeletal muscle. Improvements associated to gastric bypass and link to insulin resistance remission

months after RYGB, the structural organization of LDs had improved in 5 of the 10 obese subjects, while the structural organization and content of mitochondria had improved in 7 of 10 subjects. Subjects with a good IS recovery seemed to have a higher degree of mitochondrial structure improvement than poor IS responders. Three of 5 lean sedentary controls presented impaired mitochondrial dynamics, but no clear alterations in LD structure.

Our results indicate that physical inactivity leads to alterations in mitochondrial dynamics preceding impairments in LD quality and, RYGB-induced improvement of mitochondrial dynamics is a prerequisite for skeletal muscle LD quality and IS improvement. We propose that mitochondrial dynamics are key regulators of metabolic health.

Kristensen, MD; Petersen, SM; Møller, KE; Lund, MT; Hansen, M; Helge, JW; Dela, F; Prats, C

Obesity-induced insulin resistance (IR) has been proposed to be caused by lipotoxicity arising from ectopic lipid depositions and mitochondrial dysfunction in skeletal muscle. We hypothesized that the quality more than the quantity of intramuscular lipid deposition and mitochondrial network organization is key in IR development.

The aim of the present project therefore was to compare the amount and structural quality of lipid droplets (LDs) and mitochondrial network in skeletal muscle from morbid obese normoglycemic (NGT, n=5, BMI= 42.1 +/-1.1) and type 2 diabetic patients (n=5, BMI=43.4 +/-2.9) with lean NGT (n=5, BMI=22.0 +/-0.7), and furthermore relate these features to improvements in insulin sensitivity (IS) during a Roux-en-Y gastric bypass (RYGB)-induced weight loss.

The structural organization of both LDs and mitochondrial networks were severely disrupted in all obese subjects before the intervention. Four


22 Genomic, transcriptomic, and metabolic variations shape the complex adaptation landscape of bacteria to varying environmental conditions

Max Conway

University of Cambridge Computer Laboratory

lucidating the genotype-phenotype relation paves the way for the prediction of such effects, but methods for characterizing the relationship between multiple environmental factors are still lacking. Here, we tackle the problem of extracting meaningful information from collections of environmental conditions, by integrating the multiple omic levels on which the bacterial response is measured. To this end, we model a large compendium of growth conditions as a multiplex network consisting of transcriptomic and fluxomic layers, and we propose a multi-omic network approach to infer similarity of growth conditions by integrating layers of the multiplex network.

Each node of the network represents a single condition, while edges are similarities between conditions, as measured by phenotypic and transcriptomic properties on different layers of the network.

We then fuse these layers into one network, therefore capturing a global network of conditions and the associated similarities across two omic levels.

We apply this multi-omic fusion to an updated genome-scale reconstruction of E. Coli that includes underground metabolism and new gene-protein-reaction associations.

Our method can be readily used to evaluate and cross-compare different collections of conditions among different species. Acquiring multi-omic information on the topology of the space of experimental conditions makes it possible to infer the position and to build condition-specific models of untested or incomplete profiles for which experimental data is not available.


23 Identification of novel genetic determinants of erythrocyte membrane fatty acid composition among Greenlandic Inuit

Samples from 2,626 Greenlandic Inuit were genotyped on the MetaboChip and enriched for variants with imputation. Applying a linear mixed model we identified six independent association signals. Novel loci were identified on chromosomes 5 and 11 showing the strongest association with oleic acid (rs76430747 in ACSL6, beta (SE): -0.386 (0.034), p=1.8x10-28) and docosahexaenoic acid (rs6035106 in DTD1, 0.137 (0.025), p=6.4x10-8), respectively. For a missense variant (rs80356779) in CPT1A, we identified a number of novel FA associations, the strongest with 11-eicosenoic acid (0.473 (0.035), p=2.6x10-38), and for variants in FADS2 (rs174570), LPCAT3 (rs2110073), and CERS4 (rs11881630) we replicated known FA associations.

Moreover, we observed metabolic implications of the ACSL6 (rs76430747) and CPT1A (rs80356779) variants, which were both associated with altered HbA1c (0.051 (0.013), p=5.6x10-6 and -0.034 (0.016), p=3.1x10-4, respectively), and the latter variant was also associated with reduced insulin resistance (HOMA-IR, -0.193 (0.050), p=3.8x10-6), and measures of decreased body size, including weight (-2.676 (0.523), p=2.4x10-7).

In conclusion, we have identified novel genetic determinants of FA composition in phospholipids in erythrocyte membranes, and have shown examples of links between genetic variants associated with altered FA levels and changes in metabolic traits.

Mette K. Andersen1*, Emil Jørsboe2*, Camilla H. Sandholt1, Niels Grarup1, Marit E. Jørgensen3, Nils J. Færgeman4, Peter Bjerregaard5, Oluf Pedersen1, Ida Moltke2,6, Torben Hansen1,7, Anders Albrechtsen2

1 Section for Metabolic Genetics, The Novo Nordisk Foundation Center for Basic Metabolic Research, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, DK 2 The Bioinformatics Centre, Dept. of Biology, University of Copenhagen, Copenhagen, DK 3 Steno Diabetes Center, Gentofte, Denmark.4 Villum Center for Bioanalytical Sciences, Dept. of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, DK5 National Institute of Public Health, University of Southern Denmark, Copenhagen, DK 6 Dept. of Human Genetics, University of Chicago, Chicago, Illinois, USA7 Faculty of Health Sciences, University of Southern Denmark, Odense, DK

Fatty acids (FAs) are important for normal body function, and perturbation of FA balance has been linked to metabolic disturbances. The individual levels of FAs are affected by diet, lifestyle, and genetic variation. We aimed to improve the understanding of the regulation of FAs, by identifying genetic loci associated with inter-individual differences in 22 FAs in erythrocyte membranes.


24 Anabolic compounds during human catabolic and inactive conditions

Rittig N1, Bach E1, Thomsen HH1, Møller AB2, Johannsen M3, Jensen E4, Serena A4, Jørgensen OJ1, Richelsen B1, Jessen N1, 2, and Møller N1

1 Department of Internal Medicine and Endocrinology (MEA) and Medical Research Laboratory, Aarhus University Hospital, Nørrebrogade 44, DK-8000 Aarhus C, DK 2 Research Laboratory for Biochemical Pathology, Institute for Clinical Medicine, Aarhus University Hospital, Nørrebrogade 44, DK-8000 Aarhus C, DK 3 Section for Forensic Chemistry, Bioanalytical unit, Aarhus University, Palle Juul-Jensens Boulevard 99, 8200 Aarhus N, DK 4 Arla Foods Ingredients Group P/S, Sønderhøj 10-12, 8260 Viby J, DK

BackgroundProtein beverages rich in leucine or its keto-metabolite β-hydroxy-β-methylbutyrate (HMB) are especially anabolic in combination with exercise. It is uncertain if these effects include catabolic and inactive conditions. AimsTo investigate i) whether intravenous amino acid supplementation has anabolic effects during an acute inflammatory state (endotoxin/LPS administration) at rest and ii) to compare leucine-rich whey protein (WH) with isocaloric carbohydrate- (CHO), soy protein (SOY), and soy protein +3 g HMB (HMB) during fasting-induced catabolic and inactive conditions.

Methods Two trials were conducted using eight healthy lean male subjects in a randomized crossover trial. We applied the forearm model using isotopic phenylalanine tracers to express muscle metabolism during the different interventions. Muscle biopsy samples and indirect calorimetry were used to investigate muscle tissue signaling and estimating energy expenditure (EE) and oxidation rates.

ResultsAmino acid supplementation improved whole-body and muscle protein kinetics during endotoxin-induced inflammatory conditions. The four nutritional beverages all improved muscle protein kinetics following 36 hours of fasting, but most abundantly so for WH and HMB. Muscle protein phosphorylation of mammalian target of rapamycin (mTOR) and its downstream targets eukaryotic translation factor 4E-binding protein 1 (4EBP1) and ribosomal S6 kinase 1 (S6) were distinctly increased during WH consumption (p<0.05). Both WH and HMB had higher EE, higher lipid oxidation rates, and lower glucose oxidation rates compared with CHO. ConclusionProtein supplementation has anabolic effects during different catabolic and inactive conditions and compounds containing WH and/or HMB seem to have superior anabolic effects on muscle but trough distinct mechanisms.


25 High intensity interval training improves glycaemic control and pancreatic β cell function of type 2 diabetes patients

ResultsIn the T2D-group, glycaemic control as determined by average fasting venous glucose concentration (p = 0.01), end point 2-hour OGTT (p = 0.04) and glycosylated haemoglobin (p = 0.04) were significantly reduced. Pancreatic homeostasis as determined by homeostatic model assessment of insulin resistance (HOMA-IR) and HOMA β cell function (HOMA-%β) were both significantly ameliorated (p = 0.03 and p = 0.03, respectively). Whole body insulin sensitivity as determined by the disposition index (DI) was significantly increased (p = 0.03). During OGTT, the glucose continuum was significantly reduced at -15 (p = 0.03), 30 (p = 0.03) and 120 min (p = 0.03) and at -10 (p = 0.003) and 0 min (p = 0.003) with an additional improvement (p = 0.03) of its 1st phase (30 min) area under curve (AUC). Significant abdominal fat mass losses were seen in both groups (T2D: p = 0.004 and CON: p = 0.02) corresponding to a percentage change of -17.84%±5.02 and -9.66%±3.07, respectively.

ConclusionThese results demonstrate that HIIT improves overall glycaemic control and pancreatic β cell function in T2D patients. Additionally, both groups experienced abdominal fat mass losses. These findings demonstrate that HIIT is a health beneficial exercise strategy in T2D patients.

Søren Møller Madsen1, Anne Cathrine Thorup1, Kristian Overgaard2, Per Bendix Jeppesen1

1 Department of Endocrinology and Internal Medicine, Aarhus Sygehus THG, Aarhus University Hospital, Aarhus C, DK2 Section of Sport Science, Dept. of Public Health, Aarhus University, Aarhus C, DKE-mail: [email protected]

Physical activity improves the regulation of glucose homeostasis in both type 2 diabetes (T2D) patients and healthy individuals, but the effect on pancreatic β cell function is unknown. We investigated glycaemic control, pancreatic function and total fat mass before and after 8 weeks of low volume high intensity interval training (HIIT) on cycle ergometer in T2D patients and matched healthy control individuals. Study design/methodElderly (56 yrs±2), non-active T2D patients (n = 10) and matched (52 yrs±2) healthy controls (CON) (n = 13) exercised 3 times (10×60 sec. HIIT) a week over an 8 week period on a cycle ergometer. Participants underwent a 2-hour oral glucose tolerance test (OGTT). On a separate day, resting blood pressure measurement was conducted followed by an incremental maximal oxygen uptake (O2max) cycle ergometer test. Finally, a whole body dual X-ray absorptiometry (DXA) was performed. After 8 weeks of training, the same measurements were performed.


26 Transcriptional regulators of metabolic adaptation in muscle are robustly induced by acute exercise in type 2 diabetics. No evidence of exercise resistance

Rugivan Sabaratnam1, 2, Andreas J. Pedersen, Tilde Eskildsen, Jonas Kristensen, Jørgen F. Wojtaszewski, Kurt Højlund

1 Section of Molecular Diabetes and Metabolism, Institute of Molecular Medicine and Institute of Clinical Research, University of Southern Denmark, Odense, DK2 Department of Endocrinology, Odense University Hospital Odense, Denmark, J. B. Winslovs Vej 21. 3, 5000 Odense, [email protected]

Background and aimsType 2 diabetes (T2D) is characterized by insulin resistance. An impaired response to exercise and recovery may contribute to insulin resistance in T2D. We hypothesized that key transcriptional regulators of metabolic adaptation in skeletal muscle show an attenuated response to exercise in patients with T2D.

Material and methodsWe investigated the effect of acute endurance exercise (60 min, 70% VO2max) on the mRNA expression (quantitative RT-PCR) of the NR4A subfamily and other key transcription factors and protein abundance/phosphorylation of mitogen-activated protein kinases (MAPKs), such as p38 and ERK1/2 in skeletal muscle biopsies from T2D patients (n=13) and weight-matched controls (n=14).

ResultsAcute exercise robustly increased the expression of the NR4A family (~4-40-fold) as well as ATF3,

CTGF, CYR61, EGR1, JUNB, SIK1, PPARA, and PPARG (~1.5-10 fold) with no differences between the groups. In recovery from exercise, the expression of PGC-1α also increased (~6-fold), and the expression of NR4A1 (~7-fold) and NR4A3 (~70-fold) increased further compared to baseline. However, most transcripts known as early immediate genes decreased or even returned to baseline levels in recovery. No differences in recovery from exercise were seen between the groups. Acute exercise increased protein content (ERK1) and phosphorylation of p38 and ERK1/2 in both groups.

ConclusionIn conclusion, acute exercise elicits pronounced increases in expression of key regulators of muscle metabolic adaptation and phosphorylation of MAPKs in T2D patients and weight-matched controls with no evidence of exercise resistance in patients with T2D.


27 Are the effects of adiposity loci modified by cardiorespiratory fitness among Danish adults?

weight was correlated with adiposity, whereas VO2max per kg lean mass was independent of adiposity

ResultsThe interactions between VOβinteractionβinteractionper kg lean mass and FTO rs1558902 (BMI GRS/ body fat percentage GRS) (ββinteractionβinteraction2max per kg body weight and FTO rs1558902 (BMI GRS/ body fat percentage GRS) on BMI (BMI/ body fat percentage) were significant (βinteraction=-0.03 kg/m2, p=0.02 (=-0.0036 kg/m

2, p=0.004/ =-0.01 kg/m2, p=0.03)

but we did not find an interaction between VO2max

interaction=-0.0001 kg/m2, p=0.94 (=-0.0003 kg/m2, p=0.81/

=-0.07 kg/m2, p=0.30).

ConclusionsWhile we find a significant interaction between adiposity loci and VO2 2 22max per kg body weight, the lack of interaction with VOmax per kg lean mass suggests that the interaction is not explained by VOmax per se but confounded by body adiposity when scaling VOmax by body weight.

Theresia M. Schnurr1, Camilla H. Sandholt, Anette P. Gjesing, Anna Jonsson, Christian T. Have, Daniel Witte, Marit E. Jørgensen, Mette Aadahl, Allan Linneberg, Niels Grarup, Oluf Pedersen, Tuomas O. Kilpeläinen, Torben Hansen

1 NNF-CBMR Center for Basic Metabolic Research, University of Copenhagen Universitetsparken 1, 1st floor, DIKU building, 210 Copenhagen, [email protected]

AimThe present study aims to examine whether the effect of FTO gene and genetic risk scores for BMI and body fat percentage on adiposity are modified by cardiorespiratory fitness in Danish adults.

Materials and methodsWe performed a meta-analysis of two population-based cohorts, Health2006 (n=2586) and ADDITION-PRO (n=716), where information on BMI, body fat percentage, aerobic fitness, and common genetic variants known to associate with adiposity were available. Two weighted genetic risk scores (GRS) were created using 97 independent variants known to be associated with BMI (BMI GRS) and 12 independent variants shown to be associated with body fat percentage (body fat percentage GRS). Linear regression models were used to test for FTO × VOaerobic fitness as VO22VO. The analyses were adjusted for age and gender. 2max and GRS x VO2max interactions to determine whether the effects of adiposity loci on adiposity are modified by aerobic fitness. All analysis were conducted for max per kg body weight and VOmax per kg lean body mass since 2max per kg body


28 Dissecting skeletal muscle mechanical stress x AMPK signalling interactions using phosphoproteomics

A. B. Madsena, B. Parkerb, E. A. Richtera, D. E. Jamesb and T. E. Jensena

a Dpt. of nutrition, exercise and sports, Univ. of Copenhagen, DK b Charles Perkins Centre, The University of Sydney, Australia [email protected]

During various exercise modalities, mechanical stress and AMPK have been proposed to activate diverse molecular signalling responses and phenotypic adaptations. Despite many known convergence points between e.g. AMPK and mechanical stress-responsive signalling with for instance synergistic effects on glucose transport and opposing effects on the control of mTORC1 activity, there are likely to be undiscovered sites of cross-talk, the detection of which might provide important insight into skeletal muscle phenotypic regulation. To reveal novel substrates in mechanical stress x AMPK crosstalk, we performed quantitative phosphoproteomic analyses using multiplexed isobaric labelling and phosphopeptide enrichment coupled to tandem mass spectrometry on incubated mouse extensor digitorum longus muscles stimulated with the AMPK activator AICAR (2 mM, 40´), passive stretching (130 mN, 15´) or combined AICAR+stretch. A total of 14.268 phosphosites were detected.

Using the Benjamini and Hochberg method to control for multiple testing controlling for 1% false discovery rate to determine phosphosites regulated ± 1.5-fold and p<0.05, we found ~2000 phospho-

sites that changed significantly in response to AICAR, stretch or combined AICAR+stretch.

Ingenuity pathway analyses suggested multi-level regulation of e.g. insulin receptor, RhoA, PKA, PI3K, autophagy, mTOR and Rab GTPase signalling by AICAR and e.g. ILK1, Ca2+, eIF4 and p70S6K, CDC42 and actin cytoskeleton signalling by stretch, including at many previously unidentified phospho-sites. Interestingly, combining AICAR+stretch had an additive effect on some phospho-sites while it had an opposing effect on others, consistent with the observation that these stimuli are synergistic or antagonistic, depending on the endpoint studied. This work is expected to provide a valuable resource for hypothesis-generation with regards to AMPK and mechanical stress-regulation of different endpoints in skeletal muscle.


29 Detecting habitual sleep in free-living physical activity records

Tom White, Kate Westgate, Nick Wareham, Soren Brage


Introduction Sleep behaviours are implicitly captured by 24 hour physical activity monitoring, but existing algorithms to segment these signals have poor validity, are not applicable to the free-living context, or are strongly coupled to a specific device. In this work we present a general purpose sleep detection algorithm and evaluate its performance with respect to a sleep diary in signals collected from the chest, wrist and thigh.

Methods 202 UK adults wore three devices for 5 days, which measured acceleration of the chest, wrist, and thigh. The algorithm searches for the section of day that optimally discriminates between highest and lowest activity across the whole signal, by fitting a repeating binary signal with a periodicity of one day. This algorithm was evaluated for each signal independently, as well as using all three simultaneously.

Results There was a trade-off between sensitivity and specificity; chest acceleration was the most specific but least sensitive, thigh acceleration was the most sensitive but least specific, wrist acceleration was in the middle.

Signal Sensitivity Specificity

Chest .86 .97

Wrist .92 .94

Thigh .94 .89

Chest, wrist and thigh

.93 .97

Discussion and Conclusion The algorithm demonstrated strong validity against the sleep diary, and performed best when multiple signals were used. The accuracy is sufficient for implementation in large cohort studies where objective physical activity was assessed by a 24 hour protocol.


Danish Diabetes AcademyFunded by the Novo Nordisk Foundation

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Abstract book: Workshop at cambridge university€¦ · Bjørn Richelsen Effect of intake of proteins high in ketogenic amino acids (e.G. Leucine) together with resistance training