Andreas Buch Møller

Key points This is the first long‐term human clinical trial to report on effects of nicotinamide riboside (NR) on skeletal muscle mitochondrial function, content and morphology. NR supplementation decreases nicotinamide phosphoribosyltransferase (NAMPT) protein abundance in skeletal muscle. NR supplementation does not affect NAD metabolite concentrations in skeletal muscle. Respiration, distribution and quantity of muscle mitochondria are unaffected by NR. NAMPT in skeletal muscle correlates positively with oxidative phosphorylation Complex I, sirtuin 3 and succinate dehydrogenase. Abstract Preclinical evidence suggests that the nicotinamide adenine dinucleotide (NAD + ) precursor nicotinamide riboside (NR) boosts NAD + levels and improves diseases associated with mitochondrial dysfunction. We aimed to determine if dietary NR supplementation in middle‐aged, obese, insulin‐resistant men affects mitochondrial respiration, content and morphology in skeletal muscle. In a randomized, placebo‐controlled clinical trial, 40 participants received 1000 mg NR or placebo twice daily for 12 weeks. Skeletal muscle biopsies were collected before and after the intervention. Mitochondrial respiratory capacity was determined by high‐resolution respirometry on single muscle fibres. Protein abundance and mRNA expression were measured by Western blot and quantitative PCR analyses, respectively, and in a subset of the participants (placebo n = 8; NR n = 8) we quantified mitochondrial fractional area and mitochondrial morphology by laser scanning confocal microscopy. Protein levels of nicotinamide phosphoribosyltransferase (NAMPT), an essential NAD + biosynthetic enzyme in skeletal muscle, decreased by 14% with NR. However, steady‐state NAD + levels as well as gene expression and protein abundance of other NAD + biosynthetic enzymes remained unchanged. Neither respiratory capacity of skeletal muscle mitochondria nor abundance of mitochondrial associated proteins were affected by NR. Moreover, no changes in mitochondrial fractional area or network morphology were observed. Our data do not support the hypothesis that dietary NR supplementation has significant impact on skeletal muscle mitochondria in obese and insulin‐resistant men. Future studies on the effects of NR on human skeletal muscle may include both sexes and potentially provide comparisons between young and older people.

Data from transgenic animal models suggest that exercise-induced autophagy is critical for adaptation to physical training, and that Unc-51 like kinase-1 (ULK1) serves as an important regulator of autophagy. Phosphorylation of ULK1 at Ser(555) stimulates autophagy, whereas phosphorylation at Ser(757) is inhibitory. To determine whether exercise regulates ULK1 phosphorylation in humans in vivo in a nutrient-dependent manner, we examined skeletal muscle biopsies from healthy humans after 1-h cycling exercise at 50% maximal O2 uptake on two occasions: 1) during a 36-h fast, and 2) during continuous glucose infusion at 0.2 kg/h. Physical exercise increased ULK1 phosphorylation at Ser(555) and decreased lipidation of light chain 3B. ULK1 phosphorylation at Ser(555) correlated positively with AMP-activated protein kinase-α Thr(172) phosphorylation and negatively with light chain 3B lipidation. ULK1 phosphorylation at Ser(757) was not affected by exercise. Fasting increased ULK1 and p62 protein expression, but did not affect exercise-induced ULK1 phosphorylation. These data demonstrate that autophagy signaling is activated in human skeletal muscle after 60 min of exercise, independently of nutritional status, and suggest that initiation of autophagy constitutes an important physiological response to exercise in humans.

Acute inflammation, and subsequent release of bacterial products (e.g. LPS), inflammatory cytokines, and stress hormones, is catabolic, and the loss of lean body mass predicts morbidity and mortality. Lipid intermediates may reduce protein loss, but the roles of free fatty acids (FFAs) and ketone bodies during acute inflammation are unclear.We aimed to test whether infusions of 3-hydroxybutyrate (3OHB), FFAs, and saline reduce protein catabolism during exposure to LPS and Acipimox (to restrict and control endogenous lipolysis).A total of 10 healthy male subjects were randomly tested 3 times, with: 1) LPS, Acipimox (Olbetam) and saline, 2) LPS, Acipimox, and nonesterified fatty acids (Intralipid), and 3) LPS, Acipimox, and 3OHB, during a 5-h basal period and a 2-h hyperinsulinemic, euglycemic clamp. Labeled phenylalanine, tyrosine, and urea tracers were used to estimate protein kinetics, and muscle biopsies were taken for Western blot analysis of protein metabolic signaling.3OHB infusion increased 3OHB concentrations (P < 0.0005) to 3.5 mM and decreased whole-body phenylalanine-to-tyrosine degradation. Basal and insulin-stimulated net forearm phenylalanine release decreased by >70% (P < 0.005), with both appearance and phenylalanine disappearance being profoundly decreased. Phosphorylation of eukaryotic initiation factor 2α at Ser51 was increased in skeletal muscle, and S6 kinase phosphorylation at Ser235/236 tended (P = 0.074) to be decreased with 3OHB infusion (suggesting inhibition of protein synthesis), whereas no detectable effects were seen on markers of protein breakdown. Lipid infusion did not affect phenylalanine kinetics, and insulin sensitivity was unaffected by interventions.During acute inflammation, 3OHB has potent anticatabolic actions in muscle and at the whole-body level; in muscle, reduction of protein breakdown overrides inhibition of synthesis. This trial was registered at clinicaltrials.gov as NCT01752348.

Type 2 diabetes mellitus (T2DM) is associated with impaired skeletal muscle function and degeneration of the skeletal muscles. However, the mechanisms underlying the degeneration are not well described in human skeletal muscle. Here we show that skeletal muscle of T2DM patients exhibit degenerative remodeling of the extracellular matrix that is associated with a selective increase of a subpopulation of fibro-adipogenic progenitors (FAPs) marked by expression of THY1 (CD90)-the FAPCD90+. We identify platelet-derived growth factor (PDGF) as a key FAP regulator, as it promotes proliferation and collagen production at the expense of adipogenesis. FAPsCD90+ display a PDGF-mimetic phenotype, with high proliferative activity, clonogenicity, and production of extracellular matrix. FAPCD90+ proliferation was reduced by in vitro treatment with metformin. Furthermore, metformin treatment reduced FAP content in T2DM patients. These data identify a PDGF-driven conversion of a subpopulation of FAPs as a key event in the fibrosis development in T2DM muscle.

To investigate; (i) the relationship between the 5STS-test and lower extremity muscle strength and balance, and (ii) the variability of the 5STS-test in multiple sclerosis (MS) patients.22 MS patients were divided into two groups (Group A and Group B) who completed one 5STS familiarization test session and two testing sessions. In Group A, session 1 also included assessment of lower extremity muscle strength. Session 2 and 3 involved completion of two 5STS-tests and assessment of balance. In Group B, session 2 and 3 involved completion of two rounds of two 5STS-tests separated by a 30 min break.Significant correlations were found between the 5STS-test and isometric and isokinetic knee flexor and extensor muscle strength of the most affected leg (R = -0.60 to -0.77), and between the 5STS-test and balance performance (R = 0.69). Intra-assessor day-to-day variability, intra-assessor test-retest variability and intra-assessor variability within test were 25.5, 22.3, and 23.1%, respectively. Inter-assessor variability within test and inter-assessor variability were 23.4 and 5.9%, respectively.The 5STS-test is related to lower extremity muscle strength and to balance performance in MS patients. For interventional purposes, a change of >25% can be regarded as a real change.

Farup, J, Kjølhede, T, Sørensen, H, Dalgas, U, Møller, AB, Vestergaard, PF, Ringgaard, S, Bojsen-Møller, J, and Vissing, K. Muscle morphological and strength adaptations to endurance vs. resistance training. J Strength Cond Res 26(2): 398–407, 2012—Fascicle angle (FA) is suggested to increase as a result of fiber hypertrophy and furthermore to serve as the explanatory link in the discrepancy in the relative adaptations in the anatomical cross-sectional area (CSA) and fiber CSA after resistance training (RT). In contrast to RT, the effects of endurance training on FA are unclear. The purpose of this study was therefore to investigate and compare the longitudinal effects of either progressive endurance training (END, n = 7) or RT (n = 7) in young untrained men on FA, anatomical CSA, and fiber CSA. Muscle morphological measures included the assessment of vastus lateralis FA obtained by ultrasonography and anatomical CSA by magnetic resonance imaging of the thigh and fiber CSA deduced from histochemical analyses of biopsy samples from m. vastus lateralis. Functional performance measures included V̇;O2max and maximal voluntary contraction (MVC). The RT produced increases in FA by 23 ± 8% (p < 0.01), anatomical CSA of the knee extensor muscles by 9 ± 3% (p = 0.001), and fiber CSA by 19 ± 7% (p < 0.05). RT increased knee extensor MVC by 20 ± 5% (p < 0.001). END increased V̇;O2max by 10 ± 2% but did not evoke changes in FA, anatomical CSA, or in fiber CSA. In conclusion, the morphological changes induced by 10 weeks of RT support that FA does indeed serve as the explanatory link in the observed discrepancy between the changes in anatomical and fiber CSA. Contrarily, 10 weeks of endurance training did not induce changes in FA, but the lack of morphological changes from END indirectly support the fact that fiber hypertrophy and FA are interrelated.

Fasting is characterised by profound changes in energy metabolism including progressive loss of body proteins. The underlying mechanisms are however unknown and we therefore determined the effects of a 72-hour-fast on human skeletal muscle protein metabolism and activation of mammalian target of rapamycin (mTOR), a key regulator of cell growth.Eight healthy male volunteers were studied twice: in the postabsorptive state and following 72 hours of fasting. Regional muscle amino acid kinetics was measured in the forearm using amino acid tracers. Signaling to protein synthesis and breakdown were assessed in skeletal muscle biopsies obtained during non-insulin and insulin stimulated conditions on both examination days.Fasting significantly increased forearm net phenylalanine release and tended to decrease phenylalanine rate of disappearance. mTOR phosphorylation was decreased by ∼50% following fasting, together with reduced downstream phosphorylation of 4EBP1, ULK1 and rpS6. In addition, the insulin stimulated increase in mTOR and rpS6 phosphorylation was significantly reduced after fasting indicating insulin resistance in this part of the signaling pathway. Autophagy initiation is in part regulated by mTOR through ULK1 and fasting increased expression of the autophagic marker LC3B-II by ∼30%. p62 is degraded during autophagy but was increased by ∼10% during fasting making interpretation of autophagic flux problematic. MAFbx and MURF1 ubiquitin ligases remained unaltered after fasting indicating no change in protesomal protein degradation.Our results show that during fasting increased net phenylalanine release in skeletal muscle is associated to reduced mTOR activation and concomitant decreased downstream signaling to cell growth.

This case-control study was designed to investigate the gene expression profile in skeletal muscle from severely insulin resistant patients with long-standing type 2 diabetes (T2D), and to determine associated signaling pathways. Gene expression profiles were examined by whole transcriptome, strand-specific RNA-sequencing and associated signaling was determined by western blot. We identified 117 differentially expressed gene transcripts. Ingenuity Pathway Analysis related these differences to abnormal muscle morphology and mitochondrial dysfunction. Despite a ~5-fold difference in plasma insulin, we did not observe any difference in phosphorylation of AKT or AS160, although other insulin-sensitive cascades, as mTOR/4EBP1, had retained their sensitivity. Autophagy-related gene (ATG14, RB1CC1/FIP200, GABARAPL1, SQSTM1/p62, and WIPI1) and protein (LC3BII, SQSTM1/p62 and ATG5) expression were decreased in skeletal muscle from the patients, and this was associated with a trend to increased phosphorylation of the insulin-sensitive regulatory transcription factor FOXO3a. These data show that gene expression is highly altered and related to mitochondrial dysfunction and abnormal morphology in skeletal muscle from severely insulin resistant patients with T2D, and that this is associated with decreased expression of autophagy-related genes and proteins. We speculate that prolonged treatment with high doses of insulin may suppress autophagy thereby generating a vicious cycle maintaining insulin resistance.

Abstract: Astroglia‐rich cultures derived from brains of newborn rats or mice use a transport system for the uptake of creatine. The uptake system is saturable, Na + ‐dependent, and highly specific for creatine and Na + . Kinetic studies on rat cells revealed a K m value for creatine of 45 μ M , a V max of 17 nmol X h −1 X (mg of protein) −1 , and a K m value of 55 m M for Na + . The carrier is competitively inhibited by guanidinopropionate ( K i = 15 μ M ). No such transport system was found in neuron‐rich primary cultures from embryonic rat brain. It is hypothesized that creatine transport is an astroglial rather than a neuronal function.

The endocrine system participates in the regulation of the immune and neural systems and therefore hormonal factors probably play an important role in the development and course of multiple sclerosis (MS). Specifically, the hypothalamic-pituitary-adrenal (HPA) system seems crucial because (a) the inflammatory response is accompanied by HPA activation; (b) animal models with an inherited HPA defect are prone to developing experimental autoimmune encephalitis; and (c) most important, corticosteroids are still the most widely used treatment. We administered a recently developed neuroendocrine function test that combines dexamethasone suppression (1.5 mg orally at 2300 h) and corticotropin-releasing hormone (CRH) stimulation (100 micrograms i.v. at 1500 h the following day) and measured the response of plasma cortisol and corticotrophin (ACTH) secretion in 19 patients with an acute exacerbation of MS. These patients had a significantly higher mean plasma cortisol response than age-matched controls (peak minus baseline; 48.1 +/- 10.5 ng/ml [mean +/- SEM] versus 19.8 +/- 4.2 ng/ml; p < 0.05), but the corresponding ACTH values for the two groups were indistinguishable (13.4 +/- 1.4 pg/ml [mean +/- SEM] versus 11.3 +/- 1.4 pg/ml; n.s.). The response range in the patients was broader and we identified six patients with excessive cortisol release (peak minus baseline: 100.5 +/- 14.4 ng/ml [mean +/- SEM]), whereas four patients failed to respond at all. The hormonal response patterns were not related to previous treatments with corticosteroids or other immunosuppressants or to psychopathological features. These results point to a heterogeneity of HPA system function, most likely at the corticosteroid receptor level, which has clinical implications for all those treatments that affect the HPA system and the course of MS.

Background Direct measurement of whole body maximal oxygen consumption (VO2-max test) is considered the gold standard when assessing cardiorespiratory fitness. Nonetheless, the validity and reliability of the test have not been examined in persons with multiple sclerosis (PwMS). Objective To investigate the validity and reliability of VO2-max measurements in PwMS, and additionally to compare these measures to those of healthy controls. Methods Twenty PwMS completed two incremental VO2-max tests on a leg cycling ergometer. Test validity was analyzed based on the first VO2-max test in the total sample and in patient subgroups based on Expanded Disability Status Scale (EDSS) scores (EDSS ≤ 2.5, n = 10 and EDSS ≥ 3.0, n = 10) by evaluation of the primary VO2 plateau criterion and four common secondary validity criteria. Data from 20 age- and gender-matched healthy controls were used for comparison. The second VO2-max test was used to establish day-to-day reliability. Results In PwMS 40% were able to achieve the primary validity criterion for VO2-max measurements, while 65–95% were able to achieve the secondary criteria. This corresponded to the age-matched healthy controls. Strong correlations were found between measurements of VO2-max and between the validity criteria from test 1 compared to test 2 in PwMS. MS disability level did not influence criteria attainment. The variability analysis exhibited a 95% prediction interval of −238 to 201 mL·min−1 (−9.8 to 8.1%) for the difference between the two measurements of VO2-max. Conclusion In mild to moderately impaired PwMS less than half achieve the primary validity criterion when performing a VO2-max test, but the high reliability and the better achievement of the secondary criteria implies that a valid test of VO2-max can be performed, at a level corresponding to that of healthy controls. The day-to-day variation implies that a change of more than 10% in VO2-max is required to be interpreted as a real change.

Tumor necrosis factor-α (TNF-α) has widespread metabolic actions. Systemic TNF-α administration, however, generates a complex hormonal and metabolic response. Our study was designed to test whether regional, placebo-controlled TNF-α infusion directly affects insulin resistance and protein breakdown. We studied eight healthy volunteers once with bilateral femoral vein and artery catheters during a 3-h basal period and a 3-h hyperinsulinemic-euglycemic clamp. One artery was perfused with saline and one with TNF-α. During the clamp, TNF-α perfusion increased glucose arteriovenous differences (0.91 ± 0.17 vs. 0.74 ± 0.15 mmol/L, P = 0.012) and leg glucose uptake rates. Net phenylalanine release was increased by TNF-α perfusion with concomitant increases in appearance and disappearance rates. Free fatty acid kinetics was not affected by TNF-α, whereas interleukin-6 (IL-6) release increased. Insulin and protein signaling in muscle biopsies was not affected by TNF-α. TNF-α directly increased net muscle protein loss, which may contribute to cachexia and general protein loss during severe illness. The finding of increased insulin sensitivity, which could relate to IL-6, is of major clinical interest and may concurrently act to provide adequate tissue fuel supply and contribute to the occurrence of systemic hypoglycemia. This distinct metabolic feature places TNF-α among the rare insulin mimetics of human origin.

Increased availability of lipids may conserve muscle protein during catabolic stress. Our study was designed to define 1) intracellular mechanisms leading to increased lipolysis and 2) whether this scenario is associated with decreased amino acid and urea fluxes, and decreased muscle amino acid release in obese subjects under basal and fasting conditions. We therefore studied nine lean and nine obese subjects twice, after 12 and 72 h of fasting, using measurements of mRNA and protein expression and phosphorylation of lipolytic and protein metabolic signaling molecules in fat and muscle together with whole body and forearm tracer techniques. Obese subjects displayed increased whole body lipolysis, decreased urea production rates, and decreased forearm muscle protein breakdown per 100 ml of forearm tissue, differences that persisted after 72 h of fasting. Lipolysis per fat mass unit was reduced in obese subjects and, correspondingly, adipose tissue hormone-sensitive lipase (HSL) phosphorylation and mRNA and protein levels of the adipose triglyceride lipase (ATGL) coactivator CGI58 were decreased. Fasting resulted in higher HSL phosphorylations and lower protein levels of the ATGL inhibitor G0S2. Muscle protein expressions of mammalian target of rapamycin (mTOR) and 4EBP1 were lower in obese subjects, and MuRf1 mRNA was higher with fasting in lean but not obese subjects. Phosphorylation and signaling of mTOR decreased with fasting in both groups, whereas ULK1 protein and mRNA levels increased. In summary, obese subjects exhibit increased lipolysis due to a large fat mass with blunted prolipolytic signaling, together with decreased urea and amino acid fluxes both in the basal and 72-h fasted state; this is compatible with preservation of muscle and whole body protein.

Introduction: The majority of young women use oral contraceptives (OCs). Use of OCs has been associated with lower myofibrillar protein and tendon collagen synthesis rates, but it is unknown whether OCs will limit the adaptive response of myotendinous tissue to resistance training. Design and Methods: Fourteen healthy untrained young regular OC users (24 ± 1 years, fat% 32 ± 1, 35 ± 2 ml⋅min-1⋅kg-1) and 14 NOC users (non-OC, controls) (24 ± 1 years, fat% 32 ± 2, 34 ± 2 ml⋅min-1⋅kg-1) performed a 10-week supervised lower extremity progressive resistance training program. Before and after the intervention biopsies from the vastus lateralis muscle and the patellar tendon were obtained. Muscle (quadriceps) and tendon cross-sectional area (CSA) was determined by magnetic resonance imaging (MRI) scans, and muscle fiber CSA was determined by histochemistry. Maximal isometric knee extension strength was assessed by dynamometry while 1 repetition maximum (RM) was determined during knee extension. Results: Training enhanced CSA in both muscle (p < 0.001) and tendon (p < 0.01). A trend toward a greater increase in muscle CSA was observed for OC (11%) compared to NOC (8%) (interaction p = 0.06). Analysis of mean muscle fiber type CSA showed a trend toward an increase in type II muscle fiber area in both groups (p = 0.11, interaction p = 0.98), whereas type I muscle fiber CSA increased in the OC group (n = 9, 3821 ± 197 to 4490 ± 313 mm2, p < 0.05), but not in NOC (n = 7, 4020 ± 348 to 3777 ± 354 mm2, p = 0.40) (interaction p < 0.05). Post hoc analyses indicated that the effect of OCs on muscle mass increase was induced by the OC-users (n = 7), who used OCs containing 30 μg ethinyl estradiol (EE), whereas the response in users taking OCs with 20 μg EE (n = 7) did not differ from NOC. Both the OC and NOC group experienced an increase in maximal knee strength (p < 0.001) and 1RM leg extension (p < 0.001) after the training period with no difference between groups. Conclusion: Use of OCs during a 10-week supervised progressive resistance training program was associated with a trend toward a greater increase in muscle mass and a significantly greater increase in type I muscle fiber area compared to controls. Yet, use of OCs did not influence the overall increase in muscle strength related to training.

NAD+ is a co-factor and substrate for enzymes maintaining energy homeostasis. Nicotinamide phosphoribosyltransferase (NAMPT) controls NAD+ synthesis, and in skeletal muscle, NAD+ is essential for muscle integrity. However, the underlying molecular mechanisms by which NAD+ synthesis affects muscle health remain poorly understood. Thus, the objective of the current study was to delineate the role of NAMPT-mediated NAD+ biosynthesis in skeletal muscle development and function.To determine the role of Nampt in muscle development and function, we generated skeletal muscle-specific Nampt KO (SMNKO) mice. We performed a comprehensive phenotypic characterization of the SMNKO mice, including metabolic measurements, histological examinations, and RNA sequencing analyses of skeletal muscle from SMNKO mice and WT littermates.SMNKO mice were smaller, with phenotypic changes in skeletal muscle, including reduced fiber area and increased number of centralized nuclei. The majority of SMNKO mice died prematurely. Transcriptomic analysis identified that the gene encoding the mitochondrial permeability transition pore (mPTP) regulator Cyclophilin D (Ppif) was upregulated in skeletal muscle of SMNKO mice from 2 weeks of age, with associated increased sensitivity of mitochondria to the Ca2+-stimulated mPTP opening. Treatment of SMNKO mice with the Cyclophilin D inhibitor, Cyclosporine A, increased membrane integrity, decreased the number of centralized nuclei, and increased survival.Our study demonstrates that NAMPT is crucial for maintaining cellular Ca2+ homeostasis and skeletal muscle development, which is vital for juvenile survival.

BACKGROUNDDuring aging, there is a functional decline in the pool of muscle stem cells (MuSCs) that influences the functional and regenerative capacity of skeletal muscle. Preclinical evidence has suggested that nicotinamide riboside (NR) and pterostilbene (PT) can improve muscle regeneration, e.g., by increasing MuSC function. The objective of this study was to investigate if supplementation with NR and PT (NRPT) promotes skeletal muscle regeneration after muscle injury in elderly individuals by improved recruitment of MuSCs.METHODSThirty-two elderly individuals (55-80 years of age) were randomized to daily supplementation with either NRPT (1,000 mg NR and 200 mg PT) or matched placebo. Two weeks after initiation of supplementation, skeletal muscle injury was induced by electrically induced eccentric muscle work. Skeletal muscle biopsies were obtained before, 2 hours after, and 2, 8, and 30 days after injury.RESULTSA substantial skeletal muscle injury was induced by the protocol and associated with release of myoglobin and creatine kinase, muscle soreness, tissue edema, and a decrease in muscle strength. MuSC content, proliferation, and cell size revealed a large demand for recruitment after injury, but this was not affected by NRPT. Furthermore, histological analyses of muscle fiber area, central nuclei, and embryonic myosin heavy chain showed no NRPT supplementation effect.CONCLUSIONDaily supplementation with 1,000 mg NR and 200 mg PT is safe but does not improve recruitment of the MuSC pool or other measures of muscle recovery in response to injury or subsequent regeneration in elderly individuals.TRIAL REGISTRATIONClinicalTrials.gov NCT03754842.FUNDINGNovo Nordisk Foundation (NNF17OC0027242) and Novo Nordisk Foundation CBMR.

A physiological feedback system exists between hepatocytes and the alpha cells, termed the liver-alpha cell axis and refers to the relationship between amino acid-stimulated glucagon secretion and glucagon-stimulated amino acid catabolism. Several reports indicate that non-alcoholic fatty liver disease (NAFLD) disrupts the liver-alpha cell axis, because of impaired glucagon receptor signaling (glucagon resistance). However, no experimental test exists to assess glucagon resistance in humans. The objective was to develop an experimental test to determine glucagon sensitivity with respect to amino acid and glucose metabolism in humans. The proposed glucagon sensitivity test (comprising two elements: 1) i.v. injection of 0.2 mg glucagon and 2) infusion of mixed amino acids 331 mg/hour/kg) is based on nine pilot studies which are presented. Calculation of a proposed glucagon sensitivity index with respect to amino acid catabolism is also described. Secondly, we describe a complete study protocol (GLUSENTIC) according to which the glucagon sensitivity test will be applied in a cross-sectional study currently taking place. 65 participants including 20 individuals with a BMI 18.6-25 kg/m2, 30 individuals with a BMI ≥ 25-40 kg/m2, and 15 individuals with type 1 diabetes with a BMI between 18.6 and 40 kg/m2 will be included. Participants will be grouped according to their degree of hepatic steatosis measured by whole-liver magnetic resonance imaging (MRI). The primary outcome measure will be differences in the glucagon sensitivity index between individuals with and without hepatic steatosis. Developing a glucagon sensitivity test and index may provide insight into the physiological and pathophysiological mechanism of glucagon action and glucagon-based therapies.

Protein-rich beverages are widely used clinically to preserve muscle protein and improve physical performance. Beverages with high contents of leucine or its keto-metabolite β-hydroxy-β-methylbutyrate (HMB) are especially anabolic in muscle, but it is uncertain whether this also applies to catabolic conditions such as fasting and whether common or separate intracellular signaling cascades are involved.To compare a specific leucine-rich whey protein beverage (LWH) with isocaloric carbohydrate- (CHO), soy protein (SOY), and soy protein +3 g HMB (HMB) during fasting-induced catabolic conditions.Eight healthy lean male subjects underwent four interventions (LWH, CHO, SOY, and HMB) using a randomized crossover design. Each trial included a 36 h fast and consisted of a 3 h basal fasting period and a 4 h 'sipping' period.Forearm net balances of phenylalanine (NBphe, measure of net protein loss) improved for all groups (p < 0.05), but more prominently so for LWH and HMB compared with SOY (p < 0.05). 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 LWH consumption (p < 0.05). The ratio between autophagy protein microtubule-associated protein 1 light chain-3β II and I (LC3II/LC3I, a measure of autophagy activity) was decreased during LWH and SOY intake compared with the fasting period (p < 0.05).LWH and HMB have superior anabolic effects on muscle protein kinetics after 36 h of fasting, and LWH distinctly activates the mTOR pathway. These novel findings suggest that leucine-rich whey protein and/or HMB are specifically beneficial during fasting-induced catabolic conditions.

Recent studies demonstrate that adaptations to white adipose tissue (WAT) are important components of the beneficial effects of exercise training on metabolic health. Exercise training favorably alters the phenotype of subcutaneous inguinal WAT (iWAT) in male mice, including decreasing fat mass, improving mitochondrial function, inducing beiging, and stimulating the secretion of adipokines. In this study, we find that despite performing more voluntary wheel running compared with males, these adaptations do not occur in the iWAT of female mice. Consistent with sex-specific adaptations, we report that mRNA expression of androgen receptor coactivators is upregulated in iWAT from trained male mice and that testosterone treatment of primary adipocytes derived from the iWAT of male, but not female mice, phenocopies exercise-induced metabolic adaptations. Sex specificity also occurs in the secretome profile, as we identify cysteine-rich secretory protein 1 (Crisp1) as a novel adipokine that is only secreted from male iWAT in response to exercise. Crisp1 expression is upregulated by testosterone and functions to increase glucose and fatty acid uptake. Our finding that adaptations to iWAT with exercise training are dramatically greater in male mice has potential clinical implications for understanding the different metabolic response to exercise training in males and females and demonstrates the importance of investigating both sexes in studies of adipose tissue biology.

Enhanced secretion of glucagon-like peptide 1 (GLP-1) seems to be essential for improved postprandial β-cell function after Roux-en-Y gastric bypass (RYGB) but is less studied after sleeve gastrectomy (SG). Moreover, the role of the other major incretin hormone, glucose-dependent insulinotropic polypeptide (GIP), is relatively unexplored after bariatric surgery. We studied the effects of separate and combined GLP-1 receptor (GLP-1R) and GIP receptor (GIPR) blockade during mixed-meal tests in unoperated (CON), SG-operated, and RYGB-operated people with no history of diabetes. Postprandial GLP-1 concentrations were highest after RYGB but also higher after SG compared with CON. In contrast, postprandial GIP concentrations were lowest after RYGB. The effect of GLP-1R versus GIPR blockade differed between groups. GLP-1R blockade reduced β-cell glucose sensitivity and increased or tended to increase postprandial glucose responses in the surgical groups but had no effect in CON. GIPR blockade reduced β-cell glucose sensitivity and increased or tended to increase postprandial glucose responses in the CON and SG groups but had no effect in the RYGB group. Our results support that GIP is the most important incretin hormone in unoperated people, whereas GLP-1 and GIP are equally important after SG, and GLP-1 is the most important incretin hormone after RYGB.

Sacubitril/valsartan is a neprilysin-inhibitor/angiotensin II receptor blocker used for the treatment of heart failure. Recently, a post-hoc analysis of a 3-year randomized controlled trial showed improved glycaemic control with sacubitril/valsartan in patients with heart failure and type 2 diabetes. We previously reported that sacubitril/valsartan combined with a dipeptidyl peptidase-4 inhibitor increases active glucagon-like peptide-1 (GLP-1) in healthy individuals. We now hypothesized that administration of sacubitril/valsartan with or without a dipeptidyl peptidase-4 inhibitor would lower postprandial glucose concentrations (primary outcome) in patients with type 2 diabetes via increased active GLP-1.We performed a crossover trial in 12 patients with obesity and type 2 diabetes. A mixed meal was ingested following five respective interventions: (a) a single dose of sacubitril/valsartan; (b) sitagliptin; (c) sacubitril/valsartan + sitagliptin; (d) control (no treatment); and (e) valsartan alone. Glucose, gut and pancreatic hormone responses were measured.Postprandial plasma glucose increased by 57% (incremental area under the curve 0-240 min) (p = .0003) and increased peak plasma glucose by 1.7 mM (95% CI: 0.6-2.9) (p = .003) after sacubitril/valsartan compared with control, whereas postprandial glucose levels did not change significantly after sacubitril/valsartan + sitagliptin. Glucagon, GLP-1 and C-peptide concentrations increased after sacubitril/valsartan, but insulin and glucose-dependent insulinotropic polypeptide did not change.The glucose-lowering effects of long-term sacubitril/valsartan treatment reported in patients with heart failure and type 2 diabetes may not depend on changes in entero-pancreatic hormones. Neprilysin inhibition results in hyperglucagonaemia and this may explain the worsen glucose tolerance observed in this study.gov (NCT03893526).

Fatty liver disease has mainly been characterized under fasting conditions. However, as the liver is essential for postprandial homeostasis, identifying postprandial disturbances may be important. Here, we investigated postprandial changes in markers of metabolic dysfunction between healthy individuals, obese individuals with non-alcoholic fatty liver disease (NAFLD) and patients with cirrhosis. We included individuals with biopsy-proven NAFLD (n = 9, mean age 50 years, mean BMI 35 kg/m2 , no/mild fibrosis), cirrhosis with hepatic steatosis (n = 10, age 62 years, BMI 32 kg/m2 , CHILD A/B) and healthy controls (n = 10, age 23, BMI 25 kg/m2 ), randomized 1:1 to fasting or standardized mixed meal test (postprandial). None of the patients randomized to mixed meal test had type 2 diabetes (T2D). Peripheral blood was collected for 120 min. After 60 min, a transjugular liver biopsy and liver vein blood was taken. Plasma levels of glucose, insulin, C-peptide, glucagon, and fibroblast growth factor 21 (FGF21) were measured. Postprandial peak glucose and C-peptide were significantly increased in NAFLD, and cirrhosis compared with healthy. Patients with NAFLD and cirrhosis had hyperglucagonemia as a potential sign of glucagon resistance. FGF21 was increased in NAFLD and cirrhosis independent of sampling from the liver vein versus peripheral blood. Glucagon levels were higher in the liver vein compared with peripheral blood. Patients with NAFLD and cirrhosis without T2D showed impaired glucose tolerance, hyperinsulinemia, and hyperglucagonemia after a meal compared to healthy individual. Postprandial characterization of patients with NAFLD may be important to capture their metabolic health.

Background: Heat sensitivity (HS) is reported by 58% of all persons with multiple sclerosis (MS), causing symptom exacerbation possibly limiting exercise participation. Objective: The purpose of this study was to test the hypotheses that (a) a relationship between exercise-induced changes in core–temperature (C temp ) and changes in symptom intensity exists, and (b) that resistance exercise (RE), as a consequence of a minor increase in core temperature, will induce a lesser worsening of symptoms than endurance exercise (EE) in HS persons with MS. Methods: On two separate days, 16 HS persons with MS randomly completed a session of RE and EE, or EE and RE, respectively. Testing was conducted pre, post and one hour after exercise and consisted of Visual Analogue Scale (VAS) scoring (fatigue, spasticity, pain, strength, walking and balance), the 5-time sit-to-stand (5STS), the Multiple Sclerosis Functional Composite (MSFC) and Body Sway. Composite scores describing average subjective symptom intensity (SI) and total number of symptoms (SN) were calculated from VAS scores. Results: C temp (0.9±0.4°C vs 0.3±0.1°C, p&lt;0.001), SI (1.7±1.9 cm vs 0.6±1.5 cm, p&lt;0.05) and SN (1.6±1.9 vs 0.6±2.1, p&lt;0.05) increased significantly more during EE than RE. Changes in C temp correlated to changes in SI ( r=0.50, p&lt;0.01). No differences were observed in 5STS, MSFC and Body Sway scores after EE when compared to RE. Conclusion: An exercise-induced increase in C temp is associated with increased number and severity of perceived symptoms in HS persons with MS. Based on these findings it is expected that HS persons with MS do tolerate RE better than EE.

Regular physical exercise has undisputed health benefits in the prevention and the treatment of many diseases. Understanding the mechanisms that regulate adaptations to exercise training therefore has obvious clinical perspectives. Several lines of evidence suggest that the AMP-activated protein kinase (AMPK) has a central role as a master metabolic regulator in skeletal muscle. Exercise is a potent activator of AMPK, and AMPK signaling can play a key part in regulating protein turnover during and after exercise training.

Endurance exercise training induces adaptations in metabolically active organs, but adaptations in human subcutaneous adipose tissue (scAT) remains incompletely understood. On the basis of animal studies, we hypothesized that endurance exercise training would increase the expression of proteins involved in lipolysis and glucose uptake in scAT. To test these hypotheses, 19 young and healthy males were randomized to either endurance exercise training (TR; age 18–24 yr; BMI 19.0–25.4 kg/m 2 ) or a nonexercising control group (CON; age 21–35 yr; BMI 20.5–28.8 kg/m 2 ). Abdominal subcutaneous fat biopsies and blood were obtained at rest before and after intervention. By using Western blotting and PCR, we determined expression of lipid droplet-associated proteins, various proteins involved in substrate metabolism, and mRNA abundance of cell surface G protein-coupled receptors (GPCRs). Adipose tissue insulin sensitivity was determined from fasting plasma insulin and nonesterified fatty acids (adipose tissue insulin resistance index; Adipo-IR). Adipo-IR improved in TR compared with CON ( P = 0.03). This was accompanied by increased insulin receptor (IR) protein expression in scAT with a 1.54-fold (SD 0.79) change from baseline in TR vs. 0.85 (SD 0.30) in CON ( P = 0.007). Additionally, hexokinase II (HKII) and succinate dehydrogenase complex subunit A (SDHA) protein increased in TR compared with CON ( P = 0.006 and P = 0.04, respectively). We did not observe changes in lipid droplet-associated proteins or mRNA abundance of GPCRs. Collectively, 10 weeks of endurance exercise training improved adipose tissue insulin sensitivity, which was accompanied by increased IR, HKII, and SDHA protein expression in scAT. We suggest that these adaptations contribute to an improved metabolic flexibility. NEW &amp; NOTEWORTHY This study is the first to investigate the molecular adaptations in human subcutaneous adipose tissue (scAT) after endurance exercise training compared with a nonexercising control group. We show that endurance exercise training improves insulin sensitivity in human scAT, and this is accompanied by increased expression of insulin receptor, hexokinase II, and succinate dehydrogenase complex subunit A. Collectively, our data suggest that endurance exercise training induces molecular adaptations in human scAT, which may contribute to an improved metabolic flexibility.

Objective This study aimed to present vocal extent measure (VEM), demonstrate its changes with phonomicrosurgical treatment in patients with vocal fold polyps (VFPs), and to compare its performance to that of established vocal parameters. Study Design This is an individual cohort study. Methods Microlaryngoscopic ablation was executed in 61 patients with manifestation of VFP (28 male, 33 female; 45 ± 13 years [mean ± SD]). Analysis of treatment outcome was based on pre- and postoperative voice function diagnostics and videolaryngostroboscopy. Examination instruments were: auditory-perceptual voice assessment (roughness, breathiness, and overall hoarseness [RBH]-status), voice range profile (VRP), acoustic-aerodynamic analysis, and patients' self-assessment of voice using the voice handicap index. The VEM, a parameter not yet commonly established in phoniatric diagnostics, was calculated from area and shape of the VRP to be compared with the dysphonia severity index (DSI) concerning diagnostic suitability. Results All polyps were completely excised. Three months postoperatively, mucosal wave propagation had recovered. All subjective and most objective acoustic and aerodynamic parameters showed highly significant improvement. The VHI-9i-score decreased from 15 ± 8 to 6 ± 7 points. The average total vocal range extended by 4 ± 5 semitones, the mean speaking pitch decreased by 1 ± 2 semitones. The DSI increased on average from 2.6 ± 2.1 to 4.0 ± 2.2, VEM from 83 ± 28 to 107 ± 21 (P < 0.01). VEM and DSI correlated significantly with each other (rs = 0.65; P < 0.01). Conclusion Phonomicrosurgery of VFP is an objectively and subjectively satisfactory therapy for voice improvement. The VEM represents a comprehensible and easy-to-use unidimensional measure for objective VRP evaluation. This positive measure of vocal function seems to be a compelling diagnostic addition for objective quantification of vocal performance.

Animal models clearly illustrate that the maintenance of skeletal muscle mass depends on the function and interaction of a heterogeneous population of resident and infiltrating mononuclear cells. Several lines of evidence suggest that mononuclear cells also play a role in muscle wasting in humans, and targeting these cells may open new treatment options for intervention or prevention in sarcopenia. Methodological and ethical constraints have perturbed exploration of the cellular characteristics and function of mononuclear cells in human skeletal muscle. Thus, investigations of cellular phenotypes often depend on immunohistochemical analysis of small tissue samples obtained by needle biopsies, which do not match the deep phenotyping of mononuclear cells obtained from animal models. Here, we have developed a protocol for fluorescence-activated cell sorting (FACS), based on single-cell RNA-sequencing data, for quantifying and characterizing mononuclear cell populations in human skeletal muscle. Muscle stem cells, fibro-adipogenic progenitors, and two subsets of macrophages (CD11c +/– ) are present in needle biopsies in comparable quantities per milligram tissue to open surgical biopsies. We find that direct cell isolation is preferable due to a substantial shift in transcriptome when using preculture before the FACS procedure. Finally, in vitro validation of the cellular phenotype of muscle stem cells, fibro-adipogenic progenitors, and macrophages confirms population-specific traits. This study demonstrates that mononuclear cell populations can be quantified and subsequently analyzed from needle biopsy material and opens the perspective for future clinical studies of cellular mechanisms in muscle wasting.

Abstract. Microalbuminuria in non‐insulin‐dependent diabetes (NIDDM) is a strong predictor of increased mortality. The major causes of death are cardiovascular, whereas end‐stage renal failure is of low frequency. To define kidney function and the presence of some assumed cardiovascular risk factors, we compared a group of 19 microalbuminuric NIDDM patients (M), of mean age (±SD) 65 ± 4 years, and known duration of diabetes 8 ± 7 years, with 19 randomly selected matched normoalbuminuric patients (N). Seven macroalbuminuric patients (P) were also studied. Glomerular filtration rate (GFR) did not differ between N and M, whereas kidney volume was increased in M (260.3 ± 54.1 ml 1.73 m −2 ) compared to N (220.4 ± 44.8 ml 1.73 m −2 ; P =0.018). The frequency of cardiac disease increased with increasing albuminuria. Glycaemic control did not differ between the groups, but fasting plasma C‐peptide levels increased from 2.8 ± 1.1 μg l −1 in N, to 3.7 ± 1.7 μg l −1 in M ( P =0.08), and to 4.2 ± 1.9 μg l −1 ( P= 0.03) in P. The lipoprotein profile showed no significant differences, although the LDL cholesterol /HDL cholesterol (LDL‐C/HDL‐C) ratio tended to rise. A significant correlation was found between C‐peptide and LDL‐C/HDL‐C ( r =0.5; P &lt; 10 −3 ). In conclusion, GFR was not increased, and did not differ between N and M, whereas kidney volume was enhanced in M. Several assumed cardiovascular risk factors showed values of M intermediate between those of N and P.

The acute-phase response is a catabolic event involving increased waste of amino-nitrogen (N) via hepatic urea synthesis, despite an increased need for amino-N incorporation into acute-phase proteins. This study aimed to clarify the regulation of N elimination via urea during different phases of the tumor necrosis factor-α (TNF-α)-induced acute-phase response in rats. We used four methods to study the regulation of urea synthesis: We examined urea cycle enzyme mRNA levels in liver tissue, the hepatocyte urea cycle enzyme proteins, the in vivo capacity of urea-N synthesis (CUNS), and known humoral regulators of CUNS at 1, 3, 24, and 72 h after TNF-α injection (25 μg/kg iv rrTNF-α) in rats. Serum acute-phase proteins and their liver mRNA levels were also measured. The urea cycle enzyme mRNA levels acutely decreased and then gradually normalized, whereas the urea cycle enzyme proteins remained essentially unchanged over time. The CUNS rose after 3 h and then normalized. The acute-phase response was fully activated at 24 h with markedly increased serum levels of the acute-phase proteins. TNF-α acutely upregulated the CUNS. Later, despite the fully established 24-h acute-phase response and the decreased activity of the urea cycle enzyme genes, there was no change in the urea cycle enzyme proteins or the CUNS. Thus in no phase after the initiation of the acute-phase response was in vivo urea synthesis orchestrated in combination with acute-phase protein synthesis so as to limit N waste.

Accumulating evidence suggests that chronic exposure to lipopolysaccharide (LPS, endotoxin) may create a constant low-grade inflammation, leading to insulin resistance and diabetes. All previous human studies assessing the metabolic actions of LPS have used systemic administration, making discrimination between direct and indirect effects impossible. We sought to define the direct, placebo-controlled effects of LPS on insulin resistance and protein and lipid metabolism in the infused human leg without systemic interference from cytokines and stress hormones. This was a randomized, placebo-controlled, single-blinded study. We studied 8 healthy volunteers with bilateral femoral vein and artery catheters during a 3-hour basal and 3-hour hyperinsulinemic-euglycemic clamp period with bilateral muscle biopsies in each period during infusion with saline and LPS. Overall, LPS perfusion significantly decreased leg glucose uptake, and during the clamp LPS decreased glucose arteriovenous differences (0.65 ± 0.07 mmol/L vs 0.73 ± 0.08 mmol/L). Net palmitate release was increased by LPS, and secondary post hoc testing indicated increased palmitate isotopic dilution, although primary ANOVA tests did not reveal significant dilution. Leg blood flows, phenylalanine, lactate kinetics, cytokines, and intramyocellular insulin signaling were not affected by LPS. LPS thus directly inhibits insulin-stimulated glucose uptake and increases palmitate release in the perfused human leg without detectable effects on amino acid metabolism. These data strongly suggest that the primary metabolic effect of LPS is increased lipolysis and muscle insulin resistance, which, together with secondary insulin resistance, caused by systemic cytokine and stress hormone release may lead to overt glucose intolerance and diabetes.

Voice range profile (VRP) and evaluation using the dysphonia severity index (DSI) represent essentials of instrument-based objective voice diagnostics and are implemented in different standardized registration programs. The respective measurement results, however, show differences. The aim of the study was to prove these differences statistically and to develop a new parameter, the Vocal Extent Measure (VEM), which is not influenced by the measurement program. VRPs of 97 subjects were recorded by two examiners using the established registration programs DiVAS (XION medical) and LingWAVES (WEVOSYS) simultaneously. The VEM was developed on the basis of VRP area and perimeter. All 194 VRP files were analyzed for various parameters and gender independence. The registration programs exhibited significant differences in several vocal parameters. A significant gender influence for DSI was found with DiVAS (<mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" id="M1"><mml:mi>p</mml:mi><mml:mo>&lt;</mml:mo><mml:mn fontstyle="italic">0.01</mml:mn></mml:math>), but not with LingWAVES. The VEM quantified the dynamic performance and frequency range by a unidimensional, interval-scaled value without unit, mostly between 0 and 120. This novel parameter represents an intelligible and user-friendly positive measure of vocal function, allows simple and stable VRP description, and seems to be suitable for quantification of vocal capacity. In contrast to DSI, the VEM proved to be less susceptible to registration program and gender.

Immobilization of the lower limbs promotes a catabolic state that reduces muscle mass, whereas physical training promotes an anabolic state that increases muscle mass. Understanding the molecular mechanisms underlying this is of clinical interest, as loss of muscle mass is a major complication to critical illness in humans. To determine the molecular regulation of protein synthesis and degradation during muscle loss and hypertrophy, we examined skeletal muscle biopsies from healthy human subjects after 2 weeks unilateral immobilization of a lower limb and during 6 weeks of physical rehabilitation. We have previously shown that cross-sectional area of the knee muscle-extensors decreased by ~10% during immobilization and was completely restored during rehabilitation. Here we provide novel data to suggest that autophagy is an important underlying mechanism involved in regulation of muscle mass. Protein expression of MuRF1 and ATROGIN-1 did not change during the study, indicating that the recruitment of substrates to the proteasomes was unaltered. Phosphorylation of mTORat Ser2448 did not change during the study, and neither did phosphorylation of the mTORC1 substrates 4EBP1 Thr37/46 and p70S6K Thr389, suggesting that this pathway does not suppress protein synthesis during muscle wasting. Protein levels of p62 and ULK1 increased during immobilization and returned to baseline levels during rehabilitation. Same pattern was observed for FOXO3a phosphorylation at Ser318/321, suggesting transcriptional activation during immobilization and inactivation during rehabilitation. To investigate this further, we analyzed mRNA expression of seven autophagy-related genes controlled by FOXO3a. Five of these (p62, LC3B, BECLIN-1, ATG12, and BNIP3) increased during immobilization and returned to baseline during rehabilitation. In conclusion, immobilization of a lower limb increases autophagy-related gene and protein expression in human skeletal muscle in a pattern that mirrors FOXO3a phosphorylation. These findings could imply that FOXO3a dependent transcriptional regulation of autophagy is involved in the regulation of muscle mass in humans.

&lt;p dir="ltr"&gt;&lt;b&gt;Describe your practice setting and location.&lt;/b&gt;&lt;/p&gt;&lt;p dir="ltr"&gt;The Clinic for Athletes With Type 1 Diabetes (CATOD) team is based at the Steno Diabetes Center Aarhus, Aarhus University Hospital, in Aarhus, Denmark.&lt;/p&gt;&lt;p&gt;&lt;br&gt;&lt;/p&gt;&lt;p dir="ltr"&gt;&lt;b&gt;Describe the specific quality gap addressed through the initiative.&lt;/b&gt;&lt;/p&gt;&lt;p dir="ltr"&gt;The goal of the clinic is to help athletes with type 1 diabetes exercise at the same level as their healthy peers so they can compete and achieve high rank positions in competitions. This is done by assisting them in improving their diabetes management skills in relation to physical training and competitions, which also supports their efforts to enhance their sporting performance. There has been a lack of existing mechanisms through which athletes with type 1 diabetes can network. Within this clinic, athletes with type 1 diabetes are brought together, allowing them to share peer support, tips, and tricks, which they have found very valuable.&lt;/p&gt;

&lt;p dir="ltr"&gt;&lt;b&gt;Describe your practice setting and location.&lt;/b&gt;&lt;/p&gt;&lt;p dir="ltr"&gt;The Clinic for Athletes With Type 1 Diabetes (CATOD) team is based at the Steno Diabetes Center Aarhus, Aarhus University Hospital, in Aarhus, Denmark.&lt;/p&gt;&lt;p&gt;&lt;br&gt;&lt;/p&gt;&lt;p dir="ltr"&gt;&lt;b&gt;Describe the specific quality gap addressed through the initiative.&lt;/b&gt;&lt;/p&gt;&lt;p dir="ltr"&gt;The goal of the clinic is to help athletes with type 1 diabetes exercise at the same level as their healthy peers so they can compete and achieve high rank positions in competitions. This is done by assisting them in improving their diabetes management skills in relation to physical training and competitions, which also supports their efforts to enhance their sporting performance. There has been a lack of existing mechanisms through which athletes with type 1 diabetes can network. Within this clinic, athletes with type 1 diabetes are brought together, allowing them to share peer support, tips, and tricks, which they have found very valuable.&lt;/p&gt;

To evaluate whether the moderately elevated human growth hormone concentration, seen in insulin dependent diabetic patients, has any impact on lipoproteins, human growth hormone was given to nondiabetic persons in doses which would bring their plasma human growth hormone concentration up in the same level as seen in insulin dependent diabetic patients. After one week of treatment with human growth hormone we found total plasma triglyceride to be significantly raised (0.98 mmol/l +/- 0.28 mmol/l (mean +/- SD) before versus 1.27 mmol/l +/- 0.38 mmol/l (mean +/- SD) after treatment). Very low density lipoprotein (VLDL) was separated into two fractions (VLDL-1 and VLDL-2) of which VLDL-2 is regarded as a VLDL-remnant which is suggested to be of importance for development of atherosclerosis. After one week of human growth hormone treatment there were no changes in VLDL-1 concentrations whereas a significant raise in VLDL-2 triglyceride and VLDL-2 cholesterol was seen.

Repeated periodization of carbohydrate (CHO) intake using a diet‐exercise strategy called the sleep‐low model can potentially induce mitochondrial biogenesis and improve endurance performance in endurance‐trained individuals. However, more studies are needed to confirm the performance‐related effects and to investigate the sustained effects on maximal fat oxidation (MFO) rate and proteins involved in intramuscular lipid metabolism. Thirteen endurance‐trained males (age 23‐44 years; O 2 ‐max, 63.9 ± 4.6 mL·kg −1 ·min −1 ) were randomized into two groups: sleep‐low (LOW‐CHO) or high CHO availability (HIGH‐CHO) in three weekly training blocks over 4 weeks. The acute metabolic response was investigated during 60 minutes of exercise within the last 3 weeks of the intervention. Pre‐ and post‐intervention, 30‐minute time‐trial performance was investigated after a 90‐minute pre‐load, which as a novel approach included nine intense intervals (and estimation of MFO). Additionally, muscle biopsies ( v. lateralis ) were obtained to investigate expression of proteins involved in intramuscular lipid metabolism using Western blotting. During acute exercise, average fat oxidation rate was ~36% higher in LOW‐CHO compared to HIGH‐CHO ( P = .03). This did not translate into sustained effects on MFO. Time‐trial performance increased equally in both groups (overall time effect: P = .005). We observed no effect on intramuscular proteins involved in lipolysis (ATGL, G0S2, CGI‐58, HSL) or fatty acid transport and β‐oxidation (CD‐36 and HAD, respectively). In conclusion, the sleep‐low model did not induce sustained effects on MFO, endurance performance, or proteins involved in intramuscular lipid metabolism when compared to HIGH‐CHO. Our study therefore questions the transferability of acute effects of the sleep‐low model to superior sustained adaptations.

In a previous study we investigated the analgesic efficacy of metamizol. After laparoscopic operations, in particular, the reduction of postoperative opioid requirements within the first 24 h after surgery attained clinical relevance (-67%). In the present study we investigated the analgesic efficacy of supplementary diclofenac.86 patients, scheduled for minor orthopaedic surgery, laparoscopic cholecystectomy or resection of the thyroid gland, participated in a doubleblind, randomised, placebo-controlled study. The setting was comparable to our previous study, apart from the supplementary administration of diclofenac. Before induction of anaesthesia, verum-treated patients received a diclofenac suppository (100 mg), in addition to metamizol (1 g/100 ml NaCl 0.9% intravenous over 15 min). These infusions were repeated at 6h and 12h. In addition to the third infusion, the patients received a further diclofenac suppository (100 mg). Cumulated doses of buprenorphine (PCA, patient-controlled analgesia), pain scores (0-10), blood pressure, pulse and side effects were recorded during the first 6 h and again at 24 h.All verum-treated patients had significantly less pain immediately after surgery and required lower cumulated doses of buprenorphine during the first 24 h after operation (laparoscopic cholecystectomy -33%, minor orthopaedic surgery -73%, resection of thyroid gland -60%).Combination of metamizol and diclofenac cause a clinically relevant reduction in opioid requirements, in particular after minor orthopaedic surgery and resection of the thyroid gland. There is no need for supplementary diclofenac following laparoscopic surgery.

Acute exercise increases autophagic signaling through Unc-51 like kinase-1 (ULK1) in human skeletal muscle during both anabolic and catabolic conditions. The aim of the present study was to investigate if changes in ULK1 Ser555 phosphorylation during exercise are reflected by changes in phosphorylation of a newly identified ULK1 substrate (ATG14 Ser29) and to elucidate the involvement of circulatory hormones in the regulation of autophagy in human skeletal muscle. We show that 1 h of cycling exercise increases ATG14 Ser29 phosphorylation during both hyperinsulinemic euglycemic and euinsulinemic euglycemic conditions. This could suggest that counterregulatory hormones stimulate autophagy in skeletal muscle, as circulating concentrations of these hormones are highly elevated during exercise. Furthermore, ATG14 Ser29 correlated positively with ULK1 phosphorylation, suggesting that ULK1 Ser555 (activating site) phosphorylation reflects ULK1 kinase activity. In a separate series of experiments, we show that insulin stimulates ULK1 phosphorylation at Ser757 (inhibitory site) in both hypoglycemic and euglycemic conditions, suggesting that counterregulatory hormones (such as epinephrine, norepinephrine, growth hormone, and glucagon) have limited effects on autophagy signaling in human skeletal muscle. In conclusion, 1 h of cycling exercise increases phosphorylation of ATG14 at Ser29 in a pattern that mirrors ULK1 phosphorylation at Ser555. Moreover, insulin effects on autophagy signaling in human skeletal muscle are independent of hypoglycemic and euglycemic conditions.NEW & NOTEWORTHY Autophagy signaling is regulated in a hierarchical order by exercise, insulin, and counterregulatory hormones. Exercise-induced autophagy signaling is stimulated by local factors in skeletal muscle rather than circulatory hormones. Unc-51 like kinase-1 (ULK1) phosphorylation at Ser555 reflects ULK1 kinase activity.

The metabolic consequences after Roux-en-Y gastric bypass (RYGB) and sleeve gastrectomy (SG) are often studied using a liquid mixed meal. However, liquid meals may not be representative of the patients' everyday diet. We therefore examined postprandial glucose and gut hormone responses using mixed meals differing only with respect to meal texture.Twelve RYGB-operated, 12 SG-operated, and 12 unoperated individuals (controls) were enrolled in the study. Participants were matched on age, sex, and body mass index. In randomized order, each participant underwent a liquid and a solid 4-h mixed meal test on separate days. The meals were isocaloric (309 kcal), and with identical macronutrient composition (47 E% carbohydrate, 18 E% protein, 32 E% fat, and 3 E% dietary fibers). The liquid meal was blended to create a smooth liquid texture while the other meal retained its solid components.Postprandial glucose concentrations (peak and incremental area under curve, iAUC) did not differ between the two meal textures in any group. In the control group, peak C-peptide was higher after the liquid meal compared with the solid meal (p = 0.04), whereas iAUCs of C-peptide were similar between the two meals in all groups. Peak of glucagon-like peptide-1 (GLP-1) was higher after the liquid meal compared with the solid meal in RYGB- and SG-operated individuals (RYGB p = 0.02; SG p < 0.01), but iAUC of GLP-1 did not differ between meal textures within any group. Peak of glucose-dependent insulin tropic polypeptide (GIP) was higher after the liquid meal in the SG and control groups (SG p = 0.02; controls p < 0.01), but iAUCs of GIP were equal between meals. There were no differences in total AUC of ghrelin between the liquid and solid meals within any of the groups.A liquid and a solid meal with identical macronutrient composition result in similar postprandial glucose responses, both in operated and unoperated individuals. Small differences were observed for the postprandial peaks of C-peptide, GLP-1, and GIP concentrations. Overall, a liquid meal is suitable for evaluating glucose tolerance, β-cell function, and gut hormones responses, both after RYGB and SG and in unoperated individuals.[www.clinicaltrials.gov], identifier [NCT04082923].

Patients with chronic kidney disease (CKD) have a high risk of bone fractures. A circadian rhythmicity in turnover and mineralization of bone appears to be of importance for bone health. In CKD disturbances in the circadian rhythm of various functions has been demonstrated and indeed the circadian rhythm in the mineral metabolism is disturbed. The aim of the present study was to compare the circadian rhythm of bone turnover markers in ten patients with CKD to ten healthy controls. Bone turnover markers (C-terminal telopeptide of type I collagen, tartrate-resistant acid phosphatase 5b, N-terminal propeptide of type I procollagen, bone alkaline phosphatase and osteocalcin) were measured every third hour for 24 h. All bone turnover markers displayed a significant circadian rhythm in both groups and there were no significant differences in the rhythmicity between the two groups (no group*time interaction). As expected, due to the reduced renal clearance, the overall level of C-terminal telopeptide of type I collagen and osteocalcin was higher in CKD compared to the healthy controls. The present study suggests that disturbances in the circadian rhythm of bone turnover do not explain the metabolic bone disease and increased risk of fractures in CKD.

In the present work we investigate the analgesic combination of the pyrazole-derivative metamizol and the non-steroidal anti-inflammatory drug diclofenac after trauma surgery with respect to the postoperative opioid consumption (via patient-controlled analgesia (PCA)) in the first 24 h after surgery, and the reduction of pain in the immediate postoperative period. In a double-blind randomized placebo controlled study, 48 patients, scheduled for minor trauma surgery received either metamizol (three doses of 1 g intravenously) and diclofenac (two doses of 100 mg suppository) or placebo beginning immediately preoperatively and repeated postoperatively. Totally the verum group received metamizol 3 g and diclofenac 200 mg. Postoperative pain intensity was measured by the numeric rating scale (NRS). All patients were allowed to order levomethadone from a PCA-pump for 24 h after surgery. Pain scores and opioid consumption were evaluated in the first six postoperative hours, 12 and 24 h after end of surgery. The patients receiving verum had significantly less pain at all points in time except for 2 h and ordered a significantly lower cumulated dose of levomethadone during the first 24 h postoperatively than placebo treated subjects (opioid-sparing effect after 24 h: −74%). There were no significant differences in the incidence of side effects between the two groups. Perioperative application of metamizol and diclofenac results in better pain relief and significantly lowers opioid requirements in patients after minor trauma surgery.

Objective Acute and chronic inflammatory and metabolic responses are generated by lipopolysaccharide (LPS) during acute illness and in the pathogenesis of the metabolic syndrome, type 2 diabetes and cardiovascular disease, but whether these responses depend on intact pituitary release of hormones are not clearly identified. We compared the metabolic effects of LPS in hypopituitary patients (HPs) (in the absence of growth hormone (GH) and ACTH responses) and healthy control subjects (CTR) (with normal pituitary hormone responses). Design Single-blind randomized. Methods We compared the effects of LPS on glucose, protein and lipid metabolism in eight HP and eight matched CTR twice during 4-h basal and 2-h hyperinsulinemic–euglycemic clamp conditions with muscle and fat biopsies in each period during infusion with saline or LPS. Results LPS increased cortisol and GH levels in CTR but not in HP. Also, it increased whole-body palmitate fluxes (3-fold) and decreased palmitate-specific activity (SA) 40–50% in CTR, but not in HP. G(0)/G(1) Switch Gene 2 (G0S2 – an inhibitor of lipolysis) adipose tissue (AT) mRNA was decreased in CTR. Although LPS increased phenylalanine fluxes significantly more in CTR, there was no difference in glucose metabolism between groups and intramyocellular insulin signaling was unaltered in both groups. Conclusions LPS increased indices of lipolysis and amino acid/protein fluxes significantly more in CTR compared with HP and decreased adipocyte G0S2 mRNA only in CTR. Thus, in humans intact pituitary function and appropriate cortisol and GH release are crucial components of the metabolic response to LPS.

TNF-α generates inflammatory responses and insulin resistance, lipolysis, and protein breakdown. It is unclear whether these changes depend on intact hypothalamo-pituitary stress hormone responses to trigger the release of cortisol and growth hormone.To define differential effects of TNF-α on glucose, protein, and lipid metabolism in hypopituitary patients (without intact hypothalamo-pituitary axis) and healthy controls.Randomized, placebo controlled, single-blinded. Setting, Participants, and Intervention: We studied eight hypopituitary (HP) patients and eight matched control subjects [control volunteers (CTR)] twice during 4-h basal and 2-h hyperinsulinemic clamp conditions with isotope dilution during infusion of saline or TNF-α(12 ng/kg/h) for 6 h.Phenylalanine, urea, palmitate, and glucose fluxes and fat biopsies in basal and clamp periods.TNF-α infusion significantly increased cortisol and GH levels in CTR but not in HP. TNF-α increased phenylalanine fluxes in both groups, with the increase being significantly greater in CTR, and raised urea flux by 40 % in CTR without any alteration in HP. Endogenous glucose production (EGP) was elevated in CTR compared to HP after TNF-α administration, whereas insulin sensitivity remained similarly unaffected in both groups. TNF-α increased whole body palmitate fluxes and decreased palmitate specific activity in CTR, but not in HP without statistical difference between groups. We did not detect significant effects TNF-α on lipase expression or regulation in fat.TNF-α increased both urea and amino acid fluxes and EGP significantly more in CTR compared to HP, suggesting that increases in endogenous cortisol and GH release are significant components of the metabolic response to TNF-α.

Instrument-assisted measuring procedures expand the options within phoniatric diagnostics by quantifying the condition of the voice. The aim of this study was to examine objective treatment-associated changes of the recently developed vocal extent measure (VEM) and the established dysphonia severity index (DSI) in relation to subjective tools, i.e., self-evaluation via voice handicap index (VHI-12) and external evaluation via auditory-perceptual assessment of hoarseness ( H ). The findings for H (3 raters’ group assessment), VHI-12, DSI, and VEM in 152 patients of both sexes (age range 16–75 years), taken before and 3 months after phonosurgery or vocal exercises, were compared and correlated. Posttherapeutically, all of the recorded parameters improved (<mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" id="M1"><mml:mrow><mml:mi>p</mml:mi><mml:mo>&lt;</mml:mo><mml:mn>0.001</mml:mn></mml:mrow></mml:math>). The degree of H reduced on average by 0.5, the VHI-12 score sank by 5 points, while DSI and VEM rose by 1.5 and 19, respectively. The correlations of these changes were significant but showed gradual differences between H and VHI-12 ( r = 0.3), H and DSI ( r = −0.3), and H and VEM ( r = −0.4). We conclude that all investigated parameters are adequate to verify therapeutic outcomes but represent different dimensions of the voice. However, changes in the degree of H as gold standard were best recognized with the new VEM.

Abstract. In addition to hyperglycaemia, derangement of metabolic and hormonal control may play an important role in the development of microvascular complications in diabetes. Little, however, is known about the impact of insulin pump treatment on metabolic and hormonal parameters. In a 6-month prospective randomized study in insulin-dependent diabetics we therefore investigated the effects of continuous subcutaneous insulin infusion by pump (10 patients) and conventional insulin treatment (10 patients) on the 24-h profiles of blood glucose, glycerol, lactate, 3-hydroxybutyrate, insulin, glucagon and growth hormone by measuring the respective concentrations every 2 h. We found that average blood glucose levels and HbA 1c were significantly lower in the group treated by continuous subcutaneous insulin infusion as compared with the group on conventional insulin treatment. Furthermore, we observed an improvement in diurnal levels of lactate and 3-hydroxybutyrate in the pumptreated group which was not seen in the conventionally treated group. A slight increment in alanine was seen in the group treated with insulin pump. Serum growth hormone, glycerol, plasma free insulin as well as the daily insulin supply were unchanged and identical in the two groups. It is noteworthy that in the pump group, the decrease in blood glucose and 3-hydroxybutyrate takes place concomitantly with a significant suppression of glucagon.

ABSTRACT Aging and type 2 diabetes mellitus (T2DM) are associated with impaired skeletal muscle function and degeneration of the skeletal muscle microenvironment. However, the origin and mechanisms underlying the degeneration are not well described in human skeletal muscle. Here we show that skeletal muscles of T2DM patients exhibit pathological degenerative remodeling of the extracellular matrix that was associated with a selective increase of a subpopulation of fibro-adipogenic progenitors (FAPs) marked by expression of THY1 (CD90) - the FAP CD90+ . We identified Platelet-derived growth factor (PDGF) signaling as key regulator of human FAP biology, as it promotes proliferation and collagen production at the expense of adipogenesis, an effect accompanied with a metabolic shift towards glycolytic lactate fermentation. FAPs CD90+ showed a PDGF-mimetic phenotype, with high proliferative activity and clonogenicity, increased production of extracellular matrix production and enhanced glycolysis. Importantly, the pathogenic phenotype of T2DM FAP CD90+ was reduced by treatment with the anti-diabetic drug Metformin. These data identify PDGF-driven conversion of a sub-population of FAPs as a key event in the pathogenic accumulation of extracellular matrix in T2DM muscles.

AbstractWe have studied the long-term effects (9 months) in plasma lipoprotein concentrations during continuous subcutaneous insulin infusion (CSII) (n=11, six females, five males) and compared these changes to conventional insulin therapy (CIT) (n=12, six females, six males). The two groups were allocated to CSII or CIT randomly, and were comparable as regards lipoprotein values at the start of the study. There were initially normal total plasma cholesterol values in both groups (CSII group: mean plasma cholesterol 3.77±0.57 mmo1/1, CIT group: mean plasma cholesterol 4.37±0.55 mmol/1, x±SD). Further, there were normal total plasma triglyceride values at the start of the study (CSII group: mean plasma triglyceride 0.86±0.23 mmo1/1, CIT group: mean plasma triglyceride 0.84±0.26 mmo1/1, x±SD). There were no alterations seen in total plasma cholesterol and total plasma triglyceride in either groups during a 9 months observation period. In the same period no changes in LDL and HDL levels were registered. The very low density lipoprotein (VLDL) was separated into VLDL-1 and VLDL-2 by its binding to heparin-sepharose columns. It was found that CSII treatment for 9 months resulted in a decline in VLDL-2-triglyceride values (0.18±0.07 mmo1/1 before versus 0.10±0.07 mmo1/1 after, p<0.05, x±SD) which was not seen in the CIT group. Decline in VLDL-2-triglyceride might delay the development of late diabetic manifestations.Key Words: atherosclerosisbeta-very-low-density lipoproteindiabetic angiopathydiabetic large vessel diseasediabetic macroangiopathy

Elderly women are more vulnerable to muscle disuse atrophy than men, due to lower muscle mass. Thus, identification of preventive strategies is of principal interest. We studied effects of training and timing of protein-rich meals after exercise sessions, on skeletal muscle mass in healthy postmenopausal women. Following 2 weeks of one-legged immobilization, a protein-rich meal was ingested either immediately (T0; n = 9, 56 ± 2 years) or 2 hours (T2; n = 8, 57 ± 3 years) after termination of resistance exercise sessions for 6 weeks. Muscle mass, strength, and power were assessed before, after 2 weeks of lower limb immobilization and after 2 and 6 weeks of combined resistance training and diet intervention. Immobilization reduced muscle mass (~10%), strength (~23%), and power (~26%), but was returned to baseline levels after 2 weeks of training. Improvements in muscle mass and strength did not differ between groups. T0, but not T2, increased their total daily protein intake during the training period. In conclusion, only 2 weeks of supervised resistance training counteracted reductions in muscle mass and strength in postmenopausal women after 2 weeks of immobilization. Timing of protein-rich meals in the hours after resistance exercise did not have any impact on the effects of training.

Abstract The human liver is dynamic organ with minute to hourly adaptions in response to feeding. Patients with non-alcoholic fatty liver disease (NAFLD) and cirrhosis have altered transcriptomic features compared to controls but how and if food intake affects such is unknown in humans. Our aim was to investigate the hepatic transcriptome at both fasting and postprandial states in patients with NAFLD, cirrhosis, and healthy controls and secondly to develop a browsable resource enabling easy and unrestricted access to such data. We obtained liver tissue by transjugular liver biopsies from patients with NAFLD (n = 9, mean age 49 (16 SD) y, BMI 35 (5) kg/m 2 ), cirrhosis (n = 9, age 61 (11) y, BMI 32 (5) kg/m 2 ) and healthy controls (n = 10, age 25 (3) y, BMI 23 (3) kg/m 2 ). The hepatic transcriptome was sequenced using NGS and evaluated in bioinformatic analyses to assess differentially expressed genes (DEG) and gene ontology biological processes (GOBP). We identified 553 DEG between healthy controls and patients with NAFLD, 5527 DEG between healthy controls and patients with cirrhosis, and 3898 DEG in NAFLD compared with cirrhosis. A hitherto uncharacterized gene (MET proto-oncogene) was differentially expressed in human NAFLD and cirrhosis. The hepatic transcriptome changed significantly during a standardized meal and these changes were blunted in patients with NAFLD and cirrhosis. GOBP analyses revealed an increase in pro-inflammatory and pro-fibrotic genes in NAFLD and cirrhosis, as well as a decrease in genes related to metabolism. Data were made browsable using two web-based apps. The hepatic transcriptome is differentially regulated by a standardized meal in healthy individuals compared to patients with fatty liver disease.

Postprandial hypoglycemia is a complication of Roux-en-Y gastric bypass (RYGB), but the effects of postprandial exercise and meal glycemic index (GI) on postprandial glucose and glucoregulatory hormone responses are unknown. Ten RYGB-operated and 10 age and weight-matched unoperated women completed four test days in random order ingesting mixed meals with high GI (HGI, GI = 93) or low GI (LGI, GI = 54), but matched on energy and macronutrient content. Ten minutes after meal completion, participants rested or cycled for 30 min at 70% of maximum oxygen uptake (V̇o2max). Blood was collected for 4 h. Postprandial exercise did not lower plasma nadir glucose in RYGB after HGI (HGI/rest 3.7 ± 0.5 vs. HGI/Ex 4.1 ± 0.4 mmol/L, P = 0.070). Replacing HGI with LGI meals raised glucose nadir in RYGB (LGI/rest 4.1 ± 0.5 mmol/L, P = 0.034) and reduced glucose excursions (Δpeak-nadir) but less so in RYGB (-14% [95% CI: -27; -1]) compared with controls (-33% [-51; -14]). Insulin responses mirrored glucose concentrations. Glucagon-like peptide 1 (GLP-1) responses were greater in RYGB versus controls, and higher with HGI versus LGI. Glucose-dependent insulinotropic polypeptide (GIP) responses were greater after HGI versus LGI in both groups. Postexercise glucagon responses were lower in RYGB than controls, and noradrenaline responses tended to be lower in RYGB, whereas adrenaline responses were similar between groups. In conclusion, moderate intensity cycling shortly after meal intake did not increase the risk of postprandial hypoglycemia after RYGB. The low GI meal increased nadir glucose and reduced glucose excursions compared with the high GI meal. RYGB participants had lower postexercise glucagon responses compared with controls.NEW & NOTEWORTHY We investigate the effect of moderate exercise after a high or a low glycemic index meal on nadir glucose and glucoregulatory hormones in gastric bypass-operated individuals and in matched unoperated controls. Cycling shortly after meal intake did not increase the risk of hypoglycemia in operated individuals. The low glycemic index meal increased glucose nadir and reduced excursions compared with the high glycemic index meal. Operated individuals had lower postexercise glucagon responses compared with controls.

At some vaccination centres, it seems to be a clinical observation, that there is a reduction in the number of attacks of herpes labialis following yellow fever vaccination. We therefore conducted a double blind, prospective, randomized study to evaluate the efficacy of yellow fever vaccination against recurrent herpes labialis. Twenty-four patients with culture proven herpes labialis were allocated to either yellow fever vaccination or placebo (saline), with 12 persons in each group. After vaccination/ placebo the patients were followed for one year. The patients returned a letter every other month with information concerning the number of attacks during the period. Comparison of the data from the two groups after one year revealed no significant difference in the number of attacks between the two groups. In conclusion evaluation of the data from this study does not confirm the clinical observation that yellow fever vaccination may act as prophylaxis against herpes labialis.

Recent studies demonstrate that adaptations to white adipose tissue are important components of the beneficial effects of exercise training on metabolic health. Exercise training favorably alters the phenotype of subcutaneous inguinal white adipose tissue (iWAT) in male mice including decreasing fat mass, improving mitochondrial function, inducing beiging and stimulating the secretion of adipokines. Here, we find that despite performing more voluntary wheel running compared to males, these adaptations do not occur in the iWAT of female mice. Consistent with sex-specific adaptations, we report that mRNA expression of androgen receptor co-activators are upregulated in iWAT from trained male mice, and that testosterone treatment of primary adipocytes derived from the iWAT of male, but not female mice, phenocopies exercise-induced metabolic adaptations. Sex-specificity also occurs in the secretome profile, as we identify Cysteine Rich Secretory Protein 1(&lt;i&gt;Crisp1&lt;/i&gt;) as a novel adipokine that is only secreted from male iWAT in response to exercise. &lt;i&gt;Crisp1&lt;/i&gt; expression is upregulated by testosterone and functions to increase glucose and fatty acid uptake. Our finding that adaptations to iWAT with exercise training are dramatically greater in male mice has potential clinical implications for understanding the different metabolic response to exercise training in males and females, and demonstrates the importance of investigating both sexes in studies of adipose tissue biology.

Abstract Background Maintenance and regeneration of functional skeletal muscle are dependent on a sufficient pool of muscle stem cells (MuSCs). During ageing there is a functional decline in this cellular pool which influences the regenerative capacity of skeletal muscle. Preclinical evidence have suggested that Nicotinamide Riboside (NR) and Pterostilbene (PT) can improve muscle regeneration e.g. by increasing MuSC function. The objective of the present study was to investigate if NRPT supplementation promotes skeletal muscle regeneration after muscle injury in elderly humans by improved recruitment of MuSCs. Methods In a randomized, double-blinded, placebo-controlled trial, 32 elderly men and women (55-80 yr) received daily supplementation with either NRPT (1000 mg NR + 200 mg PT) or matched placebo. Two weeks after initiation of supplementation, a skeletal muscle injury was applied in the vastus lateralis part of the quadriceps femoris muscle by electrically induced eccentric muscle work in a dynamometer. Skeletal muscle biopsies were obtained pre, 2h, 2, 8, and 30 days post injury. The main outcome of the study was change in MuSC content 8 days post injury. Results 31 enrolled subjects completed the entire protocol. The muscle work induced a substantial skeletal muscle injury in the study participants and was associated with release of myoglobin and creatine kinase, muscle soreness, tissue edema, and a decrease in muscle strength. MuSC content increased by 107% 8 days post injury (p= 0.0002) but with no effect of NRPT supplementation (p=0.58 for supplementation effect). MuSC proliferation and cell size revealed a large demand for recruitment post injury but was not affected by NRPT. Furthermore, histological analyses of muscle fiber area, internal nuclei and embryonic Myosin Heavy Chain showed no effect of NRPT supplementation. Conclusion Daily supplementation with 1000 mg NR + 200 mg PT is safe but does not improve recruitment of the MuSC pool or other measures of muscle recovery in response to injury or subsequent regeneration in elderly subjects.

Gluco-regulatory disturbances such as hepatic insulin resistance, hyperinsulinemia and prediabetes are commonly present in patients with nonalcoholic fatty liver disease (NAFLD) and those individuals may over time develop full-blown type 2 diabetes. Chronic liver diseases­ such as NAFLD and autoimmune liver diseases (AILDs) are heterogenous but may affect glucose-metabolism similarly. It is, however, unknown if AILDs—such as primary biliary cholangitis (PBC) —display gluco-regulatory impairments. We therefore investigated glucose and hormonal responses during a 75 g oral glucose tolerance test (OGTT) in patients with biopsy-verified, non-cirrhotic PBC (n = 9, age 55 ± y (mean ± sd) , BMI 31 ± 6 kg/m2 (mean ± sd)) , NAFLD (n = 6, age 38 ± 17 y, BMI 31 ± 4 kg/m2) and healthy controls (n = 8, age 23 ± 3 y, BMI 23 ± 2 kg/m2) . None of the participants had diabetes. In the PBC group, 3 had NAFLD. Fasting glucose, c-peptide and insulin levels were significantly increased in PBC and NAFLD compared with healthy controls ([mean (95 % CI) ]; glucose (mM) 5.6 (4.7-6.7) , 5.7 (5.2-6.1) , 4.7 (3.9-5.6) ; c-peptide (pM) 993 (556-1773) , 1334 (1036-1719) , 483 (268-869) ; insulin (pM) 98 (33-298) , 166 (103-267) , 43 (14-136) ; respectively) . Hepatic insulin resistance (reflected by fasting homeostasis model assessment of insulin resistance (HOMA-IR)) was present in PBC (mean 4.0 (95 % CI 1.2-13.9)) and NAFLD (7.0 (4.1-11.9)) but not in healthy controls (1.5 (0.4-5.4)) . There was no significant difference in glucose levels between the groups. Beta-cell secretion (c-peptide) was significantly increased in PBC and NAFLD. Insulin responses were higher in PBC and NAFLD compared with healthy but only reached statistical significance in NAFLD. Our data suggest that patients with PBC have gluco-regulatory disturbances including hepatic insulin resistance and impaired beta-cell function. Metabolic dysfunction of PBC may be underestimated and warrant further investigation. Disclosure A.H. Jensen: None. H. Ytting: Other Relationship; Gilead Sciences, Inc. J. Grandt: None. M.P. Werge: None. E.B. Rashu: None. L.E. Hetland: None. A. Junker: None. L. Hobolth: None. C. Mortensen: None. F. Tofteng: None. M. Vyberg: None. R. Serizawa: Consultant; Merck Sharp &amp; Dohme Corp. L. Gluud: Advisory Panel; Novo Nordisk. Consultant; Pfizer Inc. Research Support; Alexion Pharmaceuticals, Inc., Gilead Sciences, Inc., Novo Nordisk, Sobi. N.J. Wewer Albrechtsen: Research Support; Mercodia AB, Novo Nordisk, Regeneron Pharmaceuticals Inc. Speaker’s Bureau; Merck &amp; Co., Inc., Mercodia AB. Funding Nicolai J. Wewer Albrechtsen were financed by NNF Excellence Emerging Investigator Grant – Endocrinology and Metabolism (Application No. NNF19OC0055001) , EFSD Future Leader Award (NNF21SA0072746) and DFF Sapere Aude

Glucagon is essential for glucose control and increased levels of glucagon (hyperglucagonemia) observed in patients with type 2 diabetes contribute to their hyperglycemia. Recently, hyperglucagonemia has also been found in individuals with non-alcoholic fatty liver disease (NAFLD) as well as impaired actions of glucagon on amino acid catabolism. Whether glucagon actions on hepatic glucose production are impaired is unknown. We investigated the acute effects of a single bolus of glucagon (0.2mg) on glucose dynamics in 18 normoglycemic individuals (age: 51±3 years, BMI; 31± 0.8kg/m2, hepatic fat content: 20±2%, fasting glucose: 5.5±0.1mM) with magnetic resonance imaging verified NAFLD and 22 controls (age: 38±3 years, BMI; 24± 0.8kg/m2, hepatic fat content: 4±0.1%, fasting glucose: 5.0±0.1mM) . On a separate day, a mixture of amino acids (14 g/L; 331 mg/min/kg body weight) was infused intravenously for 45min to evaluate the actions of endogenous glucagon on glucose dynamics. Glucose levels (see figure) were significantly increased in individuals with NAFLD 60min after the glucagon bolus and during the amino acid infusion with a maximal difference of 0.5mM 30min into the infusion. These data suggest that the actions of glucagon on hepatic glucose production are not impaired by NAFLD. Therefore, the hyperglucagonemia in patients with NAFLD may constitute a diabetogenic risk factor. Disclosure S.Kjeldsen: None. H.Vilstrup: None. F.V.Schiødt: Advisory Panel; Novo Nordisk. A.Møller: None. E.B.Rashu: None. L.Gluud: Advisory Panel; Novo Nordisk, Consultant; Pfizer Inc., Research Support; Alexion Pharmaceuticals, Inc., Gilead Sciences, Inc., Novo Nordisk, Sobi. S.B.Haugaard: None. J.J.Holst: Advisory Panel; Novo Nordisk, Board Member; Antag Therapeutics, Bainan Biotech. J.Rungby: Advisory Panel; Abbott, Boehringer Ingelheim International GmbH, Speaker’s Bureau; AstraZeneca, Bayer AG, Novo Nordisk, Pfizer Inc. N.J.Wewer albrechtsen: Research Support; Mercodia AB, Novo Nordisk, Regeneron Pharmaceuticals Inc., Speaker’s Bureau; Merck &amp; Co., Inc., Mercodia AB. N.J.Jensen: None. M.Nilsson: None. N.Heinz: None. J.D.Nybing: None. F.H.Linden: None. E.Høgh-schmidt: n/a. M.P.Boesen: None. S.Madsbad: None. Funding NNF Excellence Emerging Investigator Grant – Endocrinology and Metabolism (Application No. NNF19OC0055001) , EFSD Future Leader Award (NNF21SA0072746) and DFF Sapere Aude.

Abstract Introduction A physiological feedback system exists between hepatocytes and the alpha cells termed the liver-alpha cell axis and signifies the role between amino acid-stimulated glucagon secretion and glucagon-stimulated amino acid catabolism. Several reports indicate that metabolic diseases such as non-alcoholic fatty liver disease (NAFLD) disrupts this feedback system, because of impaired glucagon receptor (GCGR) signaling (glucagon resistance). However, no experimental test exists to assess glucagon resistance in humans. Objective To develop and evaluate a test for measuring glucagon sensitivity towards amino acid and glucose metabolism in humans. Methods and analysis The study protocol is based on several pilot studies presented in this paper. The study will include 65 participants including 20 individuals with a BMI 18.6-25 kg/m 2 , 30 individuals with a BMI ≥25-40 kg/m 2 , and 15 individuals with type 1 diabetes with a BMI between 18.6-40 kg/m 2 . Participants will be grouped according to their percentage of hepatic steatosis measured by whole-liver magnetic resonance imaging (MRI). The primary outcome measure will be differences in a novel ‘glucagon sensitivity’ index between individuals with and without hepatic steatosis (&lt;5.6 % vs ≥5.6 %) without diabetes. Secondary outcomes include between-group differences regarding the glucagon-alanine-index, incremental and decremental area under the curve (AUC) and association analyses between hepatic steatosis and glucagon sensitivity. This report describes the design of the cross-sectional study currently taking place at Copenhagen University hospital Bispebjerg and Frederiksberg. Results These data will be published in peer-reviewed scientific journals and presented at scientific conferences. Ethics and dissemination The study was approved by the scientific-ethical committee of the Capital region of Denmark (H-20023717) and registered with Danish Data protection Agency (P-2021-39) and ClinicalTrials.gov ( NCT04907721 ). Written and oral consent will be obtained from all participants, and the study will adhere to the principles of the Declaration of Helsinki. Strengths and limitations of this study The glucagon sensitivity test is based on several pilot experiments Liver fat is based on whole-liver imaging and not region of interest (ROI) The glucagon sensitivity test may be limited to assess glucagon sensitivity towards amino acid catabolism and glucose production The glucagon sensitivity test does not use amino acid or glucose tracers which expands the generalizability of such test but also may impair its accuracy

This double-blind cross-over study was performed to investigate whether the lipoproteins in plasma were different on furosemide (Lasix Retard) and thiazide (hydrochlorthiazide) treatment in patients suffering from type II diabetes. Twenty-four patients were randomly allocated to either furosemide-hydrochlorthiazide (LR-HCT) or HCT-LR treatment. The treatment period was 12 months: 6 months on each sequence. After inclusion, the patients were seen every second month. Laboratory data were recorded at each visit. The only significant treatment effect was observed for high-density-lipoprotein3 cholesterol concentration (HDL3 cholesterol concentration), which was higher when patients were on furosemide therapy (p less than 0.05). We conclude from the present study that blood-glucose HbA1c, and the concentration of lipoproteins connected to development of atherosclerosis is unaffected whether type II diabetes patients are treated with HCT or furosemide.

In the present study lipoprotein fractions were studied in clinically well-characterized patients with type 2 diabetes. Apart from the usually isolated fractions (VLDL, LDL and HDL) a subfraction of VLDL was studied. This VLDL-subfraction (VLDL-2) has a similar chemical composition to beta-VLDL which has been suggested to be of importance in the development of atherosclerosis. The results shows that obese type 2 diabetic women carry substantial amounts of VLDL-2 (greater than 100% above the level of comparable non-diabetic women), (2p less than 0.01 for cholesterol and triglyceride amounts in VLDL-2). Furthermore obese diabetic women had total plasma cholesterol values 27% higher than non-diabetic obese women (2p less than 0.05). Total plasma triglyceride values were found to be significantly higher in diabetic women compared with non-diabetic women (2p less than 0.01). LDL-cholesterol was found significantly raised in obese diabetic women compared with the corresponding group of non-diabetic women (2p less than 0.05). HDL-cholesterol showed lower values in ideal-weight diabetic women than in ideal-weight non-diabetic women (2p less than 0.05). In summary, the results show that obese women with type 2 diabetes carry high concentrations of lipoprotein fractions which have been incriminated as being of importance in the development of atherosclerosis. This is in accordance with the sex ratio of atherosclerotic disease among diabetic patients compared to the sex ratio in non-diabetic patients.

Recent studies demonstrate that adaptations to white adipose tissue are important components of the beneficial effects of exercise training on metabolic health. Exercise training favorably alters the phenotype of subcutaneous inguinal white adipose tissue (iWAT) in male mice including decreasing fat mass, improving mitochondrial function, inducing beiging and stimulating the secretion of adipokines. Here, we find that despite performing more voluntary wheel running compared to males, these adaptations do not occur in the iWAT of female mice. Consistent with sex-specific adaptations, we report that mRNA expression of androgen receptor co-activators are upregulated in iWAT from trained male mice, and that testosterone treatment of primary adipocytes derived from the iWAT of male, but not female mice, phenocopies exercise-induced metabolic adaptations. Sex-specificity also occurs in the secretome profile, as we identify Cysteine Rich Secretory Protein 1(&lt;i&gt;Crisp1&lt;/i&gt;) as a novel adipokine that is only secreted from male iWAT in response to exercise. &lt;i&gt;Crisp1&lt;/i&gt; expression is upregulated by testosterone and functions to increase glucose and fatty acid uptake. Our finding that adaptations to iWAT with exercise training are dramatically greater in male mice has potential clinical implications for understanding the different metabolic response to exercise training in males and females, and demonstrates the importance of investigating both sexes in studies of adipose tissue biology.