Dose-response effects on HbA1c and bodyweight reduction of survodutide, a dual glucagon/GLP-1 receptor agonist, compared with placebo and open-label semaglutide in people with type 2 diabetes: a randomised clinical trial.

AIMS/HYPOTHESIS: The aim of this study was to assess the dose-response effects of the subcutaneous glucagon receptor/glucagon-like peptide-1 receptor dual agonist survodutide (BI 456906) on HbA1c levels and bodyweight reduction. METHODS: This Phase II, multicentre, randomised, double-blind, parallel-group, placebo-controlled study, conducted in clinical research centres, assessed survodutide in participants aged 18-75 years with type 2 diabetes, an HbA1c level of 53-86 mmol/mol (7.0-10.0%) and a BMI of 25-50 kg/m2 on a background of metformin therapy. Participants were randomised via interactive response technology to receive survodutide (up to 0.3, 0.9, 1.8 or 2.7 mg once weekly [qw; dose group (DG) 1-4, respectively] or 1.2 or 1.8 mg twice weekly [DG 5 and 6, respectively]), placebo or semaglutide (up to 1.0 mg qw). Participants and all those involved in the trial conduct/analysis were blinded; the semaglutide arm was open-label. The primary endpoint was absolute change from baseline in HbA1c after 16 weeks' treatment. The key secondary endpoint was relative change from baseline in bodyweight after 16 weeks' treatment. RESULTS: A total of 413 participants were randomised (DG1, n=50; DG2, n=50; DG3, n=52; DG4, n=50; DG5, n=51; DG6, n=50; semaglutide, n=50; placebo, n=60). The full analysis set comprised 411 treated participants (DG6, n=49; placebo, n=59). Adjusted mean (95% CI) HbA1c decreased from baseline (mean ± SD 64.7±9.2 mmol/mol [8.07±0.84%] after 16 weeks' treatment: DG1 (n=41), -9.92 mmol/mol (-12.27, -7.56; -0.91% [-1.12, -0.69]); DG2 (n=46), -15.95 mmol/mol (-18.27, -13.63; -1.46% [-1.67, -1.25]); DG3 (n=36), -18.72 mmol/mol (-21.15, -16.29; -1.71% [-1.94, -1.49]); DG4 (n=33), -17.01 mmol/mol (-19.59, -14.43; -1.56% [-1.79, -1.32]); DG5 (n=44), -17.84 mmol/mol (-20.18, -15.51; -1.63% [-1.85, -1.42]); DG6 (n=36), -18.38 mmol/mol (-20.90, -15.87; -1.68% [-1.91, -1.45]). The mean reduction in HbA1c was similar with low-dose survodutide (DG2: -15.95 mmol/mol [-1.46%]; n=46) and semaglutide (-16.07 mmol/mol [-1.47%]; n=45). Mean (95% CI) bodyweight decreased dose-dependently up to -8.7% (-10.1, -7.3; DG6, n=37); survodutide ≥1.8 mg qw produced greater bodyweight reductions than semaglutide (-5.3% [-6.6, -4.1]; n=45). Adverse events (AEs) were reported for 77.8% of survodutide-treated participants (mainly gastrointestinal), 52.5% receiving placebo and 52.0% receiving semaglutide. CONCLUSIONS/INTERPRETATION: Survodutide reduced HbA1c levels and bodyweight after 16 weeks' treatment in participants with type 2 diabetes. Dose-related gastrointestinal AEs could be mitigated with slower dose escalations. TRIAL REGISTRATION: ClinicalTrials.gov NCT04153929 and EudraCT 2019-002390-60. FUNDING: Boehringer Ingelheim Pharma GmbH & Co. KG, Ingelheim, Germany.

Acylated- and unacylated ghrelin during an oral glucose tolerance test in humans at risk for type 2 diabetes mellitus.

BACKGROUND/OBJECTIVES: The orexigenic peptide hormone ghrelin has been implicated in the pathophysiology of obesity and type 2 diabetes mellitus through its effects on nutrient homeostasis. Ghrelin is subject to a unique post-translational acyl modification regulating its biochemical activity. SUBJECTS/METHODS: In this study we aimed to investigate the relation of acylated (AcG) as well as unacylated ghrelin (UnG) with body weight and insulin resistance in the fasting (n = 545) and post-oral glucose tolerance test (oGTT) state (n = 245) in a metabolically well characterized cohort covering a broad range of BMI (17.95 kg/m²-76.25 kg/m²). RESULTS: Fasting AcG (median 94.2 pg/ml) and UnG (median 175.3 pg/ml) were negatively and the AcG/UnG ratio was positively correlated with BMI (all p < 0.0001). Insulin sensitivity (ISI) correlated positively with AcG (p = 0.0014) and UnG (p = 0.0004) but not with the AcG/UnG ratio. In a multivariate analysis, including ISI and BMI, only BMI, but not ISI was independently associated with AcG and UnG concentrations. Significant changes of AcG and UnG concentrations were detectable after oGTT stimulation, with slight decreases after 30 min and increases after 90-120 min. Subject stratification into BMI-divergent groups revealed more pronounced AcG increases in the two groups with BMI < 40 kg/m². CONCLUSION: Our data demonstrate lower concentrations for both AcG and UnG with increasing BMI as well as an increased proportion of the biologically active, acylated form of ghrelin giving point to pharmacologic intervention in ghrelin acylation and/or increase in UnG for treatment of obesity despite decreased absolute AcG levels.

A randomized Phase I study of the safety, tolerability, pharmacokinetics and pharmacodynamics of BI 456906, a dual glucagon receptor/glucagon-like peptide-1 receptor agonist, in healthy Japanese men with overweight/obesity.

AIM: To report a Phase I study of subcutaneous glucagon receptor (GCGR)/glucagon-like peptide-1 receptor (GLP-1R) dual agonist BI 456906 versus placebo in healthy Japanese men with overweight/obesity. MATERIALS AND METHODS: We investigated multiple rising doses of BI 456906 escalated over 16 weeks (maximum doses: 1.8 mg once weekly [dose group {DG} 1], 4.8 mg once weekly [DG 2] and 2.4 mg twice weekly [DG 3]) in Japanese men with a body mass index of 23 to 40 kg/m2 . RESULTS: Thirty-six participants were treated (n = 9 per DG and placebo). Overall, 10 participants (37.0%) treated with BI 456906 withdrew from dose escalation due to adverse events (amylase increase, n = 1; decreased appetite, n = 9), and the proportion of participants was higher in DG 2 (n = 6, 66.7%) versus DGs 1 and 3 (both n = 2, 22.2%). No participants receiving placebo withdrew from dose escalation. BI 456906 exposure increased with dose and dose escalation in each DG. Treatment with BI 456906 decreased placebo-corrected bodyweight after 16 weeks (placebo +1.06%): DG 1, -5.57%; DG 2, -12.37%; DG 3, -9.62%. Paracetamol absorption decreased in Week 1 for DGs 2 and 3, indicating transient delayed gastric emptying. BI 456906 reduced plasma alanine and glucagon levels, indicating indirect target engagement at GCGRs and GLP-1Rs. Drug-related adverse events were reported for all participants receiving BI 456906 and four receiving placebo, the most frequent being decreased appetite (n = 24, 66.7%). CONCLUSIONS: BI 456906 showed no unexpected tolerability concerns and it reduced placebo-corrected bodyweight by up to 12.37% in Japanese men with overweight/obesity after 16 weeks of treatment.

Phase I studies of the safety, tolerability, pharmacokinetics and pharmacodynamics of the dual glucagon receptor/glucagon-like peptide-1 receptor agonist BI 456906.

AIM: To report two phase I studies of the novel subcutaneous glucagon-like peptide-1 receptor/glucagon receptor (GLP-1R/GCGR) dual agonist BI 456906 versus placebo in healthy volunteers and people with overweight/obesity. MATERIALS AND METHODS: A phase Ia study (NCT03175211) investigated single rising doses (SRDs) of BI 456906 in 24 males with a body mass index (BMI) of 20-<30 kg/m2 . A phase Ib study (NCT03591718) investigated multiple rising doses (MRDs) of BI 456906 (escalated over 6 [Part A] or 16 [Part B] weeks) in 125 adults with a BMI of 27-40 kg/m2 . RESULTS: In the SRD study (N = 24), mean body weight decreased with increasing BI 456906 dose. In the MRD study, the maximum decreases in placebo-corrected mean body weight were at week 6 (-5.79%, dosage schedule [DS] 1; Part A) and week 16 (-13.8%, DS7; Part B). BI 456906 reduced plasma amino acids and glucagon, indicating target engagement at GCGRs and GLP-1Rs. Drug-related adverse events (AEs) increased with BI 456906 dose. The most frequent drug-related AE with SRDs was decreased appetite (n = 9, 50.0%), and two subjects (8.3%) did not complete the trial because of AEs (nausea and vomiting). During MRD Part A (N = 80), 10 subjects (12.5%) discontinued BI 456906, most commonly because of a cardiac or vascular AE (n = 6, 7.5%); during Part B (N = 45), eight subjects (17.8%) discontinued BI 456906, mainly because of AEs (n = 6, 13.3%), most commonly gastrointestinal disorders. CONCLUSIONS: BI 456906 produced a placebo-corrected body weight loss of 13.8% (week 16), highlighting its potential to promote clinically meaningful body weight loss in people with overweight/obesity.

Protein kinase Cδ regulates nuclear export of FOXO1 through phosphorylation of the chaperone 14-3-3ζ.

AIMS/HYPOTHESIS: Forkhead box protein O1 (FOXO1) is a transcription factor essential for beta cell fate. Protein kinase B-dependent phosphorylation of FOXO1 at S256 (P-FOXO1) enables its binding to 14-3-3 dimers and nuclear export. Dephosphorylated FOXO1 enters nuclei and activates pro-apoptotic genes. Since our previous observations suggest that protein kinase C delta (PKCδ) induces nuclear accumulation of FOXO1, the underlying mechanism was examined. METHODS: In human islets, genetically modified mice and INS-1E cells apoptosis was assessed by TUNEL staining. Subcellular translocation of proteins was examined by confocal microscopy and signalling pathways were analysed by western blotting and overlay assay. RESULTS: In PKCδ-overexpressing (PKCδ-tg) mouse islet cells and INS-1E cells FOXO1 accumulated in nuclei, surprisingly, as P-FOXO1. PKCδ-tg decelerated IGF-1-dependent stimulation of nuclear export, indicating that changes in export caused nuclear retention of P-FOXO1. Nuclear accumulation of P-FOXO1 was accompanied by increased phosphorylation of 14-3-3ζ at S58 and reduced dimerisation of 14-3-3ζ. Palmitic acid further augmented phosphorylation of 14-3-3ζ and triggered nuclear accumulation of FOXO1 in both INS-1E and human islet cells. Furthermore, the overexpression of a phosphomimicking mutant of 14-3-3ζ (S58D) enhanced nuclear FOXO1. In accordance with the nuclear accumulation of P-FOXO1, PKCδ overexpression alone did not increase apoptotic cell death. Additionally, insulin secretion and glucose homeostasis in PKCδ-overexpressing mice remained unaffected. CONCLUSIONS/INTERPRETATION: These results suggest that PKCδ-mediated phosphorylation of 14-3-3ζ contributes to the nuclear retention of FOXO1, even when FOXO1 is phosphorylated as under non-stress conditions. P-FOXO1 does not induce pro-apoptotic genes, but may rather exert beneficial effects on beta cells.

Perspectives in weight control in diabetes - Survodutide.

Glucagon-like peptide-1 receptor (GLP-1R) agonists are approved treatments for Type 2 diabetes mellitus, with liraglutide and semaglutide also approved for the treatment of obesity. The natural gut hormone oxyntomodulin is a weak dual agonist of the glucagon receptor (GCGR) and GLP-1R. Development of poly-agonists mimicking oxyntomodulin, such as the novel dual GCGR/GLP-1R agonist survodutide, represents an important step towards a more effective treatment for people with Type 2 diabetes mellitus and obesity. Survodutide is a 29-amino acid peptide derived from glucagon, with the incorporation of potent GLP-1 activities. It contains a C18 diacid which mediates binding to albumin, thereby prolonging the half-life to enable once-weekly subcutaneous dosing. The utilisation of GCGR agonism aims to enhance body weight-lowering effects by increasing energy expenditure in addition to the anorectic action of GLP-1R agonists. Glucose-lowering efficacy of survodutide has been demonstrated in a Phase II trial in patients with Type 2 diabetes mellitus and obesity and was associated with clinically meaningful body weight loss. These data highlight the potential of dual GCGR/GLP-1R agonism for reducing glycated haemoglobin and body weight in patients with Type 2 diabetes mellitus, and for greater therapeutic efficacy compared with GLP-1R agonism alone.

Glucagon and GLP-1 receptor dual agonist survodutide for obesity: a randomised, double-blind, placebo-controlled, dose-finding phase 2 trial.

BACKGROUND: Obesity is a widespread and chronic condition that requires long-term management; research into additional targets to improve treatment outcomes remains a priority. This study aimed to investigate the safety, tolerability, and efficacy of glucagon receptor-GLP-1 receptor dual agonist survodutide (BI 456906) in obesity management. METHODS: In this randomised, double-blind, placebo-controlled, dose-finding phase 2 trial conducted in 43 centres in 12 countries, we enrolled participants (aged 18-75 years, BMI ≥27 kg/m2, without diabetes) and randomly assigned them by interactive response technology (1:1:1:1:1; stratified by sex) to subcutaneous survodutide (0·6, 2·4, 3·6, or 4·8 mg) or placebo once-weekly for 46 weeks (20 weeks dose escalation; 26 weeks dose maintenance). The primary endpoint was the percentage change in bodyweight from baseline to week 46. Primary analysis included the modified intention-to-treat population (defined as all randomly assigned patients who received at least one dose of trial medication and who had analysable data for at least one efficacy endpoint) and was based on the dose assigned at randomisation (planned treatment), including all data censored for COVID-19-related discontinuations; the sensitivity analysis was based on the actual dose received during maintenance phase (actual treatment) and included on-treatment data. Safety analysis included all participants who received at least one dose of study drug. The trial is registered with ClinicalTrials.gov (NCT04667377) and EudraCT (2020-002479-37). FINDINGS: Between March 30, 2021, and Nov 11, 2021, we enrolled 387 participants; 386 (100%) participants were treated (0·6 mg, n=77; 2·4 mg, n=78; 3·6 mg, n=77; 4·8 mg, n=77; placebo n=77) and 233 (60·4%) of 386 completed the 46-week treatment period (187 [61%] of 309 receiving survodutide; 46 [60%] of 77 receiving placebo). When analysed according to planned treatment, mean (95% CI) changes in bodyweight from baseline to week 46 were -6·2% (-8·3 to -4·1; 0·6 mg); -12·5% (-14·5 to -10·5; 2·4 mg); -13·2% (-15·3 to -11·2; 3·6 mg); -14·9% (-16·9 to -13·0; 4·8 mg); -2·8% (-4·9 to -0·7; placebo). Adverse events occurred in 281 (91%) of 309 survodutide recipients and 58 (75%) of 77 placebo recipients; these were primarily gastrointestinal in 232 (75%) of 309 survodutide recipients and 32 (42%) of 77 placebo recipients. INTERPRETATION: All tested survodutide doses were tolerated, and dose-dependently reduced bodyweight. FUNDING: Boehringer Ingelheim.

Prevalence of Overweight and Obesity, Its Complications, and Progression in a 10-Year Follow-Up in the Gutenberg Health Study (GHS).

INTRODUCTION: Overweight and obesity lead to numerous complications and their treatment. The associated costs represent a health and sociopolitical burden. Therefore, the development of overweight and obesity is of great importance for health policy. METHODS: The Gutenberg Health Study (GHS), a population-based observational study of individuals aged 35-74 years in the city of Mainz and the district of Mainz-Bingen, examined current data on the prevalence and development of overweight and obesity and their association with concomitant diseases and medication use. RESULTS: Among men, 48.1% were overweight and 26.3% had obesity. Among women, these proportions were 32.1% and 24.1%, respectively. Elevated body mass index (BMI) was associated with numerous complications, particularly insulin resistance and type 2 diabetes, arterial hypertension, elevated triglycerides and low HDL cholesterol, and cardiovascular disease. Accordingly, medications to treat these conditions were used significantly more often in individuals with elevated BMI. During the 10-year observation period, mean weight increased in the population. Both men and women had a moderate but significant increase in BMI compared to men and women of the same age at baseline. Individual weight changes over the 10-year observation period, on the other hand, were age-dependent. In the two younger age decades, weight gain was observed, while in the oldest age decade, mean body weight decreased. CONCLUSION: These current data confirm that overweight and obesity are associated with relevant complications and that these complications lead to significant use of appropriate medications. The study also suggests that there is a significant trend toward increased prevalence of obesity (BMI ≥30) over the 10-year period.

Obesity-related inflammatory protein signature in cardiovascular clinical outcomes: results from the Gutenberg Health Study.

OBJECTIVE: The objective of this study was to investigate whether an obesity-related inflammatory protein signature (OIPS) is associated with adverse cardiovascular events. METHODS: The Olink Target 96 Inflammation panel was performed in 6662 participants from the population-based Gutenberg Health Study (GHS). The OIPS was selected by a logistic regression model, and its association with cardiovascular outcomes was evaluated by Cox regression analysis. The GHS-derived OIPS was externally validated in the MyoVasc study. RESULTS: The identified OIPS entailed 21 proteins involved in chemokine activity, tumor necrosis factor (TNF) receptor binding, and growth factor receptor binding. The signature revealed a novel positive association of axis inhibition protein 1 with obesity. The OIPS was associated with increased risk of all-cause and cardiac deaths, major adverse cardiovascular events, and incident coronary artery disease, independent of clinical covariates and established risk instruments. A BMI-stratified analysis confirmed the association of OIPS with increased death in those with obesity and overweight and with increased risk for coronary artery disease in those with obesity. The association of OIPS with increased risk of all-cause and cardiac deaths was validated in the MyoVasc cohort. CONCLUSIONS: The OIPS showed a significant association with adverse clinical outcomes, particularly in those with overweight and obesity, and represents a promising tool for identifying patients at higher risk for worse cardiovascular outcomes.

Toll-like receptors 2 and 4 impair insulin-mediated brain activity by interleukin-6 and osteopontin and alter sleep architecture.

Impaired insulin action in the brain represents an early step in the progression toward type 2 diabetes, and elevated levels of saturated free fatty acids are known to impair insulin action in prediabetic subjects. One potential mediator that links fatty acids to inflammation and insulin resistance is the Toll-like receptor (TLR) family. Therefore, C3H/HeJ/TLR2-KO (TLR2/4-deficient) mice were fed a high-fat diet (HFD), and insulin action in the brain as well as cortical and locomotor activity was analyzed by using telemetric implants. TLR2/4-deficient mice were protected from HFD-induced glucose intolerance and insulin resistance in the brain and displayed an improvement in cortical and locomotor activity that was not observed in C3H/HeJ mice. Sleep recordings revealed a 42% increase in rapid eye movement sleep in the deficient mice during daytime, and these mice spent 41% more time awake during the night period. Treatment of control mice with a neutralizing IL-6 antibody improved insulin action in the brain as well as cortical activity and diminished osteopontin protein to levels of the TLR2/4-deficient mice. Together, our data suggest that the lack of functional TLR2/4 protects mice from a fat-mediated impairment in insulin action, brain activity, locomotion, and sleep architecture by an IL-6/osteopontin-dependent mechanism.

Stratification of obesity phenotypes to optimize future therapy (SOPHIA).

INTRODUCTION: Obesity is considered a poor lifestyle choice. 'Obesity' is not a sufficient definition for patients, any more than 'cancer' or 'arthritis' would be. A major obstacle is the lack of understanding of pathogenesis. The disease of obesity is considered homogenous, while response to treatment is thought of as heterogeneous. This can change if pathogenesis, risk profiles for complications, and treatment responses are viewed within the context of obesity consisting of several subsets of disease. AREAS COVERED: The European Union-funded Innovative Medicine Initiative project Stratification of Obesity Phenotypes to Optimize Future Obesity Therapy is part of a momentum shift. Operational variables are being used to develop tests and therapies which may allow the prediction of risk of obesities and the prediction of response to obesity treatments. However, changing stakeholder perspectives on obesity may require more than high-quality data and analysis. EXPERT OPINION: For patients to benefit, clinicians need to integrate evidence-based treatments and payers need to reimburse the management of the disease of obesity. This will generate commercial opportunities for industry. We need to involve stakeholders (patients, clinicians, regulators, payer, patient organisations) to create a shared value for mutual gain.

Leptin affects insulin action in astrocytes and impairs insulin-mediated physical activity.

BACKGROUND/AIMS: Impaired insulin action is an early event in the pathogenesis of obesity and type 2-diabetes, and among the metabolic confounders in obese, hyperleptinaemia is constantly present; however its impact on insulin action in the brain and locomotor activity is unknown. METHODS: We examined insulin action by Western Blot analysis and glycogen synthesis in primary astrocytes and brain tissue and detected locomotion in C57BL/6 mice. The insulin-mediated desire to move was evaluated in healthy volunteers and correlated to leptin levels. RESULTS: Leptin treatment led to a significant decrease in insulin-mediated phosphorylation of the insulin receptor and Akt473 which was accompanied by a decline in glycogen synthesis in primary astrocytes and significantly decreased insulin-induced phosphorylation of the insulin receptor and insulin receptor substrate-2 in brain tissues of mice. Intracerebroventricular insulin failed to promote locomotion in the presence of elevated leptin levels. Lean human subjects reported an increase in the desire to move following insulin which failed in obese and there was an inverse correlation between the insulin-mediated desire to move and leptin levels. CONCLUSIONS: Our data suggest a crosstalk of leptin and insulin in the brain which leads to a decline in locomotor activity. This might represent a molecular mechanism in obese to inhibit physical activity.

Insulin-induced serine phosphorylation of IRS-2 via ERK1/2 and mTOR: studies on the function of Ser675 and Ser907.

The identity of specific serine phosphorylation residues of insulin receptor substrate (IRS)-2 and their impact on insulin signal transduction are largely unknown. Ser(675) and Ser(907) of mouse IRS-2 are adjacent to PI 3-kinase or Grb2 binding domains, respectively. Using monoclonal phosphosite-specific antibodies, we demonstrated the phosphorylation of both serines after stimulation of Fao hepatoma cells with insulin, anisomycin, or phorbol esters. Phosphorylation of both sites was a late and prolonged event during insulin treatment and was also detected in liver tissue of insulin-treated as well as refed mice. Inhibition and siRNA-mediated knockdown of ERK1/2 indicated that the insulin-induced phosphorylation of Ser(907) was ERK dependent. Phosphorylation of Ser(907) did not prevent the insulin-induced association of IRS-2 with Grb2, but phosphorylation of the adjacent Tyr(911) was proved to be crucial in HEK 293 cells expressing IRS-2 Ala mutants. The insulin-induced phosphorylation of Ser(675) was prevented by inhibition and siRNA-mediated knockdown of mTOR but not of p70(S6K1). Mutation of Ser(675) to Ala did not affect downstream insulin signaling but increased the half-life of the protein, suggesting an involvement of phospho-Ser(675) in an accelerated degradation of IRS-2. Moreover, the insulin-induced degradation of IRS-2 was blocked by inhibition of mTOR. We conclude that the two novel insulin-dependent serine phosphorylation sites of IRS-2 were not involved in the regulation of the adjacent PI 3-kinase and Grb2 binding domains but might be implicated in the ERK- and mTOR-mediated negative feedback control.

Challenges in tackling energy expenditure as obesity therapy: From preclinical models to clinical application.

BACKGROUND: A chronic imbalance of energy intake and energy expenditure results in excess fat storage. The obesity often caused by this overweight is detrimental to the health of millions of people. Understanding both sides of the energy balance equation and their counter-regulatory mechanisms is critical to the development of effective therapies to treat this epidemic. SCOPE OF REVIEW: Behaviors surrounding ingestion have been reviewed extensively. This review focuses more specifically on energy expenditure regarding bodyweight control, with a particular emphasis on the organs and attractive metabolic processes known to reduce bodyweight. Moreover, previous and current attempts at anti-obesity strategies focusing on energy expenditure are highlighted. Precise measurements of energy expenditure, which consist of cellular, animal, and human models, as well as measurements of their translatability, are required to provide the most effective therapies. MAJOR CONCLUSIONS: A precise understanding of the components surrounding energy expenditure, including tailored approaches based on genetic, biomarker, or physical characteristics, must be integrated into future anti-obesity treatments. Further comprehensive investigations are required to define suitable treatments, especially because the complex nature of the human perspective remains poorly understood.

Enforced expression of protein kinase C in skeletal muscle causes physical inactivity, fatty liver and insulin resistance in the brain.

Among the multitude of dysregulated signalling mechanisms that comprise insulin resistance in divergent organs, the primary events in the development of type 2 diabetes are not well established. As protein kinase C (PKC) activation is consistently present in skeletal muscle of obese and insulin resistant subjects, we generated a transgenic mouse model that overexpresses constitutively active PKC-beta(2) in skeletal muscle to test whether activation of PKC is sufficient to cause an aversive whole-body phenotype. Upon this genetic modification, increased serine phosphorylation in Irs1 was observed and followed by impaired (3)H-deoxy-glucose uptake and muscle glycogen content, and transgenic mice exhibited insulin and glucose intolerance as they age. Muscle histochemistry revealed an increase in lipid deposition (intramyocellular lipids), and transgenic mice displayed impaired expression of transcriptional regulators of genes involved in fatty acid oxidation (peroxisome proliferator-activated receptor-gamma, PGC-1beta, acyl-CoA oxidase) and lipolysis (hormone-sensitive lipase). In this regard, muscle of transgenic mice exhibited a reduced capacity to oxidize palmitate and contained less mitochondria as determined by citrate synthase activity. Moreover, the phenotype included a profound decrease in the daily running distance, intra-abdominal and hepatic fat accumulation and impaired insulin action in the brain. Together, our data suggest that activation of a classical PKC in skeletal muscle as present in the pre-diabetic state is sufficient to cause disturbances in whole-body glucose and lipid metabolism followed by profound alterations in oxidative capacity, ectopic fat deposition and physical activity.

Acute regulation of metabolic genes and insulin receptor substrates in the liver of mice by one single bout of treadmill exercise.

Acute exercise performance represents a major metabolic challenge for the skeletal muscle, but also for the liver as the most important source of energy. However the molecular adaptation of the liver to one single bout of exercise is largely unknown. C57BL/6 mice performed a 60 min treadmill run at high aerobic intensity. Liver, soleus and white gastrocnemius muscle were removed immediately after exercise. The single bout of exercise resulted in a very rapid and pronounced induction of hepatic metabolic enzymes and regulators of metabolism or transcription: glucose-6-phosphatase (G6Pase; 3-fold), pyruvate dehydrogenase kinase-4 (PDK4; 4.8-fold), angiopoietin-like 4 (2.1-fold), insulin receptor substrate (IRS)-2 (5.1-fold), peroxisome proliferator activated receptor-gamma coactivator 1alpha (PGC-1alpha; 3-fold). In soleus and white gastrocnemius muscle the up-regulation of IRS-2 and PDK4 was less pronounced compared with the liver and no significant induction of PGC-1alpha could be detected at this early time point. Activation of AMPK was found in both liver and white gastrocnemius muscle as phosphorylation of Thr-172. The induction of endogenous insulin secretion by a glucose load directly after the exercise bout resulted in a significantly higher PKB/Akt phosphorylation in the liver of exercised mice. The markedly enhanced IRS-2 protein amount, and presumably reduced serine/threonine phosphorylation of the IRS proteins induced by the acute exercise could be responsible for this enhanced action of insulin. In conclusion, acute exercise induced a rapid and pronounced transcriptional adaptation in the liver, and regulated hepatic IRS proteins leading to improved cellular insulin signal transduction.

The Gly385(388)Arg Polymorphism of the FGFR4 Receptor Regulates Hepatic Lipogenesis Under Healthy Diet.

CONTEXT: The effect of a lifestyle intervention to reduce liver fat content in nonalcoholic fatty liver disease in humans is influenced by genetics. We hypothesized that the amino acid exchange in human Gly388Arg (mouse homolog: Gly385Arg) in fibroblast growth factor receptor 4 (FGFR4), which regulates bile acid, lipid, and glucose metabolism, could determine hepatic lipid accumulation and insulin sensitivity. Mechanisms of this substitution were studied in mice under normal chow and high-fat diets. DESIGN: In humans, the Gly388Arg polymorphism was studied for its relationship with changes in liver fat content and insulin sensitivity during 9 months of a lifestyle intervention. We also studied a knock-in mouse strain with an Arg385 allele introduced into the murine FGFR4 gene under normal chow and high-fat diets. RESULTS: In humans, the FGFR4 Arg388 allele was not associated with liver fat content or insulin sensitivity in subjects who were overweight and obese before lifestyle intervention. However, it was associated with less decrease in liver fat content and less increase in insulin sensitivity during the intervention. In mice receiving normal chow, the FGFR4 Arg385 allele was associated with elevated hepatic triglyceride content, altered hepatic lipid composition, and increased hepatic expression of genes inducing de novo lipogenesis and glycolysis. Body fat mass and distribution, glucose tolerance, and insulin sensitivity were unaltered. The FGFR4 Arg385 allele had no effect on glucose or lipid metabolism under the high-fat diet. CONCLUSION: Our data indicate that the FGFR4 Arg388(385) allele affects hepatic lipid and glucose metabolism specifically during healthy caloric intake.

IGF-1 protects against dexamethasone-induced cell death in insulin secreting INS-1 cells independent of AKT/PKB phosphorylation.

Appropriate insulin secretion depends on beta-cell mass that is determined by the balance between cell proliferation and death. IGF-1 stimulates proliferation and protects against apoptosis. In contrast, glucocorticoids promote cell death. In this study we examined molecular interactions of the glucocorticoid dexamethasone (dexa) with IGF-1 signalling pathways in insulin secreting INS-1 cells. IGF-1 (50 ng/ml) increased the growth rate and stimulated BrdU incorporation, while dexa (100 nmol/l) inhibited cell growth, BrdU incorporation and induced apoptosis. Dexa-induced cell death was partially antagonized by IGF-1. This protection was further increased by LY294002 (10 micromol/l), an inhibitor of PI3 kinase. In contrast, MAP kinase inhibitor PD98059 (10 micromol/l) significantly reduced the protective effect of IGF-1. The analysis of signalling pathways by Western blotting revealed that dexa increased IRS-2 protein abundance while the expression of PI3K, PKB and ERK remained unchanged. Despite increased IRS-2 protein,IRS-2 tyrosine phosphorylation stimulated by IGF-1 was inhibited by dexa. Dexa treatment reduced basal PKB phosphorylation. However, IGF-1-mediated stimulation of PKB phosphorylation was not affected by dexa, but ERK phosphorylation was reduced. LY294002 restored IGF-1-induced ERK phosphorylation. These data suggest that dexa induces apoptosis in INS-1 cells by inhibiting phosphorylation of IRS-2, PKB and ERK. IGF-1 counteracts dexa-mediated apoptosis in the presence of reduced PKB but increased ERK phosphorylation.

Effect of insulin detemir, compared to human insulin, on 3T3-L1 adipogenesis.

Insulin detemir (DET) represents a myristic acid (MA)-coupled insulin derivative with protracted action due to reversible albumin binding. As compared to human insulin (HI), DET provokes no or only minor body weight gain in vivo. Therefore, we compared DET's and HI's adipogenic effects. 3T3-L1 preadipocytes were differentiated with 5 nmol/l HI, 5 nmol/l DET (=DET(equimolar)), or 20 nmol/l DET (=DET(equipotent); equipotent in terms of the reported metabolic potency in vitro). Due to differentiation-suppressive effects, albumin was excluded from the studies. During the induction period, only HI allowed clonal expansion. Moreover, HI induced a 200-fold increase in specific glycerol-3-phosphate dehydrogenase activity, whereas DET(equimolar) and DET(equipotent) were markedly less adipogenic (P