Insulin Sensitivity Glucose Uptake Research Articles

Activation of GFRAL+ neurons induces hypothermia and glucoregulatory responses associated with nausea and torpor

GFRAL-expressing neurons actuate aversion and nausea, are targets for obesity treatment, and may mediate metformin effects by long-term GDF15-GFRAL agonism. Whether GFRAL+ neurons acutely regulate glucose and energy homeostasis is, however, underexplored. Here, we report that cell-specific activation of GFRAL+ neurons using a variety of techniques causes a torpor-like state, including hypothermia, the release of stress hormones, a shift from glucose to lipid oxidation, and impaired insulin sensitivity, glucose tolerance, and skeletal muscle glucose uptake but augmented glucose uptake in visceral fat. Metabolomic analysis of blood and transcriptomics of muscle and fat indicate alterations in ketogenesis, insulin signaling, adipose tissue differentiation and mitogenesis, and energy fluxes. Our findings indicate that acute GFRAL+ neuron activation induces endocrine and gluco- and thermoregulatory responses associated with nausea and torpor. While chronic activation of GFRAL signaling promotes weight loss in obesity, these results show that acute activation of GFRAL+ neurons causes hypothermia and hyperglycemia.

OR33-04 Mechanistic Dissection Of Circadian-Specific Glucocorticoid Effects On Exercise Tolerance And Muscle Glucose Utilization

Abstract Disclosure: A. Prabakaran: None. M. Wintzinger: None. K. Piczer: None. K. Miz: None. A. Walton: None. H. Durumutla: None. M. Quattrocelli: None. Glucocorticoids (GCs) are circadian hormones regulating metabolism by activating the glucocorticoid receptor (GR) as pleiotropic transcription factor. Timing of exogenous GC drug regimens is emerging as a key determinant of their metabolic effects. This was shown by the surprising finding that once-weekly GC intermittence reverses many of the pro-obesogenic side effects of once-daily GC dosing in mice and humans. However, despite the intrinsic circadian nature of this signaling, the impact of time-of-day of exogenous GC dosing, i.e. chrono-pharmacology, remains remarkably unknown for metabolic physiology and exercise tolerance. Relevance of this question for humans is supported by the initial findings of increased lean mass and motor function with circadian-restricted prednisone intermittence in a recent pilot clinical trial in patients with genetic myopathies. However, mechanisms to support significance of these effects for metabolic diseases and aging remain unknown. We studied 12-week-long intermittent regimens of the exogenous GC prednisone in mice in conditions of diet-induced obesity or advanced aging, comparing dosing at light-phase start (ZT0) versus dark-phase start (ZT12). We found that, compared to dark-phase, light-phase dosing enhanced the regimen-driven increase in body-wide lean mass, muscle mass and muscle function in both obese and aged mice. Intriguingly, experiments with adiponectin-knockout mice showed that these effects were dependent on adiponectin, which promotes muscle insulin sensitivity. We then used tissue-specific inducible knockout models to dissect non-muscle versus muscle-autonomous effects of GC chrono-pharmacology. We found that the GC effects on total and high-molecular-weight adiponectin upregulation were dependent on regimen-specific engagement of the adipocyte-specific GR on the adiponectin gene promoter. Complementary to adipose-driven adiponectin production, we found that adiponectin receptor AdipoR1 upregulation in muscle depended on circadian-specific engagement of the myocyte-specific GR on the AdipoR1 promoter. Consistent with the adiponectin action on muscle insulin sensitivity, ablation of myocyte-specific GR blocked the regimen-driven effects on insulin-sensitive muscle glucose uptake and muscle mass. Furthermore, we found that light-phase GC dosing engaged the muscle GR for a non-canonical interaction with the clock factor BMAL1 to upregulate the mitochondrial regulator PGC1alpha. Indeed, we found that the regimen-driven increase in glucose oxidation and amino acid biogenesis from TCA cycle intermediates in muscle was dependent on the myocyte-specific PGC1alpha. In summary, we present here novel circadian and molecular mechanisms reconverting glucocorticoid drugs from deleterious to re-energizing agents for potential chrono-treatment of metabolic conditions and aging. Presentation: Sunday, June 18, 2023

Contribution of skeletal muscle-specific microRNA-133b to insulin resistance in heart failure.

Insulin resistance (IR) is one of the hallmarks of heart failure (HF). Abnormalities in skeletal muscle (SM) metabolism have been identified in patients with HF. However, the underlying mechanisms of IR development in SM in HF are poorly understood. Herein, we hypothesize that HF upregulates miR-133b in SM and in turn alters glucose metabolism and the propensity toward IR. Mitochondria isolated from SM of mice with HF induced by transverse aortic constriction (TAC) showed lower respiration and downregulation of muscle-specific components of the tricarboxylic acid (TCA) cycle, AMP deaminase 1 (AMPD1), and fumarate compared with those from control animals. RNA-Seq and subsequent qPCR validation confirmed upregulation of SM-specific microRNA (miRNA), miR-133b, in TAC versus sham animals. miR-133b overexpression alone resulted in significantly lower mitochondrial respiration, cellular glucose uptake, and glycolysis along with lower ATP production and cellular energy reserve compared with the scramble (Scr) in C2C12 cells. miR-133b binds to the 3'-untranslated region (UTR) of KLF15, the transcription factor for the insulin-sensitive glucose transporter, GLUT4. Overexpression of miR-133b lowers GLUT4 and lowers pAkt in presence of insulin in C2C12 cells. Finally, lowering miR-133b in primary skeletal myocytes isolated from TAC mice using antagomir-133b reversed the changes in KLF15, GLUT4, and AMPD1 compared with the scramble-transfected myocytes. Taken together, these data demonstrate a role for SM miR-133b in altered glucose metabolism in HF and suggest the therapeutic potential in HF to improve glucose uptake and glycolysis by restoring GLUT4 abundance. The data uncover a novel mechanism for IR and ultimately SM metabolic abnormalities in patients with HF.NEW & NOTEWORTHY Heart failure is associated with systemic insulin resistance and abnormalities in glucose metabolism but the underlying mechanisms are poorly understood. In the skeletal muscle, the major peripheral site of glucose utilization, we observe an increase in miR-133b in heart failure mice, which reduces the insulin-sensitive glucose transporter (GLUT4), glucose uptake, and metabolism in C2C12 and in myocytes. The antagomir for miR-133b restores GLUT4 protein and markers of metabolism in skeletal myocytes from heart failure mice demonstrating that miR-133b is an exciting target for systemic insulin resistance in heart failure and an important player in the cross talk between the heart and the periphery in the heart failure syndrome.

Cocoa Flavanol Supplementation and the Effect on Insulin Resistance in Females Who Are Overweight or Obese: A Randomized, Placebo-Controlled Trial.

There is interest in the impact that dietary interventions can have on preventing the transition from insulin resistance to type 2 diabetes, including a suggestion that the bioactive components of cocoa may enhance fasting insulin sensitivity. However, a role for cocoa flavanols (CF) in reducing insulin resistance in the insulin-stimulated state, an important risk factor for cardiovascular disease, is unresolved. This study investigated whether CF consumption improved whole-body insulin-mediated glucose uptake ('M') in females with overweight/obesity, using a randomized, double-blinded, placebo-controlled, parallel-group design. Thirty-two premenopausal females (19-49 years; 27-35 kg·m-2) with elevated HOMA-IR (HOMA-IR >1.5) supplemented their habitual diet with two servings/day of a high-flavanol cocoa drink (HFC; 609 mg CF/serving; n = 16) or low-flavanol cocoa drink (LFC; 13 mg CF/serving; n = 16) for 4 weeks. Assessment of HOMA-IR and 'M' during a 3-h, 60 mIU insulin·m-2·min-1 euglycemic clamp was performed before and after the intervention. Data are the mean (SD). Changes to HOMA-IR (HFC -0.003 (0.57); LFC -0.0402 (0.86)) and 'M' (HFC 0.99 (7.62); LFC -1.32 (4.88) µmol·kg-1·min-1) after the intervention were not different between groups. Four weeks' consumption of ~1.2 g CF/day did not improve indices of fasting insulin sensitivity or insulin-mediated glucose uptake. A recommendation for dietary supplementation with cocoa flavanols to improve glycemic control is therefore not established.

824-P: Dual Amylin and Calcitonin Receptor Agonists Improve Insulin Sensitivity and Increase Tissue-Specific Glucose Uptake Independent of Weight Loss

Dual amylin and calcitonin receptor agonists (DACRAs) are known to induce a significant weight loss and improve glucose tolerance and glucose control in rats. However, it is unknown if DACRAs has an insulin sensitizing effect beyond that induced by weight loss and if DACRAs affect tissue specific glucose uptake. Additionally, clamp studies with DACRAs have not previously been performed in diabetic rats. Pre-diabetic ZDSD and diabetic ZDF rats were treated with the DACRA KBP-088 (1.5 nmol//kg, s.c.) for 12 days, and a hyperinsulinemic clamp was conducted on day 13, 24 h post-dosing. In ZDSD rats a hyperinsulinemic euglycemic clamp was carried out, while a hyperinsulinemic isoglycemic clamp was carried out in ZDF rats. In addition, specific glucose uptake was assessed during a hyperinsulinemic isoglycemic clamp in ZDF rats. In ZDSD rats, KBP-088 treatment resulted in a significant reduction in body weight and fasting blood glucose, and improved insulin sensitivity by increasing the glucose infusion rate (GIR) (vehicle: 8.6, KBP-088: 12 mg glucose/kg/min) . In ZDF rats, KBP-088 significantly reduced fasting blood glucose and improved insulin sensitivity (GIR, vehicle: 1.5, KBP-088: 16.5 mg glucose/kg/min) , independent on weight loss. In both studies, KBP-088 increased the rate of glucose clearance, likely be increasing glucose storage, but without altering the endogenous glucose production. Direct assessment of tissue specific glucose uptake showed, that KBP-088 significantly increased glucose uptake in both muscles and adipose tissues when compared to vehicle. In summary, KBP-088 significantly improved insulin sensitivity in both pre-diabetic and diabetic rats and markedly increased tissue specific glucose uptake. Importantly, KBP-088 has an insulin sensitizing effect independent of weight loss, highlighting DACRAs as promising agents for treatment of diabetes. Disclosure A.T.Larsen: Employee; Nordic Bioscience. N.Sonne: None. E.Bredtoft: None. M.A.Karsdal: Employee; Nordic Bioscience A/S, Nordic Bioscience A/S, Nordic Bioscience A/S. K.Henriksen: Employee; Nordic Bioscience A/S, Stock/Shareholder; Nordic Bioscience A/S.

Acute bioenergetic insulin sensitivity of skeletal muscle cells: ATP-demand-provoked glycolysis contributes to stimulation of ATP supply

Skeletal muscle takes up glucose in an insulin-sensitive manner and is thus important for the maintenance of blood glucose homeostasis. Insulin resistance during development of type 2 diabetes is associated with decreased ATP synthesis, but the causality of this association is controversial. In this paper, we report real-time oxygen uptake and medium acidification data that we use to quantify acute insulin effects on intracellular ATP supply and ATP demand in rat and human skeletal muscle cells. We demonstrate that insulin increases overall cellular ATP supply by stimulating the rate of glycolytic ATP synthesis. Stimulation is immediate and achieved directly by increased glycolytic capacity, and indirectly by elevated ATP demand from protein synthesis. Raised glycolytic capacity does not result from augmented glucose uptake. Notably, insulin-sensitive glucose uptake is increased synergistically by nitrite. While nitrite has a similar stimulatory effect on glycolytic ATP supply as insulin, it does not amplify insulin stimulation. These data highlight the multifarious nature of acute bioenergetic insulin sensitivity of skeletal muscle cells, and are thus important for the interpretation of changes in energy metabolism that are seen in insulin-resistant muscle.

Genes controlling skeletal muscle glucose uptake and their regulation by endurance and resistance exercise.

Exercise improves the insulin sensitivity of glucose uptake in skeletal muscle. Due tothat, exercise has become a cornerstone treatment for type 2 diabetes mellitus (T2DM). The mechanisms by which exercise improves skeletal muscle insulin sensitivity are, however, incompletely understood. We conducted a systematic review to identify all genes whose gain or loss of function alters skeletal muscle glucose uptake. We subsequently cross-referenced these genes with recently generated data sets on exercise-induced gene expression and signaling. Our search revealed 176 muscle glucose-uptake genes, meaning that their genetic manipulation altered glucose uptake in skeletal muscle. Notably, exercise regulates the expression or phosphorylation of more than 50% of the glucose-uptake genes or their protein products. This included many genes that previously have not been associated with exercise-induced insulin sensitivity. Interestingly, endurance and resistance exercise triggered some common but mostly unique changes in expression and phosphorylation of glucose-uptake genes or their protein products. Collectively, our work provides a resource of potentially new molecular effectors that play a role in the incompletely understood regulation of muscle insulin sensitivity by exercise.

Probing Insulin Sensitivity with Metabolically Competent Human Stem Cell-Derived White Adipose Tissue Microphysiological Systems.

Impaired white adipose tissue (WAT) function has been recognized as a critical early event in obesity-driven disorders, but high buoyancy, fragility, and heterogeneity of primary adipocytes have largely prevented their use in drug discovery efforts highlighting the need for human stem cell-based approaches. Here, human stem cells are utilized to derive metabolically functional 3D adipose tissue (iADIPO) in a microphysiological system (MPS). Surprisingly, previously reported WAT differentiation approaches create insulin resistant WAT ill-suited for type-2 diabetes mellitus drug discovery. Using three independent insulin sensitivity assays, i.e., glucose and fatty acid uptake and suppression of lipolysis, as the functional readouts new differentiation conditions yielding hormonally responsive iADIPO are derived. Through concomitant optimization of an iADIPO-MPS, it is abled to obtain WAT with more unilocular and significantly larger (≈40%) lipid droplets compared to iADIPO in 2D culture, increased insulin responsiveness of glucose uptake (≈2-3 fold), fatty acid uptake (≈3-6 fold), and ≈40% suppressing of stimulated lipolysis giving a dynamic range that is competent to current in vivo and ex vivo models, allowing to identify both insulin sensitizers and desensitizers.

Effects of a novel polyphenol-rich plant extract on body composition, inflammation, insulin sensitivity, and glucose homeostasis in obese mice.

The worldwide prevalence of obesity, metabolic syndrome and type 2 diabetes (T2D) is reaching epidemic proportions that urge the development of new management strategies. Totum-63 is a novel, plant-based polyphenol-rich active principle that has been shown to reduce body weight, fasting glycemia, glucose intolerance, and fatty liver index in obese subjects with prediabetes. Here, we investigated the effects and underlying mechanism(s) of Totum-63 on metabolic homeostasis in insulin-resistant obese mice. Male C57Bl6/J mice were fed a high-fat diet for 12 weeks followed by supplementation with Totum-63 for 4 weeks. The effects on whole-body energy and metabolic homeostasis, as well as on tissue-specific inflammation and insulin sensitivity were assessed using a variety of immunometabolic phenotyping tools. Totum-63 decreased body weight and fat mass in obese mice, without affecting lean mass, food intake and locomotor activity, and increased fecal energy excretion and whole-body fatty acid oxidation. Totum-63 reduced fasting plasma glucose, insulin and leptin levels, and improved whole-body insulin sensitivity and peripheral glucose uptake. The expression of insulin receptor β and the insulin-induced phosphorylation of Akt/PKB were increased in liver, skeletal muscle, white adipose tissue (WAT) and brown adipose tissue (BAT). Hepatic steatosis was also decreased by Totum-63 and associated with a lower expression of genes involved in fatty acid uptake, de novo lipogenesis, inflammation, and fibrosis. Furthermore, a significant reduction in pro-inflammatory macrophages was also observed in epidydimal WAT. Finally, a potent decrease in BAT mass associated with enhanced tissue expression of thermogenic genes was found, suggesting BAT activation by Totum-63. Our results show that Totum-63 reduces inflammation and improves insulin sensitivity and glucose homeostasis in obese mice through pleiotropic effects on various metabolic organs. Altogether, plant-derived Totum-63 might constitute a promising novel nutritional supplement for alleviating metabolic dysfunctions in obese people with or without T2D.

β-catenin regulates muscle glucose transport via actin remodelling and M-cadherin binding

ObjectiveSkeletal muscle glucose disposal following a meal is mediated through insulin-stimulated movement of the GLUT4-containing vesicles to the cell surface. The highly conserved scaffold-protein β-catenin is an emerging regulator of vesicle trafficking in other tissues. Here, we investigated the involvement of β-catenin in skeletal muscle insulin-stimulated glucose transport. MethodsGlucose homeostasis and transport was investigated in inducible muscle specific β-catenin knockout (BCAT-mKO) mice. The effect of β-catenin deletion and mutation of β-catenin serine 552 on signal transduction, glucose uptake and protein–protein interactions were determined in L6-G4-myc cells, and β-catenin insulin-responsive binding partners were identified via immunoprecipitation coupled to label-free proteomics. ResultsSkeletal muscle specific deletion of β-catenin impaired whole-body insulin sensitivity and insulin-stimulated glucose uptake into muscle independent of canonical Wnt signalling. In response to insulin, β-catenin was phosphorylated at serine 552 in an Akt-dependent manner, and in L6-G4-myc cells, mutation of β-cateninS552 impaired insulin-induced actin-polymerisation, resulting in attenuated insulin-induced glucose transport and GLUT4 translocation. β-catenin was found to interact with M-cadherin in an insulin-dependent β-cateninS552-phosphorylation dependent manner, and loss of M-cadherin in L6-G4-myc cells attenuated insulin-induced actin-polymerisation and glucose transport. ConclusionsOur data suggest that β-catenin is a novel mediator of glucose transport in skeletal muscle and may contribute to insulin-induced actin-cytoskeleton remodelling to support GLUT4 translocation.

TNF-α (-308) Gene Polymorphism and Type 2 Diabetes Mellitus in Ethiopian Diabetes Patients.

BackgroundType 2 diabetes mellitus (T2DM) is a metabolic disorder resulting from insulin insufficiency or function. Predisposing factors for T2DM are mainly genetic and environmental. Genetic polymorphism of cytokines like tumor necrosis factor-alpha (TNF-α) is suggestive of interference with insulin-sensitive glucose uptake and induces insulin resistance that ultimately could lead to T2DM. In this study, we assessed the effect of TNF-α (−308) G/A gene polymorphism and its association with the development of T2DM in an Ethiopian population.MethodsAn institutional-based cross-sectional study was conducted on study subjects with T2DM and non-diabetic healthy controls. DNA was extracted and genotyping was carried out by using amplification refractory mutation system polymerase chain reaction. A genetic-polymorphism on TNF-α (−308) G/A with T2DM was evaluated by logistic regression and Student’s t-test. A P-value <0.05 was considered as statistically significant.ResultsIn the present study, we observed a significant association between T2DM and TNF-α (−308) gene polymorphism’s GG genotype [χ2 test P = 0.005, OR (95% CI) =2.667 (1.309–5.45d8)]. In contrast, no statistically significant differences were observed in the frequencies of genotypes AA and AG (χ2 test P=0.132 and 0.067, respectively). Moreover, T2DM individuals had higher concentrations of lipid profiles for those carrying the TNF-α (−308) GG genotype as compared to the control group.ConclusionTNF-α (−308) genetic polymorphism may be implicated in the genetic susceptibility for, as well as the development of T2DM and lipid metabolism in the Ethiopian population. Therefore, a large-scale study and early screening of TNF-α (−308) genetic polymorphism may help in early management and control of diabetes and related outcomes.

Elevated chemerin induces insulin resistance in human granulosa-lutein cells from polycystic ovary syndrome patients.

The insulin resistance (IR) of ovarian granulosa cells from polycystic ovary syndrome (PCOS) aggravates the abnormalities in steroidogenesis and anovulation, and chemerin is an adipokine involved in regulating adipogenesis and glucose homeostasis. The role and underlying mechanism of chemerin in developing IR of the granulosa cells from PCOS remain unclear. Plasma, follicular fluid, and human granulosa-lutein cells (hGLs) were collected from non-PCOS and patients with PCOS with or without IR. The chemerin levels were elevated in both follicular fluid and hGL samples from patients with PCOS with IR, and the hGLs from patients with PCOS with IR showed decreased insulin sensitivity and impaired glucose uptake capacity. Moreover, treatment of chemerin attenuated insulin-stimulated glucose uptake by decreasing phosphorylation of insulin receptor substrate (IRS)1/2 Tyr612, phosphorylation of protein kinase B Ser473, and membrane translocation of glucose transporter type 4 through increasing Ser307 phosphorylation of IRS1 in cultured hGLs. These effects could be abolished by small interfering RNA-mediated knockdown of chemokine-like receptor 1. Furthermore, insulin induced the expression of chemerin in hGLs. Our findings demonstrate a novel role of chemerin in the metabolic dysfunction of PCOS, which suggested that chemerin and its receptor can be further implicated as potential therapeutic targets in the future treatment of PCOS.-Li, X., Zhu, Q., Wang, W., Qi, J., He, Y., Wang, Y., Lu, Y., Wu, H., Ding, Y., Sun, Y. Elevated chemerin induces insulin resistance in human granulosa-lutein cells from polycystic ovary syndrome patients.

Fatty acid type-specific regulation of SIRT1 does not affect insulin sensitivity in human skeletal muscle.

The nicotinamide adenine dinucleotide-dependent deacetylase, sirtuin (SIRT)1, in skeletal muscle is reduced in insulin-resistant states. However, whether this is an initial mechanism responsible for mediating insulin resistance in human skeletal muscle remains to be investigated. Also, SIRT1 acts as a mitochondrial gene transcriptional regulator and is induced by a short-term, high-fat diet (HFD) in human skeletal muscle. Whether saturated or unsaturated fatty acids (FAs) in the diet are important for this is unknown. We subjected 17 healthy, young men to a eucaloric control (Con) diet and 1 of 2 hypercaloric [+75% energy (E%)] HFDs for 3 d enriched in either saturated (Sat) FA (79 E% fat; Sat) or unsaturated FA (78 E% fat; Unsat). After Sat, SIRT1 protein content and activity in skeletal muscle increased ( P < 0.05; ∼40%) while remaining unchanged after Unsat. Whole-body insulin sensitivity and insulin-stimulated leg glucose uptake were reduced ( P < 0.01; ∼20%) to a similar extent compared to Con after both HFDs. We demonstrate a novel FA type-dependent regulation of SIRT1 protein in human skeletal muscle. Moreover, regulation of SIRT1 does not seem to be an initiating factor responsible for mediating insulin resistance in human skeletal muscle.-Fritzen, A. M., Lundsgaard, A.-M., Jeppesen, J. F., Sjøberg, K. A., Høeg, L. D., Deleuran, H. H., Wojtaszewski, J. F. P., Richter, E. A., Kiens, B. Fatty acid type-specific regulation of SIRT1 does not affect insulin sensitivity in human skeletal muscle.

Coadministration of GLP-1 and Y2R Receptor Agonists Synergize to Promote Muscle Glucose Uptake and Induce Diabetic Remission

One proposed mechanism for the effects of bariatric surgery is the enhanced postprandial release of enteroendocrine L-cell hormones, such as PYY and GLP-1. Four week co-administration of Fc-conjugated GLP-1R and Y2R-selective agonists induced weight loss, reduced %HbA1c, and improved glucose tolerance in DIO and KS db/db mice compared to vehicle or monotherapy control groups. To assess the impact of this combination on glucose control, a hyperinsulinemic/euglycemic clamp was performed in KS db/db mice following two week treatment with vehicle or co-administration of GLP-1/Y2R agonists. A vehicle-treated group weight-matched to combination-treated mice was also assessed. Compared to KS db/db vehicle animals, combination-treated mice exhibited a 16-fold greater glucose infusion rate compared to only 4-fold in weight-matched controls. Glucose uptake was increased 4-fold in the skeletal muscle of combination-treated animals, but was unaltered in adipose or brain. Weight-matched controls did not exhibit increased glucose uptake in any tissue. To better characerize the mechanism of Fc-GLP-1/Fc-Y2R action on peripheral glucose metabolism, lean mice were acutely administered Fc-GLP-1 and/or Fc-Y2R, and then sacrificed to assess brain c-Fos activation. Compared to monotherapies, combination administration led to additive c-Fos activation in hindbrain regions. Strikingly, whereas monotherapy had no effect on the PVH, combination administration led to a synergistic 2-fold increase in c-Fos immunoreactivity. These findings implicate a neuronal population in the PVH as a putative driver of the antidiabetic efficacy observed following co-administration of GLP-1R and Y2R selective peptides, and may represent a novel mechanism to improve insulin sensitivity and skeletal muscle glucose uptake. Disclosure B. Boland: Employee; Self; MedImmune. V.G. Howard: Employee; Self; AstraZeneca. Stock/Shareholder; Self; Regeneron Pharmaceuticals, Inc., AstraZeneca. M. Beaton: Employee; Self; MedImmune. Employee; Spouse/Partner; MedImmune. S. Will: Employee; Self; MedImmune. Stock/Shareholder; Self; AstraZeneca. S. Oldham: Stock/Shareholder; Self; AstraZeneca. Employee; Self; MedImmune. J. Trevaskis: Employee; Self; MedImmune. Stock/Shareholder; Self; AstraZeneca. C.J. Rhodes: Stock/Shareholder; Self; AstraZeneca. J. Grimsby: Employee; Self; AstraZeneca.

Urocortin-2 Gene Transfer Effectively Treats Type 1 Diabetes in Mice

Type 1 diabetes (T1D) affects 1.3 million U.S. patients. Tight glucose control reduces microvascular complications and adverse cardiovascular events. Insulin therapy is essential but has shortcomings: a) only 1 in 3 patients achieve targeted glucose control; b) aggressive insulin use increases hypoglycemia; c) 25% of T1D patients develop insulin resistance. We previously reported that urocortin-2 (UCn2) gene transfer increases insulin sensitivity and release in mouse models of type 2 diabetes. In the present study, 3m-old male Akita mice with T1D (due to Ins2 mutation) received IV saline, AAV8. Null or AAV8.UCn2 (2x1012 gc/kg). Ten weeks after UCn2 gene transfer we saw normalization of fasting glucose, HbA1c and glucose tolerance (Table). Increased body weight and reduced water intake was seen after UCn2 gene transfer. Hyperinsulinemic euglycemic clamps showed increased insulin sensitivity and skeletal muscle glucose uptake. There was reduced nephropathy and retinopathy and increased survival (Table). Echocardiography showed improved ejection fraction (p&amp;lt;.001), end-systolic dimension (p&amp;lt;.001) and diastolic function (p&amp;lt;.0001). Data acquisition and analyses were blinded. The mechanisms for beneficial effects include increased insulin release and sensitivity. Thus, UCn2 gene transfer may be a viable therapy both for new onset T1D with detectable endogenous insulin, and for later stage T1D to reduce exogenous insulin needs. UCn2 Gene Transfer in Akita Mice (Type 1 Diabetes)Saline or AAV8.Null (n)AAV8.UCn2 (n)pPlasma UCn2 (ng/mL)1±.2 (14)12±.9 (14)&amp;lt;.0001Urine Output (ml/hr).43±.06 (8).24±.06 (8).0412hr Fasting Glucose (mg/dL)272±25 (8)106±12 (8)&amp;lt;.0001HbA1c (%)8.2±.4 (5)5.5±.4 (5)&amp;lt;.002Glucose Tolerance Test (AUC)3128±215 (8)2062±286 (8)&amp;lt;.0001Insulin Release 2h post gluc (ng/ml).15±.01 (8).29±.4 (8).004Glucose Infusion Rate (mg/kg/min)12±2.89 (6)33.6±5.3 (5).001Vast Lat Glu Uptake (µM/m/100g)8.2±.4 (7)23.2±2.6 (6)&amp;lt;.0001Glomerulus Expansion Score1.7±.04 (8)1.5±.04 (8).01Urine Albumin/Creatinine (3hr)28±3 (8)17±3 (8).01Retinal Vascular Leak (%)43±1 (5)19±5 (4).001200-day Survival (%)0 (17 of 17 died)82 (3 of 17 died)&amp;lt;.0001Mean ± SE; p from Student’s t-test (unpaired, 2 tails); AAV8, Adeno-Associated Virus Type 8 Disclosure M. Gao: None. N. Lai: None. D. Giamouridis: None. T.Y. Guo: None. B. Xia: None. Y. Kim: None. M.V. Estrada: None. V. Nguyen Huu: None. D. Skowronska-Krawczyk: None. H. Hammond: Stock/Shareholder; Self; Renova Therapeutics.

Exercise training decreases pancreatic fat content and improves beta cell function regardless of baseline glucose tolerance: a randomised controlled trial

Aims/hypothesisPancreatic fat accumulation may contribute to the development of beta cell dysfunction. Exercise training improves whole-body insulin sensitivity, but its effects on pancreatic fat content and beta cell dysfunction are unclear. The aim of this parallel-group randomised controlled trial was to evaluate the effects of exercise training on pancreatic fat and beta cell function in healthy and prediabetic or type 2 diabetic participants and to test whether the responses were similar regardless of baseline glucose tolerance.MethodsUsing newspaper announcements, a total of 97 sedentary 40–55-year-old individuals were assessed for eligibility. Prediabetes (impaired fasting glucose and/or impaired glucose tolerance) and type 2 diabetes were defined by ADA criteria. Of the screened candidates, 28 healthy men and 26 prediabetic or type 2 diabetic men and women met the inclusion criteria and were randomised into 2-week-long sprint interval or moderate-intensity continuous training programmes in a 1:1 allocation ratio using random permuted blocks. The primary outcome was pancreatic fat, which was measured by magnetic resonance spectroscopy. As secondary outcomes, beta cell function was studied using variables derived from OGTT, and whole-body insulin sensitivity and pancreatic fatty acid and glucose uptake were measured using positron emission tomography. The measurements were carried out at the Turku PET Centre, Finland. The analyses were based on an intention-to-treat principle. Given the nature of the intervention, blinding was not applicable.ResultsAt baseline, the group of prediabetic or type 2 diabetic men had a higher pancreatic fat content and impaired beta cell function compared with the healthy men, while glucose and fatty acid uptake into the pancreas was similar. Exercise training decreased pancreatic fat similarly in healthy (from 4.4% [3.0%, 6.1%] to 3.6% [2.4%, 5.2%] [mean, 95% CI]) and prediabetic or type 2 diabetic men (from 8.7% [6.0%, 11.9%] to 6.7% [4.4%, 9.6%]; p = 0.036 for time effect) without any changes in pancreatic substrate uptake (p ≥ 0.31 for time effect in both insulin-stimulated glucose and fasting state fatty acid uptake). In prediabetic or type 2 diabetic men and women, both exercise modes similarly improved variables describing beta cell function.Conclusions/interpretationTwo weeks of exercise training improves beta cell function in prediabetic or type 2 diabetic individuals and decreases pancreatic fat regardless of baseline glucose tolerance. This study shows that short-term training efficiently reduces ectopic fat within the pancreas, and exercise training may therefore reduce the risk of type 2 diabetes.Trial registrationClinicalTrials.gov NCT01344928FundingThis study was funded by the Emil Aaltonen Foundation, the European Foundation for the Study of Diabetes, the Finnish Diabetes Foundation, the Orion Research Foundation, the Academy of Finland (grants 251399, 256470, 281440, and 283319), the Ministry of Education of the State of Finland, the Paavo Nurmi Foundation, the Novo Nordisk Foundation, the Finnish Cultural Foundation, the Hospital District of Southwest Finland, the Turku University Foundation, and the Finnish Medical Foundation.

AICAR Prevents Acute Olanzapine-Induced Disturbances in Glucose Homeostasis.

Olanzapine (OLZ) is an antipsychotic drug used in the treatment of schizophrenia. Although effective in reducing psychoses, OLZ causes acute increases in blood glucose. The acute effects of OLZ on hyperglycemia are likely caused by reductions in insulin secretion, insulin resistance, and increased hepatic glucose production. 5AMP-activated protein kinase (AMPK) is an energy sensor activated during exercise that can increase insulin sensitivity and insulin-independent glucose uptake in muscle. 5-Aminoimidazole-4-carboxamide ribonucleoside (AICAR) is a pharmacologic agent that, among other effects, can activate AMPK in vivo. Conversely, hypothalamic activation of AMPK has been suggested to mediate the hyperglycemic effects of OLZ. The purpose of this investigation was to determine whether cotreatment with AICAR could prevent acute OLZ-induced hyperglycemia in lean and obese C57BL6/J mice. OLZ (5 mg/kg, i.p.) caused rapid increases in blood glucose, a blunted insulin response, and pyruvate intolerance, all of which were prevented with AICAR cotreatment in both lean and obese mice. AICAR did not affect OLZ-induced changes in whole-body substrate oxidation or energy expenditure. Peripheral injection of AICAR, but not OLZ, activated AMPK signaling in the hypothalamus. The results of the current study provide evidence that AICAR prevents OLZ-induced hyperglycemia, despite increasing hypothalamic AMPK signaling. These protective effects were associated with the preservation of whole-body insulin action and reductions in markers of liver glucose production.

Combined Hyperglycemia- and Hyperinsulinemia-Induced Insulin Resistance in Adipocytes Is Associated With Dual Signaling Defects Mediated by PKC-ζ.

A hyperglycemic and hyperinsulinemic environment characteristic of type 2 diabetes causes insulin resistance. In adipocytes, defects in both insulin sensitivity and maximum response of glucose transport have been demonstrated. To investigate the molecular mechanisms, freshly isolated rat adipocytes were incubated in control (5.6 mM glucose, no insulin) and high glucose (20 mM)/high insulin (100 nM) (HG/HI) for 18 hours to induce insulin resistance. Insulin-resistant adipocytes manifested decreased sensitivity of glucose uptake associated with defects in insulin receptor substrate (IRS)-1 Tyr phosphorylation, association of p85 subunit of phosphatidylinositol-3-kinase, Akt Ser473 and Thr308 phosphorylation, accompanied by impaired glucose transporter 4 translocation. In contrast, protein kinase C (PKC)-ζ activity was augmented by chronic HG/HI. Inhibition of PKC-ζ with a specific cell-permeable peptide reversed the signaling defects and insulin sensitivity of glucose uptake. Transfection of dominant-negative, kinase-inactive PKC-ζ blocked insulin resistance, whereas constitutively active PKC-ζ recapitulated the defects. The HG/HI incubation was associated with stimulation of IRS-1 Ser318 and Akt Thr34 phosphorylation, targets of PKC-ζ. Transfection of IRS-1 S318A and Akt T34A each partially corrected insulin signaling, whereas combined transfection of both completely normalized insulin signaling. In vivo hyperglycemia/hyperinsulinemia in rats for 48 hours similarly resulted in activation of PKC-ζ and increased phosphorylation of IRS-1 Ser318 and Akt Thr34. These data indicate that impairment of insulin signaling by chronic HG/HI is mediated by dual defects at IRS-1 and Akt mediated by PKC-ζ.

Elevated Chemerin Induces Insulin Resistance in Human Granulosa-Lutein Cells from Polycystic Ovary Syndrome Patients

Background: The insulin resistance (IR) of the ovarian granulosa cells from polycystic ovary syndrome (PCOS) aggravates the abnormalities in steroidogenesis and anovulation, and chemerin is an adipokine involved in regulating adipogenesis and glucose homeostasis. The underlying mechanism of IR and the role of chemerin in the development of IR remain to be demonstrated. Methods: The granulosa-lutein cells (hGLs) and follicular fluid from PCOS and non-PCOS patients with or without IR were collected to determine the chemerin levels and molecules pertinent to insulin signaling. The effects and molecular mechanism of chemerin on the insulin signaling were investigated in cultured hGLs with immunofluorescence staining, special enzyme-linked immunosorbent assay (ELISA), quantitative real-time PCR (qRT-PCR), western blotting, [18F]-Fluorodeoxyglucose ([18F]-FDG) incorporation and siRNA transfection respectively. Findings: The chemerin levels were elevated in both follicular fluid and hGLs samples obtained from PCOS with IR patients and the hGLs obtained from PCOS with IR patients showed decreased insulin sensitivity and impaired glucose uptake capacity. Moreover, treatment of chemerin attenuated insulin-stimulated glucose uptake by decreasing phosphorylation of Akt and membrane translocation of glucose transporter type 4 (GLUT4) through increasing Ser307 phosphorylation of insulin receptor substrate 1 (IRS1) in cultured hGLs. These effects could be abolished by siRNA-mediated knockdown of chemokine-like receptor 1 (CMKLR1). Furthermore, insulin induced the expression of chemerin in hGLs. Interpretation: Our findings demonstrate a novel role of chemerin in the metabolic dysfunction of PCOS, which suggested that chemerin and its receptor can be further implicated as potential therapeutic targets in the future treatment of PCOS. Funding Statement: This work was supported by the National Natural Science Foundation of China [grant numbers 81771648, 81571499]; the National Key RD the Shanghai Commission of Science and Technology [grant number 17DZ2271100]; the Shanghai Municipal Education Commission-Gaofeng Clinical Medicine Grant [grant number 20161413]; the Program of Shanghai Academic Research Leader in Shanghai Municipal Commission of Health and Family Planning [grant number 2017BR015]; and the Clinical Skills Improvement Project of Major Disorders, Hospital Development Center of Shanghai [grant number 16CR1022A] and Shanghai Technological Innovation Plan [grant number 18140902400]. Declaration of Interests: The authors report no conflicts of interest in this work. Ethics Approval Statement: All samples were collected with written informed consent under a protocol approved by the Ethics Committee of Renji Hospital.

Effects of thrombin on insulin signalling and glucose uptake in cultured human myotubes

BackgroundHyper-coagulability (elevated thrombin) is a feature of type 2 diabetes and contributes to an increased risk of thrombotic and vascular events. Skeletal muscle is the key peripheral tissue site of insulin resistance in type 2 diabetes. Cultured human skeletal muscle cells were used to explore the effects of thrombin on insulin signalling and glucose uptake. We hypothesized that thrombin affects insulin activity in human skeletal muscle cells which could link the hypercoagulability and insulin resistance in type 2 diabetes. MethodsHuman skeletal muscle cell cultures (myotubes) were treated with +/−5 units/ml thrombin for 6h. Insulin signalling pathway components and AMPK were examined by Western blotting. Real time PCR and glucose uptake assays were performed. ResultsThere was a significant decrease (p<0.01) in insulin mediated IRS-1 and Akt phosphorylation in response to thrombin in cultured myotubes. Diminished Akt phosphorylation was alleviated by treatment with a PKC inhibitor. Thrombin directly increased basal glucose uptake (p<0.05) that involved AMPK phosphorylation (p<0.01) and this was partly repressed by compound C (AMPK inhibitor). Thrombin also significantly increased the gene expression level of both GLUT1 and GLUT4 in cultured human skeletal muscle cells. ConclusionThrombin decreased insulin signalling in skeletal muscle cells through a PKC mediated mechanism, but did not affect the net action of insulin on glucose uptake. The direct stimulatory effect of thrombin on glucose uptake was mediated, at least in part, via an AMPK dependent mechanism. We conclude that thrombin activation results in multiple metabolic effects beyond increased thrombogenicity but does not include a decrease in insulin sensitivity (glucose uptake) in cultured human skeletal muscle cells.