Rare fully penetrant mutations in AKT2 are an established cause of monogenic disorders of glucose metabolism. Recently, a novel partial loss-of-function AKT2 coding variant (p.Pro50Thr) was identified that is nearly specific to Finns (frequency 1.1%), with the low-frequency allele associated with an increase in fasting plasma insulin level and risk of type 2 diabetes. The effects of the p.Pro50Thr AKT2 variant (p.P50T/AKT2) on insulin-stimulated glucose uptake (GU) in the whole body and in different tissues have not previously been investigated. We identified carriers (N = 20) and matched noncarriers (N = 25) for this allele in the population-based Metabolic Syndrome in Men (METSIM)study and invited these individuals back for positron emission tomography study with [18F]-fluorodeoxyglucose during euglycemic hyperinsulinemia. When we compared p.P50T/AKT2 carriers to noncarriers, we found a 39.4% reduction in whole-body GU (P = 0.006) and a 55.6% increase in the rate of endogenous glucose production (P = 0.038). We found significant reductions in GU in multiple tissues-skeletal muscle (36.4%), liver (16.1%), brown adipose (29.7%), and bone marrow (32.9%)-and increases of 16.8-19.1% in seven tested brain regions. These data demonstrate that the p.P50T substitution of AKT2 influences insulin-mediated GU in multiple insulin-sensitive tissues and may explain, at least in part, the increased risk of type 2 diabetes in p.P50T/AKT2 carriers.

Adipose tissue expansion progresses rapidly during postnatal life, influenced by both prenatal maternal factors and postnatal developmental cues. The ratio of omega-6 (n-6) relative to n-3 polyunsaturated fatty acids (PUFAs) is believed to regulate perinatal adipogenesis, but the cellular mechanisms and long-term effects are not well understood. We lowered the fetal and postnatal n-6/n-3 PUFA ratio exposure in wild-type offspring under standard maternal dietary fat amounts to test the effects of low n-6/n-3 ratios on offspring adipogenesis and adipogenic potential. Relative to wild-type pups receiving high perinatal n-6/n-3 ratios, subcutaneous adipose tissue in 14-day-old wild-type pups receiving low n-6/n-3 ratios had more adipocytes that were smaller in size; decreased Pparγ2, Fabp4, and Plin1; several lipid metabolism mRNAs; coincident hypermethylation of the PPARγ2 proximal promoter; and elevated circulating adiponectin. As adults, offspring that received low perinatal n-6/n-3 ratios were diet-induced obesity (DIO) resistant and had a lower positive energy balance and energy intake, greater lipid fuel preference and non-resting energy expenditure, one-half the body fat, and better glucose clearance. Together, the findings support a model in which low early-life n-6/n-3 ratios remodel adipose morphology to increase circulating adiponectin, resulting in a persistent adult phenotype with improved metabolic flexibility that prevents DIO.

Adipose-derived stem cells (ADSCs) play critical roles in controlling obesity-associated inflammation and metabolic disorders. Exosomes from ADSCs exert protective effects in several diseases, but their roles in obesity and related pathological conditions remain unclear. In this study, we showed that treatment of obese mice with ADSC-derived exosomes facilitated their metabolic homeostasis, including improved insulin sensitivity (27.8% improvement), reduced obesity, and alleviated hepatic steatosis. ADSC-derived exosomes drove alternatively activated M2 macrophage polarization, inflammation reduction, and beiging in white adipose tissue (WAT) of diet-induced obese mice. Mechanistically, exosomes from ADSCs transferred into macrophages to induce anti-inflammatory M2 phenotypes through the transactivation of arginase-1 by exosome-carried active STAT3. Moreover, M2 macrophages induced by ADSC-derived exosomes not only expressed high levels of tyrosine hydroxylase responsible for catecholamine release, but also promoted ADSC proliferation and lactate production, thereby favoring WAT beiging and homeostasis in response to high-fat challenge. These findings delineate a novel exosome-mediated mechanism for ADSC-macrophage cross talk that facilitates immune and metabolic homeostasis in WAT, thus providing potential therapy for obesity and diabetes.

Hypertension is considered a powerful cardiovascular risk factor and is present in up to two-thirds of patients who suffer from diabetes. In the background of an established epidemiological association between lower blood pressure (BP) and improvement in long-term clinical outcomes, several large landmark trials and analyses have attempted to examine the possible benefit of tighter BP control in patients with Type 2 diabetes mellitus. Although aggressive BP targets in patients with diabetes have been advocated for a long time, currently accepted evidence from these studies has led to a general recommendation of systolic BP <140 mmHg and diastolic BP <90 mmHg. Therapy consists of lifestyle management, including weight loss if overweight or obese, a Dietary Approaches to Stop Hypertension (DASH)-style based nutrition counselling, and reduced sodium intake. Timely initiation and subsequent titration of antihypertensive medications to achieve individualised BP goals is recommended. A therapeutic agent that acts on the renin-angiotensin-aldosterone pathway, such as an angiotensin-converting enzyme inhibitor or an angiotensin receptor blocker, should generally be included in the pharmacologic therapy for hypertension in patients with Type 2 diabetes mellitus. A multi-drug combination, particularly including a thiazide diuretic, is very often necessary and should be started early in the course of management. Finally, an accurate and standardised method of BP measurement in the outpatient setting is essential to ensure proper monitoring and gauge the effectiveness of treatment.

Glucagon-like peptide 1 receptor (GLP1R) agonists are widely used to treat diabetes. However, their function is dependent on adequate GLP1R expression, which is downregulated in diabetes. GLP1R is highly expressed on pancreatic β-cells, and activation by endogenous incretin or GLP1R agonists increases cAMP generation, which stimulates glucose-induced β-cell insulin secretion and helps maintain glucose homeostasis. We now have discovered that the highly β-cell-enriched microRNA, miR-204, directly targets the 3' UTR of GLP1R and thereby downregulates its expression in the β-cell-derived rat INS-1 cell line and primary mouse and human islets. Furthermore, in vivo deletion of miR-204 promoted islet GLP1R expression and enhanced responsiveness to GLP1R agonists, resulting in improved glucose tolerance, cAMP production, and insulin secretion as well as protection against diabetes. Since we recently identified thioredoxin-interacting protein (TXNIP) as an upstream regulator of miR-204, we also assessed whether in vivo deletion of TXNIP could mimic that of miR-204. Indeed, it also enhanced islet GLP1R expression and GLP1R agonist-induced insulin secretion and glucose tolerance. Thus, the present studies show for the first time that GLP1R is under the control of a microRNA, miR-204, and uncover a previously unappreciated link between TXNIP and incretin action.

Insulin has a narrow therapeutic index, reflected in a small margin between a dose that achieves good glycemic control and one that causes hypoglycemia. Once injected, the clearance of exogenous insulin is invariant regardless of blood glucose, aggravating the potential to cause hypoglycemia. We sought to create a "smart" insulin, one that can alter insulin clearance and hence insulin action in response to blood glucose, mitigating risk for hypoglycemia. The approach added saccharide units to insulin to create insulin analogs with affinity for both the insulin receptor (IR) and mannose receptor C-type 1 (MR), which functions to clear endogenous mannosylated proteins, a principle used to endow insulin analogs with glucose responsivity. Iteration of these efforts culminated in the discovery of MK-2640, and its in vitro and in vivo preclinical properties are detailed in this report. In glucose clamp experiments conducted in healthy dogs, as plasma glucose was lowered stepwise from 280 mg/dL to 80 mg/dL, progressively more MK-2640 was cleared via MR, reducing by ∼30% its availability for binding to the IR. In dose escalations studies in diabetic minipigs, a higher therapeutic index for MK-2640 (threefold) was observed versus regular insulin (1.3-fold).

Impaired skeletal muscle energy metabolism is linked to the pathogenesis of insulin resistance and glucose intolerance in type 2 diabetes. The contractile and metabolic properties of myofibers exhibit a high degree of heterogeneity and plasticity. The regulatory circuitry underpinning skeletal muscle energy metabolism is critically linked to exercise endurance and systemic homeostasis. Recent work has identified the Baf60 subunits of the SWI/SNF chromatin-remodeling complex as powerful regulators of the metabolic gene programs. However, their role in integrating myofiber energy metabolism with exercise endurance and metabolic physiology remains largely unknown. In this study, we conditionally inactivated Baf60a, Baf60c, or both in mature skeletal myocytes to delineate their contribution to muscle bioenergetics and metabolic physiology. Our work revealed functional redundancy between Baf60a and Baf60c in maintaining oxidative and glycolytic metabolism in skeletal myofibers and exercise endurance. Unexpectedly, mice lacking these two factors in skeletal muscle were protected from diet-induced and age-associated metabolic disorders. Transcriptional profiling analysis identified the muscle thermogenic gene program and myokine secretion as key pathways that integrate myofiber metabolism with systemic energy balance. As such, Baf60 deficiency in skeletal muscle illustrates a surprising disconnect between exercise endurance and systemic metabolic homeostasis.

Over the past 20 years, a large body of experimental and epidemiologic evidence has linked sleep duration and quality to glucose homeostasis, although the mechanistic pathways remain unclear. The aim of the current study was to determine whether genetic variation influencing both sleep and glucose regulation could underlie their functional relationship. We hypothesized that the genetic regulation of electroencephalographic (EEG) activity during non-rapid eye movement sleep, a highly heritable trait with fingerprint reproducibility, is correlated with the genetic control of metabolic traits including insulin sensitivity and β-cell function. We tested our hypotheses through univariate and bivariate heritability analyses in a three-generation pedigree with in-depth phenotyping of both sleep EEG and metabolic traits in 48 family members. Our analyses accounted for age, sex, adiposity, and the use of psychoactive medications. In univariate analyses, we found significant heritability for measures of fasting insulin sensitivity and β-cell function, for time spent in slow-wave sleep, and for EEG spectral power in the delta, theta, and sigma ranges. Bivariate heritability analyses provided the first evidence for a shared genetic control of brain activity during deep sleep and fasting insulin secretion rate.

The mitochondrial carrier uncoupling protein (UCP) 2 belongs to the family of the UCPs. Despite its name, it is now accepted that UCP2 is rather a metabolite transporter than a UCP. UCP2 can regulate oxidative stress and/or energetic metabolism. In rodents, UCP2 is involved in the control of α- and β-cell mass as well as insulin and glucagon secretion. Our aim was to determine whether the effects of UCP2 observed on β-cell mass have an embryonic origin. Thus, we used Ucp2 knockout mice. We found an increased size of the pancreas in Ucp2-/- fetuses at embryonic day 16.5, associated with a higher number of α- and β-cells. This phenotype was caused by an increase of PDX1+ progenitor cells. Perinatally, an increase in the proliferation of endocrine cells also participates in their expansion. Next, we analyzed the oxidative stress in the pancreata. We quantified an increased nuclear translocation of nuclear factor erythroid 2-related factor 2 (NRF2) in the mutant, suggesting an increased production of reactive oxygen species (ROS). Phosphorylation of AKT, an ROS target, was also activated in the Ucp2-/- pancreata. Finally, administration of the antioxidant N-acetyl-l-cysteine to Ucp2-/- pregnant mice alleviated the effect of knocking out UCP2 on pancreas development. Together, these data demonstrate that UCP2 controls pancreas development through the ROS-AKT signaling pathway.

We previously showed that the glutathione precursor, N-acetylcysteine (NAC), prevented hypoglycemia-associated autonomic failure (HAAF) and impaired activation of ventromedial hypothalamus (VMH) glucose-inhibited (GI) neurons by low glucose after recurrent hypoglycemia (RH) in nondiabetic rats. However, NAC does not normalize glucose sensing by VMH GI neurons when RH occurs during diabetes. We hypothesized that recruiting the thioredoxin (Trx) antioxidant defense system would prevent HAAF and normalize glucose sensing after RH in diabetes. To test this hypothesis, we overexpressed Trx-1 (cytosolic form of Trx) in the VMH of rats with streptozotocin (STZ)-induced type 1 diabetes. The counterregulatory response (CRR) to hypoglycemia in vivo and the activation of VMH GI neurons in low glucose using membrane potential sensitive dye in vitro was measured before and after RH. VMH Trx-1 overexpression normalized both the CRR and glucose sensing by VMH GI neurons in STZ rats. VMH Trx-1 overexpression also lowered the insulin requirement to prevent severe hyperglycemia in STZ rats. However, like NAC, VMH Trx-1 overexpression did not prevent HAAF or normalize activation of VMH GI neurons by low glucose in STZ rats after RH. We conclude that preventing HAAF in type 1 diabetes may require the recruitment of both antioxidant systems.

Cardiac insulin resistance is a key pathogenic factor for diabetic cardiomyopathy (DCM), but the mechanism remains largely unclear. We found that diabetic hearts exhibited decreased phosphorylation of total Akt and isoform Akt2 but not Akt1 in wild-type (WT) male FVB mice, which was accompanied by attenuation of Akt downstream glucose metabolic signal. All of these signal changes were not observed in metallothionein cardiac-specific transgenic (MT-TG) hearts. Furthermore, insulin-induced glucose metabolic signals were attenuated only in WT diabetic hearts. In addition, diabetic hearts exhibited increased Akt-negative regulator tribbles pseudokinase 3 (TRB3) expression only in WT mice, suggesting that MT may preserve Akt2 function via inhibiting TRB3. Moreover, MT prevented tert-butyl hydroperoxide (tBHP)-reduced insulin-stimulated Akt2 phosphorylation in MT-TG cardiomyocytes, which was abolished by specific silencing of Akt2. Specific silencing of TRB3 blocked tBHP inhibition of insulin-stimulated Akt2 phosphorylation in WT cardiomyocytes, whereas overexpression of TRB3 in MT-TG cardiomyocytes and hearts abolished MT preservation of insulin-stimulated Akt2 signals and MT prevention of DCM. Most importantly, supplementation of Zn to induce MT preserved cardiac Akt2 signals and prevented DCM. These results suggest that diabetes-inhibited cardiac Akt2 function via TRB3 upregulation leads to aberrant cardiac glucose metabolism. MT preservation of cardiac Akt2 function by inhibition of TRB3 prevents DCM.

Diabetes-induced visual dysfunction is associated with significant neuroretinal cell death. The current study was designed to investigate the role of the Protein Regulated in Development and DNA Damage Response 1 (REDD1) in diabetes-induced retinal cell death and visual dysfunction. We recently demonstrated that REDD1 protein expression was elevated in response to hyperglycemia in the retina of diabetic rodents. REDD1 is an important regulator of Akt and mammalian target of rapamycin and as such plays a key role in neuronal function and survival. In R28 retinal cells in culture, hyperglycemic conditions enhanced REDD1 protein expression concomitant with caspase activation and cell death. By contrast, in REDD1-deficient R28 cells, neither hyperglycemic conditions nor the absence of insulin in culture medium were sufficient to promote cell death. In the retinas of streptozotocin-induced diabetic mice, retinal apoptosis was dramatically elevated compared with nondiabetic controls, whereas no difference was observed in diabetic and nondiabetic REDD1-deficient mice. Electroretinogram abnormalities observed in b-wave and oscillatory potentials of diabetic wild-type mice were also absent in REDD1-deficient mice. Moreover, diabetic wild-type mice exhibited functional deficiencies in visual acuity and contrast sensitivity, whereas diabetic REDD1-deficient mice had no visual dysfunction. The results support a role for REDD1 in diabetes-induced retinal neurodegeneration.

Obesity is associated with elevated intracellular nitric oxide (NO) production, which promotes nitrosative stress in metabolic tissues such as liver and skeletal muscle, contributing to insulin resistance. The onset of obesity-associated insulin resistance is due, in part, to the compromise of hepatic autophagy, a process that leads to lysosomal degradation of cellular components. However, it is not known how NO bioactivity might impact autophagy in obesity. Here, we establish that S-nitrosoglutathione reductase (GSNOR), a major protein denitrosylase, provides a key regulatory link between inflammation and autophagy, which is disrupted in obesity and diabetes. We demonstrate that obesity promotes S-nitrosylation of lysosomal proteins in the liver, thereby impairing lysosomal enzyme activities. Moreover, in mice and humans, obesity and diabetes are accompanied by decreases in GSNOR activity, engendering nitrosative stress. In mice with a GSNOR deletion, diet-induced obesity increases lysosomal nitrosative stress and impairs autophagy in the liver, leading to hepatic insulin resistance. Conversely, liver-specific overexpression of GSNOR in obese mice markedly enhances lysosomal function and autophagy and, remarkably, improves insulin action and glucose homeostasis. Furthermore, overexpression of S-nitrosylation-resistant variants of lysosomal enzymes enhances autophagy, and pharmacologically and genetically enhancing autophagy improves hepatic insulin sensitivity in GSNOR-deficient hepatocytes. Taken together, our data indicate that obesity-induced protein S-nitrosylation is a key mechanism compromising the hepatic autophagy, contributing to hepatic insulin resistance.

The phenomenon of a discrepancy between glycated hemoglobin levels and other indicators of average glycemia may be due to many factors but can be measured as the glycation gap (GGap). This GGap is associated with differences in complications in patients with diabetes and may possibly be explained by dissimilarities in deglycation in turn leading to altered production of advanced glycation end products (AGEs). We hypothesized that variations in the level of the deglycating enzyme fructosamine-3-kinase (FN3K) might be associated with the GGap. We measured erythrocyte FN3K concentrations and enzyme activity in a population dichotomized for a large positive or negative GGap. FN3K protein was higher and we found a striking threefold greater activity (323%) at any given FN3K protein level in the erythrocytes of the negative-GGap group compared with the positive-GGap group. This was associated with lower AGE levels in the negative-GGap group (79%), lower proinflammatory adipokines (leptin-to-adiponectin ratio) (73%), and much lower prothrombotic PAI-1 levels (19%). We conclude that FN3K may play a key role in the GGap and thus diabetes complications such that FN3K may be a potential predictor of the risk of diabetes complications. Pharmacological modifications of its activity may provide a novel approach to their prevention.

We recently showed that interleukin (IL)-6-type cytokine signaling in adipocytes induces free fatty acid release from visceral adipocytes, thereby promoting obesity-induced hepatic insulin resistance and steatosis. In addition, IL-6-type cytokines may increase the release of leptin from adipocytes and by those means induce glucagon-like peptide 1 (GLP-1) secretion. We thus hypothesized that IL-6-type cytokine signaling in adipocytes may regulate insulin secretion. To this end, mice with adipocyte-specific knockout of gp130, the signal transducer protein of IL-6, were fed a high-fat diet for 12 weeks. Compared with control littermates, knockout mice showed impaired glucose tolerance and circulating leptin, GLP-1, and insulin levels were reduced. In line, leptin release from isolated adipocytes was reduced, and intestinal proprotein convertase subtilisin/kexin type 1 (Pcsk1) expression, the gene encoding PC1/3, which controls GLP-1 production, was decreased in knockout mice. Importantly, treatment with the GLP-1 receptor antagonist exendin 9-39 abolished the observed difference in glucose tolerance between control and knockout mice. Ex vivo, supernatant collected from isolated adipocytes of gp130 knockout mice blunted Pcsk1 expression and GLP-1 release from GLUTag cells. In contrast, glucose- and GLP-1-stimulated insulin secretion was not affected in islets of knockout mice. In conclusion, adipocyte-specific IL-6 signaling induces intestinal GLP-1 release to enhance insulin secretion, thereby counteracting insulin resistance in obesity.

We examined the association between plasma 25-hydroxyvitamin D [25(OH)D] concentration and islet autoimmunity (IA) and whether vitamin D gene polymorphisms modify the effect of 25(OH)D on IA risk. We followed 8,676 children at increased genetic risk of type 1 diabetes at six sites in the U.S. and Europe. We defined IA as positivity for at least one autoantibody (GADA, IAA, or IA-2A) on two or more visits. We conducted a risk set sampled nested case-control study of 376 IA case subjects and up to 3 control subjects per case subject. 25(OH)D concentration was measured on all samples prior to, and including, the first IA positive visit. Nine polymorphisms in VDR, CYP24A, CYP27B1, GC, and RXRA were analyzed as effect modifiers of 25(OH)D. Adjusting for HLA-DR-DQ and ancestry, higher childhood 25(OH)D was associated with lower IA risk (odds ratio = 0.93 for a 5 nmol/L difference; 95% CI 0.89, 0.97). Moreover, this association was modified by VDR rs7975232 (interaction P = 0.0072), where increased childhood 25(OH)D was associated with a decreasing IA risk based upon number of minor alleles: 0 (1.00; 0.93, 1.07), 1 (0.92; 0.89, 0.96), and 2 (0.86; 0.80, 0.92). Vitamin D and VDR may have a combined role in IA development in children at increased genetic risk for type 1 diabetes.