Endothelial nitric oxide (NO) signaling plays a physiological role in limiting obesity-associated insulin resistance and inflammation. This study was undertaken to investigate whether this NO effect involves polarization of macrophages toward an anti-inflammatory M2 phenotype. Mice with transgenic endothelial NO synthase overexpression were protected against high-fat diet (HFD)-induced hepatic inflammation and insulin resistance, and this effect was associated with reduced proinflammatory M1 and increased anti-inflammatory M2 activation of Kupffer cells. In cell culture studies, exposure of macrophages to endothelial NO similarly reduced inflammatory (M1) and increased anti-inflammatory (M2) gene expression. Similar effects were induced by macrophage overexpression of vasodilator-stimulated phosphoprotein (VASP), a key downstream mediator of intracellular NO signaling. Conversely, VASP deficiency induced proinflammatory M1 macrophage activation, and the transplantation of bone marrow from VASP-deficient donor mice into normal recipients caused hepatic inflammation and insulin resistance resembling that induced in normal mice by consumption of an HFD. These data suggest that proinflammatory macrophage M1 activation and macrophage-mediated inflammation are tonically inhibited by NO → VASP signal transduction, and that reduced NO → VASP signaling is involved in the effect of HFD feeding to induce M1 activation of Kupffer cells and associated hepatic inflammation. Our data implicate endothelial NO → VASP signaling as a physiological determinant of macrophage polarization and show that signaling via this pathway is required to prevent hepatic inflammation and insulin resistance.

Metabolic syndrome (MetS) doubles the risk of adverse cardiovascular events. Glucagon-like peptide 1 (GLP-1) receptor agonists induce weight loss, increase insulin secretion, and improve glucose tolerance. Studies in healthy animals suggest cardioprotective properties of GLP-1 receptor agonists, perhaps partially mediated by improved sarco-endoplasmic reticulum Ca(2+) ATPase (SERCA) activity. We examined the acute effect of GLP-1 receptor agonists on coronary smooth muscle cells (CSM) enzymatically isolated from lean, healthy Ossabaw miniature swine. Intracellular Ca(2+) handling was interrogated with fura-2. The GLP-1 receptor agonist exenatide activated SERCA but did not alter other Ca(2+) transporters. Further, we tested the hypothesis that chronic, in vivo treatment with GLP-1 receptor agonist AC3174 would attenuate coronary artery disease (CAD) in swine with MetS. MetS was induced in 20 swine by 6 months' feeding of a hypercaloric, atherogenic diet. Swine were then randomized (n = 10/group) into placebo or AC3174 treatment groups and continued the diet for an additional 6 months. AC3174 treatment attenuated weight gain, increased insulin secretion, and improved glucose tolerance. Intravascular ultrasound and histology showed no effect of AC3174 on CAD. MetS abolished SERCA activation by GLP-1 receptor agonists. We conclude that MetS confers vascular resistance to GLP-1 receptor agonists, partially through impaired cellular signaling steps involving SERCA.

Mitochondrial ADP transport may represent a convergence point unifying two prominent working models for the development of insulin resistance, as reactive lipids (specifically palmitoyl-CoA [P-CoA]) can inhibit ADP transport and subsequently increase mitochondrial reactive oxygen species emissions. In the current study, we aimed to determine if exercise training in humans diminished P-CoA attenuation of mitochondrial ADP respiratory sensitivity. Six weeks of exercise training increased whole-body glucose homeostasis and skeletal muscle Akt signaling and reduced markers of oxidative stress without reducing maximal mitochondrial H2O2 emissions. To ascertain if enhanced mitochondrial ADP transport contributed to the improvement in the in vivo oxidative state, we determined mitochondrial ADP sensitivity in the presence and absence of P-CoA. In the absence of P-CoA, exercise training reduced mitochondrial ADP sensitivity. In contrast, exercise training increased mitochondrial ADP sensitivity with P-CoA present. We further show that P-CoA noncompetitively inhibits mitochondrial ADP transport and the ability of ADP to attenuate mitochondrial H2O2 emission. Altogether, the current data provide a potential mechanism for how P-CoA contributes to insulin resistance and highlight the ability of exercise training to diminish P-CoA attenuation in mitochondrial ADP transport.

Dietary polyphenols protect against metabolic syndrome, despite limited absorption and digestion, raising questions about their mechanism of action. We hypothesized that one mechanism may involve the gut microbiota. To test this hypothesis, C57BL/6J mice were fed a high-fat diet (HFD) containing 1% Concord grape polyphenols (GP). Relative to vehicle controls, GP attenuated several effects of HFD feeding, including weight gain, adiposity, serum inflammatory markers (tumor necrosis factor [TNF]α, interleukin [IL]-6, and lipopolysaccharide), and glucose intolerance. GP lowered intestinal expression of inflammatory markers (TNFα, IL-6, inducible nitric oxide synthase) and a gene for glucose absorption (Glut2). GP increased intestinal expression of genes involved in barrier function (occludin) and limiting triglyceride storage (fasting-induced adipocyte factor). GP also increased intestinal gene expression of proglucagon, a precursor of proteins that promote insulin production and gut barrier integrity. 16S rRNA gene sequencing and quantitative PCR of cecal and fecal samples demonstrated that GP dramatically increased the growth of Akkermansia muciniphila and decreased the proportion of Firmicutes to Bacteroidetes, consistent with prior reports that similar changes in microbial community structure can protect from diet-induced obesity and metabolic disease. These data suggest that GP act in the intestine to modify gut microbial community structure, resulting in lower intestinal and systemic inflammation and improved metabolic outcomes. The gut microbiota may thus provide the missing link in the mechanism of action of poorly absorbed dietary polyphenols.

Islet autoantibodies detected at disease onset in patients with type 1 diabetes are signs of an autoimmune destruction of the insulin-producing β-cells. To further investigate the genetic determinants of autoantibody positivity, we performed dense immune-focused genotyping on the Immunochip array and tested for association with seven disease-specific autoantibodies in a large cross-sectional cohort of 6,160 type 1 diabetes-affected siblings. The genetic association with positivity for GAD autoantibodies (GADAs), IA2 antigen (IA-2A), zinc transporter 8, thyroid peroxidase, gastric parietal cells (PCAs), tissue transglutaminase, and 21-hydroxylase was tested using a linear mixed-model regression approach to simultaneously control for population structure and family relatedness. Four loci were associated with autoantibody positivity at genome-wide significance. Positivity for GADA was associated with 3q28/LPP, for IA-2A with 1q23/FCRL3 and 11q13/RELA, and for PCAs with 2q24/IFIH1. The 3q28 locus showed association after only 3 years duration and might therefore be a marker of persistent GADA positivity. The 1q23, 11q13, and 2q24 loci were associated with autoantibodies close to diabetes onset and constitute candidates for early screening. Major susceptibility loci for islet autoantibodies are separate from type 1 diabetes risk, which may have consequences for intervention strategies to reduce autoimmunity.

Perilipin 5 (PLIN5) is a lipid droplet protein and is highly expressed in oxidative tissue. Expression of the PLIN5 gene is regulated by peroxisome proliferator-activated receptor-α, fasting, and exercise. However, the effect of increased muscle PLIN5 expression on whole-body energy homeostasis remains unclear. To examine this, we developed a mouse line with skeletal muscle PLIN5 overexpression (MCK-Plin5). We show that MCK-Plin5 mice have increased energy metabolism and accumulate more intramyocellular triacylglycerol but have normal glucose and insulin tolerance. MCK-Plin5 mice fed high-fat chow manifest lower expression of inflammatory markers in their liver and increased expression of "browning" factors in adipose tissue. This muscle-driven phenotype is, at least in part, mediated by myokines; the MCK-Plin5 mice have 80-fold higher FGF21 gene expression in muscle and increased serum FGF21 concentration. The increase in FGF21 occurs mainly in muscles with a predominance of fast-twitch fibers, suggesting that fiber type-specific lipid storage may be part of the mechanism conferring metabolic protection in MCK-Plin5 mice. In conclusion, upregulating the PLIN5 level in skeletal muscle drives expression of the FGF21 gene in fast-twitch fibers and is metabolically protective. These findings provide insight into the physiology of PLIN5 and the potential contribution of its upregulation during exercise.

Intestine contributes to energy homeostasis through the absorption, metabolism, and transfer of nutrients to the organism. We demonstrated previously that hepatocyte nuclear receptor-4α (HNF-4α) controls intestinal epithelium homeostasis and intestinal absorption of dietary lipids. HNF-4γ, the other HNF-4 form highly expressed in intestine, is much less studied. In HNF-4γ knockout mice, we detect an exaggerated insulin peak and improvement in glucose tolerance during oral but not intraperitoneal glucose tolerance tests, highlighting the involvement of intestine. Moreover, the enteroendocrine L-type cell lineage is modified, as assessed by the increased expression of transcription factors Isl1, Foxa1/2, and Hnf4a, leading to an increase of both GLP-1-positive cell number and basal and stimulated GLP-1 plasma levels potentiating the glucose-stimulated insulin secretion. Using the GLP-1 antagonist exendin (9-39), we demonstrate a direct effect of GLP-1 on improved glucose tolerance. GLP-1 exerts a trophic effect on pancreatic β-cells, and we report an increase of the β-cell fraction correlated with an augmented number of proliferative islet cells and with resistance to streptozotocin-induced diabetes. In conclusion, the loss of HNF-4γ improves glucose homeostasis through a modulation of the enteroendocrine cell lineage.

In the nearly 100 years since the discovery of therapeutic insulin, significant research efforts have been directed at finding the underlying cause of type 1 diabetes (T1D) and developing a "cure" for the disease. While progress has clearly been made toward each of these goals, neither vision has been fulfilled. With increasing pressure from both public and private funders of diabetes research, growing impatience of those with T1D at the lack of practical discoveries, increased competition for research funds, uncertainties on the reproducibility of published scientific data, and questions regarding the value of animal models, the current research environment has become extraordinarily difficult to traverse from the perspective of investigators. As a result, there is an increasing pressure toward performance of what might be considered "safe" research, where the aim is to affirm existing dogmas rather than to pioneer efforts involving unconventional thought. Psychologists refer to this practice as "observational bias" while cartoonists label the process the "streetlight effect." In this Perspective, we consider notions in T1D research that should be subject to bold question and provide additional concepts, many somewhat orphan to research efforts, whose investigation could lead to a means for truly identifying the cause of and a cure for T1D.

Recent increases in the prevalence of obesity and type 2 diabetes mellitus (T2DM) in modern societies have been paralleled by reductions in the time their denizens spend asleep. Epidemiological studies have shown that disturbed sleep-comprising short, low-quality, and mistimed sleep-increases the risk of metabolic diseases, especially obesity and T2DM. Supporting a causal role of disturbed sleep, experimental animal and human studies have found that sleep loss can impair metabolic control and body weight regulation. Possible mechanisms for the observed changes comprise sleep loss-induced changes in appetite-signaling hormones (e.g., higher levels of the hunger-promoting hormone ghrelin) or hedonic brain responses, altered responses of peripheral tissues to metabolic signals, and changes in energy intake and expenditure. Even though the overall consensus is that sleep loss leads to metabolic perturbations promoting the development of obesity and T2DM, experimental evidence supporting the validity of this view has been inconsistent. This Perspective aims at discussing molecular to behavioral factors through which short, low-quality, and mistimed sleep may threaten metabolic public health. In this context, possible factors that may determine the extent to which poor sleep patterns increase the risk of metabolic pathologies within and across generations will be discussed (e.g., timing and genetics).

Previous studies suggest that diabetes impairs hematopoietic stem cell (HSC) mobilization in response to granulocyte colony-stimulating factor (G-CSF). In this study, we tested whether the CXCR4 antagonist plerixafor, differently from G-CSF, is effective in mobilizing HSCs in patients with diabetes. In a prospective study, individuals with and without diabetes (n = 10/group) were administered plerixafor to compare CD34(+) HSC mobilization; plerixafor was equally able to mobilize CD34(+) HSCs in the two groups, whereas in historical data, G-CSF was less effective in patients with diabetes. In a retrospective autologous transplantation study conducted on 706 patients, diabetes was associated with poorer mobilization in patients who received G-CSF with/without chemotherapy, whereas it was not in patients who received G-CSF plus plerixafor. Similarly in an allogeneic transplantation study (n = 335), diabetes was associated with poorer mobilization in patients who received G-CSF. Patients with diabetes who received G-CSF without plerixafor had a lower probability of reaching >50/μL CD34(+) HSCs, independent from confounding variables. In conclusion, diabetes negatively impacted HSC mobilization after G-CSF with or without chemotherapy but had no effect on mobilization induced by G-CSF with plerixafor. This finding has major implications for the care of patients with diabetes undergoing stem cell mobilization and transplantation and for the vascular regenerative potential of bone marrow stem cells.

Diabetes diagnostic therapy and research would strongly benefit from noninvasive accurate imaging of the functional β-cells in the pancreas. Here, we developed an analysis of functional β-cell mass (BCM) by measuring manganese (Mn(2+)) uptake kinetics into glucose-stimulated β-cells by T1-weighted in vivo Mn(2+)-mediated MRI (MnMRI) in C57Bl/6J mice. Weekly MRI analysis during the diabetes progression in mice fed a high-fat/high-sucrose diet (HFD) showed increased Mn(2+)-signals in the pancreas of the HFD-fed mice during the compensation phase, when glucose tolerance and glucose-stimulated insulin secretion (GSIS) were improved and BCM was increased compared with normal diet-fed mice. The increased signal was only transient; from the 4th week on, MRI signals decreased significantly in the HFD group, and the reduced MRI signal in HFD mice persisted over the whole 12-week experimental period, which again correlated with both impaired glucose tolerance and GSIS, although BCM remained unchanged. Rapid and significantly decreased MRI signals were confirmed in diabetic mice after streptozotocin (STZ) injection. No long-term effects of Mn(2+) on glucose tolerance were observed. Our optimized MnMRI protocol fulfills the requirements of noninvasive MRI analysis and detects already small changes in the functional BCM.

Diabetes affects bone marrow (BM) structure and impairs mobilization of stem cells (SCs) into peripheral blood (PB). This amplifies multiorgan complications because BMSCs promote vascular repair. Because diabetes skews macrophage phenotypes and BM macrophages (BMMΦ) prevent SC mobilization, we hypothesized that excess BMMΦ contribute to diabetic SC mobilopathy. We show that patients with diabetes have increased M1 macrophages, whereas diabetic mice have increased CD169(+) BMMΦ with SC-retaining activity. Depletion of BMMΦ restored SC mobilization in diabetic mice. We found that CD169 labels M1 macrophages and that conditioned medium (CM) from M1 macrophages, but not from M0 and M2 macrophages, induced chemokine (C-X-C motif) ligand 12 (CXCL12) expression by mesenchymal stem/stromal cells. In silico data mining and in vitro validation identified oncostatin M (OSM) as the soluble mediator contained in M1 CM that induces CXCL12 expression via a mitogen-activated protein kinase kinase-p38-signal transducer and activator of a transcription 3-dependent pathway. In diabetic mice, OSM neutralization prevented CXCL12 induction and improved granulocyte-colony stimulating factor and ischemia-induced mobilization, SC homing to ischemic muscles, and vascular recovery. In patients with diabetes, BM plasma OSM levels were higher and correlated with the BM-to-PB SC ratio. In conclusion, BMMΦ prevent SC mobilization by OSM secretion, and OSM antagonism is a strategy to restore BM function in diabetes, which can translate into protection mediated by BMSCs.

One of the most common complications of diabetes is diabetic foot ulcer. Diabetic ulcers do not heal easily due to diabetic neuropathy and reduced blood flow, and nonhealing ulcers may progress to gangrene, which necessitates amputation of the patient's foot. This study attempted to develop a new cell-based therapy for nonhealing diabetic ulcers using a full-thickness skin defect in a rat model of type 2 diabetes and obesity. Allogeneic adipose-derived stem cells (ASCs) were harvested from the inguinal fat of normal rats, and ASC sheets were created using cell sheet technology and transplanted into full-thickness skin defects in Zucker diabetic fatty rats. The results indicate that the transplantation of ASC sheets combined with artificial skin accelerated wound healing and vascularization, with significant differences observed 2 weeks after treatment. The ASC sheets secreted large amounts of several angiogenic growth factors in vitro, and transplanted ASCs were observed in perivascular regions and incorporated into the newly constructed vessel structures in vivo. These results suggest that ASC sheets accelerate wound healing both directly and indirectly in this diabetic wound-healing model. In conclusion, allogeneic ASC sheets exhibit potential as a new therapeutic strategy for the treatment of diabetic ulcers.