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AbstractThe "Integrated Physiology of the Exocrine and Endocrine Compartments in Pancreatic Diseases" Workshop was a 1.5-day scientific conference at the National Institutes of Health (Bethesda, MD) that engaged clinical and basic science investigators interested in diseases of the pancreas. This report summarizes the workshop proceedings. The goal of the workshop was to forge connections and identify gaps in knowledge that could guide future research directions. Presentations were segregated into 6 major themes, including (a) Pancreas Anatomy and Physiology; (b) Diabetes in the Setting of Exocrine Disease; (c) Metabolic Influences on the Exocrine Pancreas; (d) Genetic Drivers of Pancreatic Diseases; (e) Tools for Integrated Pancreatic Analysis; and (f) Implications of Exocrine-Endocrine Crosstalk. For each theme, there were multiple presentations followed by panel discussions on specific topics relevant to each area of research; these are summarized herein. Significantly, the discussions resulted in the identification of research gaps and opportunities for the field to address. In general, it was concluded that as a pancreas research community, we must more thoughtfully integrate our current knowledge of the normal physiology as well as the disease mechanisms that underlie endocrine and exocrine disorders so that there is a better understanding of the interplay between these compartments.

Aims/hypothesisT cell protein tyrosine phosphatase (TCPTP, encoded by PTPN2) regulates cytokine-induced pancreatic beta cell apoptosis and may contribute to the pathogenesis of type 1 diabetes. However, the role of TCPTP in pancreatic endocrine function and insulin secretion remains largely unknown.MethodsTo investigate the endocrine role of pancreatic TCPTP we generated mice with pancreas Ptpn2/TCPTP deletion (panc-TCPTP KO).ResultsWhen fed regular chow, panc-TCPTP KO and control mice exhibited comparable glucose tolerance. However, when challenged with prolonged high fat feeding panc-TCPTP KO mice exhibited impaired glucose tolerance and attenuated glucose-stimulated insulin secretion (GSIS). The defect in GSIS was recapitulated in primary islets ex vivo and after TCPTP pharmacological inhibition or lentiviral-mediated TCPTP knockdown in the glucose-responsive MIN6 beta cells, consistent with this being cell autonomous. Reconstitution of TCPTP in knockdown cells reversed the defect in GSIS demonstrating that the defect was a direct consequence of TCPTP deficiency. The reduced insulin secretion in TCPTP knockdown MIN6 beta cells was associated with decreased insulin content and glucose sensing. Furthermore, TCPTP deficiency led to enhanced tyrosyl phosphorylation of signal transducer and activator of transcription 1 and 3 (STAT 1/3), and substrate trapping studies in MIN6 beta cells identified STAT 1/3 as TCPTP substrates. STAT3 pharmacological inhibition and small interfering RNA-mediated STAT3 knockdown in TCPTP deficient cells restored GSIS to control levels, indicating that the effects of TCPTP deficiency were mediated, at least in part, through enhanced STAT3 phosphorylation and signalling.Conclusions/interpretationThese studies identify a novel role for TCPTP in insulin secretion and uncover STAT3 as a physiologically relevant target for TCPTP in the endocrine pancreas.

Introduction: The enhanced β-cell senescence that accompanies insulin resistance and aging contributes to cellular dysfunction and loss of transcriptional identity leading to type 2 diabetes (T2D). While senescence is among the 12 recognized hallmarks of aging, its relation to other hallmarks including altered nutrient sensing (insulin/IGF1 pathway) in β-cells is not fully understood. We previously reported that an increased expression of IGF1R in mouse and human β-cells is a marker of older β-cells; however, its contribution to age-related dysfunction and cellular senescence remains to be determined. Methods: In this study, we explored the direct role of IGF1R in β-cell function and senescence using two independent mouse models with decreased IGF1/IGF1R signaling: a) Ames Dwarf mice (Dwarf +/+), which lack growth hormone and therefore have reduced circulating levels of IGF1, and b) inducible β-cell-specific IGF1R knockdown (βIgf1rKD) mice. Results: Compared to Dwarf+/- mice, Dwarf+/+ mice had lower body and pancreas weight, lower circulating IGF1 and insulin levels, and lower IGF1R and p21Cip1 protein expression in β-cells, suggesting the suppression of senescence. Adult βIgf1rKD mice showed improved glucose clearance and glucose-induced insulin secretion, accompanied by decreased p21Cip1 protein expression in β-cells. RNA-Seq of islets isolated from these βIgf1rKD mice revealed the restoration of three signaling pathways known to be downregulated by aging: sulfide oxidation, autophagy, and mTOR signaling. Additionally, deletion of IGF1R in mouse β-cells increased transcription of genes important for maintaining β-cell identity and function, such as Mafa, Nkx6.1, and Kcnj11, while decreasing senescence-related genes, such as Cdkn2a, Il1b, and Serpine 1. Decreased senescence and improved insulin-secretory function of β-cells were also evident when the βIgf1rKD mice were fed a high-fat diet (HFD; 60% kcal from fat, for 5 weeks). Discussion: These results suggest that IGF1R signaling plays a causal role in aging-induced β-cell dysfunction. Our data also demonstrate a relationship between decreased IGF1R signaling and suppressed cellular senescence in pancreatic β-cells. Future studies can further our understanding of the interaction between senescence and aging, developing interventions that restore β-cell function and identity, therefore preventing the progression to T2D. [ABSTRACT FROM AUTHOR]

Healthy pancreatic β-cells adapt to systemic insulin resistance to maintain normal blood glucose levels, and a failure of this adaptation leads to type 2 diabetes in humans. While genome-wide association studies have uncovered genetic variants that are associated with type 2 diabetes, it is still insufficient to explain the high prevalence of this disease. Epigenetics is the study of gene expression changes that do not involve DNA sequence alterations such as DNA methylation, histone modification, and non-coding RNAs. Over the last decade, a large number of studies have reported on the role of epigenetics in β-cell biology. In this review, we summarize the epigenetic mechanisms in β-cell adaptation and type 2 diabetes, including alterations in three-dimensional chromatin structure and RNA modifications. [ABSTRACT FROM AUTHOR]

Although it is well established that type 2 diabetes (T2D) is generally due to the progressive loss of β-cell insulin secretion against a background of insulin resistance, the actual correlation of reduced β-cell mass to its defective function continues to be debated. There is evidence that a compensatory increase in β-cell mass, and the consequent insulin secretion, can effectively cope with states of insulin resistance, until hyperglycemia supervenes. Recent data strongly indicate that the mechanisms by which islets are able to compensate in response to insulin resistance in peripheral tissues is secondary to hyperplasia, as well as the activation of multiple cellular machineries with diverse functions. Importantly, islet cells exhibit plasticity in altering their endocrine commitment; for example, by switching from secretion of glucagon to secretion of insulin and back (transdifferentiation) or from an active secretory state to a nonsecretory quiescent state (dedifferentiation) and back. Lineage tracing (a method used to track each cell though its differentiation process) has demonstrated these potentials in murine models. A limitation to drawing conclusions from human islet research is that most studies are derived from human autopsy and/or organ donor samples, which lack in vivo functional and metabolic profiling. In this review, we specifically focus on evidence of islet plasticity in humans-from the normal state, progressing to insulin resistance to overt T2D-to explain the seemingly contradictory results from different cross-sectional studies in the literature. We hope the discussion on this intriguing scenario will provide a forum for the scientific community to better understand the disease and in the long term pave the way for personalized therapies. [ABSTRACT FROM AUTHOR]

Abstract: Cannabinoid 1 receptors (CB1Rs) are expressed in peripheral tissues, including islets of Langerhans, where their function(s) is under scrutiny. Using mouse β‐cell lines, human islets and CB1R‐null (CB1R−/−) mice, we have now investigated the role of CB1Rs in modulating β‐cell function and glucose responsiveness. Synthetic CB1R agonists diminished GLP‐1‐mediated cAMP accumulation and insulin secretion as well as glucose‐stimulated insulin secretion in mouse β‐cell lines and human islets. In addition, silencing CB1R in mouse β cells resulted in an increased expression of pro‐insulin, glucokinase (GCK) and glucose transporter 2 (GLUT2), but this increase was lost in β cells lacking insulin receptor. Furthermore, CB1R−/− mice had increased pro‐insulin, GCK and GLUT2 expression in β cells. Our results suggest that CB1R signalling in pancreatic islets may be harnessed to improve β‐cell glucose responsiveness and preserve their function. Thus, our findings further support that blocking peripheral CB1Rs would be beneficial to β‐cell function in type 2 diabetes. [ABSTRACT FROM AUTHOR]

This is the third in a series of Perspectives on intracellular signaling pathways coupled to proliferation in pancreatic β-cells. We contrast the large knowledge base in rodent β-cells with the more limited human database. With the increasing incidence of type 1 diabetes and the recognition that type 2 diabetes is also due in part to a deficiency of functioning β-cells, there is great urgency to identify therapeutic approaches to expand human β-cell numbers. Therapeutic approaches might include stem cell differentiation, transdifferentiation, or expansion of cadaver islets or residual endogenous β-cells. In these Perspectives, we focus on β-cell proliferation. Past Perspectives reviewed fundamental cell cycle regulation and its upstream regulation by insulin/IGF signaling via phosphatidylinositol-3 kinase/ mammalian target of rapamycin signaling, glucose, glycogen synthase kinase-3 and liver kinase B1, protein kinase Cζ, calcium-calcineurin-nuclear factor of activated T cells, epidermal growth factor/platelet-derived growth factor family members, Wnt/β-catenin, leptin, and estrogen and progesterone. Here, we emphasize Janus kinase/signal transducers and activators of transcription, Ras/Raf/extracellular signal-related kinase, cadherins and integrins, G-protein-coupled receptors, and transforming growth factor b signaling. We hope these three Perspectives will serve to introduce these pathways to new researchers and will encourage additional investigators to focus on understanding how to harness key intracellular signaling pathways for therapeutic human β-cell regeneration for diabetes. [ABSTRACT FROM AUTHOR]

Type 2 diabetes (T2D) is associated with insulin resistance in target tissues including the β-cell, leading to significant β-cell loss and secretory dysfunction. T2D is also associated with aging, and the underlying mechanisms that increase susceptibility of an individual to develop the disease implicate epigenetics: interactions between susceptible loci and the environment. In this review, we discuss the effects of aging on β-cell function and adaptation, besides the significance of mitochondria in islet bioenergetics and epigenome. We highlight three important modulators of the islet epigenome, namely: metabolites, hormones, and the nutritional state. Unraveling the signaling pathways that regulate the islet epigenome during aging will help to better understand the development of disease progression and to design novel therapies for diabetes prevention. [ABSTRACT FROM AUTHOR]

Insulin resistance, hyperinsulinemia, and hyperproinsulinemia occur early in the pathogenesis of type 2 diabetes (T2D). Elevated levels of proinsulin and proinsulin intermediates are markers of β-cell dysfunction and are strongly associated with development of T2D in humans. However, the mechanism(s) underlying β-cell dysfunction leading to hyperproinsulinemia is poorly understood. Here, we show that disruption of insulin receptor (IR) expression in β cells has a direct impact on the expression of the convertase enzyme carboxypeptidase E (CPE) by inhibition of the eukaryotic translation initiation factor 4 gamma 1 translation initiation complex scaffolding protein that is mediated by the key transcription factors pancreatic and duodenal homeobox 1 and sterol regulatory element-binding protein 1, together leading to poor proinsulin processing. Reexpression of IR or restoring CPE expression each independently reverses the phenotype. Our results reveal the identity of key players that establish a previously unknown link between insulin signaling, translation initiation, and proinsulin processing, and provide previously unidentified mechanistic insight into the development of hyperproinsulinemia in insulinresistant states. [ABSTRACT FROM AUTHOR]

Patients with type 2 diabetes (T2D) often exhibit hyperglucago-nemia despite hyperglycemia, implicating defective α-cell function. Although endoplasmic reticulum (ER) stress has been suggested to underlie b-cell dysfunction in T2D, its role in α-cell biology remains unclear. X-box binding protein 1 (XBP1) is a transcription factor that plays a crucial role in the unfolded protein response (UPR), and its deficiency in β-cells has been reported to impair insulin secretion, leading to glucose intolerance. To evaluate the role of XBP1 in α-cells, we created complementary in vivo (α-cell-specific XBP1 knockout [αXBPKO] mice) and in vitro (stable XBP1 knockdown α-cell line [αXBPKD]) models. The αXBPKO mice exhibited glucose intolerance, mild insulin resistance, and an inability to suppress glucagon secretion after glucose stimulation. αXBPKD cells exhibited activation of inositol-requiring enzyme 1, an upstream activator of XBP1, leading to phosphorylation of Jun NH2-terminal kinase. Interestingly, insulin treatment of αXBPKD cells reduced tyrosine phosphorylation of insulin receptor substrate 1 (IRS1) (pY896) and phosphorylation of Akt while enhancing serine phosphorylation (pS307) of IRS1. Consequently, the αXBPKD cells exhibited blunted suppression of glucagon secretion after insulin treatment in the presence of high glucose. Together, these data indicate that XBP1 deficiency in pancreatic α-cells induces altered insulin signaling and dysfunctional glucagon secretion. [ABSTRACT FROM AUTHOR]

Insulin resistance, reduced β-cell mass, and hyperglucagonemia are consistent features in type 2 diabetes mellitus (T2DM). We used pancreas and islets from humans with T2DM to examine the regulation of insulin signaling and cell-cycle control of islet cells. We observed reduced β-cell mass and increased a-cell mass in the Type 2 diabetic pancreas. Confocal microscopy, real-time PCR and western blotting analyses revealed increased expression of PCNA and down-regulation of p27-Kip1 and altered expression of insulin receptors, insulin receptor substrate-2 and phosphorylated BAD. To investigate the mechanisms underlying these findings, we examined a mouse model of insulin resistance in β-cells - which also exhibits reduced β-cell mass, the β-cell-specific insulin receptor knockout (bIRKO). Freshly isolated islets and β-cell lines derived from bIRKO mice exhibited poor cell-cycle progression, nuclear restriction of FoxO1 and reduced expression of cell-cycle proteins favoring growth arrest. Re-expression of insulin receptors in bIRKO β-cells reversed the defects and promoted cell cycle progression and proliferation implying a role for insulin-signaling in β-cell growth. These data provide evidence that human b- and α-cells can enter the cell-cycle, but proliferation of β-cells in T2DM fails due to G1-to-S phase arrest secondary to defective insulin signaling. Activation of insulin signaling, FoxO1 and proteins in β-cell-cycle progression are attractive therapeutic targets to enhance β-cell regeneration in the treatment of T2DM. [ABSTRACT FROM AUTHOR]

OBJECTIVE--A major determinant of the progression from insulin resistance to the development of overt type 2 diabetes is a failure to mount an appropriate compensatory β-cell hyperplastic response to maintain normoglycemia. We undertook the present study to directly explore the significance of the cell cycle protein cyclin D2 in the expansion of β-cell mass in two different models of insulin resistance. RESEARCH DESIGN AND METHODS--We created compound knockouts by crossing mice deficient in cyclin D2 (D2KO) with either the insulin receptor substrate 1 knockout (IRS1KO) mice or the insulin receptor liver-specific knockout mice (LIRKO), neither of which develops overt diabetes on its own because of robust compensatory β-cell hyperplasia. We phenotyped the double knockouts and used RT-qPCR and immunohistochemistry to examine β-cell mass. RESULTS--Both compound knockouts, D2KO/LIRKO and D2KO/IRS1KO, exhibited insulin resistance and hyperinsulinemia and an absence of compensatory β-cell hyperplasia. However, the diabetic D2KO/LIRKO group rapidly succumbed early compared with a relatively normal lifespan in the glucose-intolerant D2KO/IRS1KO mice. CONCLUSIONS--This study provides direct genetic evidence that cyclin D2 is essential for the expansion of β-cell mass in response to a spectrum of insulin resistance and points to the cell-cycle protein as a potential therapeutic target that can be harnessed for preventing and curing type 2 diabetes. Diabetes 59:987-996, 2010 [ABSTRACT FROM AUTHOR]

Islet β-cells express both insulin receptors and insulin-signaling proteins. Recent evidence from rodents in vivo and from islets isolated from rodents or humans suggests that the insulin signaling pathway is physiologically important for glucose sensing. We evaluated whether insulin regulates β-cell function in healthy humans in vivo. Glucose-induced insulin secretion was assessed in healthy humans following 4-h saline (low insulin/sham clamp) or isoglycemic-hyperinsulinemic (high insulin) clamps using B28- Asp insulin that could be immunologically distinguished from endogenous insulin. Insulin and C-peptide clearance were evaluated to understand the impact of hyperinsulinemia on estimates of β-cell function. Preexposure to exogenous insulin increased the endogenous insulin secretory response to glucose by ≈40%. C-peptide response also increased, although not to the level predicted by insulin. Insulin clearance was not saturated at hyperin- sulinemia, but metabolic clearance of C-peptide, assessed by infusion of stable isotope-labeled C-peptide, increased modestly during hyperinsulinemic clamp. These studies demonstrate that insulin potentiates glucose-stimulated insulin secretion in vivo in healthy humans. In addition, hyperinsulinemia increases C-peptide clearance, which may lead to modest underestimation of β-cell secretory response when using these methods during prolonged dynamic testing. [ABSTRACT FROM AUTHOR]

Summary: β cell dysfunction is a central component of the pathogenesis of type 2 diabetes. Using oligonucleotide microarrays and real-time PCR of pancreatic islets isolated from humans with type 2 diabetes versus normal glucose-tolerant controls, we identified multiple changes in expression of genes known to be important in β cell function, including major decreases in expression of HNF4α, insulin receptor, IRS2, Akt2, and several glucose-metabolic-pathway genes. There was also a 90% decrease in expression of the transcription factor ARNT. Reducing ARNT levels in Min6 cells with small interfering RNA (siRNA) resulted in markedly impaired glucose-stimulated insulin release and changes in gene expression similar to those in human type 2 islets. Likewise, β cell-specific ARNT knockout mice exhibited abnormal glucose tolerance, impaired insulin secretion, and changes in islet gene expression that mimicked those in human diabetic islets. Together, these data suggest an important role for decreased ARNT and altered gene expression in the impaired islet function of human type 2 diabetes. [Copyright &y& Elsevier]

Mice with deletion of insulin receptor substrate (IRS)-1 (IRS-1 knockout [KO] mice) show mild insulin resistance and defective glucose-stimulated insulin secretion and reduced insulin synthesis. To further define the role of IRS-1 in islet function, we examined the insulin secretory defect in the knockouts using freshly isolated islets and primary beta-cells. IRS-1 KO beta-cells exhibited a significantly shorter increase in intracellular free Ca(2+) concentration ([Ca(2+)](i)) than controls when briefly stimulated with glucose or glyceraldehyde and when l-arginine was used to potentiate the stimulatory effect of glucose. These changes were paralleled by a lower number of exocytotic events in the KO beta-cells in response to the same secretagogues, indicating reduced insulin secretion. Furthermore, the normal oscillations in intracellular Ca(2+) and O(2) consumption after glucose stimulation were dampened in freshly isolated KO islets. Semiquantitative RT-PCR showed a dramatically reduced islet expression of sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA)-2b and -3 in the mutants. These data provide evidence that IRS-1 modulation of insulin secretion is associated with Ca(2+) signaling and expression of SERCA-2b and -3 genes in pancreatic islets and provides a direct link between insulin resistance and defective insulin secretion. [ABSTRACT FROM AUTHOR]

Summary objective The development of type 2 diabetes is linked to insulin resistance coupled with a failure of pancreatic β-cells to compensate by adequate insulin secretion. design Here, we review studies obtained from genetically engineered mice that provide novel insights into the pathophysiology of insulin resistance. results Knockout models with monogenic impairment in insulin action have highlighted the potential role for insulin signalling molecules in insulin resistance at a tissue-specific level. Polygenic models have strengthened the idea that minor defects in insulin secretion and insulin action, when combined, can lead to diabetes, emphasizing the importance of interactions of different genetic loci in the production of diabetes. Knockout models with tissue-specific alterations in glucose or lipid metabolism have dissected the individual contributions of insulin-responsive organs to glucose homeostasis. They have demonstrated the central role of fat as an endocrine tissue in the maintenance of insulin sensitivity and the development of insulin resistance. Finally, these models have shown the potential role of impaired insulin action in pancreatic β-cells and brain in the development of insulin deficiency and obesity. [ABSTRACT FROM AUTHOR]

Investigates evidences related to the circulating islet growth factor in insulin-resistant states. Evidence of insulin resistance in obesity and non-insulin-dependent diabetes; Role of beta cells in insulin resistance compensation; Employment of normal islets in mice.

Type 2 diabetes is a polygenic disease characterized by defects in both insulin secretion and insulin action. We have previously reported that isolated insulin resistance in muscle by a tissue-specific insulin receptor knockout (MIRKO mouse) is not sufficient to alter glucose homeostasis, whereas beta-cell-specific insulin receptor knockout (betaIRKO) mice manifest severe progressive glucose intolerance due to loss of glucose-stimulated acute-phase insulin release. To explore the interaction between insulin resistance in muscle and altered insulin secretion, we created a double tissue-specific insulin receptor knockout in these tissues. Surprisingly, betaIRKO-MIRKO mice show an improvement rather than a deterioration of glucose tolerance when compared to betaIRKO mice. This is due to improved glucose-stimulated acute insulin release and redistribution of substrates with increased glucose uptake in adipose tissue and liver in vivo, without a significant decrease in muscle glucose uptake. Thus, insulin resistance in muscle leads to improved glucose-stimulated first-phase insulin secretion from beta-cells and shunting of substrates to nonmuscle tissues, collectively leading to improved glucose tolerance. These data suggest that muscle, either via changes in substrate availability or by acting as an endocrine tissue, communicates with and regulates insulin sensitivity in other tissues. [ABSTRACT FROM AUTHOR]

The isolation of high-quality RNA from endocrine pancreas sections represents a considerable challenge largely due to the high ribonuclease levels. Laser capture microdissection (LCM) of mammalian islets, in association with RNA extraction protocols, has emerged as a feasible approach to characterizing their genetic and proteomic profiles. However, a validated protocol to obtain high-quality RNA from LCM-derived human pancreas specimens that is appropriate for next-generation sequencing analysis is still lacking. In this study, we applied four methods (Picopure extraction kit, Qiazol protocol, Qiazol + Clean-up kit, and RNeasy Microkit + Carrier) to extract RNA from human islets obtained from both non-diabetic individuals and patients with type 2 diabetes who had undergone partial pancreatectomy, as well as handpicked islets from both non-diabetic and diabetic organ donors. The yield and purity of total RNA were determined by 260/280 absorbance using Nanodrop 100 and the RNA integrity number with a bioanalyzer. The results indicated that among the four methods, the RNeasy MicroKit + Carrier (Qiagen) provides the highest yield and purity. [ABSTRACT FROM AUTHOR]

Pancreatic β cell dysfunction is a key feature of type 2 diabetes, and novel regulators of insulin secretion are desirable. Here, we report that succinate receptor 1 (SUCNR1) is expressed in β cells and is upregulated in hyperglycemic states in mice and humans. We found that succinate acted as a hormone-like metabolite and stimulated insulin secretion via a SUCNR1-GqPKC–dependent mechanism in human β cells. Mice with β cell–specific Sucnr1 deficiency exhibited impaired glucose tolerance and insulin secretion on a high-fat diet, indicating that SUCNR1 is essential for preserving insulin secretion in diet-induced insulin resistance. Patients with impaired glucose tolerance showed an enhanced nutrition-related succinate response, which correlates with the potentiation of insulin secretion during intravenous glucose administration. These data demonstrate that the succinate/SUCNR1 axis is activated by high glucose and identify a GPCR-mediated amplifying pathway for insulin secretion relevant to the hyperinsulinemia of prediabetic states. [ABSTRACT FROM AUTHOR]

Summary Investigation of cell-cycle kinetics in mammalian pancreatic β cells has mostly focused on transition from the quiescent (G0) to G1 phase. Here, we report that centromere protein A (CENP-A), which is required for chromosome segregation during the M-phase, is necessary for adaptive β cell proliferation. Receptor-mediated insulin signaling promotes DNA-binding activity of FoxM1 to regulate expression of CENP-A and polo-like kinase-1 (PLK1) by modulating cyclin-dependent kinase-1/2. CENP-A deposition at the centromere is augmented by PLK1 to promote mitosis, while knocking down CENP-A limits β cell proliferation and survival. CENP-A deficiency in β cells leads to impaired adaptive proliferation in response to pregnancy, acute and chronic insulin resistance, and aging in mice. Insulin-stimulated CENP-A/PLK1 protein expression is blunted in islets from patients with type 2 diabetes. These data implicate the insulin-FoxM1/PLK1/CENP-A pathway-regulated mitotic cell-cycle progression as an essential component in the β cell adaptation to delay and/or prevent progression to diabetes. [ABSTRACT FROM AUTHOR]

Type 2 diabetes is a complex and heterogeneous disease caused by both environmental and genetic factors. The pathophysiology of the common polygenic form of type 2 diabetes results from early-onset insulin resistance coupled with functional defects in insulin secretion by pancreatic islet cells. However, the precise factors leading to this disease are not known. In contrast, the monogenic form of diabetes, maturity-onset diabetes of the young (MODY), primarily results from defects that affect the functioning of islet cells.

Insulin resistance is a major hallmark in the development of type II diabetes, which is characterized by the failure of insulin to promote glucose uptake in muscle and to suppress glucose production in liver. The serine-threonine kinase Akt (PKB) is a principal target of insulin signaling that inhibits hepatic glucose output when glucose is available from food. Here we show that TRB3, a mammalian homolog of Drosophila tribbles, functions as a negative modulator of Akt. TRB3 expression is induced in liver under fasting conditions, and TRB3 disrupts insulin signaling by binding directly to Akt and blocking activation of the kinase. Amounts of TRB3 RNA and protein were increased in livers of db/db diabetic mice compared with those in wild-type mice. Hepatic overexpression of TRB3 in amounts comparable to those in db/db mice promoted hyperglycemia and glucose intolerance. Our results suggest that, by interfering with Akt activation, TRB3 contributes to insulin resistance in individuals with susceptibility to type II diabetes. [ABSTRACT FROM AUTHOR]

Dysregulated glucagon secretion deteriorates glycemic control in type 1 and type 2 diabetes. Although insulin is known to regulate glucagon secretion via its cognate receptor (insulin receptor, INSR) in pancreatic alpha cells, the role of downstream proteins and signaling pathways underlying insulin's activities are not fully defined. Using in vivo (knockout) and in vitro (knockdown) studies targeting insulin receptor substrate (IRS) proteins, we compared the relative roles of IRS1 and IRS2 in regulating alpha cell function. Alpha cell-specific IRS1-knockout mice exhibited glucose intolerance and inappropriate glucagon suppression during glucose tolerance tests. In contrast, alpha cell-specific IRS2-knockout animals manifested normal glucose tolerance and suppression of glucagon secretion after glucose administration. Alpha cell lines with stable IRS1 knockdown could not repress glucagon mRNA expression and exhibited a reduction in phosphorylation of AKT Ser/Thr kinase (AKT, at Ser-473 and Thr-308). AlphaIRS1KD cells also displayed suppressed global protein translation, including reduced glucagon expression, impaired cytoplasmic Ca2+ response, and mitochondrial dysfunction. This was supported by the identification of novel IRS1-specific downstream target genes, Trpc3 and Cartpt, that are associated with glucagon regulation in alpha cells. These results provide evidence that IRS1, rather than IRS2, is a dominant regulator of pancreatic alpha cell function. [ABSTRACT FROM AUTHOR]

Analogs of the incretin hormones Gip and Glp-1 are used to treat type 2 diabetes and obesity. Findings in experimental models suggest that manipulating several hormones simultaneously may be more effective. To identify small molecules that increase the number of incretin-expressing cells, we established a high-throughput in vivo chemical screen by using the gip promoter to drive the expression of luciferase in zebrafish. All hits increased the numbers of neurogenin 3-expressing enteroendocrine progenitors, Gip-expressing K-cells, and Glp-1-expressing L-cells. One of the hits, a dual-specificity tyrosine phosphorylation-regulated kinase (DYRK) inhibitor, additionally decreased glucose levels in both larval and juvenile fish. Knock-down experiments indicated that nfatc4 , a downstream mediator of DYRKs, regulates incretin+ cell number in zebrafish, and that Dyrk1b regulates Glp-1 expression in an enteroendocrine cell line. DYRK inhibition also increased the number of incretin-expressing cells in diabetic mice, suggesting a conserved reinforcement of the enteroendocrine system, with possible implications for diabetes. [Display omitted] • In vivo screen identifies small molecules that increase the number of incretin+ cells • A GnRH agonist and DGAT1 and DYRK inhibitors boost differentiation of incretin+ cells • DYRK1A/B and downstream NFATC4 regulate the differentiation of incretin+ cells • DYRK inhibitors improve glucose control in both zebrafish and diabetic mice Analogs of the incretin hormones Gip and Glp-1 are typically used to treat diabetes. To identify factors directly increasing the number of incretin-expressing cells, Chu et al. establish a zebrafish screen and identify multiple hits, including DYRK inhibitors that increase incretins and improve glucose control in zebrafish and diabetic mice. [ABSTRACT FROM AUTHOR]

Insufficient insulin secretion and reduced pancreatic beta cell mass are hallmarks of type 2 diabetes (T2DM). Here, we confirm that a previously identified polymorphism (rs2295490/Q84R) in exon 2 of the pseudokinase-encoding gene tribbles 3 (TRB3) is associated with an increased risk for T2DM in 2 populations of people of mixed European descent. Carriers of the 84R allele had substantially reduced plasma levels of C-peptide, the product of proinsulin processing to insulin, suggesting a role for TRB3 in beta cell function. Overexpression of TRB3 84R in mouse beta cells, human islet cells, and the murine beta cell line MIN6 revealed reduced insulin exocytosis, associated with a marked reduction in docked insulin granules visualized by electron microscopy. Conversely, knockdown of TRB3 in MIN6 cells restored insulin secretion and expression of exocytosis genes. Further analysis in MIN6 cells demonstrated that TRB3 interacted with the transcription factor ATF4 and that this complex acted as a competitive inhibitor of cAMP response element-binding (CREB) transcription factor in the regulation of key exocytosis genes. In addition, the 84R TRB3 variant exhibited greater protein stability than wild-type TRB3 and increased binding affinity to Akt. Mice overexpressing TRB3 84R in beta cells displayed decreased beta cell mass, associated with reduced proliferation and enhanced apoptosis rates. These data link a missense polymorphism in human TRB3 to impaired insulin exocytosis and thus increased risk for T2DM. [ABSTRACT FROM AUTHOR]

Hypoxia-inducible factor-1alpha (HIF-1alpha) is a transcription factor that regulates cellular stress responses. While the levels of HIF-1alpha protein are tightly regulated, recent studies suggest that it can be active under normoxic conditions. We hypothesized that HIF-1alpha is required for normal beta cell function and reserve and that dysregulation may contribute to the pathogenesis of type 2 diabetes (T2D). Here we show that HIF-1alpha protein is present at low levels in mouse and human normoxic beta cells and islets. Decreased levels of HIF-1alpha impaired glucose-stimulated ATP generation and beta cell function. C57BL/6 mice with beta cell-specific Hif1a disruption (referred to herein as beta-Hif1a-null mice) exhibited glucose intolerance, beta cell dysfunction, and developed severe glucose intolerance on a high-fat diet. Increasing HIF-1alpha levels by inhibiting its degradation through iron chelation markedly improved insulin secretion and glucose tolerance in control mice fed a high-fat diet but not in beta-Hif1a-null mice. Increasing HIF-1alpha levels markedly increased expression of ARNT and other genes in human T2D islets and improved their function. Further analysis indicated that HIF-1alpha was bound to the Arnt promoter in a mouse beta cell line, suggesting direct regulation. Taken together, these findings suggest an important role for HIF-1alpha in beta cell reserve and regulation of ARNT expression and demonstrate that HIF-1alpha is a potential therapeutic target for the beta cell dysfunction of T2D. [ABSTRACT FROM AUTHOR]

The transcriptional coactivator PPARgamma coactivator 1alpha (PGC-1alpha) is a strong activator of mitochondrial biogenesis and oxidative metabolism. While expression of PGC-1alpha and many of its mitochondrial target genes are decreased in the skeletal muscle of patients with type 2 diabetes, no causal relationship between decreased PGC-1alpha expression and abnormal glucose metabolism has been established. To address this question, we generated skeletal muscle-specific PGC-1alpha knockout mice (MKOs), which developed significantly impaired glucose tolerance but showed normal peripheral insulin sensitivity. Surprisingly, MKOs had expanded pancreatic beta cell mass, but markedly reduced plasma insulin levels, in both fed and fasted conditions. Muscle tissue from MKOs showed increased expression of several proinflammatory genes, and these mice also had elevated levels of the circulating IL-6. We further demonstrated that IL-6 treatment of isolated mouse islets suppressed glucose-stimulated insulin secretion. These data clearly illustrate a causal role for muscle PGC-1alpha in maintenance of glucose homeostasis and highlight an unexpected cytokine-mediated crosstalk between skeletal muscle and pancreatic islets. [ABSTRACT FROM AUTHOR]

The insulin receptor substrate-2 (Irs2) branch of the insulin/IGF signaling system coordinates peripheral insulin action and pancreatic beta cell function, so mice lacking Irs2 display similarities to humans with type 2 diabetes. Here we show that beta cell-specific expression of Irs2 at a low or a high level delivered a graded physiologic response that promoted beta cell growth, survival, and insulin secretion that prevented diabetes in Irs2-/- mice, obese mice, and streptozotocin-treated mice; and that upon transplantation, the transgenic islets cured diabetes more effectively than WT islets. Thus, pharmacological approaches that promote Irs2 expression in beta cells, especially specific cAMP agonists, could be rational treatments for beta cell failure and diabetes. [ABSTRACT FROM AUTHOR]