Islet Beta Cell Function Research Articles

Galectin‐3 deficiency protects pancreatic islet cells from cytokine‐triggered apoptosis in vitro

Beta cell apoptosis is a hallmark of diabetes. Since we have previously shown that galectin-3 deficient (LGALS3(-/-) ) mice are relatively resistant to diabetes induction, the aim of this study was to examine whether beta cell apoptosis depends on the presence of galectin-3 and to delineate the underlying mechanism. Deficiency of galectin-3, either hereditary or induced through application of chemical inhibitors, β-lactose or TD139, supported survival and function of islet beta cells compromised by TNF-α + IFN-γ + IL-1β stimulus. Similarly, inhibition of galectin-3 by β-lactose or TD139 reduced cytokine-triggered apoptosis of beta cells, leading to conclusion that endogenous galectin-3 propagates beta apoptosis in the presence of an inflammatory milieu. Exploring apoptosis-related molecules expression in primary islet cells before and after treatment with cytokines we found that galectin-3 ablation affected the expression of major components of mitochondrial apoptotic pathway, such as BAX, caspase-9, Apaf, SMAC, caspase-3, and AIF. In contrast, anti-apoptotic molecules Bcl-2 and Bcl-XL were up-regulated in LGALS3(-/-) islet cells when compared to wild-type (WT) counterparts (C57BL/6), resulting in increased ratio of anti-apoptotic versus pro-apoptotic molecules. However, Fas-triggered apoptotic pathway as well as extracellular signal-regulated kinase 1/2 (ERK1/2) was not influenced by LGALS-3 deletion. All together, these results point to an important role of endogenous galectin-3 in beta cell apoptosis in the inflammatory milieu that occurs during diabetes pathogenesis and implicates impairment of mitochondrial apoptotic pathway as a key event in protection from beta cell apoptosis in the absence of galectin-3.

The Impact of a Family History of Type 2 Diabetes on Insulin Secretion and Insulin Sensitivity in Individuals With Varying Glucose Tolerance

IntroductionThis study was performed to investigate the impact of a family history of type 2 diabetes (T2DM) on insulin resistance and beta-cell dysfunction in populations with varying glucose tolerance. MethodsAmong the total of 142 participants, 73 subjects with no family history of T2DM (FH−) included 42 with normal glucose tolerance (NGT/FH−) and 31 with impaired glucose tolerance (IGT/FH−); and 69 first-degree relatives of patients with T2DM (FH+) included 36 with NGT (NGT/FH+) and 33 with IGT (IGT/FH+). Insulin resistance was evaluated by Insulin Sensitivity Index (ISI) based on the euglycemic hyperinsulinemic clamp. Islet beta-cell function was assessed by disposition index (DI) for the acute insulin response to glucose (AIRg) using intravenous glucose tolerance test. Metabolic data were compared between groups after adjustment for age, sex, body mass index and waist-to-hip ratio. ResultsThe NGT/FH+ group showed lower level of ISI (P=0.023) than the NGT/FH− group, whereas no difference was found in AIRg or DI between these 2 subgroups. In the FH− individuals, both ISI and DI of the IGT/FH− group decreased compared with the NGT/FH− group (both P<0.05). In the FH+ individuals, no difference was found in ISI between the IGT/FH+ and NGT/FH+ groups, whereas the IGT/FH+ group had a lower level of AIRg and DI than the NGT/FH+ group (both P<0.0001). ConclusionsThis study showed that the pathophysiological changes were different between individuals with and without a family history of T2DM during the glucose tolerance aggravation.

Evidence of safety and efficacy of two forms of neonatal cell transplants

On behalf of DOL and LCT A clinical trial in 14 adult type 1 diabetics intra‐peritoneal alginate microencapsulated neonatal porcine islets transplants commenced in NZ over 2 years ago and the last patient has just completed the required 12 months of follow up.To meet regulatory requirements for xenotransplantation, an elaborate program on xenovirology has been developed on one unique herd of pigs that was free from all conventional pathogens and was qualified as “null” pigs, i.e. they do not have a transmittable pig endogenous retrovirus (PERV). These pigs are the animal founders for the DPF donor herd. A comprehensive program for patients’ microbiology follow‐up was also developed. This program includes assays developed for monitoring potential infection with PERV and other potentially xenotic pathogens.All of the patients selected had unaware hypoglycaemia. The dose of islets used varied from 5000/kg (n = 4) 10 000/kg (n = 4) 15 000/kg (n = 4), and 20 000/kg (n = 2). The patients experience little if any side effects and there was no evidence of xenosis. Unaware hypoglycaemia was lessened in all groups but more so in the 10 000/kg group. Insulin dose was lowered and HbA1c improved in some but not all subjects. The patients with maximum insulin dose drop received 5000/kg dose. There was no clear cut relationship of return of hypoglycaemia awareness and glucagon or adrenergic responses to induced hypoglycaemia. There was direct laboratory evidence of islet beta cell function. Clinical follow up at 2 years of some patients indicates persistence of the benefits. A further clinical trial involving a different dosing schedule and some technical improvements is in progress.Choroid plexus (CP) ependymal epithelium secretes a wide variety of neurotrophic and neural support molecules. Micro‐encapsulated CP delivered intra‐cerebrally into rat models of stroke, Huntington's disease and Parkinson's disease show remarkable functional and histological recovery. In the quinolinic acid primate model of Huntington's and the MTPP model of Parkinson's functional recovery and relevant histological evidence of neuro‐regeneration were seen. The treatment was safe. A clinical trial of this form of treatment in humans with Parkinson's is being undertaken.Conclusions: Xenotransplantation of microencapsulated islets and other cell types into human and non‐human primates from a biocertified herd of pigs prepared under GMP conditions can be conducted with safety and efficacy. Further improvements in clinical outcomes are expected with technical advances.

The IL-1 Receptor Antagonist Anakinra Enhances Survival and Function of Human Islets during Culture: Implications in Clinical Islet Transplantation

Beta-cell replacement by human islet transplantation is a promising approach for treatment of type 1 diabetes (T1D). However, it is currently limited by low number of available pancreatic donors and loss of islets during isolation, pre-transplant culture, and following transplantation. Cytokines such as IL-1β have been implicated in beta-cell apoptosis in both T1D and T2D. Anakinra (Kineret) is an interleukin-1 (IL-1) receptor antagonist that competitively inhibits the binding of IL-1 proteins to their membrane receptor and thus blocks the biologic activity of naturally produced IL-1α and IL-1β. Anakinra is currently in clinical use for treatment of IL-1β-mediated inflammation in rheumatoid arthritis. Interestingly, recent studies have shown that Anakinra improves glycemic control in animal models of T2D. In this study, we tested whether treatment with Anakinra could enhance survival and function of isolated human islets during pre-transplant culture as a potential approach to improve quality and/or quantity of isolated islets for transplantation. Freshly isolated islets from five cadaveric human donors were cultured in CMRL (5 mM glucose, 37°C, 10% fetal bovine serum) without or with Anakinra (10 μg/ml) for 2 or 4 days. Beta-cell apoptosis, mass, function, and islet beta/alpha cell ratio were assessed in human islets before and following culture. As expected, culture of human islets resulted in an increase in the proportion of apoptotic (TUNEL-positive) islet beta cells in a time-dependent manner as assessed by immunostaining for insulin and TUNEL (d0: 2.2 ± 1.3%; d2: 4.3 ± 1.5; d4: 6.1 ± 2.0, P< 0.05). Treatment with Anakinra markedly reduced beta-cell apoptosis in cultured human islets (+An (d2): 2.4 ± 1.2; +An (d4): 3.1 ± 1.6, P< 0.05). Reduced beta-cell apoptosis was associated with a decrease in caspase-3 activation in beta cells in Anakinra-treated islets compared to non-treated cultured islets. This decrease in beta-cell apoptosis in Anakinra-treated human islets resulted in an increase in the islet beta/alpha cell ratio (-An (d4): 1.8 ± 0.2 vs +An (d4): 2.1 ± 0.2; P< 0.05) and beta-cell mass (beta-cell mass (% d0): -An (d4): 57 ± 3.4% vs +An (d4): 69 ± 3.8%, P< 0.05). Furthermore, Anakinra-treated islets had markedly enhanced beta-cell function compared to non-treated cultured islets manifested as increased insulin response to elevated glucose (-An (d2): 2.4 ± 0.2 vs +An (d2): 3.5 ± 0.4 fold; -An (d4): 2.1 ± 0.2 vs +An (d4): 3.3 ± 0.2 fold, P< 0.05) and increased islet insulin content. In summary, these studies suggest that blocking IL-1β receptor in isolated human islets reduces beta-cell death, preserves beta-cell mass, and enhances beta-cell function during pre-transplant islet culture. Treatment with IL-1 receptor antagonists may therefore provide a new approach to enhance islet beta-cell survival and function during pre-transplant culture thereby increase the success rate of clinical islet transplantation.

Sustained Beta-Cell Dysfunction but Normalized Islet Mass in Aged Thrombospondin-1 Deficient Mice

Pancreatic islet endothelial cells have in recent years been shown to support beta-cell mass and function by paracrine interactions. Recently, we identified an islets endothelial-specific glycoprotein, thrombospondin-1 (TSP-1), that showed to be of importance for islet angiogenesis and beta-cell function in young mice. The present study aimed to investigate long-term consequences for islet morphology and beta-cell function of TSP-1 deficiency. Islet and beta-cell mass were observed increased at 10–12 weeks of age in TSP-1 deficient mice, but were normalized before 16 weeks of age when compared to wild-type controls. Islet vascularity was normal in 10–12 and 16-week-old TSP-1 deficient animals, whereas islets of one-year-old animals lacking TSP-1 were hypervascular. Beta-cell dysfunction in TSP-1 deficient animals was present at similar magnitudes between 10–12 and 52 weeks of age, as evaluated by glucose tolerance tests. The insulin secretion capacity in vivo of islets in one-year-old TSP-1 deficient animals was only ∼15% of that in wild-type animals. Using a transplantation model, we reconstituted TSP-1 in adult TSP-deficient islets. In contrast to neonatal TSP-1 deficient islets that we previously reported to regain function after TSP-1 reconstitution, adult islets failed to recover. We conclude that TSP-1 deficiency in islets causes changing vascular and endocrine morphological alterations postnatally, but is coupled to a chronic beta-cell dysfunction. The beta-cell dysfunction induced by TSP-1 deficiency is irreversible if not substituted early in life.

Assessing Replication and Beta Cell Function in Adenovirally-transduced Isolated Rodent Islets

Glucose homeostasis is primarily controlled by the endocrine hormones insulin and glucagon, secreted from the pancreatic beta and alpha cells, respectively. Functional beta cell mass is determined by the anatomical beta cell mass as well as the ability of the beta cells to respond to a nutrient load. A loss of functional beta cell mass is central to both major forms of diabetes 1-3. Whereas the declining functional beta cell mass results from an autoimmune attack in type 1 diabetes, in type 2 diabetes, this decrement develops from both an inability of beta cells to secrete insulin appropriately and the destruction of beta cells from a cadre of mechanisms. Thus, efforts to restore functional beta cell mass are paramount to the better treatment of and potential cures for diabetes. Efforts are underway to identify molecular pathways that can be exploited to stimulate the replication and enhance the function of beta cells. Ideally, therapeutic targets would improve both beta cell growth and function. Perhaps more important though is to identify whether a strategy that stimulates beta cell growth comes at the cost of impairing beta cell function (such as with some oncogenes) and vice versa. By systematically suppressing or overexpressing the expression of target genes in isolated rat islets, one can identify potential therapeutic targets for increasing functional beta cell mass 4-6. Adenoviral vectors can be employed to efficiently overexpress or knockdown proteins in isolated rat islets 4,7-15. Here, we present a method to manipulate gene expression utilizing adenoviral transduction and assess islet replication and beta cell function in isolated rat islets (Figure 1). This method has been used previously to identify novel targets that modulate beta cell replication or function 5,6,8,9,16,17.

Assessing Replication and Beta Cell Function in Adenovirally-transduced Isolated Rodent Islets

Glucose homeostasis is primarily controlled by the endocrine hormones insulin and glucagon, secreted from the pancreatic beta and alpha cells, respectively. Functional beta cell mass is determined by the anatomical beta cell mass as well as the ability of the beta cells to respond to a nutrient load. A loss of functional beta cell mass is central to both major forms of diabetes (1-3). Whereas the declining functional beta cell mass results from an autoimmune attack in type 1 diabetes, in type 2 diabetes, this decrement develops from both an inability of beta cells to secrete insulin appropriately and the destruction of beta cells from a cadre of mechanisms. Thus, efforts to restore functional beta cell mass are paramount to the better treatment of and potential cures for diabetes. Efforts are underway to identify molecular pathways that can be exploited to stimulate the replication and enhance the function of beta cells. Ideally, therapeutic targets would improve both beta cell growth and function. Perhaps more important though is to identify whether a strategy that stimulates beta cell growth comes at the cost of impairing beta cell function (such as with some oncogenes) and vice versa. By systematically suppressing or overexpressing the expression of target genes in isolated rat islets, one can identify potential therapeutic targets for increasing functional beta cell mass (4-6). Adenoviral vectors can be employed to efficiently overexpress or knockdown proteins in isolated rat islets (4,7-15). Here, we present a method to manipulate gene expression utilizing adenoviral transduction and assess islet replication and beta cell function in isolated rat islets (Figure 1). This method has been used previously to identify novel targets that modulate beta cell replication or function (5,6,8,9,16,17).

Control of beta cell function and proliferation in mice stimulated by small-molecule glucokinase activator under various conditions

We investigated changes in the expression of genes involved in beta cell function and proliferation in mouse islets stimulated with glucokinase activator (GKA) in order to elucidate the mechanisms by which GKA stimulates beta cell function and proliferation. Islets isolated from mice were used to investigate changes in the expression of genes related to beta cell function and proliferation stimulated by GKA. In addition, Irs2 knockout (Irs2 (-/-)) mice on a high-fat diet or a high-fat diet containing GKA were used to investigate the effects of GKA on beta cell proliferation in vivo. In wild-type mice, Irs2 and Pdx1 expression was increased by GKA. In Irs2 (-/-) mice, GKA administration increased the glucose-stimulated secretion of insulin and Pdx1 expression, but not beta cell proliferation. It was particularly noteworthy that oxidative stress inhibited the upregulation of the Irs2 and Pdx1 genes induced by GKA. Moreover, whereas neither GKA alone nor exendin-4 alone upregulated the expression of Irs2 and Pdx1 in the islets of db/db mice, prior administration of exendin-4 to the mice caused GKA to increase the expression of these genes. GKA-stimulated IRS2 production affected beta cell proliferation but not beta cell function. Oxidative stress diminished the effects of GKA on the changes in expression of genes involved in beta cell function and proliferation. A combination of GKA and an incretin-related agent might therefore be effective in therapy.

Long term effects of the implantation of autologous bone marrow mononuclear cells for type 2 diabetes mellitus

Previous studies have shown that several types of stem cells can differentiate into insulin-secreting islet beta-cells and that these cells can reduce blood glucose in some trials, but there has been no report of a long-term follow-up. We assessed the long-term effects of the use of autologous bone marrow mononuclear cells in the treatment of type 2 diabetes mellitus (T2DM). Based on the willingness to receive implantation of bone marrow mononuclear cells, One hundred and eighteen patients with T2DM were divided into two groups; the patients in group I were treated with autologous bone marrow mononuclear cells and patients in group II were treated with insulin intensification therapy. Mononuclear cells from bone marrow were injected back into the patient's pancreas via a catheter. Patients were followed-up after the operation at monthly intervals for the first 3 months and thereafter every 3 months for the next 33 months, the occurrence of any side effects and the results of laboratory examinations were evaluated. There were no reported acute or chronic side effects in group I and both the HbA1c and C-peptide in group I patients were significantly better than either pretherapy values or group II patients during the follow-up period. These data suggested that the implantation of autologous bone marrow mononuclear cells for the treatment of T2DM is safe and effective. This therapy can partially restore the function of islet beta-cells and maintain blood glucose homeostasis in a longer time.

Islet-enriched gene expression and glucose-induced insulin secretion in human and mouse islets

Aims/hypothesisOur understanding of the transcription factors that control the development and function of rodent islet beta cells is advancing rapidly, yet less is known of the role they play in similar processes in human islets.MethodsTo characterise the abundance and regulation of key proteins involved in glucose-regulated insulin secretion in human islets, we examined the expression of MAFA, MAFB, GLUT2 (also known as SLC2A2), βGK (also known as GCK) and PDX1 in isolated, highly purified human islets with an intact insulin secretory pattern. We also assessed these features in islets from two different mouse strains (C57BL/6J and FVB).ResultsCompared with mouse islets, human islets secreted more insulin at baseline glucose (5.6 mmol/l), but less upon stimulation with high glucose (16.7 mmol/l) or high glucose plus 3-isobutyl-1-methyl-xanthine. Human islets had relatively more MAFB than PDX1 mRNA, while mouse islets had relatively more Pdx1 than Mafb mRNA. However, v-maf musculoaponeurotic fibrosarcoma oncogene homologue (MAF) B protein was found in human islet alpha and beta cells. This is unusual as this regulator is only produced in islet alpha cells in adult mice. The expression of insulin, MAFA, βGK and PDX1 was not glucose-regulated in human islets with an intact insulin secretory pattern.Conclusions/interpretationOur results suggest that human islets have a distinctive distribution and function of key regulators of the glucose-stimulated insulin secretion pathway, emphasising the urgent need to understand the processes that regulate human islet beta cell function.Electronic supplementary materialThe online version of this article (doi:10.1007/s00125-011-2369-0) contains peer-reviewed but unedited supplementary material, which is available to authorised users.

PS18 - 90. Role of tPA in islet amyloid formation and beta cell (dys)function in type 2 diabetes mellitus (DM2)

The protease tPA (tissue type plasminogen activator) is activated by fibrin and degrades blood cloths and extracellular matrix (ECM). In addition to fibrin, amyloid fibrils of the Alzheimer Aβ protein and human Islet amyloid polypeptide (hIAPP, fibril-forming protein of amyloid in pancreatic islets of DM2 patients) can activate tPA in vitro.

Effect of aging on islet beta-cell function and its mechanisms in Wistar rats

Type 2 diabetes mellitus is characterized by islet β-cell dysfunction and its incidence increases with age. However, the mechanisms underlying the effect of aging on islet β-cell function are not fully understood. We characterized β-cell function in 4-month-old (young), 14-month-old (adult), and 24-month-old (old) male Wistar rats, and found that islet β-cell function decreased gradually with age. Old rats displayed oral glucose intolerance and exhibited a decrease in glucose-stimulated insulin release (GSIR) and palmitic acid-stimulated insulin release (PSIR). Furthermore, total superoxide dismutase (T-SOD), CuZn superoxide dismutase (CuZn-SOD), and glutathione peroxidase (GSH-Px) activity decreased, whereas serum malondialdehyde (MDA) levels increased in the older rats. Moreover, we detected a significant reduction in β-cell proliferation and an increase in the frequency of apoptotic β-cells in the islets of rats in the old group. Finally, Anxa1 expression in the islets of old rats was significantly upregulated. These data provide new insights into the development of age-related β-cell dysfunction in rats.

Outcome of intensive integrated intervention in participants with impaired glucose regulation in China

This study investigated the outcomes and identified influencing factors of intensive integrated intervention over 2 years in Chinese patients with impaired glucose regulation (IGR). Adults in Beijing, China, were screened for IGR using the 75 g oral glucose tolerance test. Participants with IGR received lifestyle and health education; those who still had IGR after 1 year were randomly assigned to either a routine care group or to an intensive integrated intervention group. Of 2344 adults screened, 463 had IGR. Of these, 210 adults had IGR after 1 year and were therefore recruited and randomized to an intensive integrated intervention group (n=106) or a control group (n=104). The percentage of patients who reached the set targets of plasma glucose, blood pressure, body mass index, or triglycerides was significantly higher in the intensive integrated intervention group. None of the patients within the intensive integrated intervention group progressed to diabetes, whereas eight (9.3%) cases of the control group developed type 2 diabetes mellitus (T2DM). Logistic regression analysis showed that both an increase in waist circumference and systolic blood pressure (SBP) were positively correlated with the development of T2DM, whereas improvement in islet beta cell function was negatively correlated with the development of T2DM. Intensive integrated intervention may significantly decrease the conversion rate of IGR to T2DM, and increase the conversion ratio to normal glucose tolerance. The increase of waist circumference or SBP and the deterioration of islet beta cell function may be important risk factors for progression from prediabetes to diabetes.

Obesity in the pathogenesis of type 2 diabetes

Obesity, and especially visceral adiposity, escalates the development of insulin resistance and type 2 diabetes. Excess adipose tissue contributes to a chronic increase in circulating fatty acids reducing the usage of glucose as a source of cellular energy. Excess fatty acids also result in increased deposition of fat in muscle and liver, and increased metabolites such as diacylglycerol and ceramide which activate isoforms of protein kinase C that impede cellular insulin signalling. Chronically raised lipid levels also impair islet beta cell function, acting in conjuction with insulin resistance to aggravate hyperglycaemia. The detrimental effects of several adipokines such as TNFα, IL6 and RBP4, which are produced in excess by an increased adipose mass, and reduced production of adiponectin are further mechanisms through which obesity potentiates the development of type 2 diabetes.

Glucose and inflammation control islet vascular density and beta-cell function in NOD mice: control of islet vasculature and vascular endothelial growth factor by glucose.

OBJECTIVEβ-Cell and islet endothelial cell destruction occurs during the progression of type 1 diabetes, but, paradoxically, β-cell proliferation is increased during this period. Altered glucose tolerance may affect β-cell mass and its association with endothelial cells. Our objective was to study the effects of glucose and inflammation on islet vascularity and on β function, mass, and insulin in immunologically tolerant anti-CD3 monoclonal antibody (mAb)-treated and prediabetic NOD mice.RESEARCH DESIGN AND METHODSThe effects of phloridzin or glucose injections on β-cells and endothelial cells were tested in prediabetic and previously diabetic NOD mice treated with anti-CD3 mAbs. Glucose tolerance, immunofluorescence staining, and examination of islet cultures ex vivo were evaluated.RESULTSIslet endothelial cell density decreased in NOD mice and failed to recover after anti-CD3 mAb treatment despite baseline euglycemia. Glucose treatment of anti-CD3 mAb–treated mice showed increased islet vascular density and increased insulin content, which was associated with improved glucose tolerance. The increase in the vascular area was dependent on islet inflammation. Increased islet endothelial cell density was associated with increased production of vascular endothelial growth factor (VEGF) by islets from NOD mice. This response was recapitulated ex vivo by the transfer of supernatants from NOD islets cultured in high-glucose levels.CONCLUSIONSOur results demonstrate a novel role for glucose and inflammation in the control of islet vasculature and insulin content of β-cells in prediabetic and anti-CD3–treated NOD mice. VEGF production by the islets is affected by glucose levels and is imparted by soluble factors released by inflamed islets.

Epistasis of Transcriptomes Reveals Synergism between Transcriptional Activators Hnf1α and Hnf4α

The transcription of individual genes is determined by combinatorial interactions between DNA–binding transcription factors. The current challenge is to understand how such combinatorial interactions regulate broad genetic programs that underlie cellular functions and disease. The transcription factors Hnf1α and Hnf4α control pancreatic islet β-cell function and growth, and mutations in their genes cause closely related forms of diabetes. We have now exploited genetic epistasis to examine how Hnf1α and Hnf4α functionally interact in pancreatic islets. Expression profiling in islets from either Hnf1a+/− or pancreas-specific Hnf4a mutant mice showed that the two transcription factors regulate a strikingly similar set of genes. We integrated expression and genomic binding studies and show that the shared transcriptional phenotype of these two mutant models is linked to common direct targets, rather than to known effects of Hnf1α on Hnf4a gene transcription. Epistasis analysis with transcriptomes of single- and double-mutant islets revealed that Hnf1α and Hnf4α regulate common targets synergistically. Hnf1α binding in Hnf4a-deficient islets was decreased in selected targets, but remained unaltered in others, thus suggesting that the mechanisms for synergistic regulation are gene-specific. These findings provide an in vivo strategy to study combinatorial gene regulation and reveal how Hnf1α and Hnf4α control a common islet-cell regulatory program that is defective in human monogenic diabetes.

Cholesterol efflux via ATP-binding cassette transporter A1 (ABCA1) and cholesterol uptake via the LDL receptor influences cholesterol-induced impairment of beta cell function in mice

Cellular cholesterol accumulation is an emerging mechanism for beta cell dysfunction in type 2 diabetes. Absence of the cholesterol transporter ATP-binding cassette transporter A1 (ABCA1) results in increased islet cholesterol and impaired insulin secretion, indicating that impaired cholesterol efflux leads to beta cell dysfunction. In this study, we aimed to determine the role of the LDL receptor (LDLr) in islet cholesterol uptake and to assess the contributions of cholesterol uptake compared with efflux to islet cholesterol levels. Islet cholesterol and beta cell function were assessed in mice lacking LDLr (Ldlr(-/-)), or apolipoprotein E (Apoe(-/-)), as well as in mice with beta-cell-specific deficiency of Abca1 crossed to Ldlr(-/-) mice. Hypercholesterolaemia resulted in increased islet cholesterol levels and decreased beta cell function in Apoe(-/-) mice but not in Ldlr(-/-) mice, suggesting that the LDL receptor is required for cholesterol uptake leading to cholesterol-induced beta cell dysfunction. Interestingly, when wild-type islets with functional LDL receptors were transplanted into diabetic, hypercholesterolaemic mice, islet graft function was normal compared with Ldlr(-/-) islets, suggesting that compensatory mechanisms can maintain islet cholesterol homeostasis in a hypercholesterolaemic environment. Indeed, transplanted wild-type islets had increased Abca1 expression. However, lack of the Ldlr did not protect Abca1(-/-) mice from islet cholesterol accumulation, suggesting that cholesterol efflux is the critical regulator of cholesterol levels in islets. Our data indicate that islet cholesterol levels and beta cell function are strongly influenced by LDLr-mediated uptake of cholesterol into beta cells. Cholesterol efflux mediated by ABCA1, however, can compensate in hypercholesterolaemia to regulate islet cholesterol levels in vivo.

α Cell–Specific Men1 Ablation Triggers the Transdifferentiation of Glucagon-Expressing Cells and Insulinoma Development

The tumor suppressor menin is recognized as a key regulator of pancreatic islet development, proliferation, and beta-cell function, whereas its role in alpha cells remains poorly understood. The purpose of the current study was to address this issue in relation to islet tumor histogenesis. We generated alpha cell-specific Men1 mutant mice with Cre/loxP technology and carried out analyses of pancreatic lesions developed in the mutant mice during aging. We showed that, despite the alpha-cell specificity of the GluCre transgene, both glucagonomas and a large amount of insulinomas developed in mutant mice older than 6 months, accompanied by mixed islet tumors. Interestingly, the cells sharing characteristics of both alpha and beta cells were identified shortly after the appearance of menin-deficient alpha cells but well before the tumor onset. Using a genetic cell lineage tracing analysis, we demonstrated that insulinoma cells were directly derived from transdifferentiating glucagon-expressing cells. Furthermore, our data indicated that the expression of Pdx1, MafA, Pax4, and Ngn3 did not seem to be required for the initiation of this transdifferentiation. Our work shows cell transdifferentiation as a novel mechanism involved in islet tumor development and provides evidence showing that menin regulates the plasticity of differentiated pancreatic alpha cells in vivo, shedding new light on the mechanisms of islet tumorigenesis.

Small G proteins in islet beta-cell function.

Glucose-stimulated insulin secretion from the islet beta-cell involves a sequence of metabolic events and an interplay between a wide range of signaling pathways leading to the generation of second messengers (e.g., cyclic nucleotides, adenine and guanine nucleotides, soluble lipid messengers) and mobilization of calcium ions. Consequent to the generation of necessary signals, the insulin-laden secretory granules are transported from distal sites to the plasma membrane for fusion and release of their cargo into the circulation. The secretory granule transport underlies precise changes in cytoskeletal architecture involving a well-coordinated cross-talk between various signaling proteins, including small molecular mass GTP-binding proteins (G proteins) and their respective effector proteins. The purpose of this article is to provide an overview of current understanding of the identity of small G proteins (e.g., Cdc42, Rac1, and ARF-6) and their corresponding regulatory factors (e.g., GDP/GTP-exchange factors, GDP-dissociation inhibitors) in the pancreatic beta-cell. Plausible mechanisms underlying regulation of these signaling proteins by insulin secretagogues are also discussed. In addition to their positive modulatory roles, certain small G proteins also contribute to the metabolic dysfunction and demise of the islet beta-cell seen in in vitro and in vivo models of impaired insulin secretion and diabetes. Emerging evidence also suggests significant insulin secretory abnormalities in small G protein knockout animals, further emphasizing vital roles for these proteins in normal health and function of the islet beta-cell. Potential significance of these experimental observations from multiple laboratories and possible avenues for future research in this area of islet research are highlighted.

Cooperative Transcriptional Regulation of the Essential Pancreatic Islet Gene NeuroD1 (Beta2) by Nkx2.2 and Neurogenin 3

Nkx2.2 and NeuroD1 are two critical regulators of pancreatic beta cell development. Nkx2.2 is a homeodomain transcription factor that is essential for islet cell type specification and mature beta cell function. NeuroD1 is a basic helix-loop-helix transcription factor that is critical for islet beta cell maturation and maintenance. Although both proteins influence beta cell development directly downstream of the endocrine progenitor factor, neurogenin3 (Ngn3), a connection between the two proteins in the regulation of beta cell fate and function has yet to be established. In this study, we demonstrate that Nkx2.2 transcriptional activity is required to facilitate the activation of NeuroD1 by Ngn3. Furthermore, Nkx2.2 is necessary to maintain high levels of NeuroD1 expression in developing mouse and zebrafish islets and in mature beta cells. Interestingly, Nkx2.2 regulates NeuroD1 through two independent promoter elements, one that is bound and activated directly by Nkx2.2 and one that appears to be regulated by Nkx2.2 through an indirect mechanism. Together, these findings suggest that Nkx2.2 coordinately activates NeuroD1 with Ngn3 within the endocrine progenitor cell and also plays a role in the maintenance of NeuroD1 expression to regulate beta cell function in the mature islet. Collectively, these findings further define the conserved regulatory networks involved in islet beta cell formation and function.