Search

Islet allotransplantation offers a promising cell therapy for type 1 diabetes, but challenges such as limited donor availability and immunosuppression persist. Microencapsulation of islets in polymer-coated alginate microcapsules is a favored strategy for immune protection and maintaining islet viability. This study introduces Poly [2-(methacryloyloxy)ethyl]trimethylammonium chloride (PMETAC) as an innovative coating material for microcapsules. PMETAC enhances biocompatibility and durability, marking a significant advancement in islet encapsulation. Our approach combines alginate with PMETAC to create Langerhans islet microcapsules, simplifying material composition and preparation and ultimately lowering costs and increasing clinical applicability. Our comprehensive evaluation of the stability (including osmotic stability, thermal stability, and culture condition stability) and cytotoxicity of a novel microencapsulation system based on alginate-PMETAC-alginate offers insights into its potential application in islet immunoisolation strategies. Microcapsules with PMETAC content ranging from 0.01 to 1% are explored in the current work. The results indicate that the coatings made with 0.4% PMETAC show the most promising outcomes, remaining stable in the mentioned tests and exhibiting the required permeability. It was shown that the islets encapsulated in this manner retain viability and functional activity. Thus, alginate microcapsules coated with 0.4% PMETAC are suitable for further animal trials. While our findings are promising, further studies, including animal testing, will be necessary to evaluate the clinical applicability of our encapsulation method. [ABSTRACT FROM AUTHOR]

Insulin resistance contributes to the development of various diseases, including type 2 diabetes and gestational diabetes. Even though gestational diabetes is specific to pregnancy, it can result in long-term glucose intolerance and type 2 diabetes after delivery. Given the substantial health and economic burdens associated with diabetes, it is imperative to better understand the mechanisms leading to insulin resistance and type 2 diabetes so that treatments targeted at reversing symptoms can be developed. Considering that the endocrine cells of the pancreas (islets of Langerhans) largely contribute to the pathogenesis of diabetes (beta-cell insufficiency and dysfunction), the elucidation of the various mechanisms of endocrine cell plasticity is important to understand. By better defining these mechanisms, targeted therapeutics can be developed to reverse symptoms of beta-cell deficiency and insulin resistance in diabetes. Animal models play an important role in better understanding these mechanisms, as techniques for in vivo imaging of endocrine cells in the pancreas are limited. Therefore, this review article will discuss the available rodent models of gestational and type 2 diabetes that are characterized by endocrine cell impairments in the pancreas, discuss the models with a comparison to human diabetes, and explore the potential mechanisms of endocrine cell plasticity that contribute to these phenotypes, as these mechanisms could ultimately be used to reverse blood glucose dysregulation in diabetes. [ABSTRACT FROM AUTHOR]

Background: Lovebird (Agapornis personatus) is a monotypic species of bird of the lovebird genus in the parrot family Psittaculidae and order Psittaciformes. Objectives: The present study was designed to investigate the histology and immunohistochemistry of the pancreas in the lovebird. Methods: Totally, three adult birds were used. The pancreas was assessed using histological and immunofluorescent staining to detect insulin, glucagon, somatostatin, pancreatic polypeptide (PP) and neuropeptide Y (NY). Results: The exocrine pancreas was composed of pyramidal acinar cells with zymogen granules at the apical cytoplasm. The endocrine pancreas was identified as large alpha, small beta and mixed islets of Langerhans. No intercalated duct was observed. Alpha cells with a density of 28.55% were the most numerous cell type, which were populated throughout the large islets, especially at the periphery. The beta cells with a density of 15.78% were accumulated mostly at the periphery of islets. The delta cells exhibited 17.81% intensity. Despite their lower density, the distribution of delta cells was like that of A cells throughout the islets. PP and NY cells were distinguished with densities of 14.69% and 20.63%, respectively. Conclusions: Although the arrangement of acinar cells, ductal systems and endocrine islets reflects patterns observed in various avian species, the absence of intercalated duct, the presence of three types of Langerhans islets as alpha, beta and mixed islets and the high expression of NY in the islets were some unique features observed in the current study. These findings contribute to the broader understanding of avian pancreas histology. [ABSTRACT FROM AUTHOR]

Background: Fish oil, which is regarded as the primary source of omega-3 fatty acids, has been long studied for its potential as an antidiabetic therapy. However, its protective ability against insulin resistance and pancreatic islet alteration remains unclear and controversial. Aim: To investigate the beneficial effects of fish oil consumption on the progression of insulin resistance and pancreatic islet dysfunction in a rat model of diabetes. Methods: Diabetic rats model (n = 30) were divided into five groups and received; 1) NS injection + NS oral (normal control); 2) NS injection + 3 g/kg fish oil (fish oil control); 3) streptozotocin (STZ) injection + NS oral [diabetes control (DC)]; 4) STZ injection + 1 g/kg fish oil (DFO1); and 5) STZ injection + 3 g/kg fish oil (DFO3). Fasting blood insulin was analyzed by commercial rat insulin enzyme-linked immunosorbent assay; meanwhile, the determination of insulin sensitivity was calculated by homeostatic model assessment of insulin resistance (HOMA-IR) and homeostatic model assessment of beta-cell function. A histological study was conducted on pancreas tissue using H and E staining. Results: Fish oil supplementation reduced hyperglycemia and ameliorated HOMA-IR in STZ-induced animal models indicating that fish oil supplementation improved insulin sensitivity. Furthermore, animals treated with fish oil at a dose of 3 g/kg (DFO3) showed an enhancement in pancreatic islets, which was displayed by less abnormal structures than DC animals. This could imply that the administration of fish oil, especially rich in bioactive omega-3 fatty acids effectively inhibits insulin resistance and restore islet of Langerhans alteration in rats injected with STZ. Conclusion: Thus, the current study suggested that fish oil supplementation could support the treatment of diabetes but should not be considered as an alternative therapy. [ABSTRACT FROM AUTHOR]

Islets of Langerhans release peptide hormones in controlled amounts and patterns to ensure proper maintenance of blood glucose levels. The overall release of the hormones is shaped by external factors and by autocrine and paracrine interactions occurring within the islets. To better understand what controls the secretion of islet-secreted peptides, and how these processes go awry in diabetes, methods to monitor the release of multiple hormones simultaneously are needed. While antibody-based assays are typically used, they are most often applied to quantification of a single hormone. Mass spectrometry (MS), on the other hand, is well suited for quantifying multiple hormones simultaneously but typically requires time-consuming separation steps with biological samples. In this report, response surface methodology was used to identify a set of optimal solid-phase extraction (SPE) conditions for the islet-secreted peptides, insulin, C-peptide, glucagon, and somatostatin. The optimized SPE method was used with multiple reaction monitoring and isotopically labeled standards to quantify secretion levels. Calibrations were linear from 0.5 to 50 nM with < 15% RSD peak area ratios. A microfluidic system was used to perfuse 30 human islets with different glucose conditions, and fractions were collected every 2 min for SPE-MS analysis. Results showed the release dynamics of the individual peptides, as well as patterns, such as positively and negatively correlated release and oscillations. This rapid SPE-MS method is expected to be useful for examining other peptide and small-molecule secretions from islets and could be applied to a number of other biological systems for investigating cellular communication. [ABSTRACT FROM AUTHOR]

The secretion of insulin from β‐cells in the islet of Langerhans is governed by a series of metabolic and electrical events, which can fail during the progression of type 2 diabetes (T2D). β‐cells are electrically coupled via connexin‐36 (Cx36) gap junction channels, which coordinates the pulsatile dynamics of [Ca2+] and insulin release across the islet. Factors such as pro‐inflammatory cytokines and free fatty acids disrupt gap junction coupling under in vitro conditions. Here we test whether gap junction coupling and coordinated [Ca2+] dynamics are disrupted in T2D, and whether recovery of gap junction coupling can recover islet function. We examine islets from donors with T2D, from db/db mice, and islets treated with pro‐inflammatory cytokines (TNF‐α, IL‐1β, IFN‐ɣ) or free fatty acids (palmitate). We modulate gap junction coupling using Cx36 over‐expression or pharmacological activation via modafinil. We also develop a peptide mimetic (S293) of the c‐terminal regulatory site of Cx36 designed to compete against its phosphorylation. Cx36 gap junction permeability and [Ca2+] dynamics were disrupted in islets from both human donors with T2D and db/db mice, and in islets treated with pro‐inflammatory cytokines or palmitate. Cx36 over‐expression, modafinil treatment and S293 peptide all enhanced Cx36 gap junction coupling and protected against declines in coordinated [Ca2+] dynamics. Cx36 over‐expression and S293 peptide also reduced apoptosis induced by pro‐inflammatory cytokines. Critically, S293 peptide rescued gap junction coupling and [Ca2+] dynamics in islets from both db/db mice and a sub‐set of T2D donors. Thus, recovering or enhancing Cx36 gap junction coupling can improve islet function in diabetes. Key points: Connexin‐36 (Cx36) gap junction permeability and associated coordination of [Ca2+] dynamics is diminished in human type 2 diabetes (T2D) and mouse models of T2D.Enhancing Cx36 gap junction permeability protects against disruptions to the coordination of [Ca2+] dynamics.A novel peptide mimetic of the Cx36 c‐terminal regulatory region protects against declines in Cx36 gap junction permeability.Pharmacological elevation in Cx36 or Cx36 peptide mimetic recovers [Ca2+] dynamics and glucose‐stimulated insulin secretion in human T2D and mouse models of T2D. [ABSTRACT FROM AUTHOR]

Ion channels play an important role for regulation of the exocrine and the endocrine pancreas. This review focuses on the Ca2+-regulated K+ channel KCa3.1, encoded by the KCNN4 gene, which is present in both parts of the pancreas. In the islets of Langerhans, KCa3.1 channels are involved in the regulation of membrane potential oscillations characterizing nutrient-stimulated islet activity. Channel upregulation is induced by gluco- or lipotoxic conditions and might contribute to micro-inflammation and impaired insulin release in type 2 diabetes mellitus as well as to diabetes-associated renal and vascular complications. In the exocrine pancreas KCa3.1 channels are expressed in acinar and ductal cells. They are thought to play a role for anion secretion during digestion but their physiological role has not been fully elucidated yet. Pancreatic carcinoma, especially pancreatic ductal adenocarcinoma (PDAC), is associated with drastic overexpression of KCa3.1. For pharmacological targeting of KCa3.1 channels, we are discussing the possible benefits KCa3.1 channel inhibitors might provide in the context of diabetes mellitus and pancreatic cancer, respectively. We are also giving a perspective for the use of a fluorescently labeled derivative of the KCa3.1 blocker senicapoc as a tool to monitor channel distribution in pancreatic tissue. In summary, modulating KCa3.1 channel activity is a useful strategy for exo-and endocrine pancreatic disease but further studies are needed to evaluate its clinical suitability. [ABSTRACT FROM AUTHOR]

Objectives: A lifelong gluten‐free (GF) diet ameliorates autoimmune diabetes in non‐obese diabetic (NOD) mice and most likely in humans. Besides diabetes, NOD mice develop focal sialadenitis, as seen in Sjögren's syndrome (SS). In humans, type 1 diabetes (T1D) is also linked to SS. Here, we investigated whether a lifelong GF diet influences the immune cell infiltration in the salivary glands and pancreatic islets in NOD mice. Methods: NOD mice were fed a lifelong (i.e. 13 weeks) GF or gluten‐containing standard (STD) diet. Insulitis and sialadenitis were scored on H&E‐stained paraffin‐embedded sections of pancreas and submandibular glands. Immune cell specificity and distribution were investigated immunohistochemically. Results: There were fewer CD68+ and CD4+ cells in submandibular gland areas with focal sialadenitis as well as reduced insulitis and fewer VEGFR2+ cells in pancreatic islets in mice on GF versus STD diet. The degree of sialadenitis was not significantly lower in GF mice, but sialadenitis and insulitis correlated strongly. Lung weight was lower in GF mice. Conclusion: In NOD mice, a lifelong GF diet reduces infiltration of monocytes/macrophages and T cells in salivary glands and inflammation in pancreatic islets, possibly by reducing VEGFR2, indicating that the linked autoimmune diseases, T1D and SS, may be alleviated by a GF diet. [ABSTRACT FROM AUTHOR]

The success of pancreas islet isolation largely depends on donor characteristics, including extracellular matrix composition of which collagen is the main element. We hypothesized that isolation yields are proportional to collagen digestion percentage, and aimed to determine a threshold that predicts isolation success. The amount of pancreas collagen (I‐V) was determined using colorimetry prior to and after the digestion process in 52 human islet isolations. Collagen I‐V and VI were also assessed histologically. We identified a collagen digestion threshold of ≥ 60% as an independent factor beyond which an islet preparation has a ninefold increased odds of yielding ≥ 250 000 islet equivalents (IEQ) (P = 0.009) and a sixfold increased odds of being transplanted (P = 0.015). Preparations with ≥ 60% collagen digestion (n = 35) yielded 283 017 ± 164 214 IEQ versus 180 142 ± 85 397 in the < 60% collagen digestion group (n = 17) (P = 0.016); respectively 62.9% versus 29.4% of those were transplanted (P = 0.024). Common donor characteristics, initial collagen content, enzyme blend, and digestion times were not associated with collagen digestion percentage variations. Donor age positively correlated with the amount of collagen VI (P = 0.013). There was no difference in islet graft survival between high and low digestion groups. We determined that a 60% pancreas collagen digestion is the threshold beyond which an islet isolation is likely to be successful and transplanted. [ABSTRACT FROM AUTHOR]

A recent study aims at investigating the immunohistochemical and histochemical analysis of adult goat's pancreatic tissue. Histological parts assembled from their pancreas specimens are stained with special and general stains, otherwise recorded and examined with its image software using the Dino-eye piece camera. The endocrine part mainly resides in the Langerhans islet, which is well connected to extra-blood vessels & exocrine ducts. Every islet was constructed microscopically with several cells (alpha and beta) that varied in size, shape, and color. The pancreas is one of the unique endocrine glands, it has endocrine and exocrine secretions. The Islet of Langerhans constitute the endocrine component of the pancreas, its secretory has definite employment for life. A boundary was created by the collagen fibers between the exocrine and endocrine parts. There were oval, circular, or irregular islets based on their scale on its long axis, the form, and were graded into small, medium, and huge. The specific islet cells have specific characteristics and secrete the specific hormone. The Alpha and beta, were the two main cell types and few delta cells were found in Langerhans islets, these different islet cells synthesize and secrete different hormones which need for the biological system to achieve satisfactory metabolic control over blood sugar. The region distribution, the proportional (relative) frequency, and the appearance of glucagon (A-cells)- & glucose-dependent insulinotropic peptide(B-cells)-secreted cells in the endocrine and exocrine pancreas of goat were studied using the immunohistochemical method. Glucagon immunopositive A-cells were principally present in the center and mantle regions of Langerhans islets, whereas glucose-dependent insulinotropic peptide immunopositive B-cells were found in the periphery, center, and mantle of pancreatic islets. Moreover, some A- and B-cells were present as only single cells or clusters of (2 - 3) immunopositive cells in the exocrine parenchyma and the pancreatic duct epithelial cells. The presence, region distribution, and proportional or (relative) frequency of A-&B-cells in goat's pancreas have been analyzed firstly. The immunopositivity and distribution of endocrine cells in the goat pancreas were specified to be partially variable from that of other mammalian species like sheep. [ABSTRACT FROM AUTHOR]

Fifteen Kestrel were used in this study regardless to their sex. The results study showed that the pancreas of adult Kestrel (Falco tinnunculus) lies under the right side of the peritoneal cavity, and situated between the descending part and ascending part of the duodenum and has pale pinkish to white pinkish colored and consisted of 3 lobes dorsal (lobus pancreatic dorsalis), middle (middle pancreatic lobus) and ventral (lobus pancreatis ventralis) with well-developed interlobar connections made it difficult to distinguish between the 3 lobes. Histologically the pancreas of adult Kestrel (Falco tinnunculus) was covered by very thin layer of connective tissue with mesothelial cells, Connective tissue septa extended from capsule into parenchyma of pancreas dividing it in to many lobules. These septa were very thin and some of them carried blood vessels distributed all over the pancreas. The exocrine portion was constituted from serous acini with pyramidal shaped cells which had dark rounded nucleus in the middle part of the cell. Acidophilic granules found in the apices of cells. Centro acinar cells found as one or two nuclei in the center of the pancreatic acini. The endocrine portion (islets of Langerhans) was formed from clusters of endocrine cells in shape of oval or rounded pale structures with different sizes. The small one has mean diameter of 40.02±0.9 μm while the large one has diameter of 126.3±3.8 μm. Few endocrine cells were seen distributed as single cells among the pancreatic acini. [ABSTRACT FROM AUTHOR]

Key points: The pancreatic islets of Langerhans maintain glucose homeostasis through insulin secretion, where insulin secretion dynamics are regulated by intracellular Ca2+ signalling and electrical coupling of the insulin producing β‐cells in the islet.We have previously shown that cytokines decrease β‐cell coupling and that compounds which increase cAMP can increase coupling.In both mouse and human islets exendin‐4, which increases cAMP, protected against cytokine‐induced decreases in coupling and in mouse islets preserved glucose‐stimulated calcium signalling by increasing connexin36 gap junction levels on the plasma membrane.Our data indicate that protein kinase A regulates β‐cell coupling through a fast mechanism, such as channel gating or membrane organization, while Epac2 regulates slower mechanisms of regulation, such as gap junction turnover.Increases in β‐cell coupling with exendin‐4 may protect against cytokine‐mediated β‐cell death as well as preserve insulin secretion dynamics during the development of diabetes. The pancreatic islets of Langerhans maintain glucose homeostasis. Insulin secretion from islet β‐cells is driven by glucose metabolism, depolarization of the cell membrane and an influx of calcium, which initiates the release of insulin. Gap junctions composed of connexin36 (Cx36) electrically couple β‐cells, regulating calcium signalling and insulin secretion dynamics. Cx36 coupling is decreased in pre‐diabetic mice, suggesting a role for altered coupling in diabetes. Our previous work has shown that pro‐inflammatory cytokines decrease Cx36 coupling and that compounds which increase cAMP can increase Cx36 coupling. The goal of this study was to determine if exendin‐4, which increases cAMP, can protect against cytokine‐induced decreases in Cx36 coupling and altered islet function. In both mouse and human islets, exendin‐4 protected against cytokine‐induced decreases in coupling and preserved glucose‐stimulated calcium signalling. Exendin‐4 also protected against protein kinase Cδ‐mediated decreases in Cx36 coupling. Exendin‐4 preserved coupling in mouse islets by preserving Cx36 levels on the plasma membrane. Exendin‐4 regulated Cx36 coupling via both protein kinase A (PKA)‐ and Epac2‐mediated mechanisms in cytokine‐treated islets. In mouse islets, modulating Epac2 had a greater impact in mediating Cx36 coupling, while in human islets modulating PKA had a greater impact on Cx36 coupling. Our data indicate that PKA regulates Cx36 coupling through a fast mechanism, such as channel gating, while Epac2 regulates slower mechanisms of regulation, such as Cx36 turnover in the membrane. Increases in Cx36 coupling with exendin‐4 may protect against cytokine‐mediated β‐cell dysfunction to insulin secretion dynamics during the development of diabetes. Key points: The pancreatic islets of Langerhans maintain glucose homeostasis through insulin secretion, where insulin secretion dynamics are regulated by intracellular Ca2+ signalling and electrical coupling of the insulin producing β‐cells in the islet.We have previously shown that cytokines decrease β‐cell coupling and that compounds which increase cAMP can increase coupling.In both mouse and human islets exendin‐4, which increases cAMP, protected against cytokine‐induced decreases in coupling and in mouse islets preserved glucose‐stimulated calcium signalling by increasing connexin36 gap junction levels on the plasma membrane.Our data indicate that protein kinase A regulates β‐cell coupling through a fast mechanism, such as channel gating or membrane organization, while Epac2 regulates slower mechanisms of regulation, such as gap junction turnover.Increases in β‐cell coupling with exendin‐4 may protect against cytokine‐mediated β‐cell death as well as preserve insulin secretion dynamics during the development of diabetes. [ABSTRACT FROM AUTHOR]

The islet of Langerhans plays a key role in glucose homeostasis through regulated secretion of the hormones insulin and glucagon. Islet research has focused on the insulin‐secreting β‐cells, even though aberrant glucagon secretion from α‐cells also contributes to the aetiology of diabetes. Despite its importance, the mechanisms controlling glucagon secretion remain controversial. Proper α‐cell function requires the islet milieu, where β‐ and δ‐cells drive and constrain α‐cell dynamics. The response of glucagon to glucose is similar between isolated islets and that measured in vivo, so it appears that the glucose dependence requires only islet‐intrinsic factors and not input from blood flow or the nervous system. Elevated intracellular free Ca2+ is needed for α‐cell exocytosis, but interpreting Ca2+ data is tricky since it is heterogeneous among α‐cells at all physiological glucose levels. Total Ca2+ activity in α‐cells increases slightly with glucose, so Ca2+may serve a permissive, rather than regulatory, role in glucagon secretion. On the other hand, cAMP is a more promising candidate for controlling glucagon secretion and is itself driven by paracrine signalling from β‐ and δ‐cells. Another pathway, juxtacrine signalling through the α‐cell EphA receptors, stimulated by β‐cell ephrin ligands, leads to a tonic inhibition of glucagon secretion. We discuss potential combinations of Ca2+, cAMP, paracrine and juxtacrine factors in the regulation of glucagon secretion, focusing on recent data in the literature that might unify the field towards a quantitative understanding of α‐cell function. [ABSTRACT FROM AUTHOR]

INTRODUCTION: This study explored the adaptive changes in pancreatic islets by assessing the maternal pancreatic β-cell mass (PβCM), islets size, number, shape, distribution and vascularity using the anti-GAD65 Ab as a marker in pregnant mice. MATERIALS AND METHODS: Fourty female Swiss-Webster mice were randomly divided into two age matched groups of 20 animals each, pregnant and control (non-pregnant) groups. The pregnant mice were sacrificed at Day 19 of gestation and the control group was sacrificed 19 days after observation in the same environmental conditions. Specimens of pancreata were fixed in 10% formal-saline, and processed for light microscopy. Paraffin sections were stained with chromogen DAB. Image scope (APERIO) with Image J software was used for morphometric study. RESULTS: Islets cells showed specific reaction with GAD65 Ab. There was an increase in the cellularity and vascularity of the PβCM. A significant increase in the islets area of the pregnant group ranging between 32-723 µm² in comparison with the control group that ranged between 5-210 µm², the islet size and numbers also increased; there were numerous newly formed islets and coalescence of adjacent islets, positive reactivity of β-cells toward anti- GAD65 during pregnancy, the mean of positive granulation in PβCM in pregnant group was 8273.5±SE 831.35 while in control group was 34±SE 0.2. CONCLUSION: The changes for alteration in PβCM such as increase in cells number, size, positive anti-GAD65 reactivity and their vascularity during pregnancy is related to the adaptation for hormonal and nutritional demands of both foetus and mother. [ABSTRACT FROM AUTHOR]

Acrylamide (AA) is a well-known industrial monomer with carcinogenic, mutagenic, neurotoxic and endocrine disruptive effects on living organisms. AA has been the subject of renewed interest owing to its presence in various food products. We investigated the potential adverse effects of oral AA treatment on the endocrine pancreas of juvenile rats using histochemical, immunohistochemical, stereological and biochemical methods. Thirty juvenile male Wistar rats were divided into one control and two AA treatment groups: one treated with 25 mg/kg AA and the other treated with 50 mg/kg AA for 21 days. We found a significant decrease in β-cell mass. The significant decrease in β-cell optical density and unchanged blood glucose levels indicate that normoglycemia in AA treated rats may result from intensive exocytosis of insulin-containing secretory granules. By contrast with β-cells, we observed increased α-cell mass. The slight increase in α-cell cytoplasmic volume suggests retention of glucagon in α-cells, which is consistent with the significant increase in α-cell optical density for AA treated animals. The number of islets of Langerhans did not change significantly in AA treated groups. Our findings suggest that AA treatment causes decreased β-cell mass and moderate α-cell mass increase in the islets of Langerhans of juvenile male Wistar rats. [ABSTRACT FROM AUTHOR]

Objective To investigate the effect and potential mechanism of glucagon-like peptide-1 (GLP-1) receptor agonist liraglutide on fibroblast growth factor-21 (FGF-21) production in mouse pancreatic β cell line Min6. Methods Min6 cells were incubated for 24 h with liraglutide in the presence or absence of the GLP-1 receptor antagonist exendin (9-39). The expressions of FGF-21 mRNA and protein were detected by RT-PCR and western blotting analyses. Supernatant FGF-21 levels were measured by ELISA. In the mean time, after Min6 cells were incubated with 10 nmol/L liraglutide and/or 5 μg/ml neutralizing anti-FGF-21 IgG for 24 h, supernatant insulin levels were measured by ELISA. Different group comparisons were performed using one-way variance analysis and LSD-t test. Results Compared with control group, 1, 10 and 100 nmol/L liraglutide dose-dependently upreglulated the expression levels of FGF-21 mRNA (1.26±0.24, 1.52±0.41 and 1.21±0.26 vs 1, F=3.523, P=0.021) and protein (1.61±0.27, 1.70±0.45 and 1.27±0.10 vs 1, F=5.052, P=0.006), and increased the levels of FGF-21 secretion [(0.21± 0.05), (0.25±0.08) and (0.26±0.07) vs (0.19±0.06) μg/mg, F=16.44,P=0.007] in Min6 cells. The above effects of 10 nmol/L liraglutide could be eliminated by addition of exendin (9-39) [mRNA level, 0.85±0.15 vs 1.31±0.16, t=4.133, P=0.026; protein level, 1.27±0.13 vs 1.74±0.06, t=10.55, P=0.009; secretion level, (0.23±0.11) vs (0.26±0.10) μg/mg protein, t=4.159,P=0.025]. Moreover, compared with control group, 10 nmol/L liraglutide stimulated insulin secretion in Min6 cells [(1.22±0.47) vs (0.55±0.23) μg/mg protein, t= 3.19, P=0.024], which could be abolished by blocking FGF-21 action with anti-FGF-21 IgG [(0.67±0.28) vs (1.22±0.47) μg/mg protein, t=4.371, P=0.007]. Conclusion Liraglutide enhances FGF-21 production in mouse pancreatic β cell line Min6 in a GLP-1 receptor-dependent manner. FGF-21 may be involved in the stimulating effect of liraglutide on insulin secretion from Min6 cells in an autocrine or paracrine fashion. [ABSTRACT FROM AUTHOR]

• Nuclear and C-terminus PTHrP are important for islet development and insulin secretion. • Nuclear and C-terminus PTHrP regulate normal calcium response of islets. • PTHrP Δ/Δ mice have less glucagon than control mice. Parathyroid hormone-related protein (PTHrP) is a pleiotropic hormone essential for morphogenesis, tissue differentiation, as well as cell regulation and function. PTHrP is expressed by pancreatic beta cells which are responsible for insulin secretion. Previous studies have reported that N-terminal PTHrP stimulated proliferation in beta cells in rodents. We have developed a knockin mouse model (PTHrP Δ/Δ) lacking the C-terminal and nuclear localization sequence (NLS) of PTHrP. These mice die at ∼day 5, are severely stunted in growth, weigh 54% less than control mice at day 1–2 and eventually fail to grow. PTHrP Δ/Δ mice are also hypoinsulinemic and hypoglycemic yet have nutrient intake proportional to size. To characterize the pancreatic islets in these mice, islets (∼10–20) were isolated from 2 to 5 day-old-mice using collagenase digestion. Islets from PTHrP Δ/Δ mice were smaller in size but secreted more insulin than littermate controls. PTHrP Δ/Δ and control mice islets were exposed to various glucose concentrations and intracellular calcium, the trigger for insulin release, was elevated for glucose concentrations of 8–20 mM. Immunofluorescence staining showed less glucagon-stained area in islets from PTHrP Δ/Δ mice (∼250 µm2) compared to islets from control mice (∼900 µm2), and ELISA confirmed there was reduced glucagon content. These data collectively demonstrate increased insulin secretion and reduced glucagon at the islet level, which may contribute to the observed hypoglycemia and early death in PTHrP Δ/Δ mice. Thus, the C-terminus and NLS of PTHrP are crucial to life, including regulation of glucose homeostasis and islet function. [ABSTRACT FROM AUTHOR]

The pathophysiology of diabetes is complex and recent research put focus on the pancreatic islets of Langerhans and the insulin-secreting beta cells as central in the development of the disease. MicroRNAs (miRNAs), the small non-coding RNAs regulating post-transcriptional gene expression, are significant regulators of beta cell function. One of the most abundant miRNAs in the islets is miR-375. This review focus on the role of miR-375 in beta cell function, including effects in development and differentiation, proliferation and regulation of insulin secretion. It also discusses the regulation of miR-375 expression, miR-375 as a potential circulating biomarker in type 1 and type 2 diabetes, and the need for the beta cell to keep expression of miR-375 within optimal levels. The summed picture of miR-375 is a miRNA with multiple functions with importance in the formation of beta cell identity, control of beta cell mass and regulation of insulin secretion. [ABSTRACT FROM AUTHOR]

In pancreatic β cells, muscarinic cholinergic receptor M3 (M3R) stimulates glucose-induced secretion of insulin. Regulator of G-protein signaling (RGS) proteins are critical modulators of GPCR activity, yet their role in β cells remains largely unknown. R7 subfamily RGS proteins are stabilized by the G-protein subunit Gβ5, such that the knockout of the Gnb5 gene results in degradation of all R7 subunits. We found that Gnb5 knockout in mice or in the insulin-secreting MIN6 cell line almost completely eliminates insulinotropic activity of M3R. Moreover, overexpression of Gβ5-RGS7 strongly promotes M3R-stimulated insulin secretion. Examination of this noncanonical mechanism in Gnb5-/- MIN6 cells showed that cAMP, diacylglycerol, or Ca2+ levels were not significantly affected. There was no reduction in the amplitude of free Ca2+ responses in islets from the Gnb5-/- mice, but the frequency of Ca2+ oscillations induced by cholinergic agonist was lowered by more than 30%. Ablation of Gnb5 impaired M3R-stimulated phosphorylation of ERK1/2. Stimulation of the ERK pathway in Gnb5-/- cells by epidermal growth factor restored M3R-stimulated insulin release to near normal levels. Identification of the novel role of Gβ5-R7 in insulin secretion may lead to a new therapeutic approach for improving pancreatic β-cell function. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are cellular, short, non-coding ribonucleotides acting as endogenous posttranscriptional repressors following incorporation in the RNA-induced silencing complex. Despite being chemically and mechanistically very similar, miRNAs exert a multitude of different cellular effects by acting on mRNA species, whose gene-products partake in a wide array of processes. Here, the aim was to review the knowledge of miRNA expression and action in the islet of Langerhans. We have focused on: 1) physiological consequences of islet or beta cell specific inhibition of miRNA processing, 2) mechanisms regulating processing of miRNAs in islet cells, 3) presence and function of miRNAs in alpha versus beta cells – the two main cell types of islets, and 4) miRNA mediators of beta cell decompensation. It is clear that miRNAs regulate pancreatic islet development, maturation, and function in vivo . Moreover, processing of miRNAs appears to be altered by obesity, diabetes, and aging. A number of miRNAs (such as miR-7, miR-21, miR-29, miR-34a, miR-212/miR-132, miR-184, miR-200 and miR-375) are involved in mediating beta cell dysfunction and/or compensation induced by hyperglycemia, oxidative stress, cytotoxic cytokines, and in rodent models of fetal metabolic programming prediabetes and overt diabetes. Studies of human type 2 diabetic islets underline that these miRNA families could have important roles also in human type 2 diabetes. Furthermore, there is a genuine gap of knowledge regarding miRNA expression and function in pancreatic alpha cells. Progress in this area would be enhanced by improved in vitro alpha cell models and better tools for islet cell sorting. [ABSTRACT FROM AUTHOR]

Human enteroviruses (HEV), especially coxsackievirus serotype B (CVB) and echovirus (E), have been associated with diseases of both the exocrine and endocrine pancreas, but so far evidence on HEV infection in human pancreas has been reported only in islets and ductal cells. This study aimed to investigate the capability of echovirus strains to infect human exocrine and endocrine pancreatic cells. Infection of explanted human islets and exocrine cells with seven field strains of E6 caused cytopathic effect, virus titer increase and production of HEV protein VP1 in both cell types. Virus particles were found in islets and acinar cells infected with E6. No cytopathic effect or infectious progeny production was observed in exocrine cells exposed to the beta cell-tropic strains of E16 and E30. Endocrine cells responded to E6, E16 and E30 by upregulating the transcription of interferon-induced with helicase C domain 1 (IF1H1), 2'-5'-oligoadenylate synthetase 1 (OAS1), interferon-β (IFN-β), chemokine (C-X-C motif) ligand 10 (CXCL10) and chemokine (C-C motif) ligand 5 (CCL5). Echovirus 6, but not E16 or E30, led to increased transcription of these genes in exocrine cells. These data demonstrate for the first time that human exocrine cells represent a target for E6 infection and suggest that certain HEV serotypes can replicate in human pancreatic exocrine cells, while the pancreatic endocrine cells are permissive to a wider range of HEV. [ABSTRACT FROM AUTHOR]

Donation after circulatory death ( DCD) donors are increasingly being used as a source of pancreas allografts for vascularized organ and islet transplantation. We provide practice guidelines aiming to increase DCD pancreas utilization. We review risk assessment and donor selection criteria. We report suggested factors in donor and recipient clinical management and provide an overview of the activities and outcomes of vascularized pancreas and islet transplantation. [ABSTRACT FROM AUTHOR]

Type 1 diabetes (T1D) is caused by autoimmune destruction of islet of Langerhans β-cells. P2X7 receptors (P2X7R) modulate proinflammatory immune responses by binding extracellular ATP, a classic ‘danger signal’. Here, we evaluated whether the P2X7R has a role in T1D development. P2X7 −/− mice are resistant to TD1 induction by streptozotocin (STZ) treatment, with no increase in blood glucose, decrease in insulin-positive cells, and pancreatic islet reduction, compared to WT (C57BL/6) mice. Also, the levels of proinflammatory mediators (IL-1β, IFN-γ and NO) did not increase after STZ treatment in P2X7 −/− animals, with reduced infiltration of CD4 + , CD8 + , B220 + , CD11b + and CD11c + cells in the pancreatic lymph nodes. Treatment with a P2X7 antagonist mimicked the effect of P2X7 knockout, preventing STZ-induced diabetes. Our results show that the absence of the P2X7R provides resistance in the induction of diabetes in this model, and suggest that therapy targeting the P2X7R may be useful against clinical T1D. [ABSTRACT FROM AUTHOR]

The development and plasticity of the endocrine pancreas responds to both the intrauterine and postnatal exposome in a constant attempt to predict and respond to alterations in nutritional availability and metabolic requirements. Both under- and over-nutrition in utero, or exposure to adverse environmental pollutants or maternal behaviors, can each lead to altered β-cell or function at birth, and a subsequent mismatch in pancreatic hormonal demands and secretory capacity postnatally. This can be further exacerbated by metabolic stress postnatally such as from obesity or pregnancy, resulting in an increased risk of gestational diabetes, type 2 diabetes, and even type 1 diabetes. This review will discuss evidence identifying the cellular pathways in early life whereby the plasticity of the endocrine pancreatic can become pathologically limited. By necessity, much of this evidence has been gained from animal models, although extrapolation to human fetal development is possible from the fetal growth trajectory and study of the newborn. Cellular limitations to plasticity include the balance between β-cell proliferation and apoptosis, the appearance of β-cell oxidative stress, impaired glucose-stimulated insulin secretion, and sensitivity to circulating cytokines and responsiveness to programmed death receptor-1. Evidence suggests that many of the cellular pathways responsible for limiting β-cell plasticity are related to paracrine interactions within the islets of Langerhans. [ABSTRACT FROM AUTHOR]

Amyloid can be detected in the islets of Langerhans in a majority of type 2 diabetic patients. These deposits have been associated with β-cell death, thereby furthering diabetes progression. Islet amyloid polypeptide (IAPP) amyloidogenicity is quite variable among animal species, and studying this variability could further our understanding of the mechanisms involved in the aggregation process. Thus, the general aim of this study was to identify IAPP isoforms in different animal species and characterize their propensity to form fibrillar aggregates. A library of 23 peptides (fragment 8–32) was designed to study the amyloid formation usingin silicoanalysis andin vitroassays. Amyloid formation was impeded when the NFLVH motif found in segment 8–20 was substituted by DFLGR or KFLIR segments. A 29P, 14K and 18R substitution were often present in non-amyloidogenic sequences. Non-amyloidogenic sequences were obtained fromLeontopithecus rosalia, Tursiops truncatusandVicugna pacos. Fragment peptides from 34 species were amyloidogenic. To conclude, this project advances our knowledge on the comparative pathogenesis of amyloidosis in type II diabetes. It is conceivable that the additional information gained may help point towards new therapeutic strategies for diabetes patients. [ABSTRACT FROM AUTHOR]

Cytoplasmic ATP and Ca2+ are implicated in current models of glucose's control of glucagon and insulin secretion from pancreatic α- and β-cells, respectively, but little is known about ATP and its relation to Ca + in α-cells. We therefore expressed the fluorescent ATP biosensor Perceval in mouse pancreatic islets and loaded them with a Ca2+ indicator. With total internal reflection fluorescence microscopy, we recorded subplasma membrane concentrations of Ca2+ and ATP ([Ca2+] pm; [ATP]pm) in superficial α- and β-cells of intact islets and related signaling to glucagon and insulin secretion by immunoassay. Consistent with ATP's controlling glucagon and insulin secretion during hypo- and hyperglycemia, respectively, the dose-response relationship for glucose-induced [ATP]pm generation was left shifted in α-cells compared to β-cells. Both cell types showed [Ca2+]pm and [ATP]pm oscillations in opposite phase, probably reflecting energy-consuming Ca2+ transport. Although pulsatile insulin and glucagon release are in opposite phase, [Ca2+]pm synchronized in the same phase between α- and β-cells. This paradox can be explained by the overriding of Ca2+ stimulation by paracrine inhibition, because somato-statin receptor blockade potently stimulated glucagon release with little effect on Ca2+. The data indicate that an α-cell-intrinsic mechanism controls glucagon in hypoglycemia and that paracrine factors shape pulsatile secretion in hyperglycemia. [ABSTRACT FROM AUTHOR]

Quantitative analysis of tissues and organs can reveal large-scale patterning as well as the impact of perturbations and aging on biological architecture. Here we develop tools for imaging of single cells in intact organs and computational approaches to assess spatial relationships in 3D. In the mouse ovary, we use nuclear volume of the oocyte to read out quiescence or growth of oocyte–somatic cell units known as follicles. This in-ovary quantification of non-growing follicle dynamics from neonate to adult fits a mathematical function, which corroborates the model of fixed oocyte reserve. Mapping approaches show that radial organization of folliculogenesis established in the newborn ovary is preserved through adulthood. By contrast, inter-follicle clustering increases during aging with different dynamics depending on size. These broadly applicable tools can reveal high dimensional phenotypes and age-related architectural changes in other organs. In the adult mouse pancreas, we find stochastic radial organization of the islets of Langerhans but evidence for localized interactions among the smallest islets. [ABSTRACT FROM AUTHOR]

Background: DPP-4 inhibitors reportedly exert effects on both alpha and beta cells, and promote the proliferation and survival of beta cells. We investigated the effects of anagliptin on structurally-impaired islets of Langerhans in streptozotocin (STZ)-treated mice, fed either a normal or a high-fat diet. Pdx-1 expression in the pancreas and serum insulin/glucagon concentrations were also examined. Findings: Anagliptin treatment significantly up-regulated pancreatic Pdx-1 expression, with elevated serum glucagon-like peptide-1 concentrations, regardless of whether the diet was normal or high-fat. However, interestingly, the beta cell regeneration, structural normalization of islets of Langerhans including alpha cell: beta cell area ratios, and serum insulin elevation, all observed with anagliptin administration in the animals fed a normal diet, were markedly suppressed in the high-fat fed group. Conclusions: High-fat diet feeding clearly weakened the regenerative effects of anagliptin on the islets of Langerhans in STZ-treated mice. Our findings suggest the importance of normalizing lipid metabolism for full manifestation of DPP-4 inhibitor effects on the islets of Langerhans. [ABSTRACT FROM AUTHOR]

Dipeptidyl peptidase 4 (DPP-4) was recently found to be expressed in human and mouse islets with different expression patterns. However, whether species-dependent expression pattern is a generalized phenomenon and whether islet DPP-4 activity is regulated are not known. This study was conducted to investigate DPP-4 localization in several different species, and to examine the impact of glucose, incretin hormones, and insulin on islet DPP-4 activity. It was shown by immuofluorescent staining that there were two distinct species-specific expression patterns of islet DPP-4. The enzyme was expressed exclusively in α-cells in human and pig islets, but primarily in β-cells in mouse and rat islets. INS-1 832/13 cells also expressed DPP-4, and inhibition of DPP-4 enhanced insulin secretion in the presence of glucagon-like peptide-1 (GLP-1) in the cells. DPP-4 activity was remarkably robust when cultured with high glucose, incretin hormones, and insulin in mouse and human islets as well as INS-1 832/13 cells and islet DPP-4 activity and expression pattern was not altered in double incretin receptor knockout mice, compared to wild type mice. We conclude that islet DPP-4 is species-specifically expressed in α-cell and β-cell dominant patterns in several species and both patterns remained robust in enzyme activity during short-term metabolic challenge. [ABSTRACT FROM AUTHOR]

Coordinated pulses of electrical activity and insulin secretion are a hallmark of the islet of Langerhans. These coordinated behaviors are lost when β cells are dissociated, which also leads to increased insulin secretion at low glucose levels. Islets without gap junctions exhibit asynchronous electrical activity similar to dispersed cells, but their secretion at low glucose levels is still clamped off, putatively by a juxtacrine mechanism. Mice lacking β cell gap junctions have near-normal average insulin levels, but are glucose intolerant due to reduced first-phase and pulsatile insulin secretion, illustrating the importance of temporal dynamics. Here, we review the quantitative data on islet synchronization and the current mathematical models that have been developed to explain these behaviors and generate greater understanding of the underlying mechanisms. [ABSTRACT FROM AUTHOR]

Background The successful isolation, purification, and culture of caprine islets has recently been reported. The present study shows arange of size distribution in caprine islet diameter from 50 to 250 μm, in which 80% of the total islet yield was comprised of small islets. Methods Caprine islets were isolated and purified. Islets were handpicked and the diameter of the islets was recorded using light microscopy. Viablility of the islets was analyzed by confocal microscopy. Insulin secretion assay was carried out and analyzed by ELISA. Results When tested at 48 h after isolation, these small islets were 29.3% more viable compared to the large-sized islets. Large islets showed a high ratio (P < 0.01) of central core necrosis (29.5% ± 1.92) whilst no significant core death was observed in small islets (2.33% ± 0.59). The annexin assay demonstrated 5.21% ± 0.97 and 7.34% ± 0.78 apoptotic death for small and large islets, respectively. During static incubation, small islets released 2.89-fold (1.39 ± 0.2 ng/IE) higher insulin level under low glucose induction (3.3 m m) and simultaneously 2.92-fold (2.95 ± 0.33 ng/IE) more insulin under high glucose condition (16.7 m m) in comparison to large islets at the same islet equivalents (P < 0.05). Conclusion The present findings evidenced the superior quality of smaller caprine islets compared to larger ones under an optimized basal maintenance condition. As it is equally important to preserve the quality of larger caprine islets, this work warrants further investigation on special culture conditions to support these islets. [ABSTRACT FROM AUTHOR]

Abstract: The retinoic-acid-related receptor family of orphan receptors (RORs) act as transcriptional activators or repressors. One of their functions involves integrated actions within circadian oscillators, particularly of the periphery. The present paper describes differential expression of the orphan receptors RORα, RORβ and RORγ and of the nuclear retinoid receptor RXRα in the pancreas and islet of rats. Immunohistochemistry of rodent islets detected nuclear receptor expression. The RORα and RORβ signals were visualised in α-cells, whereas that of RORγ was largely confined to β-cells. RXRα was expressed throughout the islets. Quantitative RT-PCR revealed circadian expression in the rat pancreas for RORγ, RORα and RXRα, but not for RORβ. Circadian expression of RORγ mRNA was verified in mouse pancreas and in rat INS-1 β cells by serum shock experiments. The results point to differential and circadian expression and thus cell-type-specific functions of RORα and RORγ in islet cells secreting glucagon or insulin. [Copyright &y& Elsevier]

Type 2 diabetes is characterized by impaired insulin secretion from pancreatic b-cells. Quantification of the islet area in addition to the insulin-positive area is important for detailed understanding of pancreatic islet histopathology. Here we show computerized automatic recognition of the islets of Langerhans as a novel high-throughput method to quantify islet histopathology. We utilized state-of-the-art tissue pattern recognition software to enable automatic recognition of islets, eliminating the need to laboriously trace islet borders by hand. After training by a histologist, the software successfully recognized even irregularly shaped islets with depleted insulin immunostaining, which were quite difficult to automatically recognize. The results from automated image analysis were highly correlated with those from manual image analysis. To establish whether this automated, rapid, and objective determination of islet area will facilitate studies of islet histopathology, we showed the beneficial effect of chronic exendin-4, a glucagon-like peptide-1 analog, treatment on islet histopathology in Zucker diabetic fatty (ZDF) rats. Automated image analysis provided qualitative and quantitative evidence that exendin-4 treatment ameliorated the loss of pancreatic insulin content and gave rise to islet hypertrophy. We also showed that glucagon-positive a-cell area was decreased significantly in ZDF rat islets with disorganized structure. This study is the first to demonstrate the utility of automatic quantification of digital images to study pancreatic islet histopathology. The proposed method will facilitate evaluations in preclinical drug efficacy studies as well as elucidation of the pathophysiology of diabetes. [ABSTRACT FROM AUTHOR]

Insulin secretion from the pancreatic β-cell is controlled by changes in membrane potential and intracellular Ca2+. The contribution of intracellular Ca2+ stores to this process is poorly understood. We have previously shown that β-cells of mice lacking one copy of the Annexin 7 gene (Anx7(+/-)) express reduced levels of IP3 receptors and defects in IP3-dependent Ca2+ signaling. To further elucidate the effect of the Anx7(+/-) mutation on signaling related to intracellular Ca2+ stores in the β-cell, we measured the effects of Ca2+ mobilizing agents on electrical activity, intracellular Ca2+ and insulin secretion in control and mutant β-cells. We found that the muscarinic agonist carbachol and the ryanodine receptor agonists caffeine and 4-chloro-m-cresol had more potent depolarizing effects on Anx7(+/-) β-cells compared to controls. Accordingly, glucose-induced insulin secretion was augmented to a greater extent by caffeine in mutant islets. Surprisingly, ryanodine receptor-mediated Ca2+ mobilization was not affected by the Anx7(+/-) mutation, suggesting that the mechanism underlying the observed differences in electrical and secretory responsiveness does not involve intracellular Ca2+ stores. Our results provide evidence that both IP3 receptors and ryanodine receptors play important roles in regulating β-cell membrane potential and insulin secretion, and that the Anx7(+/-) mutation is associated with alterations in the signaling pathways related to these receptors. [ABSTRACT FROM AUTHOR]

Understanding and controlling biological responses against artificial materials is important for the development of medical devices and therapies. Self-assembled monolayers (SAMs) of alkanethiols provide well-defined surfaces that can be manipulated by varying the terminal functional groups. Thus, SAMs have been extensively used as a platform for studying how artificial materials affect biological responses. Here, we review cell adhesion behavior in response to SAMs with various surface properties and the effects that adsorbed proteins have on subsequent cell adhesion. We also describe an application for SAMs as a substrate for culturing neural stem cells (NSCs). Substrates that induced the correct orientation of immobilized growth factors, like epidermal growth factor, improved the selection of a pure NSC population during cell expansion. In addition, we review new methodologies for using amphiphilic polymers to modify the surfaces of cells and tissues. Coating the cell surface with amphiphilic polymers that can capture and immobilize bioactive substances or cells represents a promising approach for clinical applications, particularly cellular therapies. [ABSTRACT FROM AUTHOR]

Abstract: Uncoupling Protein 2 (UCP2) is expressed in the pancreatic β-cell, where it partially uncouples the mitochondrial proton gradient, decreasing both ATP-production and glucose-stimulated insulin secretion (GSIS). Increased glucose levels up-regulate UCP2 mRNA and protein levels, but the mechanism for UCP2 up-regulation in response to increased glucose is unknown. The aim was to examine the effects of glucokinase (GK) deficiency on UCP2 mRNA levels and to characterize the interaction between UCP2 and GK with regard to glucose-stimulated insulin secretion in pancreatic islets. UCP2 mRNA expression was reduced in GK+/− islets and GK heterozygosity prevented glucose-induced up-regulation of islet UCP2 mRNA. In contrast to UCP2 protein function UCP2 mRNA regulation was not dependent on superoxide generation, but rather on products of glucose metabolism, because MnTBAP, a superoxide dismutase mimetic, did not prevent the glucose-induced up-regulation of UCP2. Glucose-stimulated insulin secretion was increased in UCP2−/− and GK+/− islets compared with GK+/− islets and UCP2 deficiency improved glucose tolerance of GK+/− mice. Accordingly, UCP2 deficiency increased ATP-levels of GK+/− mice. Thus, the compensatory down-regulation of UCP2 is involved in preserving the insulin secretory capacity of GK mutant mice and might also be implicated in limiting disease progression in MODY2 patients. [Copyright &y& Elsevier]

Diabetes in the domestic ferret ( Mustela putorius furo) has previously been described and the purpose of this study was to evaluate if the ferret could serve as a model for the study of β-cell degeneration associated with formation of islet amyloid. The nucleotide and amino acid sequence of ferret islet amyloid polypeptide (IAPP) 1-37 was identified and the synthesized peptide was studied with regards to in vitro amyloidogenicity and potential cellular toxicity in a comparative approach to human, cat and the non-amyloidogenic rat IAPP. Ferret IAPP forms amyloid-like fibrils, but with a longer lag phase than human and cat IAPP and the aggregation process was shown to reduce cell viability of cultured β-cells, but with less potency than these two amyloidogenic counterparts. Immunohistochemistry of ferret pancreas confirmed IAPP expression in the islets of Langerhans, but no islet amyloid was found in a very limited sample size of one diabetic and five healthy ferrets. Islet amyloid has never been described in ferrets, and it is not possible to determine if it is due to lack of studies/material or to the fact that the ferret's life span is too short to present with such pathology. [ABSTRACT FROM AUTHOR]

Abstract: An unresolved obstacle in transplantation of islets of Langerhans is the early graft loss caused by thrombotic reactions on the surface of islets after intraportal transplantation. We investigated a versatile method for modifying the surface of islets with liposomes carrying the anticoagulant argatroban using an amphiphilic poly(ethylene glycol)-phospholipid conjugate derivative (PEG-lipid) and DNA hybridization. Argatroban was gradually released from the liposomes on the islets, and antithrombic activity was detected in culture medium. Modified islets retained the ability to control insulin release in response to glucose concentration changes. Although we mainly examined surface modification of islets, this technique may be useful for immobilizing various types of small molecules on cells and tissues and thus may have many applications in cell therapy and regenerative medicine. [Copyright &y& Elsevier]

Abstract: ATP-sensitive potassium channels (KATP) regulate electrical activity and insulin secretion in pancreatic β-cells. When glucose concentration increases, the [ATP]/[ADP] ratio rises closing KATP channels, and the membrane potential depolarizes, triggering insulin secretion. This pivotal role of KATP channels is used not only by glucose but also by neurotransmitters, hormones and other physiological agents to modulate electrical and secretory β-cell response. In recent years, it has been demonstrated that estrogens and estrogen receptors are involved in glucose homeostasis, and that they can modulate the electrical activity and insulin secretion of pancreatic β-cells. The hormone 17β-estradiol (E2), at physiological levels, is implicated in maintaining normal insulin sensitivity for β-cell function. Long term exposure to E2 increases insulin content, insulin gene expression and insulin release via the estrogen receptor α (ERα), while rapid responses to E2 can regulate KATP channels increasing cGMP levels through the estrogen receptor β (ERβ) and type A guanylate cyclase receptor (GC-A). This review summarizes the main actions of 17β-estradiol on KATP channels and the subsequent insulin release in pancreatic β-cells. [Copyright &y& Elsevier]

Abstract: A capillary electrophoresis competitive immunoassay was developed for the simultaneous quantitation of insulin, glucagon, and islet amyloid polypeptide (IAPP) secretion from islets of Langerhans. Separation buffers and conditions were optimized for the resolution of fluorescein isothiocyanate (FITC)-labeled glucagon and IAPP immunoassay reagents, which were excited with the 488nm line of an Ar+ laser and detected at 520nm with a photomultiplier tube (PMT). Cy5-labeled insulin immunoassay reagents were excited by a 635nm laser diode module and detected at 700nm with a separate PMT. Optimum resolution was achieved with a 20mM carbonate separation buffer at pH 9.0 using a 20cm effective separation length with an electric field of 500V/cm. Limits of detection for insulin, glucagon, and IAPP were 2, 3, and 3nM, respectively. This method was used to monitor the simultaneous secretion of these peptides from as few as 14 islets after incubation in 4, 11, and 20mM glucose for 6h. For insulin and IAPP, a statistically significant increase in secretion levels was observed, while glucagon levels were significantly reduced in the 4 and 11mM glucose conditions. To further demonstrate the utility of the assay, the Ca2+-dependent secretion of these peptides was demonstrated which agreed with published reports. The ability to examine the secretion of multiple peptides may allow for the determination of regulation of secretory processes within islets of Langerhans. [Copyright &y& Elsevier]

Abstract: Transplantation of islets of Langerhans is a promising method for treating patients with insulin-dependent diabetes mellitus. The major obstacle in clinical settings is early graft loss due to inflammation triggered by blood coagulation and complement activation on the surface of the islets after intraportal transplantation. We propose a versatile method for modifying the surface of islets with the fibrinolytic enzyme urokinase and the soluble domain of the anticoagulant enzyme thrombomodulin. The surfaces of islets were modified with a poly(ethylene glycol)–phospholipid conjugate bearing a maleimide group (Mal–PEG–lipid; PEG MW=5000kDa). The Mal–PEG–lipid anchored to the cell membranes of islets, resulting in the presentation of functional maleimide groups on the islet surface. The surface was further treated with thiolated urokinase and thrombomodulin that conjugated by thiol/maleimide bonding. No practical islet volume increase was observed after surface modification, and the modifications did not impair insulin release in response to glucose stimulation. Furthermore, the activity of the immobilized urokinase and thrombomodulin was maintained. These modifications could help to improve graft survival by preventing thrombus formation on the surface of transplanted islets. [Copyright &y& Elsevier]

BACKGROUND: Bisphenol A (BPA) is a widespread endocrine-disrupting chemical used as the base compound in the manufacture of polycarbonate plastics. In humans, epidemiological evidence has associated BPA exposure in adults with higher risk of type 2 diabetes and heart disease. OBJECTIVE: We examined the action of environmentally relevant doses of BPA on glucose metabolism in mice during pregnancy and the impact of BPA exposure on these females later in life. We also investigated the consequences of in utero exposure to BPA on metabolic parameters and pancreatic function in offspring. METHODS: Pregnant mice were treated with either vehicle or BPA (10 or 100 µg/kg/day) during days 9-16 of gestation. Glucose metabolism experiments were performed on pregnant mice and their offspring. RESULTS: BPA exposure aggravated the insulin resistance produced during pregnancy and was associated with decreased glucose tolerance and increased plasma insulin, triglyceride, and leptin concentrations relative to controls. Insulin-stimulated Akt phosphorylation was reduced in skeletal muscle and liver of BPA-treated pregnant mice relative to controls. BPA exposure during gestation had long-term consequences for mothers: 4 months postpartum, treated females weighed more than untreated females and had higher plasma insulin, leptin, triglyceride, and glycerol levels and greater insulin resistance. At 6 months of age, male offspring exposed in utero had reduced glucose tolerance, increased insulin resistance, and altered blood parameters compared with offspring of untreated mothers. The islets of Langerhans from male offspring presented altered Ca2+ signaling and insulin secretion. BrdU (bromodeoxyuridine) incorporation into insulin-producing cells was reduced in the male progeny, yet β-cell mass was unchanged. CONCLUSIONS: Our findings suggest that BPA may contribute to metabolic disorders relevant to glucose homeostasis and that BPA may be a risk factor for diabetes. [ABSTRACT FROM AUTHOR]

Abstract: Glucokinase activators increase insulin release from pancreatic β-cells and hepatic glucose utilization by modifying the activity of glucokinase, a key enzyme in glucose-sensing and glycemic regulation. Sulfonylureas are antihyperglycemic agents that stimulate insulin secretion via a glucose-independent mechanism that is vulnerable to secondary failure through β-cell desensitization. The present study determined whether glucokinase activator treatment retains its glucose-lowering efficacy in male, adult, non-diabetic Sprague–Dawley rats desensitized to sulfonylurea treatment and whether glucose-lowering during chronic glucokinase activator treatment is subject to secondary failure. Animals were given food containing either glimepiride (a sulfonylurea), Compound B (3-[(1S)-2-hydroxy-1-methylethoxy]-5-[4-(methylsulfonyl)phenoxy]-N-1,3-thiazol-2-ylbenzamide, an experimental glucokinase activator), or no drug for up to 5weeks. Food containing 0.04% of either drug produced acute (within 4–8h) and significant (P <0.05) reductions in blood glucose to approximately 50% of control levels. Chronic treatment with either 0.01% or 0.04% glimepiride resulted in complete failure of glucose-lowering efficacy within 3days whereas the efficacy of Compound B was sustained throughout the entire study. Glipizide, also a sulfonylurea, had no glucose-lowering effect when given by gavage (3mg/kg) to glimepiride-desensitized animals whereas Compound B retained full glucose-lowering efficacy in glimepiride-desensitized animals. Oral glucose tolerance was significantly impaired, compared with controls, in animals treated with glimepiride for two weeks but was enhanced to a small extent in animals treated with Compound B. Compound B also significantly increased pancreatic insulin content, compared with controls. These findings suggest that Compound B has sustained glucose-lowering effects in a rat model of sulfonylurea failure. [Copyright &y& Elsevier]

Abstract: Pancreatic β-cells release insulin in appropriate amounts in order to keep blood glucose levels within physiological limits. Failure to do so leads to the most common metabolic disorder in man, diabetes mellitus. The glucose-stimulus/insulin-secretion coupling represents a sophisticated interplay between glucose and a variety of modulatory factors. These factors are provided by the blood supply (such as nutrients, vitamins, incretins etc.), the nerval innervations, cell-cell contacts as well as by paracrine and autocrine feedback loops within the pancreatic islet of Langerhans. However, the underlying mechanisms of their action remain poorly understood. In the present mini-review we discuss novel aspects of selective insulin signaling in the β-cell and novel insights into the role of higher inositol phosphates in insulin secretion. Finally we present a newly developed experimental platform that allows non-invasive and longitudinal in vivo imaging of pancreatic islet/β-cell biology at single-cell resolution. [Copyright &y& Elsevier]

Abstract: The concept of glucolipotoxicity refers to the combined, deleterious effects of elevated glucose and fatty acid levels on pancreatic beta-cell function and survival. Significant progress has been made in recent years towards a better understanding of the cellular and molecular basis of glucolipotoxicity in the beta cell. The permissive effect of elevated glucose on the detrimental actions of fatty acids stems from the influence of glucose on intracellular fatty acid metabolism, promoting the synthesis of cellular lipids. The combination of excessive levels of fatty acids and glucose therefore leads to decreased insulin secretion, impaired insulin gene expression, and beta-cell death by apoptosis, all of which probably have distinct underlying mechanisms. Recent studies from our laboratory have identified several pathways implicated in fatty acid inhibition of insulin gene expression, including the extracellular-regulated kinase (ERK1/2) pathway, the metabolic sensor Per-Arnt-Sim kinase (PASK), and the ATF6 branch of the unfolded protein response. We have also confirmed in vivo in rats that the decrease in insulin gene expression is an early defect which precedes any detectable abnormality in insulin secretion. While the role of glucolipotoxicity in humans is still debated, the inhibitory effects of chronically elevated fatty acid levels has been clearly demonstrated in several studies, at least in individuals genetically predisposed to developing type 2 diabetes. It is therefore likely that glucolipotoxicity contributes to beta-cell failure in type 2 diabetes as well as to the decline in beta-cell function observed after the onset of the disease. [Copyright &y& Elsevier]

G-protein coupled receptors (GPCRs) are targets of approximately 30% of currently marketed drugs. Over the last few years, a number of GPCRs expressed in pancreatic β-cells and activated by lipids have been discovered. GPR40 was shown to be activated by medium- to long-chain fatty acids (FAs). It has since been shown that GPR40 contributes to FA amplification of glucose-induced insulin secretion. Although some controversy still exists as to whether GPR40 agonists or antagonists should be designed as novel type 2 diabetes drugs, data obtained in our laboratory and others strongly suggest that GPR40 agonism might represent a valuable therapeutic approach. GPR119 is expressed in pancreatic β-cells and enteroendocrine L-cells, and augments circulating insulin levels both through its direct insulinotropic action on β-cells and through FA stimulation of glucagon-like peptide 1 (GLP-1) secretion. GPR120 is expressed in L-cells and was also shown to mediate FA-stimulated GLP-1 release. Finally, GPR41 and GPR43 are receptors for short-chain FAs and may indirectly regulate β-cell function via adipokine secretion. Although the discovery of these various lipid receptors opens new and exciting avenues of research for drug development, a number of questions regarding their mechanisms of action and physiological roles remain to be answered. [ABSTRACT FROM AUTHOR]

β-Cell dysfunction is an important factor in the development of hyperglycemia of type-2 diabetes mellitus, and pancreatic islet amyloidosis (IA) has been postulated to be one of the main contributors to impaired insulin secretion. The aim of this study was to evaluate the correlation of IA with metabolic parameters and its effect on islets of Langerhans remodeling and relative endocrine-cell volume in baboons. We sequenced the amylin peptide, determined the fibrillogenic propensities, and evaluated pancreatic histology, clinical and biochemical characteristics, and endocrine cell proliferation and apoptosis in 150 baboons with different metabolic status. Amylin sequence in the baboon was 92% similar to humans and showed superimposable fibrillogenic propensities. IA severity correlated with fasting plasma glucose (FPG) (r = 0.662. P < 0.001) and HbAlc (r = 0.726, P < 0.001), as well as with free fatty acid, glucagon values, decreased homeostasis model assessment (HOMA) insulin resistance, and HOMA-B. IA severity was associated with a decreased relative β-cell volume, and increased relative a-cell volume and hyperglucagonemia. These results strongly support the concept that IA and p-cell apoptosis in concert with a-cell proliferation and hypertrophy are key determinants of islets of Langerhans "dysfunctional remodeling" and hyperglycemia in the baboon, a nonhuman primate model of type-2 diabetes mellitus. The most important determinants of IA were age and FPG (R2 = 0.519, P < 0.0001), and different FPG levels were sensitive and specific to predict IA severity. Finally, a predictive model for islet amyloid severity was generated with age and FPG as required variables. [ABSTRACT FROM AUTHOR]

Ion transport activity in pancreatic α-cells was assessed by studying cell volume regulation in response to anisotonic solutions. Cell volume was measured by a video imaging method, and cells were superfused with either 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid-buffered or HCO-buffered solutions. α-Cells did not exhibit a regulatory volume increase (RVI) in response to cell shrinkage caused by hypertonic solutions. A RVI was observed, however, in cells that had first undergone a regulatory volume decrease (RVD), but only in HCO-buffered solutions. RVI was also observed in response to a HCO-buffered hypertonic solution in which the glucose concentration was increased from 4 to 20 mM. The post-RVD RVI and the glucose-induced RVI were both inhibited by 10 μM 5-( N-methyl- N-isobutyl) amiloride or 100 μM 2,2′-(1,2-ethenediyl) bis (5-isothio-cyanatobenzenesulfonic acid), but not by 10 μM benzamil nor 10 μM bumetanide. These data suggest that Na+–H+ exchangers and Cl−–HCO exchangers contribute to volume regulation in α-cells. [ABSTRACT FROM AUTHOR]

Abstract: This study was performed on the basis of previously obtained investigative gene array data concerning the over-expression of CXCL12/SDF-1 in human insulinomas versus human pancreatic islet preparations. The presence of CXCL12/SDF-1 was studied by RT-qPCR in human insulinomas (n =8) versus pancreatic islets (n =3), and was found to be significantly up-regulated in the former (p <0.012). The mRNA data were confirmed by immunostaining and confocal microscopy of human normal pancreatic islets, which showed the absence of CXCL12 protein and high expression in insulinoma tissue. Individual human insulinoma cells at cytospins stained positive for CXCL12 in the paranuclear region. These morphological data were extended by consecutive immunoblotting for cell-compartment-specific marker proteins of fractions obtained by sucrose gradient fractionation using Rin-5F insulinoma cells. CXCL12-containing fractions were positive for the membrane marker NSF but negative for SNAP-25 and appeared at a lighter density in the gradient than heavy insulin granules, suggesting packaging in specific granules different from insulin. In order to determine the biological relevance of the protein in insulinomas, we investigated the colony-forming potential of human CXCL12 stable-transfected rat Rin-5F insulinoma cells. These clones secreted human CXCL12 and contained 50–1000-fold higher copy numbers compared to its endogenous rat homologue. In colony-forming assays, these transfectant clones developed greater colony numbers, which were larger than wild-type and sham transfectants. To elucidate the mechanism of action, we identified a CXCL12 transfectant-specific increase in the pro-survival factor Mn-SOD, which is considered important for the inactivation of reactive oxygen species, thereby prolonging cell survival. These data demonstrate the importance of CXCL12 in the tumor biology of insulinoma. [Copyright &y& Elsevier]

The glucokinase (GCK) gene was one of the first candidate genes to be identified as a human “diabetes gene". Subsequently, important advances were made in understanding the impact of GCK in the regulation of glucose metabolism. Structure elucidation by crystallography provided insight into the kinetic properties of GCK. Protein interaction partners of GCK were discovered. Gene expression studies revealed new facets of the tissue distribution of GCK, including in the brain, and its regulation by insulin in the liver. Metabolic control analysis coupled to gene overexpression and knockout experiments highlighted the unique impact of GCK as a regulator of glucose metabolism. Human GCK mutants were studied biochemically to understand disease mechanisms. Drug development programs identified small molecule activators of GCK as potential antidiabetics. These advances are summarized here, with the aim of offering an integrated view of the role of GCK in the molecular physiology and medicine of glucose homeostasis. [ABSTRACT FROM AUTHOR]