Stress In Pancreatic Islets Research Articles

Identification of Compounds Protecting Pancreatic Islets against Oxidative Stress using a 3D Pseudoislet Screening Platform.

Oxidative stress leads to a lower success rate of clinical islet transplantation. Here, FDA-approved compounds are screened for their potential to decrease oxidative stress and to protect or enhance pancreatic islet viability and function. Studies are performed on in vitro "pseudoislet" spheroids, which are pre-incubated with 1280 different compounds and subjected to oxidative stress. Cell viability and oxidative stress levels are determined using a high-throughput fluorescence microscopy pipeline. Initial screening on cell viability results in 59 candidates. The top ten candidates are subsequently screened for their potential to decrease induced oxidative stress, and eight compounds efficient reduction of induced oxidative stress in both alpha and beta cells by 25-50%. After further characterization, the compound sulfisoxazole is found to be the most capable of reducing oxidative stress, also at short pre-incubation times, which is validated in primary human islets, where low oxidative stress levels and islet function are maintained. This study shows an effective screening strategy with 3D cell aggregates based on cell viability and oxidative stress, which leads to the discovery of several compounds with antioxidant capacity. The top candidate, sulfisoxazole is effective after a 30 min pre-incubation, maintains baseline islet function, and may help alleviate oxidative stress in pancreatic islets.

1725-P: The Role of miR-146a-5p on Cytokine-Induced ß-Cell Mitochondrial Dysfunction and Death

MicroRNAs are small non-coding RNAs with ~22 nucleotides in size involved in the modulation of gene expression. Previously, we observed that miR-146a-5p is upregulated during inflammatory stress in pancreatic islets and pancreatic tissue sections from T1D donors. Although others have linked overexpression of miR-146a-5p to β cell death and impaired insulin secretion, the molecular mechanisms mediating these effects remain unclear. To further understand miR-146a-5p role in β cell health and disease, we have generated stable cell lines that are either overexpressed (OE) or inhibited (ZIP) for miR-146a-5p and assessed for β cell function under inflammatory (IL-1β, IFN-γ, and TNF-α) conditions to model T1D in vitro. Treatment of OE cells with or without pro-inflammatory cytokines increased expression of cleaved caspase-3 and Annexin-V-positive cell death and reduced insulin secretion in both untreated and cytokine treatment. However, ZIP cells are protected from inflammatory stress showing decreased apoptosis and improved insulin secretion. Furthermore, ZIP cells were found to have increased mitochondrial DNA copy numbers, mitochondrial membrane potential, and basal and maximal respiration rate. Notably, cytokines-treated OE cells showed reduced basal respiration rate and ATP production compared to cytokines-treated ZIP cells. In addition, mitochondria isolated from MIN6 cells showed enriched miR-146a-5p under cytokine treatment, suggesting inflammation-mediated enrichment of miR-146a-5p. Finally, RNA sequencing analysis of OE and ZIP cells identified the RNA transcripts regulated by miR-146a-5p, including MAPT, MYRIP, NRG1, and GPX1; pathway enrichment analysis showed pathways related to insulin secretion, apoptosis, and mitochondrial function. Taken together, findings from our study show for the first time that miR-146a-5p upregulation promotes β cell dysfunction and death by suppressing mitochondrial function during T1D pathogenesis. Disclosure P.Krishnan: None. R.C.S.Branco: None. S.A.Weaver: Employee; Eli Lilly and Company. C.Lee: None. F.Syed: None. C.Evans-molina: Advisory Panel; Provention Bio, Inc., DiogenX, Avotres Inc., Neurodon, MaiCell Therapeutics, Other Relationship; Isla Technology, Bristol-Myers Squibb Company, Nimbus Therapeutics, Research Support; Lilly, Astellas Pharma Inc. Funding JDRF (2-SRA-2019-834-S-B, 2-SRA-2018-493-A-B); National Institutes of Health (DK127308, U01DK127786, UC4DK104166)

ATF5 is a regulator of ER stress and β-cell apoptosis in different mouse models of genetic- and diet-induced obesity and diabetes mellitus

Endoplasmic reticulum (ER) stress is closely associated with type 2 diabetes (T2D). Activating transcription factor 5 (ATF5) is a member of the ATF/cAMP response element binding protein (CREB) family whose levels are increased upon stress in pancreatic islets from mice. Intriguingly, ATF5 deficiency has been shown to contribute to increased ER stress and apoptosis in mouse islet micro-organs. We hypothesized that either deficiency or overexpression of ATF5 is equally deleterious for pancreatic islets in terms of ER stress and apoptosis. To test this, we used a number of in vitro and in vivo models whereby ATF5 levels were overexpressed. We also determined the regulation of ATF5 in the context of metabolic derangements by using various mouse models of obesity and T2D. Our in vitro results show that ATF5 overexpression promoted palmitic acid (PA)-induced lipotoxic apoptosis. In vivo, global ATF5 overexpression in mice was lethal and pancreas-specific ATF5 overexpressing mice exhibit increased β-cell apoptosis. Interestingly, ATF5 is downregulated in all mouse models of severe obesity and T2D used in the current study. In conclusion, a tight control on ATF5 levels might be considered when developing novel agents targeting ATF5 for prevention and treatment of metabolic diseases.

Carbonyl Posttranslational Modification Associated With Early-Onset Type 1 Diabetes Autoimmunity.

Inflammation and oxidative stress in pancreatic islets amplify the appearance of various posttranslational modifications to self-proteins. In this study, we identified a select group of carbonylated islet proteins arising before the onset of hyperglycemia in NOD mice. Of interest, we identified carbonyl modification of the prolyl-4-hydroxylase β subunit (P4Hb) that is responsible for proinsulin folding and trafficking as an autoantigen in both human and murine type 1 diabetes. We found that carbonylated P4Hb is amplified in stressed islets coincident with decreased glucose-stimulated insulin secretion and altered proinsulin-to-insulin ratios. Autoantibodies against P4Hb were detected in prediabetic NOD mice and in early human type 1 diabetes prior to the onset of anti-insulin autoimmunity. Moreover, we identify autoreactive CD4+ T-cell responses toward carbonyl-P4Hb epitopes in the circulation of patients with type 1 diabetes. Our studies provide mechanistic insight into the pathways of proinsulin metabolism and in creating autoantigenic forms of insulin in type 1 diabetes.

Antioxidant Effect of Tyr-Ala Extracted from Zein on INS-1 Cells and Type 2 Diabetes High-Fat-Diet-Induced Mice.

Type 2 diabetes mellitus (T2DM) is associated with an oxidative milieu that often leads to adverse health problems. Bioactive peptides of zein possess outstanding antioxidant activity; however, their effects on hyperglycemia-related oxidative stress remain elusive. In the present study, the dipeptide Tyr-Ala (YA), a functional peptide with typical health benefits, was applied to alleviate oxidative stress in pancreatic islets under hyperglycemic conditions. By detecting viability, antioxidant ability, and insulin secretion in INS-1 cells, YA showed excellent protection of INS-1 cells from H2O2 oxidative stress, erasing reactive oxygen species (ROS) and promoting insulin secretion. Moreover, by Western blotting, we found that YA can regulate the PI3K/Akt signaling pathway associated with glycometabolism. After establishing a T2DM mice model, we treated mice with YA and measured glucose, insulin, hemoglobin A1C (HbA1c), total cholesterol (TC), triglyceride (TG), and malonaldehyde (MDA) levels and activities of superoxide dismutase (SOD) and glutathione (GSH) from blood samples. We observed that YA could reduce the production of glucose, insulin, HbA1c, TC, TG, and MDA, in addition to enhancing the activities of SOD and GSH. YA could also repair the function of the kidneys and pancreas of T2DM mice. Along with the decline in fasting blood glucose, the oxidative stress in islets was alleviated in T2DM mice after YA administration. This may improve the health situation of diabetic patients in the future.

Regulation of metabolism by protein modifications in autoimmunity

Abstract In type 1 diabetes (T1D), inflammation and oxidative stress in pancreatic islets amplifies various post-translational modifications (PTMs) of self-proteins. The loss of immune tolerance to islet PTMs triggers an autoreactive T cell response and contributes to the destruction of insulin-producing beta cells in T1D. The major function of beta cells is to secret insulin in response to glucose uptake in order to maintain glucose homeostasis. In this study, we identify autoantibodies against to beta subunit of prolyl-4-hydroxylase (P4Hb; native and carbonylated form) and glucokinase (GK; native and citrullinated form) in both human T1D and murine models. Glucokinase serves as a glucose sensor to initiate glycolysis and insulin signaling in beta cells. P4Hb is required for the accurate folding of insulin. By mass spectrometry, six carbonyl residues and eleven citrulline residues were mapped in oxidative rhP4Hb and PAD-treated rhGK, respectively. Of interest, autoreactive CD4+ T cells to citrullinated GK epitopes are present in the circulation of T1D patients. In regards to glucose metabolism, the carbonylated-P4Hb is amplified in stressed human islets coincident with decreased glucose-stimulated insulin secretion and disturb (increase) proinsulin to insulin ratios. The citrullination alters GK biologic activity (Km) and suppresses glucose-stimulated insulin secretion. Moreover, PAD2/4 inhibitors can partially restore IFNg +IL-1b suppressed glucose stimulated insulin secretion in INS-1E beta cells. Our studies implicate the crucial enzymes, glucokinase and P4Hb, as the biomarkers, providing new insights into creating autoantigens and define the impact of metabolic PTMs on the aberrant beta cell functions of T1D. This research was supported by the Juvenile Diabetes Research Foundation (Innovative Grant to M.J.M., 2-SRA-2018-551-S-B to E.A.J.), and National Institutes of Health (DK104205-01 to K.H. and M.J.M.).

Development and Application of a Semi quantitative Scoring Method for Ultrastructural Assessment of Acute Stress in Pancreatic Islets.

Background.Pancreas and islet transplantation outcomes are negatively impacted by injury to the endocrine cells from acute stress during donor death, organ procurement, processing, and transplant procedures. Here, we report a novel electron microscopy scoring system, the Newcastle Pancreas Endocrine Stress Score (NPESS).Methods.NPESS was adapted and expanded from our previously validated method for scoring pancreatic exocrine acinar cells, yielding a 4-point scale (0–3) classifying ultrastructural pathology in endocrine cell nuclei, mitochondria, endoplasmic reticulum, cytoplasmic vacuolization, and secretory granule depletion, with a maximum additive score of 15. We applied NPESS in a cohort of deceased organ donors after brainstem (DBD) and circulatory (DCD) death with a wide range of cold ischemic times (3.6–35.9 h) including 3 donors with type 1 and 3 with type 2 diabetes to assess islets in situ (n = 30) in addition to pancreata (n = 3) pre- and postislet isolation.Results.In DBD pancreata, NPESS correlated with cold ischemic time (head: r = 0.55; P = 0.02) and mirrored exocrine score (r = 0.48; P = 0.01). When stratified by endocrine phenotype, cells with granules of heterogeneous morphology had higher scores than α, β, and δ cells (P < 0.0001). Cells of mixed endocrine-exocrine morphology were observed in association with increased NPESS (P = 0.02). Islet isolation was associated with improved NPESS (in situ: 8.39 ± 0.77 [Mean ± SD]; postisolation: 5.44 ± 0.31; P = 0.04).Conclusions.NPESS provides a robust method for semiquantitative scoring of subcellular ultrastructural changes in human pancreatic endocrine cells in situ and following islet isolation with utility for unbiased evaluation of acute stress in organ transplantation research.

Mitochondria play a key role in oxidative stress-induced pancreatic islet dysfunction after severe burns.

Severe burns are often complicated with hyperglycemia in part caused by pancreatic islet dysfunction. Previous studies have revealed that in diabetes mellitus, the pancreatic islet dysfunction is partly attributed to oxidative stress. However, the role and mechanism of oxidative stress in hyperglycemia after severe burns remain unclear. Therefore, the purpose of this study was to explore the level and mechanism of oxidative stress in pancreatic islets after severe burns and the antioxidant effect of sodium pyruvate. A 30% total body surface area full-thickness burn model was established using male C57BL/6 mice. Fasting blood glucose and glucose-stimulated insulin secretion (GSIS) 24 hours post severe burns were detected. The levels of reactive oxygen species (ROS) and mitochondrial ROS of islets were detected. The activities of complexes in the mitochondrial respiratory chain of islets were measured. The main antioxidant defense system, glutaredoxin system, and thioredoxin system-related indexes were detected, and the expression of manganese superoxide dismutase (Mn-SOD) was measured. In addition, the antioxidant activity of sodium pyruvate was evaluated post severe burns. After severe burns, fasting blood glucose levels increased, while GSIS levels decreased, with significantly elevated ROS levels of pancreatic islets. The activity of complex III decreased and the level of mitochondrial ROS increased significantly post severe burns. For the detoxification of ROS, the expressions of thioredoxin 2, thioredoxin reductase 2, and Mn-SOD located in mitochondria decreased. Sodium pyruvate reduced the level of mitochondrial ROS in islet cells and improved the GSIS of islets after severe burns. The high level of mitochondrial ROS of islets is caused by reducing the activity of complex III in mitochondrial respiratory chain, inhibiting mitochondrial thioredoxin system, and downregulating Mn-SOD post severe burns. Sodium pyruvate plays an antioxidant role post severe burns in mice islets and improves the islet function.

Glucose metabolism and pyruvate carboxylase enhance glutathione synthesis and restrict oxidative stress in pancreatic islets.

Glucose metabolism modulates the islet β cell responses to diabetogenic stress, including inflammation. Here, we probed the metabolic mechanisms that underlie the protective effect of glucose in inflammation by interrogating the metabolite profiles of primary islets from human donors and identified de novo glutathione synthesis as a prominent glucose-driven pro-survival pathway. We find that pyruvate carboxylase is required for glutathione synthesis in islets and promotes their antioxidant capacity to counter inflammation and nitrosative stress. Loss- and gain-of-function studies indicate that pyruvate carboxylase is necessary and sufficient to mediate the metabolic input from glucose into glutathione synthesis and the oxidative stress response. Altered redox metabolism and cellular capacity to replenish glutathione pools are relevant in multiple pathologies beyond obesity and diabetes. Our findings reveal a direct interplay between glucose metabolism and glutathione biosynthesis via pyruvate carboxylase. This metabolic axis may also have implications in other settings where sustaining glutathione is essential.

Caffeic acid protects mice pancreatic islets from oxidative stress induced by multi-walled carbon nanotubes (MWCNTs).

Increasing applications of carbon nanotubes (CNTs) indicate the necessity to examine their toxicity. According to previous studies, CNTs caused oxidative stress that impaired β-cell functions and reduced insulin secretion. Our previous study indicated that single-walled carbon nanotubes (SWCNTs) could induce oxidative stress in pancreatic islets. However, there is no study on the effects of multi-walled carbon nanotubes (MWCNTs) on islets and β-cells. Therefore, the present study aims to evaluate effects of MWCNTs on the oxidative stress of islets and the protective effects of caffeic acid (CA) as an antioxidant. The effects of MWCNTs and CA on islets were investigated using MTT assay, reactive oxygen species (ROS), malondialdehyde (MDA), activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), the content of glutathione (GSH) and mitochondrial membrane potential (MMP) and insulin secretion measurements. The lower viability of islet cells was dose-dependent due to the exposure to MWCNTs according to the MTT assay. Further studies revealed that MWCNTs decreased insulin secretion and MMP, induced ROS creation, increased the MDA level, and decreased activities of SOD, GSH-Px, CAT, and content of GSH. Furthermore, the pretreatment of islets with CA returned the changes. These findings indicated that MWCNTs might induce the oxidative stress of pancreatic islets occurring diabetes and protective CA effects that were mediated by the augmentation of the antioxidant defense system of islets. Our research suggested the necessity of conducting further studies on effects of MWCNTs and CA on the diabetes.

Berberine Reshapes the Balance of the Local Renin-Angiotensin System by Modulating Autophagy under Metabolic Stress in Pancreatic Islets.

Results Prolonged exposure to palmitate increased the expression of ACE and AngII type 1 receptor (ATR1) and decreased the ACE2 expression, which was partly offset by berberine. In ob/ob mice, berberine increased in tolerance to glucose, improved abnormal β-cell and α-cell distributions, upregulated ACE2 expression, and decreased autophagosomes and the expression of LC3 and SQSTM1/p62. Autophagosomes and expression of LC3 and SQSTM1/p62 were increased in ACE2KO mice. Conclusions We demonstrated that berberine may improve the pancreatic islet function by regulating local RAS-mediated autophagy under metabolic stress.

Characterization of Apoptosis, Autophagy and Oxidative Stress in Pancreatic Islets Cells and Intestinal Epithelial Cells Isolated from Equine Metabolic Syndrome (EMS) Horses.

Endocrine disorders are becoming an increasing problem in both human and veterinary medicine. In recent years, more and more horses worldwide have been suffering from equine metabolic syndrome (EMS). This metabolic disorder is characterized by pathological obesity, hyperinsulinaemia, hyperglycaemia and insulin resistance. Although metabolic disorders, including diabetes, have been extensively studied, there are still no data on the molecular effects of EMS in horses. Thus, the aim of this study was to evaluate apoptosis, oxidative stress, autophagy and microRNA (miR) expression in multipotent intestinal epithelial stem cells (IECs) and pancreatic islets (PIs) isolated post mortem form healthy and EMS diagnosed horses. Our group was the first to describe how EMS affects IEC and PI aging and senescence. First, we evaluated isolation and culture protocol for these cells and subsequently established their metabolic status in vitro. Both IECs and PIs isolated from EMS horses were characterized by increased apoptosis and senescence. Moreover, they accumulated elevated levels of reactive oxygen species (ROS). Here we have observed that autophagy/mitophagy may be a protective mechanism which allows those cells to maintain their physiological function, clear protein aggregates and remove damaged organelles. Furthermore, it may play a crucial role in reducing endoplasmic reticulum (ER) stress. This protective mechanism may help to overcome the harmful effects of ROS and provide building blocks for protein and ATP synthesis.

Effect of Seleno-L-methionine on Oxidative Stress in the Pancreatic Islets of a Short-Term Induced Diabetic Mouse Model in Insufficient Selenium Status.

The protective effects of seleno-L-methionine (SeMet) on oxidative stress in pancreatic islets were investigated with a short-term nicotinamide (NA) and streptozotocin (STZ)-induced diabetic mouse model. ICR mice were intraperitoneally injected twice with 100 mg/kg STZ and 120 mg/kg NA at a 1-d interval and were then orally administered 158 µg Se/kg SeMet with free access to a selenium-deficient diet for 5 weeks. Administration of SeMet significantly improved the levels of glycated hemoglobin (HbA1c), non-fasting and oral glucose tolerance-tested (OGTT) blood glucose, plasma adiponectin and hepatic glycogen that deteriorated by NA/STZ treatment. However, supplementary SeMet did not restore non-fasting plasma insulin levels in NA/STZ treatment group and significantly suppressed OGTT plasma insulin levels in the control group. Although SeMet significantly suppressed 8-hydroxy-2'-deoxyguanosine density in pancreatic islets, SeMet did not restore insulin density. The hepatic and pancreatic mRNA levels of glutathione peroxidase 1 (GPX1) increased by NA/STZ treatment or SeMet administration. These results suggest that although a physiological level of SeMet improves glucose tolerance by exhibiting insulin-mimetic activity in a short-term induced diabetic mouse model under insufficient Se status, the suppression of pancreatic oxidative stress with the induction GPX1 by SeMet supplementation is unlikely to restore insulin storage and secretion.

Oxidative stress increases the risk of pancreatic β cell damage in chronic renal hypertensive rats.

Hypertension often occurs in conjunction with insulin resistance. The purpose of this study was to evaluate whether sustained renal hypertension increases the risk of diabetes mellitus in rats, and to define the underlying mechanisms. Two‐kidney, one‐clip hypertensive (2K1C) rats received captopril (50 mg/kg/day), α‐lipoic acid (100 mg/kg/day), or vehicle treatment for 3 months after surgery. Blood pressure was measured by tail cuff plethysmography. Oral glucose tolerance test (OGTT), immunohistochemistry, and western blotting were performed. In addition, insulin secretion from islet cells was measured. OGTT yielded abnormal results, and the number of islet cells and the size of pancreatic β/α cells were decreased in 2K1C rats. Basal insulin levels were also reduced in the plasma. Insulin secretion from pancreatic islet cells in response to high glucose was also attenuated in 2K1C rats compared with sham rats. The levels of oxidative stress markers, including 8‐hydroxydeoxyguanosine and NADPH oxidase‐4, were increased in pancreatic tissue and pancreatic islets in 2K1C rats. The abnormalities observed in 2K1C rats were improved by captopril or α‐lipoic acid treatment. These findings indicate that sustained renal hypertension may lead to pancreatic dysfunction, increasing oxidative stress in pancreatic islets.

Palmitic acid-rich diet suppresses glucose-stimulated insulin secretion (GSIS) and induces endoplasmic reticulum (ER) stress in pancreatic islets in mice

The objective was to clarify whether dietary palmitic acid supplementation affects glucose-stimulated insulin secretion (GSIS) and the endoplasmic reticulum (ER) stress pathway in pancreatic islets in mice. Eight-week-old male C57BL/6J mice were randomly divided into three treatment diet groups: control diet, palmitic acid-supplemented diet (PAL) and oleic acid-supplemented diet (OLE). After 2 weeks of treatment, intraperitoneal glucose tolerance test and intraperitoneal insulin tolerance test were performed. GSIS was assessed by pancreatic perfusion in situ with basal (100 mg/dL) glucose followed by a high (300 mg/dL) glucose concentration. We measured mRNA levels of ER stress markers such as C/EBP homologous protein (CHOP), immunoglobulin heavy-chain binding protein (BIP) and X-box binding protein (XBP)-1 using real-time polymerase chain reaction (PCR) analyses in isolated islets. Immunohistochemical staining was also performed. Mice fed PAL showed significantly decreased glucose tolerance (p < 0.05). In the perfusion study, GSIS was significantly suppressed in the PAL group (p < 0.05). Semi-quantitative RT-PCR revealed that islet CHOP, BIP, and XBP-1 mRNA expression were significantly increased in the PAL group (p < 0.05). TUNEL-positive β-cells were not detected in all groups. Dietary palmitic acid-supplementation for 2 weeks might suppress GSIS and induce ER stress in pancreatic islets in mice, in the early stage of lipotoxicity.

12-lipoxygenase promotes obesity-induced oxidative stress in pancreatic islets.

High-fat diets lead to obesity, inflammation, and dysglycemia. 12-Lipoxygenase (12-LO) is activated by high-fat diets and catalyzes the oxygenation of cellular arachidonic acid to form proinflammatory intermediates. We hypothesized that 12-LO in the pancreatic islet is sufficient to cause dysglycemia in the setting of high-fat feeding. To test this, we generated pancreas-specific 12-LO knockout mice and studied their metabolic and molecular adaptations to high-fat diets. Whereas knockout mice and control littermates displayed identical weight gain, body fat distribution, and macrophage infiltration into fat, knockout mice exhibited greater adaptive islet hyperplasia, improved insulin secretion, and complete protection from dysglycemia. At the molecular level, 12-LO deletion resulted in increases in islet antioxidant enzymes Sod1 and Gpx1 in response to high-fat feeding. The absence or inhibition of 12-LO led to increases in nuclear Nrf2, a transcription factor responsible for activation of genes encoding antioxidant enzymes. Our data reveal a novel pathway in which islet 12-LO suppresses antioxidant enzymes and prevents the adaptive islet responses in the setting of high-fat diets.

The proapoptotic BH3-only proteins Bim and Puma are downstream of endoplasmic reticulum and mitochondrial oxidative stress in pancreatic islets in response to glucotoxicity

Apoptosis of pancreatic beta cells is a feature of type 2 diabetes and its prevention may have therapeutic benefit. High glucose concentrations induce apoptosis of islet cells, and this requires the proapoptotic Bcl-2 homology domain 3 (BH3)-only proteins Bim and Puma. We studied the stress pathways induced by glucotoxicity in beta cells that result in apoptosis. High concentrations of glucose or ribose increased expression of the transcription factor CHOP (C/EBP homologous protein) but not endoplasmic reticulum (ER) chaperones, indicating activation of proapoptotic ER stress signaling. Inhibition of ER stress prevented ribose-induced upregulation of Chop and Puma mRNA, and partially protected islets from glucotoxicity. Loss of Bim or Puma partially protected islets from the canonical ER stressor thapsigargin. The antioxidant N-acetyl-cysteine also partially protected islets from glucotoxicity. Islets deficient in both Bim and Puma, but not Bim or Puma alone, were significantly protected from killing induced by the mitochondrial reactive oxygen species donor rotenone. Our data demonstrate that high concentrations of glucose induce ER and oxidative stress, which causes cell death mediated by Bim and Puma. We observed significantly higher Bim and Puma mRNA in islets of human donors with type 2 diabetes. This indicates that inhibition of Bim and Puma, or their inducers, may prevent beta-cell destruction in type 2 diabetes.

Angiotensin II type 1 receptor antagonists prevent glucose‐induced increases in islet blood flow in rats

Objective. Renin‐angiotensin system inhibitors are reported to be beneficial in delaying the onset of diabetes mellitus. Since islet blood hyperperfusion during hyperglycaemia may be detrimental to endothelium in pancreatic islets and eventually lead to β‐cell dysfunction, we studied acute and chronic effects of angiotensin II type 1 receptor blockers (ARB) on islet blood flow before and after glucose load. Material and methods. Islet blood flow was measured using the colour microsphere method in anaesthetized Sprague‐Dawley rats before and 3 min after glucose injection. Results. Olmesartan significantly reduced blood pressure, but did not affect islet blood flow 10 min after its injection. However, pretreatment with olmesartan blunted the glucose‐induced increase in islet blood flow (62 % of control). In rats treated with olmesartan or candesartan for 4 weeks, islet blood flow was not different from untreated control, whereas the glucose‐induced increase in islet blood flow was significantly suppressed in chronically ARB‐treated rats (olmesartan 59 % of control, candesartan 64 % of control, respectively). Acute or chronic treatment with ARB did not change insulin secretion before and in response to glucose load. Pancreatic or duodenal blood flow was not affected by ARB treatment, although acute olmesartan administration reduced pancreatic blood flow after glucose load. Conclusion. ARB appears to suppress the hyperglycaemia‐induced islet hyperperfusion, which may ameliorate haemodynamic stress in pancreatic islets.

Calcineurin activity as an indicator of oxidative stress in normal islet cells and insulinoma cells

A comparative study was conducted to evaluate calcineurin activity in normal pancreatic beta cells and insulinoma cells in relation to their oxidative state. In comparison to normal islets, insulinoma cells had enhanced oxidative stress as evidenced by increased content of thiobarbituric acid reactive substances. In addition, diminished activity of calcineurin in insulinoma cells was concomitant with decreased content of reduced glutathione and glutathione peroxidase activity signifying diminished antioxidant status in these cells. Culturing insulinoma cells in presence of the calcineurin inhibitor cyclosporin A resulted in further decrease of calcineurin activity with restoration of glutathione peroxidase but without restoration of reduced glutathione levels. These results indicate that an estimate of oxidative stress in pancreatic islets and insulinoma cells can be obtained by assaying calcineurin activity.