Inhibition Of Insulin Secretion Research Articles

Comparative efficacy, toxicity, and insulin-suppressive effects of simvastatin and pravastatin in fatty acid-challenged mouse insulinoma MIN6 β-cell model

IntroductionFamilial hypercholesterolemia, the highly prevalent form of dyslipidemia, is a well-known risk factor for premature heart disease and stroke worldwide. Statins, which inhibit 3-hydroxy 3-methylglutaryl coenzyme A (HMG-CoA) reductase, are the first-choice treatment for dyslipidemias, and have been effective in reducing the risk of stroke and myocardial infarction. However, emerging evidence indicates that statins may increase the incidence of new-onset type 2 diabetes by reducing β-cell mass and function. Notably, past in vitro reports studying the effects of statins on β-cells were performed without including free fatty acids in the model. This factor should have been addressed since these agents are used to treat individuals with hyperlipidemia.MethodsHere, we used a mouse insulinoma MIN6 β-cell culture model to assess the efficacy, cytotoxicity, and insulin-suppressive effects of simvastatin and pravastatin in the presence of palmitic, linoleic, and oleic acids cocktail to mimic mixed lipids challenge in a biologically relevant setting.Results and discussionOur findings indicate that simvastatin was more effective in lowering intracellular cholesterol but was more cytotoxic as compared to pravastatin. Similarly, simvastatin exhibited a higher suppression of total insulin content and insulin secretion. Both drugs suppressed insulin secretion in phases 1 and 2, dose-dependently. No significant effect was observed on mitochondrial respiration. More importantly, elution experiments showed that insulin content diminution by simvastatin treatment was reversible, while exogenous mevalonate did not improve total insulin content. This suggests that simvastatin's influence on insulin content is independent of its specific inhibitory action on HMG-CoA reductase. In conclusion, our study identified that simvastatin was more effective in lowering intracellular cholesterol, albeit it was more toxic and suppressive of β-cells function. Notably, this suppression was found to be reversible.

Clinical significance of circulating long non-coding RNA SNHG1 in type 2 diabetes mellitus and its association with cell proliferation of pancreatic β-cell

BackgroundTo explore the association of long non-coding RNA (lncRNA) SNHG1/ miR-195 axis with type 2 diabetes mellitus (T2DM) and islet function.MethodsThe expression of SNHG1 and miR-195 was measured in T2DM patients and in healthy subjects. Correlation between indciators was evaluated using Pearson correlation analysis. INS-1 cells were used to perform the cell function assays. Insulin secretion by INS-1 was detected using ELISA. Cell counting kit-8 (CCK-8) and flow cytometry was used to detect cell proliferation and apoptosis. Luciferase report assay was to used to verify the target of SNHG1.ResultsThe expression of SNHG1 was increased and miR-195 level was decreased in the serum of T2DM patients. Both SNHG1 and miR-195 could be biomarkers for T2DM diagnosis. The fasting plasma glucose (FPG) and HbA1c were positively related to SNHG1 and negatively related to miR-195. SNHG1 inhibited insulin secretion, and cell proliferation and promoted apoptosis of INS-1 cells via binding to miR-195.ConclusionsDetection of SNHG1 and miR-195 might predict T2DM. SNHG1 could suppress proliferation and insulin secretion, but promote apoptosis of INS-1 cells via sponging miR-195.

6524 Sex-Specific Single Cell Genetic Architecture Of Human Islets With And Without Type 2 Diabetes

Abstract Disclosure: M. Qadir: None. R.M. Elgamal: None. P. Kudtarkar: None. K.J. Gaulton: None. F. Mauvais-Jarvis: None. Type 2 diabetes (T2D) is a heterogeneous disease and biological sex affects its pathogenesis including the development of adiposity, insulin resistance, and β cell failure. For example, ketosis-prone diabetes is a phenotypically-defined form of T2D with acute β cell failure and severe insulin deficiency, predominantly observed in men. However, what mechanisms drive male predominance in β cell failure is unknown. To fully understand the sex-based pathogenesis of islet failure in diabetes, the study of sex differences in human islet biology and dysfunction is essential. To decipher sex differences in human islets genetic architecture and function, we performed an orthogonal series of experiments from 15 islets of non-diabetic (ND) donors, using single cell RNAseq (scRNAseq) and single nucleus assay for transposase-accessible chromatin sequencing (snATACseq). We also leveraged 37 ND and T2D donors for scRNAseq data from the human pancreas analysis program (HPAP) dataset. We studied over 200,000 single cells spanning the accessible genome and transcriptome. It is the largest study of sex-based single-cell genomics and transcriptomics of human islets to date. We observe that in cultured human ND islets, in the absence of the in vivo hormonal environment, islet cell transcriptome, and genome accessibility across coding regions are predominantly limited to sex chromosome genes. Of particular interest are sex differences in gene accessibility and expression of the X-linked KDM6A and Y-linked KDM5D demethylase genes in female (F) and male (M) islet cells respectively. Furthermore, analysis of T2D islets revealed a greater number of autosomal differentially expressed genes (DEGs) in F (721 upregulated and 1164 downregulated, FDR < 0.01) compared to M β cells (111 upregulated and 99 downregulated, FDR < 0.01). M β cell DEGs over-representation analysis revealed suppressed insulin secretion and processing pathways, while F β cell DEGs exhibited suppression of oxidative phosphorylation and electron transport chain pathways. Thus, although, sex differences in gene expression and accessibility of cultured ND human islets are restricted to sex chromosome genes, during the transition from ND to T2D, major sex differences in autosomal DEGs appear, with a greater number of autosomal DEGs in F compared to M T2D islets. Presentation: 6/3/2024

6774 Recurrent Hypoglycemia In Malignant Insulinoma With Multiple Liver Metastases Successfully Managed With Octreotide

Abstract Disclosure: K. Kim: None. S. Mascarell: None. Introduction: Insulinoma is a rare functional pancreatic neuroendocrine tumor (pNET) with the incidence of 1-4/million person-years. Malignant insulinomas with locoregional invasion or metastases consist 10% among insulinomas. While the treatment of a single insulinoma is surgical resection, treatment of malignant insulinoma requires a comprehensive approach. Due to the low prevalence of the disease, data on therapeutic modalities of malignant insulinomas are limited. We hereby present a case of malignant insulinoma in which recurrent hypoglycemia was successfully managed with octreotide. Case: A 36 year old female diagnosed with grade III well differentiated pNET with multiple metastases to the liver and abdominal lymph nodes 1 year ago was admitted for recurrent hypoglycemic episodes. She reported progressively worsening hypoglycemia for the past 1 month. She was eating every 2 hours including her sleeping hours to prevent hypoglycemia. Due to recurrent hypoglycemia she required hospitalization with IV dextrose. After 13 hours of fasting in the ICU, hypoglycemia due to insulin hypersecretion was confirmed with blood glucose 51 mg/dL and elevated serum insulin 51.11 uIU/mL (1.9-23uIU/mL). She underwent the short octreotide test which showed suppressed insulin (5.25 uIU/mL) 6 hours after receiving 100 mcg of octreotide. She was started on octreotide 50 mcg every 12 hours and monitored for further hypoglycemic episodes. She tolerated octreotide without any side effects including hypoglycemia. She was discharged home with a continuous glucose monitor (CGM) and transitioned to intramuscular lanreotide 20mg injections every 4 weeks as an outpatient. No further hypoglycemic episodes occurred. Discussion: Few cases report treatment of hypoglycemia in malignant insulinoma with octreotide. Octreotide has the dual effect of suppressing insulin secretion and tumor growth on insulinomas that express SSTR2 receptors. However, because somatostatin agonists also inhibit the secretion of glucagon, in insulinomas without SSTR2 expression, octreotide can aggravate hypoglycemia. This was especially a concern for our patient with a large tumor burden. Thus the short octreotide test was conducted. We suggest that the octreotide test of 100 mcg SQ octreotide injection followed by hourly measurements of glucose and insulin levels is a useful and convenient test to predict efficacy while monitoring for octreotide induced hypoglycemia. Finally the use of CGM in the outpatient setting enabled successful transition to octreotide LAR which improved the patient’s quality of life greatly by enabling once a month injections. Presentation: 6/3/2024

Anekomochi glutinous rice provides low postprandial glycemic response by enhanced insulin action via GLP-1 release and vagal afferents activation

Glutinous rice (mochi rice), compared to non-glutinous rice (uruchi rice), exhibits a wide range of glycemic index (GI) values, from low to high. However, the underlying mechanisms behind the variation in GI values remain poorly understood. In this study, we aimed to identify rice cultivars with a low postprandial glycemic response and investigate the mechanisms, focusing on insulin and incretin hormones. We examined seven glutinous rice cultivars and three non-glutinous rice cultivars. We discovered that Anekomochi, a glutinous rice cultivar, has the lowest postprandial glycemic response. Anekomochi significantly enhanced glucagon-like peptide-1 (GLP-1) secretion while suppressing insulin secretion. These effects were completely blunted by inhibiting GLP-1 receptor signaling and denervating the common hepatic branch of vagal afferent nerves that are crucial for sensing intestinal GLP-1. Our findings demonstrate that Anekomochi markedly enhances insulin action via GLP-1 release and vagal afferent neural pathways, thereby leading to a lower postprandial glycemic response.

NMDA Suppresses Pancreatic ABCA1 Expression through the MEK/ERK/LXR Pathway in Pancreatic Beta Cells.

Dysfunction or loss of pancreatic β cells can cause insulin deficiency and impaired glucose regulation, resulting in conditions like type 2 diabetes. The ATP-binding cassette transporter A1 (ABCA1) plays a key role in the reverse cholesterol transport system, and its decreased expression is associated with pancreatic β cell lipotoxicity, resulting in abnormal insulin synthesis and secretion. Increased glutamate release can cause glucotoxicity in β cells, though the detailed mechanisms remain unclear. This study investigated the effect of N-methyl-D-aspartic acid (NMDA) on ABCA1 expression in INS-1 cells and primary pancreatic islets to elucidate the signaling mechanisms that suppress insulin secretion. Using Western blotting, microscopy, and biochemical analyses, we found that NMDA activated the mitogen-activated protein kinase (MEK)-dependent pathway, suppressing ABCA1 protein and mRNA expression. The MEK-specific inhibitor PD98059 restored ABCA1 promoter activity, indicating the involvement of the extracellular signal-regulated kinase (MEK/ERK) pathway. Furthermore, we identified the liver X receptor (LXR) as an effector transcription factor in NMDA regulation of ABCA1 transcription. NMDA treatment increased cholesterol and triglyceride levels while decreasing insulin secretion, even under high-glucose conditions. These effects were abrogated by treatment with PD98059. This study reveals that NMDA suppresses ABCA1 expression via the MEK/ERK/LXR pathway, providing new insights into the pathological suppression of insulin secretion in pancreatic β cells and emphasizing the importance of investigating the role of NMDA in β cell dysfunction.

Identification of small molecule glucokinase activators for the treatment of diabetes based on plants from the traditional Chinese medicine: In silico analysis

Mutations in glucokinase (GCK) can either enhance or inhibit insulin secretion, leading to different forms of diabetes, including gestational diabetes. While many glucokinase activators (GKAs) have been explored as treatments, their long-term effectiveness has often been unsatisfactory. However, recent interest has surged with the introduction of dorzagliatin and TTP399. This study investigates the efficacy of four previously studied compounds (Swertiamarin, Apigenin, Mangiferin, and Tatanan A) in activating GCK using computational methods.Initial molecular docking revealed binding affinities ranging from −6.7 to −8.6 kcal/mol. The compounds were then evaluated for drug-likeness and pharmacokinetic properties. Re-docking studies were performed for validation. Based on their favorable binding affinities and compliance with Lipinski's rule and ADMET criteria, three compounds (Swertiamarin, Apigenin, and Tatanan A) were selected for molecular dynamics (MD) simulations.MD simulations demonstrated that Swertiamarin showed excellent stability, as indicated by analyses of RMSD, RMSF, radius of gyration (Rg), hydrogen bonding, and principal component analysis (PCA). These results suggest that Swertiamarin holds promise for further investigation in in vivo and clinical settings to evaluate its potential in enhancing GCK activity and treating diabetes.This study assessed the potential of four compounds as GCK activators using molecular docking, pharmacokinetic profiling, and MD simulations. Swertiamarin, in particular, showed significant stability and adherence to drug-likeness criteria, making it a promising candidate for further research in combating diabetes.

Postprandial exercise regulates tissue-specific triglyceride uptake through angiopoietin-like proteins.

Fuel substrate switching between carbohydrates and fat is essential for maintaining metabolic homeostasis. During aerobic exercise, the predominant energy source gradually shifts from carbohydrates to fat. While it is well known that exercise mobilizes fat storage from adipose tissues, it remains largely obscure how circulating lipids are distributed tissue-specifically according to distinct energy requirements. Here, we demonstrate that aerobic exercise is linked to nutrient availability to regulate tissue-specific activities of lipoprotein lipase (LPL), the key enzyme catabolizing circulating triglyceride (TG) for tissue uptake, through the differential actions of angiopoietin-like (ANGPTL) proteins. Exercise reduced the tissue binding of ANGPTL3 protein, increasing LPL activity and TG uptake in the heart and skeletal muscle in the postprandial state specifically. Mechanistically, exercise suppressed insulin secretion, attenuating hepatic Angptl8 transcription through the PI3K/mTOR/CEBPα pathway, which is imperative for the tissue binding of its partner ANGPTL3. Constitutive expression of ANGPTL8 hampered lipid utilization and resulted in cardiac dysfunction in response to exercise. Conversely, exercise promoted the expression of ANGPTL4 in white adipose tissues, overriding the regulatory actions of ANGPTL8/ANGPTL3 in suppressing adipose LPL activity, thereby diverting circulating TG away from storage. Collectively, our findings show an overlooked bifurcated ANGPTL-LPL network that orchestrates fuel switching in response to aerobic exercise.

Directed differentiation of pancreatic δ cells from human pluripotent stem cells

Dysfunction of pancreatic δ cells contributes to the etiology of diabetes. Despite their important role, human δ cells are scarce, limiting physiological studies and drug discovery targeting δ cells. To date, no directed δ-cell differentiation method has been established. Here, we demonstrate that fibroblast growth factor (FGF) 7 promotes pancreatic endoderm/progenitor differentiation, whereas FGF2 biases cells towards the pancreatic δ-cell lineage via FGF receptor 1. We develop a differentiation method to generate δ cells from human stem cells by combining FGF2 with FGF7, which synergistically directs pancreatic lineage differentiation and modulates the expression of transcription factors and SST activators during endoderm/endocrine precursor induction. These δ cells display mature RNA profiles and fine secretory granules, secrete somatostatin in response to various stimuli, and suppress insulin secretion from in vitro co-cultured β cells and mouse β cells upon transplantation. The generation of human pancreatic δ cells from stem cells in vitro would provide an unprecedented cell source for drug discovery and cell transplantation studies in diabetes.

Inhibition of dynamin-related protein 1-filamin interaction improves systemic glucose metabolism.

Maintaining mitochondrial quality is attracting attention as a new strategy to treat diabetes and diabetic complications. We previously reported that mitochondrial hyperfission by forming a protein complex between dynamin-related protein (Drp) 1 and filamin, mediates chronic heart failure and cilnidipine, initially developed as an L/N-type Ca2+ channel blocker, improves heart failure by inhibiting Drp1-filamin protein complex. We investigated whether cilnidipine improves hyperglycaemia of various diabetic mice models. Retrospective analysis focusing on haemoglobin A1c (HbA1c) was performed in hypertensive and hyperglycaemic patients taking cilnidipine and amlodipine. After developing diabetic mice by streptozotocin (STZ) treatment, an osmotic pump including drug was implanted intraperitoneally, followed by weekly measurements of blood glucose levels. Mitochondrial morphology was analysed by electron microscopy. A Ca2+ channel-insensitive cilnidipine derivative (1,4-dihydropyridine [DHP]) was synthesized and its pharmacological effect was evaluated using obese (ob/ob) mice fed with high-fat diet (HFD). In patients, cilnidipine was superior to amlodipine in HbA1c lowering effect. Cilnidipine treatment improved systemic hyperglycaemia and mitochondrial morphological abnormalities in STZ-exposed mice, without lowering blood pressure. Cilnidipine failed to improve hyperglycaemia of ob/ob mice, with suppressing insulin secretion. 1,4-DHP improved hyperglycaemia and mitochondria abnormality in ob/ob mice fed HFD. 1,4-DHP and cilnidipine improved basal oxygen consumption rate of HepG2 cells cultured under 25 mM glucose. Inhibition of Drp1-filamin protein complex formation becomes a new strategy for type 2 diabetes treatment.

Single cell regulatory architecture of human pancreatic islets suggests sex differences in β cell function and the pathogenesis of type 2 diabetes.

Type 2 and type 1 diabetes (T2D, T1D) exhibit sex differences in insulin secretion, the mechanisms of which are unknown. We examined sex differences in human pancreatic islets from 52 donors with and without T2D combining single cell RNA-seq (scRNA-seq), single nucleus assay for transposase-accessible chromatin sequencing (snATAC-seq), hormone secretion, and bioenergetics. In nondiabetic (ND) donors, sex differences in islet cells gene accessibility and expression predominantly involved sex chromosomes. Islets from T2D donors exhibited similar sex differences in sex chromosomes differentially expressed genes (DEGs), but also exhibited sex differences in autosomal genes. Comparing β cells from T2D vs. ND donors, gene enrichment of female β cells showed suppression in mitochondrial respiration, while male β cells exhibited suppressed insulin secretion. Thus, although sex differences in gene accessibility and expression of ND β cells predominantly affect sex chromosomes, the transition to T2D reveals sex differences in autosomes highlighting mitochondrial failure in females.

Chronic Epinephrine-Induced Endoplasmic Reticulum and Oxidative Stress Impairs Pancreatic β-Cells Function and Fate.

Epinephrine influences the function of pancreatic β-cells, primarily through the α2A-adrenergic receptor (α2A-AR) on their plasma membrane. Previous studies indicate that epinephrine transiently suppresses insulin secretion, whereas prolonged exposure induces its compensatory secretion. Nonetheless, the impact of epinephrine-induced α2A-AR signaling on the survival and function of pancreatic β-cells, particularly the impact of reprogramming after their removal from sustained epinephrine stimulation, remains elusive. In the present study, we applied MIN6, a murine insulinoma cell line, with 3 days of high concentration epinephrine incubation and 2 days of standard incubation, explored cell function and activity, and analyzed relevant regulatory pathways. The results showed that chronic epinephrine incubation led to the desensitization of α2A-AR and enhanced insulin secretion. An increased number of docked insulin granules and impaired Syntaxin-2 was found after chronic epinephrine exposure. Growth curve and cell cycle analyses showed the inhibition of cell proliferation. Transcriptome analysis showed the occurrence of endoplasmic reticulum stress (ER stress) and oxidative stress, such as the presence of BiP, CHOP, IRE1, ATF4, and XBP, affecting cellular endoplasmic reticulum function and survival, along with UCP2, OPA1, PINK, and PRKN, associated with mitochondrial dysfunction. Consequently, we conclude that chronic exposure to epinephrine induces α2A-AR desensitization and leads to ER and oxidative stress, impairing protein processing and mitochondrial function, leading to modified pancreatic β-cell secretory function and cell fate.

Drosophila melanogaster Limostatin and Its Human Ortholog Promote West Nile Virus Infection.

The arbovirus West Nile virus (WNV) is a danger to global health. Spread primarily by mosquitoes, WNV causes about 2000 cases per year in the United States. The natural mosquito immune response controls viral replication so that the host survives but can still transmit the virus. Using the genetically malleable Drosophila melanogaster model, we previously dissected innate immune pathways used to control WNV infection. Specifically, we showed that insulin/IGF-1 signaling (IIS) activates a JAK/STAT-mediated immune response that reduces WNV. However, how factors that regulate IIS in insects control infection has not been identified. D. melanogaster Limostatin (Lst) encodes a peptide hormone that suppresses insulin secretion. Its mammalian ortholog, Neuromedin U (NMU), is a peptide that regulates the production and secretion of insulin from pancreatic beta cells. In this study, we used D. melanogaster and human cell culture models to investigate the roles of these insulin regulators in immune signaling. We found that D. melanogaster Lst mutants, which have elevated insulin-like peptide expression, are less susceptible to WNV infection. Increased levels of insulin-like peptides in these flies result in upregulated JAK/STAT activity, leading to protection from infection. Treatment of human cells with the insulin regulator NMU results in increased WNV replication. Further investigation of methods to target Lst in mosquitoes or NMU in mammals can improve vector control methods and may lead to improved therapeutics for human and animal infection.

Single cell regulatory architecture of human pancreatic islets suggests sex differences in β cell function and the pathogenesis of type 2 diabetes.

Biological sex affects the pathogenesis of type 2 and type 1 diabetes (T2D, T1D) including the development of β cell failure observed more often in males. The mechanisms that drive sex differences in β cell failure is unknown. Studying sex differences in islet regulation and function represent a unique avenue to understand the sex-specific heterogeneity in β cell failure in diabetes. Here, we examined sex and race differences in human pancreatic islets from up to 52 donors with and without T2D (including 37 donors from the Human Pancreas Analysis Program [HPAP] dataset) using an orthogonal series of experiments including single cell RNA-seq (scRNA-seq), single nucleus assay for transposase-accessible chromatin sequencing (snATAC-seq), dynamic hormone secretion, and bioenergetics. In cultured islets from nondiabetic (ND) donors, in the absence of the in vivo hormonal environment, sex differences in islet cell type gene accessibility and expression predominantly involved sex chromosomes. Of particular interest were sex differences in the X-linked KDM6A and Y-linked KDM5D chromatin remodelers in female and male islet cells respectively. Islets from T2D donors exhibited similar sex differences in differentially expressed genes (DEGs) from sex chromosomes. However, in contrast to islets from ND donors, islets from T2D donors exhibited major sex differences in DEGs from autosomes. Comparing β cells from T2D and ND donors revealed that females had more DEGs from autosomes compared to male β cells. Gene set enrichment analysis of female β cell DEGs showed a suppression of oxidative phosphorylation and electron transport chain pathways, while male β cell had suppressed insulin secretion pathways. Thus, although sex-specific differences in gene accessibility and expression of cultured ND human islets predominantly affect sex chromosome genes, major differences in autosomal gene expression between sexes appear during the transition to T2D and which highlight mitochondrial failure in female β cells.

Dopamine-mediated autocrine inhibition of insulin secretion

The aim of the present research was to explore the mechanisms underlying the role of dopamine in the regulation of insulin secretion in beta cells. The effect of dopamine on insulin secretion was investigated on INS 832/13 cell line upon glucose and other secretagogues stimulation. Results show that dopamine significantly inhibits insulin secretion stimulated by both glucose and other secretagogues, while it has no effect on the basal secretion. This effect requires the presence of dopamine during incubation with the various secretagogues. Both electron microscopy and immunohistochemistry indicate that in beta cells the D2 dopamine receptor is localized within the insulin granules. Blocking dopamine entry into the insulin granules by inhibiting the VMAT2 transporter with tetrabenazine causes a significant increase in ROS production. Our results confirm that dopamine plays an important role in the regulation of insulin secretion by pancreatic beta cells through a regulated and precise compartmentalization mechanisms.

A Novel Peptide COX52-69 Inhibits High Glucose-induced Insulin Secretion by Modulating BK Channel Activity.

Excessive insulin is the leading cause of metabolic syndromes besides hyperinsulinemia. Insulin-lowering therapeutic peptides have been poorly studied and warrant urgent attention. The main purpose of this study, was to introduce a novel peptide COX52-69 that was initially isolated from the porcine small intestine and possessed the ability to inhibit insulin secretion under high-glucose conditions by modulating large conductance Ca2+-activated K+ channels (BK channels) activity. Enzyme-linked immunosorbent assay results indicate that COX52-69 supressed insulin release induced by high glucose levels in pancreatic islets and animal models. Furthermore, electrophysiological data demonstrated that COX52-69 can increase BK channel currents and hyperpolarize cell membranes. Thus, cell excitability decreased, corresponding to a reduction in insulin secretion. Our study provides a novel approach to modulate high glucose-stimulated insulin secretion in patients with hyperinsulinemia.

Quercetin prevents kidney against diabetes mellitus (type 1) in rats by inhibiting TGF-β/apelin gene expression.

One of the main causes of diabetic nephropathy is oxidative stress induced by hyperglycemia. Apelin inhibits insulin secretion. Besides, renal expression of TGF-β is increased in diabetes mellitus (DM). The preventive effect of quercetin (Q) against renal functional disorders and tissue damage developed by DM in rats was assessed. Forty male Wistar rats were grouped into normal control (NC), normal + quercetin (NQ: quercetin, 50mg/kg/day by gavage), diabetic control (DC: streptozotocin, 65mg/kg, i.p.), diabetic + quercetin pretreatment (D + Qpre), and diabetic + quercetin post-treatment (D + Qpost). All samples (24-hour urine, plasma, pancreatic, and renal tissues) were obtained at the terminal of the experiment. Compared to NC and NQ groups, DM ended in elevated plasma and glucose levels, decreased plasma insulin level, kidney dysfunction, augmented levels of malondialdehyde, decreased level of reduced glutathione, reduced enzymatic activities of superoxide dismutase and catalase, elevated gene expression levels of apelin and TGF-β, also renal and pancreatic histological damages. Quercetin administration diminished entire the changes. However, the measure of improvement in the D + Qpre group was higher than that of the D + Qpost group. Quercetin prevents renal dysfunction induced by DM, which might be related to the diminution of lipid peroxidation, strengthening of antioxidant systems, and prevention of the apelin/ TGF-β signaling pathway.

Evidence of Nesfatin-1 in Mouse Intestinal Enteroendocrine Cells

Objectives: Post-prandial insulin secretion is predominantly regulated by four intestinal hormones: glucagon-like peptide 1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP), Cholecystokinin (CCK) and peptide YY (PYY). Nesfatin-1, an insulinotropic and anorexigenic secreted peptide was reported in mouse hypothalamus. Data supporting nesfatin-1 expression in intestine is minimal. Is nesfatin-1 expressed in mouse intestinal cells? Does it affect intestinal hormones known to stimulate or inhibit insulin secretion? We present STC-1 cells as viable in vitro models to study nesfatin-1’s biological action in intestine. Methods: Immunofluorescence histochemistry (IHC) of intestinal sections from male C57BL/6 mice were carried out. STC-1 cells (n=8 wells/treatment) were dose-dependently treated with nesfatin-1, followed by RT-qPCR and immunoassay (ELISA) respectively. One-Way ANOVA followed by Tukey’s multiple comparison test using GraphPad Prism software was used for data analysis. p<0.01 was considered statistically significant. Results: IHC showed NUCB2/nesfatin-1 co-localizing CCK, PYY and GLP-1 in the intestinal mucosa of mice. Static incubation of STC-1 cells with nesfatin-1 upregulated both CCK, PYY and GLP-1 mRNA expression (1 and 10nM) and secretion at 1 hr post-incubation (0.1, 1 and 10nM). Conversely, nesfatin-1 downregulated PYY mRNA expression and secretion in STC-1 cells (all doses tested). Conclusion: The current data shows cell-specific localization of nesfatin-1 in the intestinal mucosa and establishes STC-1 cells as in vitro models to study nesfatin-1. This study adds a novel function for intestinal nesfatin-1 in stimulating insulinotropic intestinal hormones (GLP-1, GIP, CCK) and inhibit PYY which is insulinostatic.

Urocortin 3 contributes to paracrine inhibition of islet alpha cells in mice.

Pancreatic alpha cell activity and glucagon secretion lower as glucose levels increase. While part of the decrease is regulated by glucose itself, paracrine signaling by their neighboring beta and delta cells also plays an important role. Somatostatin from delta cells is an important local inhibitor of alpha cells at high glucose. Additionally, urocortin 3 (UCN3) is a hormone that is co-released from beta cells with insulin and acts locally to potentiate somatostatin secretion from delta cells. UCN3 thus inhibits insulin secretion via a negative feedback loop with delta cells, but its role with respect to alpha cells and glucagon secretion is not understood. We hypothesize that the somatostatin-driven glucagon inhibition at high glucose is regulated in part by UCN3 from beta cells. Here, we use a combination of live functional Ca2+ and cAMP imaging as well as direct glucagon secretion measurement, all from alpha cells in intact mouse islets, to determine the contributions of UCN3 to alpha cell behavior. Exogenous UCN3 treatment decreased alpha cell Ca2+ and cAMP levels and inhibited glucagon release. Blocking endogenous UCN3 signaling increased alpha cell Ca2+ by 26.8 ± 7.6%, but this did not result in increased glucagon release at high glucose. Furthermore, constitutive deletion of Ucn3 did not increase Ca2+ activity or glucagon secretion relative to controls. UCN3 is thus capable of inhibiting mouse alpha cells, but, given the subtle effects of endogenous UCN3 signaling on alpha cells, we propose that UCN3-driven somatostatin may serve to regulate local paracrine glucagon levels in the islet instead of inhibiting gross systemic glucagon release.

β-Cell-selective regulation of gene expression by nitric oxide.

Nitric oxide is produced at low micromolar levels following the induction of inducible nitric oxide synthase (iNOS) and is responsible for mediating the inhibitory actions of cytokines on glucose-stimulated insulin secretion by islets of Langerhans. It is through the inhibition of mitochondrial oxidative metabolism, specifically aconitase and complex 4 of the electron transport chain, that nitric oxide inhibits insulin secretion. Nitric oxide also attenuates protein synthesis, induces DNA damage, activates DNA repair pathways, and stimulates stress responses (unfolded protein and heat shock) in β-cells. In this report, the time- and concentration-dependent effects of nitric oxide on the expression of six genes known to participate in the response of β-cells to this free radical were examined. The genes included Gadd45α (DNA repair), Puma (apoptosis), Hmox1 (antioxidant defense), Hsp70 (heat shock), Chop (UPR), and Ppargc1α (mitochondrial biogenesis). We show that nitric oxide stimulates β-cell gene expression in a narrow concentration range of ∼0.5-1 µM or levels corresponding to iNOS-derived nitric oxide. At concentrations greater than 1 µM, nitric oxide fails to stimulate gene expression in β-cells, and this is associated with the inhibition of mitochondrial oxidative metabolism. This narrow concentration range of responses is β-cell selective, as the actions of nitric oxide in non-β-cells (α-cells, mouse embryonic fibroblasts, and macrophages) are concentration dependent. Our findings suggest that β-cells respond to a narrow concentration range of nitric oxide that is consistent with the levels produced following iNOS induction, and that these concentration-dependent actions are selective for insulin-containing cells.