Catecholamine modulation of calcium currents in clonal pancreatic beta-cells

The mechanisms by which norepinephrine and epinephrine activate alpha 2-adrenergic receptors and inhibit insulin release from the pancreatic beta-cell (19, 21, 23) are not yet clear but may involve modulation at several sites. Because intracellular calcium has been implicated in the secretory process, it has been suggested that catecholamines may inhibit secretion by blocking calcium influx, thus reducing the free cytosolic calcium concentration (23). The present study examines the effects of epinephrine, norepinephrine, and clonidine on calcium current in an SV40-transformed hamster beta-cell line (HIT cells). Under voltage-clamp conditions, calcium currents were reversibly inhibited by norepinephrine, epinephrine, and clonidine in the low nanomolar range. The effects were blocked by 1) the alpha 2-antagonist yohimbine, 2) preincubation of the cells with pertussis toxin (PTX), and 3) guanosine 5'-O-(2-thiodiphosphate) (GDP beta S), the nonhydrolyzable GDP analogue that competitively inhibits the interaction of GTP with G proteins. In contrast, guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) caused irreversible blockade by catecholamines. These effects could not be overcome by adenosine 3',5'-cyclic monophosphate (cAMP), suggesting that the adenylate cyclase pathway is not involved in the G protein coupling with the channels. These studies show that catecholamines inhibit calcium currents in beta-cells through an alpha 2-adrenoreceptor PTX-sensitive G protein pathway and could inhibit insulin secretion by this mechanism.

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High dose of histone deacetylase inhibitors affects insulin secretory mechanism of pancreatic beta cell line

Endocrine Regulations ◽

10.2478/enr-2018-0004 ◽

2018 ◽

Vol 52 (1) ◽

pp. 21-26 ◽

Cited By ~ 5

Author(s):

Eiji Yamato

Keyword(s):

Insulin Secretion ◽

Beta Cell ◽

Cell Line ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Pancreatic Beta Cell ◽

High Dose ◽

Min6 Cells ◽

Beta Cell Line ◽

High Doses

Abstract Objective. Histone deacytylase inhibitors (HDACis) inhibit the deacetylation of the lysine residue of proteins, including histones, and regulate the transcription of a variety of genes. Recently, HDACis have been used clinically as anti-cancer drugs and possible anti-diabetic drugs. Even though HDACis have been proven to protect the cytokine-induced damage of pancreatic beta cells, evidence also shows that high doses of HDACis are cytotoxic. In the present study, we, therefore, investigated the eff ect of HDACis on insulin secretion in a pancreatic beta cell line. Methods. Pancreatic beta cells MIN6 were treated with selected HDACis (trichostatin A, TSA; valproic acid, VPA; and sodium butyrate, NaB) in medium supplemented with 25 mM glucose and 13% heat-inactivated fetal bovine serum (FBS) for indicated time intervals. Protein expression of Pdx1 and Mafa in MIN6 cells was demonstrated by immunohistochemistry and immunocytochemistry, expression of Pdx1 and Mafa genes was measured by quantitative RT-PCR method. Insulin release from MIN6 cells and insulin cell content were estimated by ELISA kit. Superoxide production in MIN6 cells was measured using a Total ROS/Superoxide Detection System. Results. TSA, VPA, and NaB inhibited the expression of Pdx1 and Mafa genes and their products. TSA treatment led to beta cell malfunction, characterized by enhanced insulin secretion at 3 and 9 mM glucose, but impaired insulin secretion at 15 and 25 mM glucose. Th us, TSA induced dysregulation of the insulin secretion mechanism. TSA also enhanced reactive oxygen species production in pancreatic beta cells. Conclusions. Our results showed that HDACis caused failure to suppress insulin secretion at low glucose concentrations and enhance insulin secretion at high glucose concentrations. In other words, when these HDACis are used clinically, high doses of HDACis may cause hypoglycemia in the fasting state and hyperglycemia in the fed state. When using HDACis, physicians should, therefore, be aware of the capacity of these drugs to modulate the insulin secretory capacity of pancreatic beta cells.

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PCSK9 affects expression of key surface proteins in human pancreatic beta cells through intra- and extracellular regulatory circuits

10.1101/2021.09.28.461975 ◽

2021 ◽

Author(s):

kevin Saitoski ◽

Maria Ryaboshapkina ◽

Ghaith Hamza ◽

Andrew F Jarnuczak ◽

claire berthault ◽

...

Keyword(s):

Transcription Factors ◽

Beta Cell ◽

Pancreatic Islets ◽

Cell Line ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Pancreatic Beta Cell ◽

Loss Of Function ◽

Beta Cell Line ◽

Gain And Loss

Aims/hypothesis: Proprotein convertase subtilisin/kexin 9 (PCSK9) is involved in the degradation of LDLR. However, PCSK9 can target other proteins in a cell-type specific manner. While PCSK9 has been detected in pancreatic islets, its expression in insulin-producing pancreatic beta cells is debated. Herein, we studied PCSK9 expression, regulation and function in the human pancreatic beta cell line EndoC-βH1. Methods: We assessed PCSK9 expression in mouse and human pancreatic islets, and in the pancreatic beta cell line EndoC-βH1. We also studied PCSK9 regulation by cholesterol, lipoproteins, Mevastatin, and by SREBPs transcription factors. To evaluate PCSK9 function in pancreatic beta cells, we performed PCSK9 gain-and loss-of-function experiments in EndoC-βH1 using siPCSK9 or recombinant PCSK9 treatments, respectively. Results: We demonstrate that PCSK9 is expressed and secreted by pancreatic beta cells. In EndoC-βH1 cells, PCSK9 expression is regulated by cholesterol and by SREBPs transcription factors. Importantly, PCSK9 knockdown results in multiple transcriptome, proteome and secretome deregulations and impaired insulin secretion. By gain- and loss-of- function experiments, we observed that PCSK9 regulates the expression levels of LDLR and VLDLR through an extracellular mechanism while CD36, PD-L1 and HLA-ABC are regulated through an intracellular mechanism. Conclusions/interpretation: Collectively, these results highlight PCSK9 as an important regulator of CD36, PD-L1 and HLA-ABC cell surface expression in pancreatic beta cells. Data availability: RNA-seq data have been deposited to GEO database with accession number GSE182016. Mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the following identifiers: PXD027921, PXD027911 and PXD027913.

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Pancreatic beta-cell somatostatin receptors

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1990.259.2.e216 ◽

1990 ◽

Vol 259 (2) ◽

pp. E216-E224 ◽

Cited By ~ 1

Author(s):

K. Thermos ◽

M. D. Meglasson ◽

J. Nelson ◽

K. M. Lounsbury ◽

T. Reisine

Keyword(s):

Beta Cell ◽

Physical Properties ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Pancreatic Beta Cell ◽

Hormone Secretion ◽

Isolated Islets ◽

Binding Studies ◽

Beta Cell Line ◽

Hit Cells

The characteristics of somatostatin (SRIF) receptors in rat pancreatic beta-cells were investigated using rat islets and the beta-cell line HIT-T15 (HIT). The biochemical properties of the SRIF receptors were examined with 125I-labeled des-Ala-1,Gly-2-desamino-Cys-3-[Tyr-11]- dicarba3,14-somatostatin (CGP 23996). 125I-CGP 23996 bound to SRIF receptors in HIT cells with high affinity and in a saturable manner. The binding of 125I-CGP 23996 to SRIF receptors was blocked by SRIF analogues with a rank order of potency of somatostatin 28 (SRIF-28) greater than D-Trp-8-somatostatin greater than somatostatin 14 (SRIF-14). To investigate the physical properties of the HIT cell SRIF receptor, the receptor was covalently labeled with 125I-CGP 23996 using photo-cross-linking techniques. 125I-CGP 23996 specifically labeled a protein of 55 kDa in HIT cell membranes. The size of the SRIF receptor in HIT cells is similar to the size of the SRIF receptor labeled with 125I-CGP 23996 in membranes of freshly isolated islets, suggesting that the physical properties of SRIF receptors in HIT cells and rat islet cells are similar. The binding studies suggest that beta-cells predominantly express a SRIF-28-preferring receptor. In freshly isolated islets, glucose- and arginine-stimulated insulin release was effectively blocked by SRIF-28 but not by SRIF-14. SRIF-14 did inhibit arginine-stimulated glucagon secretion from freshly isolated islets. The dissociation of the inhibitory effects of SRIF-28 and SRIF-14 on insulin and glucagon release from freshly isolated islets suggests that the two peptides act through different receptors in islets to regulate hormone secretion.

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Islet primary cilia motility controls insulin secretion

10.1101/2021.12.14.472629 ◽

2021 ◽

Author(s):

Jung Hoon Cho ◽

Zipeng A Li ◽

Lifei Zhu ◽

Brian Muegge ◽

Henry Roseman ◽

...

Keyword(s):

Insulin Secretion ◽

Beta Cell ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Primary Cilia ◽

Calcium Influx ◽

Pancreatic Beta Cell ◽

Critical Role ◽

Altered Expression ◽

Motile Cilia

Primary cilia are specialized cell-surface organelles that mediate sensory perception and, in contrast to motile cilia and flagella, are thought to lack motility function. Here we show that primary cilia in pancreatic beta cells exhibit movement that is required for glucose-dependent insulin secretion. Beta cell cilia contain motor proteins conserved from those found in classic motile cilia, and their 3D motion is dynein-driven and dependent on ATP and glucose metabolism. Inhibition of cilia motion blocks beta cell calcium influx and insulin secretion. Beta cells from humans with type 2 diabetes have altered expression of cilia motility genes. Our findings redefine primary cilia as dynamic structures possessing both sensory and motile function and establish that pancreatic beta cell cilia movement plays a critical role in controlling insulin secretion.

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TRAIL induces proliferation in rodent pancreatic beta cells via AKT activation

Journal of Molecular Endocrinology ◽

10.1530/jme-20-0037 ◽

2021 ◽

Author(s):

Sevim Kahraman ◽

Ozlem Yilmaz ◽

Hasan Ali Altunbas ◽

Ercument Dirice ◽

Ahter Dilsad Sanlioglu

Keyword(s):

Beta Cell ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Pancreatic Beta Cell ◽

Cell Mass ◽

Beta Cell Mass ◽

Min6 Cells ◽

Akt Activation ◽

Proliferative Effect ◽

Beta Cell Line

Strategies to increase functional pancreatic beta cell mass is of great interest in diabetes-related research. TNF-related apoptosis-inducing ligand (TRAIL) is well-known to promote proliferation and survival in various cell types, including vascular smooth muscle and endothelial cells. Correlation between the protective nature of TRAIL on these cells and its proliferative effect is noteworthy. TRAIL’s seemingly protective/therapeutic effect in diabetes prompted us to question whether it may act as an inducer of proliferation in pancreatic beta cells. We used rat primary islet cells and MIN6 mouse beta cell line to investigate TRAIL-induced proliferation. Cell viability and/or death was analysed by MTT, WST-1, and annexin-V/PI assays, while proliferation rates and pathways were assessed via immunocytochemical and Western blot analyses. Receptor neutralization antibodies identified the mediator receptors. Recombinant soluble TRAIL (sTRAIL) treatment led to 1.6-fold increased proliferation in insulin-positive cells in dispersed rat islets compared to the untreated group, while adenovirus-mediated overexpression of TRAIL increased the number of proliferating beta cells up to more than 6-fold. sTRAIL or adenoviral vector-mediated TRAIL overexpression induced proliferation in MIN6 cells also. TRAIL’s proliferative effect was mediated via AKT activation, which was suppressed upon specific inhibition. Neutralization of each TRAIL receptor reversed the proliferative effect to some degree, with the highest level of inhibition in death receptor 5 (DR5) blockage in MIN6 cells, and in decoy receptor 1 (DcR1) blockage in primary rat beta cells. Thus, TRAIL induces proliferation in rodent pancreatic beta cells through activation of the AKT pathway.

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1-Palmitoyl-2-Linoleoyl-3-Acetyl-rac-Glycerol Attenuates Streptozotocin-Induced Pancreatic Beta Cell Damage by Promoting Glucose Transporter 2 Endocytosis

Molecular and Cellular Biology ◽

10.1128/mcb.00157-19 ◽

2019 ◽

Vol 39 (21) ◽

Author(s):

Jimin Kim ◽

Joo Heon Kim ◽

Ki-Young Sohn ◽

Sun Young Yoon ◽

Jae Wha Kim

Keyword(s):

Beta Cell ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Glucose Transporter ◽

Cell Damage ◽

Serum Insulin ◽

Pancreatic Beta Cell ◽

Ros Generation ◽

Glucose Transporter 2 ◽

Beta Cell Line

ABSTRACT Streptozotocin (STZ) is widely used to induce diabetic rodent models. It is specifically toxic to pancreatic beta cells and causes severe destruction and dysfunction. We investigated the effect of 1-palmitoyl-2-linoleoyl-3-acetyl-rac-glycerol (PLAG) on an STZ-induced diabetic mouse model. PLAG attenuated the glucose increase and maintained serum insulin at levels similar to those seen with control mice. In pancreatic beta cell line INS-1, STZ-induced cell apoptosis and intracellular reactive oxygen species (ROS) generation were significantly reduced to nearly normal levels after PLAG treatment. Glucose transporter 2 (GLUT2) localization analyses and glucose uptake assays showed that PLAG accelerated GLUT2 internalization, which ameliorated excessive entry of glucose, as well as STZ. STZ-induced cytotoxic effects were significantly reduced in PLAG-treated groups. The biological activity of PLAG was further confirmed in GLUT2-silenced cells, and the specificity of PLAG was verified using its derivative 1-palmitoyl-2-linoleoyl-3-hydroxyl-rac-glycerol (PLH). Our results suggest that PLAG may be a useful agent for protecting beta cells in the setting of excessive glucose influx.

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Stable overexpression of the glucose-6-phosphatase catalytic subunit attenuates glucose sensitivity of insulin secretion from a mouse pancreatic beta-cell line

Journal of Endocrinology ◽

10.1677/joe.0.1640307 ◽

2000 ◽

Vol 164 (3) ◽

pp. 307-314 ◽

Cited By ~ 14

Author(s):

K Iizuka ◽

H Nakajima ◽

A Ono ◽

K Okita ◽

J Miyazaki ◽

...

Keyword(s):

Insulin Secretion ◽

Beta Cell ◽

Cell Line ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Glucose Sensor ◽

Pancreatic Beta Cell ◽

Catalytic Subunit ◽

Beta Cell Line ◽

Glucose Stimulated Insulin Secretion

Glucose-6-phosphatase (G-6-Pase) hydrolyzes glucose-6-phosphate to glucose, reciprocal with the so-called glucose sensor, glucokinase, in pancreatic beta cells. To study the role of G-6-Pase in glucose-stimulated insulin secretion from beta cells, we have introduced rat G-6-Pase catalytic subunit cDNA and have established permanent clones with 3-, 7- and 24-fold G-6-Pase activity of the mouse beta-cell line, MIN6. In these clones, glucose usage and ATP production in the presence of 5.5 or 25 mM glucose were reduced, and glucose-stimulated insulin secretion was decreased in proportion to the increased G-6-Pase activity. In addition, insulin secretory capacity in response to d-fructose and pyruvate was unchanged; however, 25 mM glucose-stimulated insulin secretion and intracellular calcium response were completely inhibited. In the clone with 24-fold G-6-Pase activity, changes in intracellular NAD(P)H autofluorescence in response to 25 mM glucose were reduced, but the changes with 20 mM fructose and 20 mM pyruvate were not altered. Stable overexpression of G-6-Pase in beta cells resulted in attenuation of the overall glucose-stimulated metabolic responses corresponding to the degree of overexpression. This particular experimental manipulation shows that the possibility exists of modulating glucose-stimulated insulin release by thoroughly altering glucose cycling at the glucokinase/G-6-Pase step.

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Paired box 6 programs essential exocytotic genes in the regulation of glucose-stimulated insulin secretion and glucose homeostasis

Science Translational Medicine ◽

10.1126/scitranslmed.abb1038 ◽

2021 ◽

Vol 13 (600) ◽

pp. eabb1038

Author(s):

Wing Yan So ◽

Wai Nam Liu ◽

Adrian Kee Keong Teo ◽

Guy A. Rutter ◽

Weiping Han

Keyword(s):

Insulin Secretion ◽

Beta Cell ◽

Pancreatic Islets ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Cell Function ◽

Pancreatic Beta Cell ◽

Beta Cell Line ◽

Pathophysiological Role ◽

Glucose Stimulated Insulin Secretion

The paired box 6 (PAX6) transcription factor is crucial for normal pancreatic islet development and function. Heterozygous mutations of PAX6 are associated with impaired insulin secretion and early-onset diabetes mellitus in humans. However, the molecular mechanism of PAX6 in controlling insulin secretion in human beta cells and its pathophysiological role in type 2 diabetes (T2D) remain ambiguous. We investigated the molecular pathway of PAX6 in the regulation of insulin secretion and the potential therapeutic value of PAX6 in T2D by using human pancreatic beta cell line EndoC-βH1, the db/db mouse model, and primary human pancreatic islets. Through loss- and gain-of-function approaches, we uncovered a mechanism by which PAX6 modulates glucose-stimulated insulin secretion (GSIS) through a cAMP response element–binding protein (CREB)/Munc18-1/2 pathway. Moreover, under diabetic conditions, beta cells and pancreatic islets displayed dampened PAX6/CREB/Munc18-1/2 pathway activity and impaired GSIS, which were reversed by PAX6 replenishment. Adeno-associated virus–mediated PAX6 overexpression in db/db mouse pancreatic beta cells led to a sustained amelioration of glycemic perturbation in vivo but did not affect insulin resistance. Our study highlights the pathophysiological role of PAX6 in T2D-associated beta cell dysfunction in humans and suggests the potential of PAX6 gene transfer in preserving and restoring beta cell function.

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Update on the Protective Molecular Pathways Improving Pancreatic Beta-Cell Dysfunction

Mediators of Inflammation ◽

10.1155/2013/750540 ◽

2013 ◽

Vol 2013 ◽

pp. 1-14 ◽

Cited By ~ 14

Author(s):

Alessandra Puddu ◽

Roberta Sanguineti ◽

François Mach ◽

Franco Dallegri ◽

Giorgio Luciano Viviani ◽

...

Keyword(s):

Beta Cell ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Cell Function ◽

Pancreatic Beta Cell ◽

Cell Mass ◽

Beta Cell Mass ◽

Beta Cell Dysfunction ◽

Molecular Pathways ◽

Cell Dysfunction

The primary function of pancreatic beta-cells is to produce and release insulin in response to increment in extracellular glucose concentrations, thus maintaining glucose homeostasis. Deficient beta-cell function can have profound metabolic consequences, leading to the development of hyperglycemia and, ultimately, diabetes mellitus. Therefore, strategies targeting the maintenance of the normal function and protecting pancreatic beta-cells from injury or death might be crucial in the treatment of diabetes. This narrative review will update evidence from the recently identified molecular regulators preserving beta-cell mass and function recovery in order to suggest potential therapeutic targets against diabetes. This review will also highlight the relevance for novel molecular pathways potentially improving beta-cell dysfunction.

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Augmented mitochondrial energy metabolism is an early response to chronic glucose stress in human pancreatic beta cells

Diabetologia ◽

10.1007/s00125-020-05275-5 ◽

2020 ◽

Vol 63 (12) ◽

pp. 2628-2640

Author(s):

Isabelle Chareyron ◽

Stefan Christen ◽

Sofia Moco ◽

Armand Valsesia ◽

Steve Lassueur ◽

...

Keyword(s):

Energy Metabolism ◽

Beta Cell ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Cell Function ◽

Blood Glucose Control ◽

Calcium Signalling ◽

Cytosolic Calcium ◽

Metabolic Changes ◽

Mitochondrial Energy Metabolism

Abstract Aims/hypothesis In islets from individuals with type 2 diabetes and in islets exposed to chronic elevated glucose, mitochondrial energy metabolism is impaired. Here, we studied early metabolic changes and mitochondrial adaptations in human beta cells during chronic glucose stress. Methods Respiration and cytosolic ATP changes were measured in human islet cell clusters after culture for 4 days in 11.1 mmol/l glucose. Metabolomics was applied to analyse intracellular metabolite changes as a result of glucose stress conditions. Alterations in beta cell function were followed using insulin secretion assays or cytosolic calcium signalling after expression of the calcium probe YC3.6 specifically in beta cells of islet clusters. Results At early stages of glucose stress, mitochondrial energy metabolism was augmented in contrast to the previously described mitochondrial dysfunction in beta cells from islets of diabetic donors. Following chronic glucose stress, mitochondrial respiration increased (by 52.4%, p < 0.001) and, as a consequence, the cytosolic ATP/ADP ratio in resting human pancreatic islet cells was elevated (by 27.8%, p < 0.05). Because of mitochondrial overactivation in the resting state, nutrient-induced beta cell activation was reduced. In addition, chronic glucose stress caused metabolic adaptations that resulted in the accumulation of intermediates of the glycolytic pathway, the pentose phosphate pathway and the TCA cycle; the most strongly augmented metabolite was glycerol 3-phosphate. The changes in metabolites observed are likely to be due to the inability of mitochondria to cope with continuous nutrient oversupply. To protect beta cells from chronic glucose stress, we inhibited mitochondrial pyruvate transport. Metabolite concentrations were partially normalised and the mitochondrial respiratory response to nutrients was markedly improved. Furthermore, stimulus–secretion coupling as assessed by cytosolic calcium signalling, was restored. Conclusion/interpretation We propose that metabolic changes and associated mitochondrial overactivation are early adaptations to glucose stress, and may reflect what happens as a result of poor blood glucose control. Inhibition of mitochondrial pyruvate transport reduces mitochondrial nutrient overload and allows beta cells to recover from chronic glucose stress.

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