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

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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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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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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Effects of High Glucose Levels and Glycated Serum on GIP Responsiveness in the Pancreatic Beta Cell Line HIT-T15

Journal of Diabetes Research ◽

10.1155/2015/326359 ◽

2015 ◽

Vol 2015 ◽

pp. 1-7 ◽

Cited By ~ 4

Author(s):

Alessandra Puddu ◽

Roberta Sanguineti ◽

Fabrizio Montecucco ◽

Giorgio Luciano Viviani

Keyword(s):

Insulin Secretion ◽

Beta Cell ◽

Cell Line ◽

High Glucose ◽

Intracellular Signaling ◽

Pancreatic Beta Cells ◽

Pancreatic Beta Cell ◽

Diabetic Patients ◽

Glucose Levels ◽

Beta Cell Line

Glucose-dependent insulinotropic peptide (GIP) is an incretin hormone produced in the gastrointestinal tract that stimulates glucose dependent insulin secretion. Impaired incretin response has been documented in diabetic patients and was mainly related to the inability of the pancreatic beta cells to secrete insulin in response to GIP. Advanced Glycation End Products (AGEs) have been shown to play an important role in pancreatic beta cell dysfunction. The aim of this study is to investigate whether the exposure to AGEs can induce GIP resistance in the pancreatic beta cell line HIT-T15. Cells were cultured for 5 days in low (CTR) or high glucose (HG) concentration in the presence of AGEs (GS) to evaluate the expression of GIP receptor (GIPR), the intracellular signaling activated by GIP, and secretion of insulin in response to GIP. The results showed that incubation with GS alone altered intracellular GIP signaling and decreased insulin secretion as compared to CTR. GS in combination with HG reduced the expression of GIPR and PI3K and abrogated GIP-induced AKT phosphorylation and GIP-stimulated insulin secretion. In conclusion, we showed that treatment with GS is associated with the loss of the insulinotropic effect of GIP in hyperglycemic conditions.

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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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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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Catecholamine modulation of calcium currents in clonal pancreatic beta-cells

AJP Cell Physiology ◽

10.1152/ajpcell.1989.257.6.c1171 ◽

1989 ◽

Vol 257 (6) ◽

pp. C1171-C1176 ◽

Cited By ~ 27

Author(s):

H. H. Keahey ◽

A. E. Boyd ◽

D. L. Kunze

Keyword(s):

Beta Cell ◽

G Protein ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Calcium Influx ◽

Pancreatic Beta Cell ◽

Cytosolic Calcium ◽

Calcium Currents ◽

Secretory Process ◽

Beta Cell Line

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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