scholarly journals Long Noncoding RNA Meg3 Regulates Mafa Expression in Mouse Beta Cells by Inactivating Rad21, Smc3 or Sin3α

2018 ◽  
Vol 45 (5) ◽  
pp. 2031-2043 ◽  
Author(s):  
Ning Wang ◽  
Yanan Zhu ◽  
Min Xie ◽  
Lintao Wang ◽  
Feiyan Jin ◽  
...  
Keyword(s):  
Beta Cell ◽  
Beta Cells ◽  
Pancreatic Beta Cells ◽  
Noncoding Rna ◽  
Islet Cells ◽  
Quantitative Polymerase Chain Reaction ◽  
Insulin Biosynthesis ◽  
Expression Levels ◽  
Beta Cell Line

Background/Aims: The main pathogenic mechanism of diabetes is a decrease in the number of islet beta cells or a decline in their function. Recent studies have shown that pancreatic long noncoding RNAs (lncRNAs) have a high degree of tissue specificity and may be involved in the maintenance of islet cells function and the development of diabetes. The aim of this study was to investigate the molecular regulatory mechanism of mouse maternal expressed gene 3 (Meg3) in insulin biosynthesis in pancreatic islets. Methods: Chromatin immunoprecipitation–quantitative polymerase chain reaction (qPCR) and RNA immunoprecipitation–qPCR were used to investigate the molecular mechanism of lncRNA Meg3 in insulin biosynthesis by regulating v-Maf musculoaponeurotic fibrosarcoma oncogene family, protein A (MafA), a mature beta cell marker in the MIN6 beta cell line. Further, the expression levels of Meg3, Ezh2, MafA, Rad21, Smc3, and Sin3α were analyzed in vivo and in vitro by RT-PCR and western blotting. Results: Intranuclear lncRNA Meg3 can bind EZH2, a methyltransferase belonging to the Polycomb repressive complex-2, in pancreatic islet cells. In addition, knockdown of Ezh2 can also inhibit the expression of MafA and Ins2, while expression levels of Rad21, Smc3, and Sin3α are upregulated, by interfering with Ezh2 or Meg3 in pancreatic beta cells. Knockdown of Meg3 resulted in the loss of EZH2 binding and H3K27 trimethylation occupancy of Rad21, Smc3, and Sin3α promoter regions. The inhibition of Rad21, Smc3, or Sin3α, which directly act on the MafA promoter, leads to upregulated expression of MafA in both MIN6 cells and mouse islets. Moreover, the synthesis and secretion of insulin were increased by inhibition of these transcription factors. Conclusions: Pancreatic lncRNA Meg3 can epigenetically regulate the expression of Rad21, Smc3, and Sin3α via EZH2-driven H3K27 methylation. By inhibiting the expression of Rad21, Smc3, or Sin3α, Meg3 promotes the expression of MafA and affects the production of insulin.

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

2018 ◽  
Vol 52 (1) ◽  
pp. 21-26 ◽  
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.

Catecholamine modulation of calcium currents in clonal pancreatic beta-cells

1989 ◽  
Vol 257 (6) ◽  
pp. C1171-C1176 ◽  
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.

Comparative toxicity of alloxan, N-alkylalloxans and ninhydrin to isolated pancreatic islets in vitro

1997 ◽  
Vol 155 (2) ◽  
pp. 283-293 ◽  
Author(s):  
A Jorns ◽  
R Munday ◽  
M Tiedge ◽  
S Lenzen
Keyword(s):  
Beta Cell ◽  
Pancreatic Islets ◽  
Beta Cells ◽  
Pancreatic Beta Cells ◽  
Ultrastructural Changes ◽  
Cell Necrosis ◽  
High Concentrations ◽  
Beta Cell Necrosis

The in vitro toxicity of the diabetogenic agent alloxan as documented by the induction of beta cell necrosis was studied in isolated ob/ob mouse pancreatic islets. The effect of alloxan has been compared with that of a number of N-alkyl alloxan derivatives and with that of the structurally related compound, ninhydrin. Alloxan and its derivatives were selectively toxic to pancreatic beta cells, with other endocrine cells and exocrine parenchymal cells being well preserved, even at high concentration. In contrast, ninhydrin was selectively toxic to pancreatic beta cells only at comparatively low concentration, destroying all islet cell types at high concentrations. The ultrastructural changes induced by all the test compounds in pancreatic beta cells in vitro were very similar to those observed during the development of alloxan diabetes in vivo. The relative toxicity of the various compounds to pancreatic beta cells in vitro was not, however, related to their ability to cause diabetes in vivo. Indeed, the non-diabetogenic substances ninhydrin, N-butylalloxan and N-isobutylalloxan were very much more toxic to isolated islets than the diabetogenic compounds alloxan and N-methylalloxan. These results suggest that the differences in diabetogenicity among alloxan derivatives are not due to intrinsic differences in the susceptibility of the pancreatic beta cells to their toxicity, but may reflect differences in distribution or metabolism. High concentrations of glucose protected islets against the harmful effects of alloxan and its derivatives, but not those of ninhydrin. Low levels of glucose, and non-carbohydrate nutrients, afforded little protection, indicating that the effect of glucose is not due to the production of reducing equivalents within the cell, 3-O-Methylglucose, which protects against alloan diabetes in vivo, did not protect against alloxan toxicity in vitro. Since 3-O-methylglucose is known to prevent uptake of alloxan by pancreatic beta cells, it appears that uptake of alloxan by the cell is not a prerequisite for the induction of beta cell necrosis.

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

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.

scholarly journals Tankyrase interacts with the allosteric site of glucokinase and inhibits its glucose-sensing function in the beta cell

2021 ◽  
Author(s):  
Nai-Wen Chi ◽  
Travis Eisemann ◽  
Tsung-Yin J Yeh ◽  
Swati Roy ◽  
Tyler J Chi ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Beta Cell ◽  
Beta Cells ◽  
Pancreatic Beta Cells ◽  
Glucose Sensor ◽  
Pancreatic Beta Cell ◽  
Glucose Sensing ◽  
Binding Motif ◽  
Allosteric Site

Insulin secretion in the pancreatic beta cell is rate-limited by glucokinase (GCK), the glucose sensor that catalyzes the first step of glucose metabolism. GCK consists of two lobes connected by a flexible hinge that allows the kinase to sample a spectrum of conformations ranging from the active, closed form to several inactive, less-compact forms. Activating GCK mutations can cause hyperinsulinemia and hypoglycemia in infants. A similar phenotype is exhibited in mice deficient in tankyrase (TNKS), prompting us to investigate whether TNKS might modulate the glucose-sensing function of GCK. We found that TNKS colocalizes and directly interacts with GCK. Their interaction is mediated by two ankyrin-repeat clusters (ARC-2 and -5) in TNKS and a tankyrase-binding motif (TBM, aa 63-68) in the GCK hinge. This interaction is conformation sensitive: human GCK variants that cause hyperglycemia (V62M) or hypoglycemia (S64Y) enhance or diminish the interaction respectively, even though they have no impact on TNKS interaction in the context of a GCK peptide (V62M) or a peptide library (S64Y). Moreover, the TNKS-GCK interaction is inhibited by high concentrations of glucose, which are known to stabilize GCK in the active (closed, glucose-avid) conformation. Conversely, glucose phosphorylation by GCK in vitro is inhibited by TNKS. To validate this in vitro inhibitory effect in the MIN6 beta cells, we showed that glucose-stimulated insulin secretion is suppressed upon stabilization of the TNKS protein and conversely is enhanced upon TNKS knockdown. Based on these findings as well as by contrasting with hexokinase-2, we propose that TNKS is a physiological GCK inhibitor in pancreatic beta cells that acts by trapping the kinase in an open (inactive) conformation.

scholarly journals Nicorandil improves diabetes and rat islet beta-cell damage induced by streptozotocin in vivo and in vitro

2004 ◽  
pp. 277-285 ◽  
Author(s):  
K Kasono ◽  
T Yasu ◽  
A Kakehashi ◽  
N Kinoshita ◽  
H Tamemoto ◽  
...  
Keyword(s):  
Beta Cell ◽  
Beta Cells ◽  
Islet Cells ◽  
Cell Damage ◽  
Neonatal Rat ◽  
Islet Beta Cells ◽  
Islet Beta Cell ◽  
Beta Cell Damage

OBJECTIVE: N-(2-hydroxyethyl)-nicotinamide nitrate (nicorandil) is a unique anti-anginal agent, reported to act as both an ATP-sensitive K(+) channel opener (PCO) and a nitric oxide donor. It also has an anti-oxidant action. We examined the effects of nicorandil on streptozotocin (STZ)-induced islet beta-cell damage both in vivo and in vitro. DESIGN AND METHODS: STZ-induced diabetic Brown Norway rats (STZ-DM) were fed with nicorandil-containing chow from day 2 (STZ-DM-N48), 3 (STZ-DM-N72), and 4 (STZ-DM-N96) to day 30. Body weight, blood glucose, and plasma insulin were measured every week. For the in vitro assay, neonatal rat islet-rich cultures were performed and cells were treated with nicorandil from 1 h before to 2 h after exposure to STZ for 30 min. Insulin secretion from islet cells was assayed after an additional 24 h of culture. We also observed the effect of nicorandil on the generation of reactive oxygen species (ROS) from rat inslinoma cells (RINm5F). RESULTS: Body weight loss and blood glucose levels of STZ-DM-N48 rats were significantly lower than those of STZ-DM rats. Immunohistochemical staining of insulin showed preservation of insulin-secreting islet beta-cells in STZ-DM-N48 rats. Nicorandil also dose-dependently recovered the insulin release from neonatal rat islet cells treated with STZ in in vitro experiments. Nicorandil did not act as a PCO on neonatal rat islet beta-cells or RINm5F cells, and did not show an inhibitory effect on poly(ADP-ribose) polymerase-1. However, the drug inhibited the production of ROS stimulated by high glucose (22.0 mmol/l) in RINm5F cells. CONCLUSIONS: These results suggested that nicorandil improves diabetes and rat islet beta-cell damage induced by STZ in vivo and in vitro. It protects islet beta-cells, at least partly, via a radical scavenging effect.

Pancreatic beta-cell somatostatin receptors

1990 ◽  
Vol 259 (2) ◽  
pp. E216-E224 ◽  
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.

scholarly journals Exogenous Ghrelin Increases Plasma Insulin Level in Diabetic Rats

Biomolecules ◽  
2020 ◽  
Vol 10 (4) ◽  
pp. 633 ◽  
Author(s):  
Haba Elabadlah ◽  
Rasheed Hameed ◽  
Crystal D’Souza ◽  
Sahar Mohsin ◽  
Ernest A. Adeghate
Keyword(s):  
Body Weight ◽  
Insulin Secretion ◽  
Amino Acid ◽  
Beta Cell ◽  
Beta Cells ◽  
Islet Cells ◽  
Insulin Level ◽  
Diabetic Rats ◽  
Pancreatic Islet Cells ◽  
Beta Cell Line

Ghrelin, a 28-amino acid peptide, is a strong growth hormone secretagogue and a regulator of food intake. In addition, ghrelin is thought to play a role in insulin secretion and in glucose homeostasis. A lot of contradictory data have been reported in the literature regarding the co-localization of ghrelin with other hormones in the islet of Langerhans, its role in insulin secretion and attenuation of type 2 diabetes mellitus. In this study, we investigate the effect of chronic ghrelin treatment on glucose, body weight and insulin level in normal and streptozotocin-induced diabetic male Wistar rats. We have also examined the distribution pattern and co-localization of ghrelin with insulin in pancreatic islet cells using immunohistochemistry and immune-electron microscopy and the ability of ghrelin to stimulate insulin release from the CRL11065 beta cell line. Control, non-diabetic groups received intraperitoneal injection of normal saline, while treated groups received intraperitoneal injection of 5 µg/kg body weight of ghrelin (amino acid chain 24–51) on a daily basis for a duration of four weeks. Our results show that the administration of ghrelin increases the number of insulin-secreting beta cells and serum insulin level in both normal and diabetic rats. We also demonstrated that ghrelin co-localizes with insulin in pancreatic islet cells and that the pattern of ghrelin distribution is altered after the onset of diabetes. Moreover, ghrelin at a dose of 10−6 M and 10−12 M increased insulin release from the CRL11065 beta cell line. In summary, ghrelin co-localizes with insulin in the secretory granules of pancreatic beta cells and enhances insulin production.

scholarly journals DNA damage induces senescent phenotypes in mouse NIT1 beta cells and human islets

2021 ◽  
Author(s):  
Gabriel Brawerman ◽  
Peter J. Thompson
Keyword(s):  
Dna Damage ◽  
Beta Cell ◽  
Beta Cells ◽  
Pancreatic Beta Cells ◽  
Transcriptional Response ◽  
Strand Break ◽  
Human Islets ◽  
Cell Senescence ◽  
Dna Double Strand Break ◽  
Beta Cell Line

Pancreatic beta cells are essential endocrine cells in the islets of Langerhans that respond to increases in blood glucose by secreting insulin and their dysfunction and death drives the development of diabetes. Beta cell senescence involving a DNA damage response (DDR) was recently shown to contribute to the pathogenesis of Type 1 Diabetes (T1D), however, a basic mechanistic understanding of how senescence develops in beta cells is lacking. Here, we investigated senescent phenotypes arising in the mouse beta cell line NIT1 derived from the T1D susceptible nonobese diabetic (NOD) mouse strain and the transcriptome response in human islets after induction of DNA damage. Sub-lethal DNA damage in NIT1 cells led to several classical hallmarks of senescence including the DDR, growth arrest, enlarged flattened morphology and a senescence-associated secretory phenotype (SASP) resembling what occurs during T1D. RNA-seq analysis on human islets with DNA double-strand break damage revealed a coordinated p53-p21 transcriptional response and upregulation of genes involved in prosurvival signaling and SASP. Taken together, these findings suggest that some of the phenotypes of mouse and human beta cell senescence during T1D can be induced by DNA damage in NIT1 cells and human islets in culture.

scholarly journals miR-212/132-Enriched Extracellular Vesicles Promote Differentiation of Induced Pluripotent Stem Cells Into Pancreatic Beta Cells

2021 ◽  
Vol 9 ◽  
Author(s):  
Chunyu Bai ◽  
Qiwei Ren ◽  
Haifeng Liu ◽  
Xiangchen Li ◽  
Weijun Guan ◽  
...  
Keyword(s):  
Stem Cells ◽  
Beta Cell ◽  
Induced Pluripotent Stem Cells ◽  
Beta Cells ◽  
Extracellular Vesicles ◽  
Pluripotent Stem Cells ◽  
Pancreatic Beta Cells ◽  
Pancreatic Beta Cell ◽  
Induced Pluripotent

Pancreatic beta cell transplantation is the ideal method for treatment of type 1 diabetes mellitus (T1DM), and the generation of beta cells from induced pluripotent stem cells (iPSCs) of patients is a promising strategy. In this study, we improved a previous strategy to produce beta cells using extracellular vesicles (EVs) derived from mature beta cells and differentiated beta cells from iPSCs (i-Beta cells), which secreted insulin under glucose stimulation in vitro and ameliorated hyperglycemia in vivo. Mechanistic analyses revealed that EV-carried microRNA (miR)-212/132 (EV-miR-212/132) directly bound to the 3′ UTR of FBW7 to prevent its translation and FBW7 combined with NGN3 to accelerate its proteasomal degradation. EV-miR-212/132 stabilized NGN3 expression to promote differentiation of endocrine cells from induced iPSCs. Moreover, NGN3 bound to PDX1 to enhance transcription of endogenous miR-212/132 and formed a positive regulatory circuit that maintained the functions of mature pancreatic beta cells.ConclusionThis study describes a novel approach for beta cell production and supports the use of iPSCs for cell replacement therapy of T1DM.

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