scholarly journals The mitochondrial Ca2+ uniporter MCU is required for normal glucose-stimulated insulin secretion in vitro and in vivo

2019 ◽  
Author(s):  
Elizabeth Haythorne ◽  
Eleni Georgiadou ◽  
Matthew T. Dickerson ◽  
Livia Lopez-Noriega ◽  
Timothy J. Pullen ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Glucose Tolerance ◽  
Mitochondrial Membrane ◽  
Inner Mitochondrial Membrane ◽  
Β Cells ◽  
Β Cell ◽  
Compensatory Mechanisms ◽  
Glucose Stimulated Insulin Secretion

AbstractMitochondrial oxidative metabolism is central to glucose-stimulated insulin secretion (GSIS). Whether Ca2+ uptake into pancreatic β-cell mitochondria potentiates or antagonises this process is still a matter of debate. Although the mitochondrial importer (MCU) complex is thought to represent the main route for Ca2+ transport across the inner mitochondrial membrane, its role in β-cells has not previously been examined in vivo. Here, we inactivated the pore-forming subunit MCUa (MCU) selectively in the β-cell in mice using Ins1Cre-mediated recombination. Glucose-stimulated mitochondrial Ca2+ accumulation, ATP production and insulin secretion were strongly (p<0.05 and p<0.01) inhibited in MCU null animals (βMCU-KO) in vitro. Interestingly, cytosolic Ca2+ concentrations increased (p<0.001) whereas mitochondrial membrane depolarisation improved in βMCU-KO animals. Male βMCU-KO mice displayed impaired in vivo insulin secretion at 5 (p<0.001) but not 15 min. post intraperitoneal (IP) injection of glucose while the opposite phenomenon was observed following an oral gavage at 5 min. Unexpectedly, glucose tolerance was improved (p<0.05) in young βMCU-KO (<12 weeks), but not older animals. We conclude that MCU is crucial for mitochondrial Ca2+ uptake in pancreatic β-cells and is required for normal GSIS. The apparent compensatory mechanisms which maintain glucose tolerance in βMCU-KO mice remain to be established.

scholarly journals Inhibition of Small Maf Function in Pancreatic β-Cells Improves Glucose Tolerance Through the Enhancement of Insulin Gene Transcription and Insulin Secretion

Endocrinology ◽  
2015 ◽  
Vol 156 (10) ◽  
pp. 3570-3580 ◽  
Author(s):  
Hiroshi Nomoto ◽  
Takuma Kondo ◽  
Hideaki Miyoshi ◽  
Akinobu Nakamura ◽  
Yoko Hida ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Glucose Tolerance ◽  
High Fat Diet ◽  
Cell Function ◽  
High Fat ◽  
Β Cells ◽  
Β Cell ◽  
Β Cell Function

The large-Maf transcription factor v-maf musculoaponeurotic fibrosarcoma oncogene homolog A (MafA) has been found to be crucial for insulin transcription and synthesis and for pancreatic β-cell function and maturation. However, insights about the effects of small Maf factors on β-cells are limited. Our goal was to elucidate the function of small-Maf factors on β-cells using an animal model of endogenous small-Maf dysfunction. Transgenic (Tg) mice with β-cell-specific expression of dominant-negative MafK (DN-MafK) experiments, which can suppress the function of all endogenous small-Mafs, were fed a high-fat diet, and their in vivo phenotypes were evaluated. Phenotypic analysis, glucose tolerance tests, morphologic examination of β-cells, and islet experiments were performed. DN-MafK-expressed MIN6 cells were also used for in vitro analysis. The results showed that DN-MafK expression inhibited endogenous small-Maf binding to insulin promoter while increasing MafA binding. DN-MafK Tg mice under high-fat diet conditions showed improved glucose metabolism compared with control mice via incremental insulin secretion, without causing changes in insulin sensitivity or MafA expression. Moreover, up-regulation of insulin and glucokinase gene expression was observed both in vivo and in vitro under DN-MafK expression. We concluded that endogenous small-Maf factors negatively regulates β-cell function by competing for MafA binding, and thus, the inhibition of small-Maf activity can improve β-cell function.

scholarly journals β-Cell-specific ablation of sirtuin 4 does not affect nutrient-stimulated insulin secretion in mice

2020 ◽  
Vol 319 (4) ◽  
pp. E805-E813
Author(s):  
Frank K. Huynh ◽  
Brett S. Peterson ◽  
Kristin A. Anderson ◽  
Zhihong Lin ◽  
Aeowynn J. Coakley ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Glucose Tolerance ◽  
Cell Function ◽  
Whole Body ◽  
Β Cells ◽  
Β Cell ◽  
Direct Role ◽  
Normal Glucose

Sirtuins are a family of proteins that regulate biological processes such as cellular stress and aging by removing posttranslational modifications (PTMs). We recently identified several novel PTMs that can be removed by sirtuin 4 (SIRT4), which is found in mitochondria. We showed that mice with a global loss of SIRT4 [SIRT4-knockout (KO) mice] developed an increase in glucose- and leucine-stimulated insulin secretion, and this was followed by accelerated age-induced glucose intolerance and insulin resistance. Because whole body SIRT4-KO mice had alterations to nutrient-stimulated insulin secretion, we hypothesized that SIRT4 plays a direct role in regulating pancreatic β-cell function. Thus, we tested whether β-cell-specific ablation of SIRT4 would recapitulate the elevated insulin secretion seen in mice with a global loss of SIRT4. Tamoxifen-inducible β-cell-specific SIRT4-KO mice were generated, and their glucose tolerance and glucose- and leucine-stimulated insulin secretion were measured over time. These mice exhibited normal glucose- and leucine-stimulated insulin secretion and maintained normal glucose tolerance even as they aged. Furthermore, 832/13 β-cells with a CRISPR/Cas9n-mediated loss of SIRT4 did not show any alterations in nutrient-stimulated insulin secretion. Despite the fact that whole body SIRT4-KO mice demonstrated an age-induced increase in glucose- and leucine-stimulated insulin secretion, our current data indicate that the loss of SIRT4 specifically in pancreatic β-cells, both in vivo and in vitro, does not have a significant impact on nutrient-stimulated insulin secretion. These data suggest that SIRT4 controls nutrient-stimulated insulin secretion during aging by acting on tissues external to the β-cell, which warrants further study.

scholarly journals Enhanced insulin secretion and improved glucose tolerance in mice with homozygous inactivation of the Na+K+2Cl− co-transporter 1

2012 ◽  
Vol 215 (1) ◽  
pp. 59-70 ◽  
Author(s):  
Saeed Alshahrani ◽  
Mauricio Di Fulvio
Keyword(s):  
Insulin Secretion ◽  
Glucose Tolerance ◽  
Cell Mass ◽  
Chloride Concentration ◽  
Cell Physiology ◽  
Β Cells ◽  
Β Cell ◽  
Functional Alleles

The intracellular chloride concentration ([Cl−]i) in β-cells plays an important role in glucose-stimulated plasma membrane depolarisation and insulin secretion. [Cl−]i is maintained above equilibrium in β-cells by the action of Cl− co-transporters of the solute carrier family 12 group A (Slc12a). β-Cells express Slc12a1 and Slc12a2, which are known as the bumetanide (BTD)-sensitive Na+-dependent K+2Cl− co-transporters 2 and 1 respectively. We show that mice lacking functional alleles of the Slc12a2 gene exhibit better fasting glycaemia, increased insulin secretion in response to glucose, and improved glucose tolerance when compared with wild-type (WT). This phenomenon correlated with increased sensitivity of β-cells to glucose in vitro and with increased β-cell mass. Further, administration of low doses of BTD to mice deficient in Slc12a2 worsened their glucose tolerance, and low concentrations of BTD directly inhibited glucose-stimulated insulin secretion from β-cells deficient in Slc12a2 but expressing intact Slc12a1 genes. Together, our results suggest for the first time that the Slc12a2 gene is not necessary for insulin secretion and that its absence increases β-cell secretory capacity. Further, impairment of insulin secretion with BTD in vivo and in vitro in islets lacking Slc12a2 genes unmasks a potential new role for Slc12a1 in β-cell physiology.

scholarly journals In vivo imaging of β-cell function reveals glucose-mediated heterogeneity of β-cell functional development

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Jia Zhao ◽  
Weijian Zong ◽  
Yiwen Zhao ◽  
Dongzhou Gou ◽  
Shenghui Liang ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
In Vivo Imaging ◽  
Cell Function ◽  
Β Cells ◽  
Β Cell ◽  
Functional Development ◽  
Β Cell Function ◽  
Glucose Stimulated Insulin Secretion

How pancreatic β-cells acquire function in vivo is a long-standing mystery due to the lack of technology to visualize β-cell function in living animals. Here, we applied a high-resolution two-photon light-sheet microscope for the first in vivo imaging of Ca2+activity of every β-cell in Tg (ins:Rcamp1.07) zebrafish. We reveal that the heterogeneity of β-cell functional development in vivo occurred as two waves propagating from the islet mantle to the core, coordinated by islet vascularization. Increasing amounts of glucose induced functional acquisition and enhancement of β-cells via activating calcineurin/nuclear factor of activated T-cells (NFAT) signaling. Conserved in mammalians, calcineurin/NFAT prompted high-glucose-stimulated insulin secretion of neonatal mouse islets cultured in vitro. However, the reduction in low-glucose-stimulated insulin secretion was dependent on optimal glucose but independent of calcineurin/NFAT. Thus, combination of optimal glucose and calcineurin activation represents a previously unexplored strategy for promoting functional maturation of stem cell-derived β-like cells in vitro.

scholarly journals Suppression of Peroxisome Proliferator-Activated Receptor γ-Coactivator-1α Normalizes the Glucolipotoxicity-Induced Decreased BETA2/NeuroD Gene Transcription and Improved Glucose Tolerance in Diabetic Rats

Endocrinology ◽  
2009 ◽  
Vol 150 (9) ◽  
pp. 4074-4083 ◽  
Author(s):  
Ji-Won Kim ◽  
Young-Hye You ◽  
Dong-Sik Ham ◽  
Jae-Hyoung Cho ◽  
Seung-Hyun Ko ◽  
...  
Keyword(s):  
Glucose Tolerance ◽  
Receptor Site ◽  
Diabetic Rats ◽  
Peroxisome Proliferator ◽  
Β Cells ◽  
Β Cell ◽  
Peroxisome Proliferator Activated Receptor ◽  
Cell Dysfunction

Abstract Peroxisome proliferator-activated receptor γ-coactivator-1α (PGC-1α) is significantly elevated in the islets of animal models of diabetes. However, the molecular mechanism has not been clarified. We investigated whether the suppression of PGC-1α expression protects against β-cell dysfunction in vivo and determined the mechanism of action of PGC-1α in β-cells. The studies were performed in glucolipotixicity-induced primary rat islets and INS-1 cells. In vitro and in vivo approaches using adenoviruses were used to evaluate the role of PGC-1α in glucolipotoxicity-associated β-cell dysfunction. The expression of PGC-1α in cultured β-cells increased gradually with glucolipotoxicity. The overexpression of PGC-1α also suppressed the expression of the insulin and β-cell E-box transcription factor (BETA2/NeuroD) genes, which was reversed by PGC-1α small interfering RNA (siRNA). BETA2/NeuroD, p300-enhanced BETA2/NeuroD, and insulin transcriptional activities were significantly suppressed by Ad-PGC-1α but were rescued by Ad-siPGC-1α. PGC-1α binding at the glucocorticoid receptor site on the BETA2/NeuroD promoter increased in the presence of PGC-1α. Ad-siPGC-1α injection through the celiac arteries of 90% pancreatectomized diabetic rats improved their glucose tolerance and maintained their fasting insulin levels. The suppression of PGC-1α expression protects the glucolipotoxicity-induced β-cell dysfunction in vivo and in vitro. A better understanding of the functions of molecules such as PGC-1α, which play key roles in intracellular fuel regulation, could herald a new era of the treatment of patients with type 2 diabetes mellitus by providing protection from glucolipotoxicity, which is an important cause of the development and progression of the disease.

scholarly journals Perilipin2 down-regulation in β cells impairs insulin secretion under nutritional stress and damages mitochondria

2020 ◽  
Author(s):  
Akansha Mishra ◽  
Siming Liu ◽  
Joseph Promes ◽  
Mikako Harata ◽  
William Sivitz ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Cell Function ◽  
Islet Cells ◽  
Cell Metabolism ◽  
Nutritional Stress ◽  
Β Cells ◽  
Β Cell ◽  
Down Regulation

Perilipin 2 (PLIN2) is the lipid droplet (LD) protein in β cells that increases under nutritional stress. Down-regulation of PLIN2 is often sufficient to reduce LD accumulation. To determine whether PLIN2 positively or negatively affects β cell function under nutritional stress, PLIN2 was down-regulated in mouse β cells, INS1 cells, and human islet cells. β cell specific deletion of PLIN2 in mice on a high fat diet reduced glucose-stimulated insulin secretion (GSIS) in vivo and in vitro. Down-regulation of PLIN2 in INS1 cells blunted GSIS after 24 h incubation with 0.2 mM palmitic acids. Down-regulation of PLIN2 in human pseudoislets cultured at 5.6 mM glucose impaired both phases of GSIS, indicating that PLIN2 is critical for GSIS. Down-regulation of PLIN2 decreased specific OXPHOS proteins in all three models and reduced oxygen consumption rates in INS1 cells and mouse islets. Moreover, we found that PLIN2 deficient INS1 cells increased the distribution of a fluorescent oleic acid analog to mitochondria and showed signs of mitochondrial stress as indicated by susceptibility to fragmentation and alterations of acyl-carnitines and glucose metabolites. Collectively, PLIN2 in β cells have an important role in preserving insulin secretion, β cell metabolism and mitochondrial function under nutritional stress.

scholarly journals Downregulation of lncRNA TUG1 Affects Apoptosis and Insulin Secretion in Mouse Pancreatic β Cells

2015 ◽  
Vol 35 (5) ◽  
pp. 1892-1904 ◽  
Author(s):  
Dan-dan Yin ◽  
Er-bao Zhang ◽  
Liang-hui You ◽  
Ning Wang ◽  
Lin-tao Wang ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Cell Function ◽  
Annexin V ◽  
Pancreatic Tissue ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Β Cells ◽  
Lncrna Tug1

Background: Increasing evidence indicates that long noncoding RNAs (IncRNAs) perform specific biological functions in diverse processes. Recent studies have reported that IncRNAs may be involved in β cell function. The aim of this study was to characterize the role of IncRNA TUG1 in mouse pancreatic β cell functioning both in vitro and in vivo. Methods: qRT-PCR analyses were performed to detect the expression of lncRNA TUG1 in different tissues. RNAi, MTT, TUNEL and Annexin V-FITC assays and western blot, GSIS, ELISA and immunochemistry analyses were performed to detect the effect of lncRNA TUG1 on cell apoptosis and insulin secretion in vitro and in vivo. Results: lncRNA TUG1 was highly expressed in pancreatic tissue compared with other organ tissues, and expression was dynamically regulated by glucose in Nit-1 cells. Knockdown of lncRNA TUG1 expression resulted in an increased apoptosis ratio and decreased insulin secretion in β cells both in vitro and in vivo . Immunochemistry analyses suggested decreased relative islet area after treatment with lncRNA TUG1 siRNA. Conclusion: Downregulation of lncRNA TUG1 expression affected apoptosis and insulin secretion in pancreatic β cells in vitro and in vivo. lncRNA TUG1 may represent a factor that regulates the function of pancreatic β cells.

scholarly journals β-Cell Differentiation from a Human Pancreatic Cell Line in Vitro and in Vivo

2001 ◽  
Vol 15 (3) ◽  
pp. 476-483 ◽  
Author(s):  
Dominique Dufayet de la Tour ◽  
Tanya Halvorsen ◽  
Carla Demeterco ◽  
Björn Tyrberg ◽  
Pamela Itkin-Ansari ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Cell Differentiation ◽  
Cell Transplantation ◽  
Cell Lines ◽  
Β Cells ◽  
Β Cell ◽  
Cell Transplantation Therapy ◽  
Transplantation Therapy

Abstract Cell transplantation therapy for diabetes is limited by an inadequate supply of cells exhibiting glucose-responsive insulin secretion. To generate an unlimited supply of human β-cells, inducibly transformed pancreatic β-cell lines have been created by expression of dominant oncogenes. The cell lines grow indefinitely but lose differentiated function. Induction of β-cell differentiation was achieved by stimulating the signaling pathways downstream of the transcription factor PDX-1, cell-cell contact, and the glucagon-like peptide (GLP-1) receptor. Synergistic activation of those pathways resulted in differentiation into functional β-cells exhibiting glucose-responsive insulin secretion in vitro. Both oncogene-expressing and oncogene-deleted cells were transplanted into nude mice and found to exhibit glucose-responsive insulin secretion in vivo. The ability to grow unlimited quantities of human β-cells is a major step toward developing a cell transplantation therapy for diabetes.

scholarly journals The HDAC Inhibitor Butyrate Impairs β Cell Function and Activates the Disallowed Gene Hexokinase I

2021 ◽  
Vol 22 (24) ◽  
pp. 13330
Author(s):  
Stephanie Bridgeman ◽  
Gaewyn Ellison ◽  
Philip Newsholme ◽  
Cyril Mamotte
Keyword(s):  
Insulin Secretion ◽  
Low Dose ◽  
Cell Function ◽  
High Dose ◽  
Hdac Inhibition ◽  
Β Cells ◽  
Β Cell ◽  
Β Cell Function

Histone deacetylase (HDAC) inhibitors such as butyrate have been reported to reduce diabetes risk and protect insulin-secreting pancreatic β cells in animal models. However, studies on insulin-secreting cells in vitro have found that butyrate treatment resulted in impaired or inappropriate insulin secretion. Our study explores the effects of butyrate on insulin secretion by BRIN BD-11 rat pancreatic β cells and examined effects on the expression of genes implicated in β cell function. Robust HDAC inhibition with 5 mM butyrate or trichostatin A for 24 h in β cells decreased basal insulin secretion and content, as well as insulin secretion in response to acute stimulation. Treatment with butyrate also increased expression of the disallowed gene hexokinase I, possibly explaining the impairment to insulin secretion, and of TXNIP, which may increase oxidative stress and β cell apoptosis. In contrast to robust HDAC inhibition (>70% after 24 h), low-dose and acute high-dose treatment with butyrate enhanced nutrient-stimulated insulin secretion. In conclusion, although protective effects of HDAC inhibition have been observed in vivo, potent HDAC inhibition impairs β cell function in vitro. The chronic low dose and acute high dose butyrate treatments may be more reflective of in vivo effects.

scholarly journals Dopamine-Mediated Autocrine Inhibitory Circuit Regulating Human Insulin Secretion in Vitro

2012 ◽  
Vol 26 (10) ◽  
pp. 1757-1772 ◽  
Author(s):  
Norman Simpson ◽  
Antonella Maffei ◽  
Matthew Freeby ◽  
Steven Burroughs ◽  
Zachary Freyberg ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Secretory Granules ◽  
Human Islets ◽  
Negative Regulator ◽  
Β Cells ◽  
Regulatory Circuit ◽  
Glucose Stimulated Insulin Secretion ◽  
Insulin Secretion In Vitro

Abstract We describe a negative feedback autocrine regulatory circuit for glucose-stimulated insulin secretion in purified human islets in vitro. Using chronoamperometry and in vitro glucose-stimulated insulin secretion measurements, evidence is provided that dopamine (DA), which is loaded into insulin-containing secretory granules by vesicular monoamine transporter type 2 in human β-cells, is released in response to glucose stimulation. DA then acts as a negative regulator of insulin secretion via its action on D2R, which are also expressed on β-cells. We found that antagonism of receptors participating in islet DA signaling generally drive increased glucose-stimulated insulin secretion. These in vitro observations may represent correlates of the in vivo metabolic changes associated with the use of atypical antipsychotics, such as increased adiposity.

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