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Sample records for interleukin-1beta-induced beta-cell apoptosis

  1. Apoptosis of beta cells in diabetes mellitus.

    PubMed

    Anuradha, Rachakatla; Saraswati, Mudigonda; Kumar, Kishore G; Rani, Surekha H

    2014-11-01

    Diabetes mellitus is a multifactorial metabolic disorder characterized by hyperglycemia. Apoptosis in beta cells has been observed in response to diverse stimuli, such as glucose, cytokines, free fatty acids, leptin, and sulfonylureas, leading to the activation of polyol, hexosamine, and diacylglycerol/protein kinase-C (DAG/PKC) pathways that mediate oxidative and nitrosative stress causing the release of different cytokines. Cytokines induce the expression of Fas and tumor necrosis factor-alpha (TNF-α) by activating the transcription factor, nuclear factor-κb, and signal transducer and activator of transcription 1 (STAT-1) in the β cells in the extrinsic pathway of apoptosis. Cytokines produced in beta cells also induce proapoptotic members of the intrinsic pathway of apoptosis. The genetic alterations in apoptosis signaling machinery and the pathogenesis of diabetes include Fas, FasL, Akt, caspases, calpain-10, and phosphatase and tensin homolog (Pten). The other gene products that are involved in diabetes are nitric oxide synthase-2 (NOS2), small ubiquitin-like modifier (SUMO), apolipoprotein CIII (ApoCIII), forkhead box protein O1 (FOXO1), and Kruppel-like zinc finger protein Gli-similar 3 (GLIS3). The gene products having antiapoptotic nature are Bcl-2 and Bcl-XL. Epigenetic mechanisms play an important role in type I and type II diabetes. Further studies on the apoptotic genes and gene products in diabetics may be helpful in pharmacogenomics and individualized treatment along with antioxidants targeting apoptosis in diabetes. PMID:25093391

  2. Foodborne Cereulide Causes Beta-Cell Dysfunction and Apoptosis

    PubMed Central

    Vangoitsenhoven, Roman; Rondas, Dieter; Crèvecoeur, Inne; D'Hertog, Wannes; Baatsen, Pieter; Masini, Matilde; Andjelkovic, Mirjana; Van Loco, Joris; Matthys, Christophe; Mathieu, Chantal; Overbergh, Lut; Van der Schueren, Bart

    2014-01-01

    Aims/Hypothesis To study the effects of cereulide, a food toxin often found at low concentrations in take-away meals, on beta-cell survival and function. Methods Cell death was quantified by Hoechst/Propidium Iodide in mouse (MIN6) and rat (INS-1E) beta-cell lines, whole mouse islets and control cell lines (HepG2 and COS-1). Beta-cell function was studied by glucose-stimulated insulin secretion (GSIS). Mechanisms of toxicity were evaluated in MIN6 cells by mRNA profiling, electron microscopy and mitochondrial function tests. Results 24 h exposure to 5 ng/ml cereulide rendered almost all MIN6, INS-1E and pancreatic islets apoptotic, whereas cell death did not increase in the control cell lines. In MIN6 cells and murine islets, GSIS capacity was lost following 24 h exposure to 0.5 ng/ml cereulide (P<0.05). Cereulide exposure induced markers of mitochondrial stress including Puma (p53 up-regulated modulator of apoptosis, P<0.05) and general pro-apoptotic signals as Chop (CCAAT/-enhancer-binding protein homologous protein). Mitochondria appeared swollen upon transmission electron microscopy, basal respiration rate was reduced by 52% (P<0.05) and reactive oxygen species increased by more than twofold (P<0.05) following 24 h exposure to 0.25 and 0.50 ng/ml cereulide, respectively. Conclusions/Interpretation Cereulide causes apoptotic beta-cell death at low concentrations and impairs beta-cell function at even lower concentrations, with mitochondrial dysfunction underlying these defects. Thus, exposure to cereulide even at concentrations too low to cause systemic effects appears deleterious to the beta-cell. PMID:25119564

  3. Angiotensin II modulates interleukin-1{beta}-induced inflammatory gene expression in vascular smooth muscle cells via interfering with ERK-NF-{kappa}B crosstalk

    SciTech Connect

    Xu, Shanqin; Zhi, Hui; Hou, Xiuyun; Jiang, Bingbing

    2011-07-08

    Highlights: {yields} We examine how angiotensin II modulates ERK-NF-{kappa}B crosstalk and gene expression. {yields} Angiotensin II suppresses IL-1{beta}-induced prolonged ERK and NF-{kappa}B activation. {yields} ERK-RSK1 signaling is required for IL-1{beta}-induced prolonged NF-{kappa}B activation. {yields} Angiotensin II modulates NF-{kappa}B responsive genes via regulating ERK-NF-{kappa}B crosstalk. {yields} ERK-NF-{kappa}B crosstalk is a novel mechanism regulating inflammatory gene expression. -- Abstract: Angiotensin II is implicated in cardiovascular diseases, which is associated with a role in increasing vascular inflammation. The present study investigated how angiotensin II modulates vascular inflammatory signaling and expression of inducible nitric oxide synthase (iNOS) and vascular cell adhesion molecule (VCAM)-1. In cultured rat aortic vascular smooth muscle cells (VSMCs), angiotensin II suppressed interleukin-1{beta}-induced prolonged phosphorylation of extracellular signal-regulated kinase (ERK) and ribosomal S6 kinase (RSK)-1, and nuclear translocation of nuclear factor (NF)-{kappa}B, leading to decreased iNOS but enhanced VCAM-1 expression, associated with an up-regulation of mitogen-activated protein kinase phosphatase-1 expression. Knock-down of RSK1 selectively down regulated interleukin-1{beta}-induced iNOS expression without influencing VCAM-1 expression. In vivo experiments showed that interleukin-1{beta}, iNOS, and VCAM-1 expression were detectable in the aortic arches of both wild-type and apolipoprotein E-deficient (ApoE{sup -/-}) mice. VCAM-1 and iNOS expression were higher in ApoE{sup -/-} than in wild type mouse aortic arches. Angiotensin II infusion (3.2 mg/kg/day, for 6 days, via subcutaneous osmotic pump) in ApoE{sup -/-} mice enhanced endothelial and adventitial VCAM-1 and iNOS expression, but reduced medial smooth muscle iNOS expression associated with reduced phosphorylation of ERK and RSK-1. These results indicate that angiotensin

  4. MST1 is a novel regulator of apoptosis in pancreatic beta-cells

    PubMed Central

    Ardestani, Amin; Khobragade, Vrushali; Yuan, Ting; Frogne, Thomas; Tao, Wufan; Oberholzer, Jose; Pattou, Francois; Conte, Julie Kerr; Maedler, Kathrin

    2014-01-01

    Apoptotic cell death is a hallmark of the loss of insulin producing beta-cells in all forms of diabetes mellitus. Current treatment fails to halt the decline in functional beta-cell mass. Strategies to prevent beta-cell apoptosis and dysfunction are urgently needed. Here, we identified Mammalian Sterile 20-like kinase 1 (MST1) as a critical regulator of apoptotic beta-cell death and function. MST1 was strongly activated in beta-cells under diabetogenic conditions and correlated with beta-cell apoptosis. MST1 specifically induced the mitochondrial-dependent pathway of apoptosis in beta-cells through up-regulation of the BH3-only protein Bim. MST1 directly phosphorylated PDX1 at Thr11, resulting in its ubiquitination, degradation and impaired insulin secretion. Mst1 deficiency completely restored normoglycemia, beta-cell function and survival in vitro and in vivo. We show MST1 as novel pro-apoptotic kinase and key mediator of apoptotic signaling and beta-cell dysfunction, which may serve as target for the development of novel therapies for diabetes. PMID:24633305

  5. Cocoa-rich diet attenuates beta cell mass loss and function in young Zucker diabetic fatty rats by preventing oxidative stress and beta cell apoptosis.

    PubMed

    Fernández-Millán, Elisa; Cordero-Herrera, Isabel; Ramos, Sonia; Escrivá, Fernando; Alvarez, Carmen; Goya, Luis; Martín, María Angeles

    2015-04-01

    We have recently shown that cocoa flavanols may have anti-diabetic potential by promoting survival and function of pancreatic beta-cells in vitro. In this work, we investigated if a cocoa-rich diet is able to preserve beta-cell mass and function in an animal model of type 2 diabetes and the mechanisms involved. Our results showed that cocoa feeding during the prediabetic state attenuates hyperglycaemia, reduces insulin resistant, and increases beta cell mass and function in obese Zucker diabetic rats. At the molecular level, cocoa-rich diet prevented beta-cell apoptosis by increasing the levels of Bcl-xL and decreasing Bax levels and caspase-3 activity. Cocoa diet enhanced the activity of endogenous antioxidant defenses, mainly glutathione peroxidase, preventing thus oxidative injury induced by the pre-diabetic condition and leading to apoptosis prevention. These findings provide the first in vivo evidence that a cocoa-rich diet may delay the loss of functional beta-cell mass and delay the progression of diabetes by preventing oxidative stress and beta-cell apoptosis.

  6. Cocoa-rich diet attenuates beta cell mass loss and function in young Zucker diabetic fatty rats by preventing oxidative stress and beta cell apoptosis.

    PubMed

    Fernández-Millán, Elisa; Cordero-Herrera, Isabel; Ramos, Sonia; Escrivá, Fernando; Alvarez, Carmen; Goya, Luis; Martín, María Angeles

    2015-04-01

    We have recently shown that cocoa flavanols may have anti-diabetic potential by promoting survival and function of pancreatic beta-cells in vitro. In this work, we investigated if a cocoa-rich diet is able to preserve beta-cell mass and function in an animal model of type 2 diabetes and the mechanisms involved. Our results showed that cocoa feeding during the prediabetic state attenuates hyperglycaemia, reduces insulin resistant, and increases beta cell mass and function in obese Zucker diabetic rats. At the molecular level, cocoa-rich diet prevented beta-cell apoptosis by increasing the levels of Bcl-xL and decreasing Bax levels and caspase-3 activity. Cocoa diet enhanced the activity of endogenous antioxidant defenses, mainly glutathione peroxidase, preventing thus oxidative injury induced by the pre-diabetic condition and leading to apoptosis prevention. These findings provide the first in vivo evidence that a cocoa-rich diet may delay the loss of functional beta-cell mass and delay the progression of diabetes by preventing oxidative stress and beta-cell apoptosis. PMID:25559866

  7. Proteasome Dysfunction Mediates High Glucose-Induced Apoptosis in Rodent Beta Cells and Human Islets

    PubMed Central

    Broca, Christophe; Varin, Elodie; Armanet, Mathieu; Tourrel-Cuzin, Cécile; Bosco, Domenico; Dalle, Stéphane; Wojtusciszyn, Anne

    2014-01-01

    The ubiquitin/proteasome system (UPS), a major cellular protein degradation machinery, plays key roles in the regulation of many cell functions. Glucotoxicity mediated by chronic hyperglycaemia is detrimental to the function and survival of pancreatic beta cells. The aim of our study was to determine whether proteasome dysfunction could be involved in beta cell apoptosis in glucotoxic conditions, and to evaluate whether such a dysfunction might be pharmacologically corrected. Therefore, UPS activity was measured in GK rats islets, INS-1E beta cells or human islets after high glucose and/or UPS inhibitor exposure. Immunoblotting was used to quantify polyubiquitinated proteins, endoplasmic reticulum (ER) stress through CHOP expression, and apoptosis through the cleavage of PARP and caspase-3, whereas total cell death was detected through histone-associated DNA fragments measurement. In vitro, we found that chronic exposure of INS-1E cells to high glucose concentrations significantly decreases the three proteasome activities by 20% and leads to caspase-3-dependent apoptosis. We showed that pharmacological blockade of UPS activity by 20% leads to apoptosis in a same way. Indeed, ER stress was involved in both conditions. These results were confirmed in human islets, and proteasome activities were also decreased in hyperglycemic GK rats islets. Moreover, we observed that a high glucose treatment hypersensitized beta cells to the apoptotic effect of proteasome inhibitors. Noteworthily, the decreased proteasome activity can be corrected with Exendin-4, which also protected against glucotoxicity-induced apoptosis. Taken together, our findings reveal an important role of proteasome activity in high glucose-induced beta cell apoptosis, potentially linking ER stress and glucotoxicity. These proteasome dysfunctions can be reversed by a GLP-1 analog. Thus, UPS may be a potent target to treat deleterious metabolic conditions leading to type 2 diabetes. PMID:24642635

  8. T cells cooperate with palmitic acid in induction of beta cell apoptosis

    PubMed Central

    Cvjetićanin, Tamara; Stojanović, Ivana; Timotijević, Gordana; Stošić-Grujičić, Stanislava; Miljković, Djordje

    2009-01-01

    Background Diabetes is characterized by progressive failure of insulin producing beta cells. It is well known that both saturated fatty acids and various products of immune cells can contribute to the reduction of beta cell viability and functionality during diabetes pathogenesis. However, their joint action on beta cells has not been investigated, so far. Therefore, we explored the possibility that leukocytes and saturated fatty acids cooperate in beta cell destruction. Results Rat pancreatic islets or insulinoma cells (RIN) were co-cultivated with concanavalin A (ConA)-stimulated rat lymph node cells (LNC), or they were treated with cell-free supernatants (Sn) obtained from ConA-stimulated spleen cells or from activated CD3+ cells, in the absence or presence of palmitic acid (PA). ConA-stimulated LNC or Sn and PA cooperated in inducing caspase-3-dependent RIN cell apoptosis. The observed effect of PA and Sn on RIN cell viability was mediated by p38 mitogen-activated protein kinase (MAPK)-signaling and was achieved through auto-destructive nitric oxide (NO) production. The cooperative effect of Sn was mimicked with the combination of interleukin-1β, interleukin-2, interleukin-6, interleukin-17, interferon-γ and tumor necrosis factor-α. Conclusion These results imply that stimulated T cells produce cytokines that cooperate with saturated free fatty acids in beta cell destruction during diabetes pathogenesis. PMID:19463182

  9. Baculovirus p35 increases pancreatic {beta}-cell resistance to apoptosis

    SciTech Connect

    Hollander, Kenneth; Bar-Chen, Michal; Efrat, Shimon . E-mail: sefrat@post.tau.ac.il

    2005-07-01

    {beta}-cells die by apoptosis in type 1 diabetes as a result of autoimmune attack mediated by cytokines, and in type 2 diabetes by various perpetrators including human islet amyloid polypeptide (hIAPP). The cascade of apoptotic events induced by cytokines and hIAPP is mediated through caspases and reactive oxygen species. The baculovirus p35 protein is a potent anti-apoptotic agent shown to be effective in a variety of species and able to inhibit a number of apoptotic pathways. Here, we aimed at determining the protective potential of p35 in {beta}-cells exposed to cytokines and hIAPP, as well as the effects of p35 on {beta}-cell function. The p35 gene was introduced into {beta}TC-tet cells, a differentiated murine {beta}-cell line capable of undergoing inducible growth-arrest. Both proliferating and growth-arrested cells expressing p35 manifested increased resistance to cytokines and hIAPP, compared with control cells, as judged by cell viability, DNA fragmentation, and caspase-3 activity assays. p35 was significantly more protective in growth-arrested, compared with proliferating, cells. No significant differences were observed in proliferation and insulin content between cells expressing p35 and control cells. In contrast, p35 manifested a perturbing effect on glucose-induced insulin secretion. These findings suggest that p35 could be incorporated as part of a multi-pronged approach of immunoprotective strategies to provide protection from recurring autoimmunity for transplanted {beta}-cells, as well as in preventive gene therapy in type 1 diabetes. p35 may also be protective from {beta}-cell damage caused by hIAPP in type 2 diabetes.

  10. Glucagon-like peptide-1 prevents methylglyoxal-induced apoptosis of beta cells through improving mitochondrial function and suppressing prolonged AMPK activation

    PubMed Central

    Chang, Tien-Jyun; Tseng, Hsing-Chi; Liu, Meng-Wei; Chang, Yi-Cheng; Hsieh, Meng-Lun; Chuang, Lee-Ming

    2016-01-01

    Accumulation of methylglyoxal (MG) contributes to glucotoxicity and mediates beta cell apoptosis. The molecular mechanism by which GLP-1 protects MG-induced beta cell apoptosis remains unclear. Metformin is a first-line drug for treating type 2 diabetes associated with AMPK activation. However, whether metformin prevents MG-induced beta cell apoptosis is controversial. Here, we explored the signaling pathway involved in the anti-apoptotic effect of GLP-1, and investigated whether metformin had an anti-apoptotic effect on beta cells. MG treatment induced apoptosis of beta cells, impaired mitochondrial function, and prolonged activation of AMP-dependent protein kinase (AMPK). The MG-induced pro-apoptotic effects were abolished by an AMPK inhibitor. Pretreatment of GLP-1 reversed MG-induced apoptosis, and mitochondrial dysfunction, and suppressed prolonged AMPK activation. Pretreatment of GLP-1 reversed AMPK activator 5-aminoimidazole-4-carboxamide riboside (AICAR)-induced apoptosis, and suppressed prolonged AMPK activation. However, metformin neither leads to beta cell apoptosis nor ameliorates MG-induced beta cell apoptosis. In parallel, GLP-1 also prevents MG-induced beta cell apoptosis through PKA and PI3K-dependent pathway. In conclusion, these data indicates GLP-1 but not metformin protects MG-induced beta cell apoptosis through improving mitochondrial function, and alleviating the prolonged AMPK activation. Whether adding GLP-1 to metformin provides better beta cell survival and delays disease progression remains to be validated. PMID:26997114

  11. Glucagon-like peptide-1 prevents methylglyoxal-induced apoptosis of beta cells through improving mitochondrial function and suppressing prolonged AMPK activation.

    PubMed

    Chang, Tien-Jyun; Tseng, Hsing-Chi; Liu, Meng-Wei; Chang, Yi-Cheng; Hsieh, Meng-Lun; Chuang, Lee-Ming

    2016-01-01

    Accumulation of methylglyoxal (MG) contributes to glucotoxicity and mediates beta cell apoptosis. The molecular mechanism by which GLP-1 protects MG-induced beta cell apoptosis remains unclear. Metformin is a first-line drug for treating type 2 diabetes associated with AMPK activation. However, whether metformin prevents MG-induced beta cell apoptosis is controversial. Here, we explored the signaling pathway involved in the anti-apoptotic effect of GLP-1, and investigated whether metformin had an anti-apoptotic effect on beta cells. MG treatment induced apoptosis of beta cells, impaired mitochondrial function, and prolonged activation of AMP-dependent protein kinase (AMPK). The MG-induced pro-apoptotic effects were abolished by an AMPK inhibitor. Pretreatment of GLP-1 reversed MG-induced apoptosis, and mitochondrial dysfunction, and suppressed prolonged AMPK activation. Pretreatment of GLP-1 reversed AMPK activator 5-aminoimidazole-4-carboxamide riboside (AICAR)-induced apoptosis, and suppressed prolonged AMPK activation. However, metformin neither leads to beta cell apoptosis nor ameliorates MG-induced beta cell apoptosis. In parallel, GLP-1 also prevents MG-induced beta cell apoptosis through PKA and PI3K-dependent pathway. In conclusion, these data indicates GLP-1 but not metformin protects MG-induced beta cell apoptosis through improving mitochondrial function, and alleviating the prolonged AMPK activation. Whether adding GLP-1 to metformin provides better beta cell survival and delays disease progression remains to be validated. PMID:26997114

  12. Nuclear orphan receptor TLX affects gene expression, proliferation and cell apoptosis in beta cells.

    PubMed

    Shi, Xiaoli; Xiong, Xiaokan; Dai, Zhe; Deng, Haohua; Sun, Li; Hu, Xuemei; Zhou, Feng; Xu, Yancheng

    Nuclear orphan receptor TLX is an essential regulator of the growth of neural stem cells. However, its exact function in pancreatic islet cells is still unknown. In the present study, gene expression profiling analysis revealed that overexpression of TLX in beta cell line MIN6 causes suppression of 176 genes and upregulation of 49 genes, including a cadre of cell cycle, cell proliferation and cell death control genes, such as Btg2, Ddit3 and Gadd45a. We next examined the effects of TLX overexpression on proliferation, apoptosis and insulin secretion in MIN6 cells. Proliferation analysis using EdU assay showed that overexpression of TLX increased percentage of EdU-positive cells. Cell cycle and apoptosis analysis revealed that overexpression of TLX in MIN6 cells resulted in higher percentage of cells exiting G1 into S-phase, and a 58.8% decrease of cell apoptosis induced by 0.5 mM palmitate. Moreover, TLX overexpression did not cause impairment of insulin secretion. Together, we conclude that TLX is among factors capable of controlling beta cell proliferation and survival, which may serve as a target for the development of novel therapies for diabetes.

  13. Interleukin-1beta induces death in chondrocyte-like ATDC5 cells through mitochondrial dysfunction and energy depletion in a reactive nitrogen and oxygen species-dependent manner.

    PubMed

    Yasuhara, Rika; Miyamoto, Yoichi; Akaike, Takaaki; Akuta, Teruo; Nakamura, Masanori; Takami, Masamichi; Morimura, Naoko; Yasu, Kayoko; Kamijo, Ryutaro

    2005-07-15

    IL-1 (interleukin-1) acts as a key mediator of the degeneration of articular cartilage in RA (rheumatoid arthritis) and OA (osteoarthritis),where chondrocyte death is observed. It is still controversial, however, whether IL-1 induces chondrocyte death. In the present study, the viability of mouse chondrocyte-like ATDC5 cells was reduced by the treatment with IL-1beta for 48 h or longer. IL-1beta augmented the expression of the catalytic gp91 subunit of NADPH oxidase, gp91phox, as well as inducible NO synthase in ATDC5 cells. Generation of nitrated guanosine and tyrosine suggested the formation of reactive nitrogen species including ONOO- (peroxynitrite), a reaction product of NO and O2-, in ATDC5 cells and rat primary chondrocytes treated with IL-1beta. Death of ATDC5 cells after IL-1beta treatment was prevented by an NADPH-oxidase inhibitor, AEBSF[4-(2-aminoethyl)benzene-sulphonyl fluoride], an NO synthase inhibitor, L-NAME (NG-nitro-L-arginine methyl ester), and a ONOO- scavenger, uric acid. The viability of ATDC5 cells was reduced by the ONOO(-)-generator 3-(4-morpholinyl)sydnonimine hydrochloride, but not by either the NO-donor 1-hydroxy-2-oxo-3-(N-methyl-2-aminopropyl)-3-methyl-1-triazene or S-nitrosoglutathione. Disruption of mitochondrial membrane potential and ATP deprivation were observed in IL-1beta-treated ATDC5 cells, both of which were restored by L-NAME, AEBSF or uric acid. On the other hand, no morphological or biochemical signs indicating apoptosis were observed in these cells. These results suggest that the death of chondrocyte-like ATDC5 cells was mediated at least in part by mitochondrial dysfunction and energy depletion through ONOO- formation after IL-1beta treatment. PMID:15784009

  14. Small-molecule suppressors of cytokine-induced beta-cell apoptosis

    PubMed Central

    Chou, Danny Hung-Chieh; Bodycombe, Nicole E.; Carrinski, Hyman A.; Lewis, Timothy A.; Clemons, Paul A.; Schreiber, Stuart L.; Wagner, Bridget K.

    2010-01-01

    Pancreatic beta-cell apoptosis is a critical event during the development of type-1 diabetes. The identification of small molecules capable of preventing cytokine-induced apoptosis could lead to avenues for therapeutic intervention. We developed a set of phenotypic cell-based assays designed to identify such small-molecule suppressors. Rat INS-1E cells were simultaneously treated with a cocktail of inflammatory cytokines and a collection of 2,240 diverse small molecules, and screened using an assay for cellular ATP levels. Forty-nine top-scoring compounds included glucocorticoids, several pyrazole derivatives, and known inhibitors of glycogen synthase kinase-3β. Two compounds were able to increase cellular ATP levels, reduce caspase-3 activity and nitrite production, and increase glucose-stimulated insulin secretion in the presence of cytokines. These results indicate that small molecules identified by this screening approach may protect beta cells from autoimmune attack, and may be good candidates for therapeutic intervention in early stages of type-1 diabetes. PMID:20550176

  15. Endoplasmic reticulum stress sensitizes pancreatic beta cells to interleukin-1β-induced apoptosis via Bim/A1 imbalance.

    PubMed

    Miani, M; Barthson, J; Colli, M L; Brozzi, F; Cnop, M; Eizirik, D L

    2013-01-01

    We have recently shown that the crosstalk between mild endoplasmic reticulum (ER) stress and low concentrations of the pro-inflammatory cytokine interleukin (IL)-1β exacerbates beta cell inflammatory responses via the IRE1α/XBP1 pathway. We presently investigated whether mild ER stress also sensitizes beta cells to cytokine-induced apoptosis. Cyclopiazonic acid (CPA)-induced ER stress enhanced the IL-1β apoptosis in INS-1E and primary rat beta cells. This was not prevented by XBP1 knockdown (KD), indicating the dissociation between the pathways leading to inflammation and cell death. Analysis of the role of pro- and anti-apoptotic proteins in cytokine-induced apoptosis indicated a central role for the pro-apoptotic BH3 (Bcl-2 homology 3)-only protein Bim (Bcl-2-interacting mediator of cell death), which was counteracted by four anti-apoptotic Bcl-2 (B-cell lymphoma-2) proteins, namely Bcl-2, Bcl-XL, Mcl-1 and A1. CPA+IL-1β-induced beta cell apoptosis was accompanied by increased expression of Bim, particularly the most pro-apoptotic variant, small isoform of Bim (BimS), and decreased expression of A1. Bim silencing protected against CPA+IL-1β-induced apoptosis, whereas A1 KD aggravated cell death. Bim inhibition protected against cell death caused by A1 silencing under all conditions studied. In conclusion, mild ER stress predisposes beta cells to the pro-apoptotic effects of IL-1β by disrupting the balance between pro- and anti-apoptotic Bcl-2 proteins. These findings link ER stress to exacerbated apoptosis during islet inflammation and provide potential mechanistic targets for beta cell protection, namely downregulation of Bim and upregulation of A1. PMID:23828564

  16. The Cytotoxic Role of Intermittent High Glucose on Apoptosis and Cell Viability in Pancreatic Beta Cells

    PubMed Central

    Zhang, Zhen; Li, Jing; Yang, Lei; Chen, Rongping; Yang, Rui; Zhang, Hua; Cai, Dehong; Chen, Hong

    2014-01-01

    Objectives. Glucose fluctuations are both strong predictor of diabetic complications and crucial factor for beta cell damages. Here we investigated the effect of intermittent high glucose (IHG) on both cell apoptosis and proliferation activity in INS-1 cells and the potential mechanisms. Methods. Cells were treated with normal glucose (5.5 mmol/L), constant high glucose (CHG) (25 mmol/L), and IHG (rotation per 24 h in 11.1 or 25 mmol/L) for 7 days. Reactive oxygen species (ROS), xanthine oxidase (XOD) level, apoptosis, cell viability, cell cycle, and expression of cyclinD1, p21, p27, and Skp2 were determined. Results. We found that IHG induced more significant apoptosis than CHG and normal glucose; intracellular ROS and XOD levels were more markedly increased in cells exposed to IHG. Cells treated with IHG showed significant decreased cell viability and increased cell proportion in G0/G1 phase. Cell cycle related proteins such as cyclinD1 and Skp2 were decreased significantly, but expressions of p27 and p21 were increased markedly. Conclusions. This study suggested that IHG plays a more toxic effect including both apoptosis-inducing and antiproliferative effects on INS-1 cells. Excessive activation of cellular stress and regulation of cyclins might be potential mechanism of impairment in INS-1 cells induced by IHG. PMID:24772447

  17. Overexpression of IRS2 in isolated pancreatic islets causes proliferation and protects human {beta}-cells from hyperglycemia-induced apoptosis

    SciTech Connect

    Mohanty, S.; Spinas, G.A.; Maedler, K.; Zuellig, R.A.; Lehmann, R.; Donath, M.Y.; Trueb, T.; Niessen, M. . E-mail: markus.niessen@usz.ch

    2005-02-01

    Studies in vivo indicate that IRS2 plays an important role in maintaining functional {beta}-cell mass. To investigate if IRS2 autonomously affects {beta}-cells, we have studied proliferation, apoptosis, and {beta}-cell function in isolated rat and human islets after overexpression of IRS2 or IRS1. We found that {beta}-cell proliferation was significantly increased in rat islets overexpressing IRS2 while IRS1 was less effective. Moreover, proliferation of a {beta}-cell line, INS-1, was decreased after repression of Irs2 expression using RNA oligonucleotides. Overexpression of IRS2 in human islets significantly decreased apoptosis of {beta}-cells, induced by 33.3 mM D-glucose. However, IRS2 did not protect cultured rat islets against apoptosis in the presence of 0.5 mM palmitic acid. Overexpression of IRS2 in isolated rat islets significantly increased basal and D-glucose-stimulated insulin secretion as determined in perifusion experiments. Therefore, IRS2 is sufficient to induce proliferation in rat islets and to protect human {beta}-cells from D-glucose-induced apoptosis. In addition, IRS2 can improve {beta}-cell function. Our results indicate that IRS2 acts autonomously in {beta}-cells in maintenance and expansion of functional {beta}-cell mass in vivo.

  18. Pancreatic-derived factor (FAM3B), a novel islet cytokine, induces apoptosis of insulin-secreting beta-cells.

    PubMed

    Cao, Xiaopei; Gao, Zhiyong; Robert, Claudia E; Greene, Scott; Xu, Gang; Xu, Weizhen; Bell, Ewan; Campbell, Don; Zhu, Yuan; Young, Robert; Trucco, Matteo; Markmann, James F; Naji, Ali; Wolf, Bryan A

    2003-09-01

    PANDER (PANcreatic DERived factor, FAM3B), a newly discovered secreted cytokine, is specifically expressed at high levels in the islets of Langerhans of the endocrine pancreas. To evaluate the role of PANDER in beta-cell function, we investigated the effects of PANDER on rat, mouse, and human pancreatic islets; the beta-TC3 cell line; and the alpha-TC cell line. PANDER protein was present in alpha- and beta-cells of pancreatic islets, insulin-secreting beta-TC3 cells, and glucagon-secreting alpha-TC cells. PANDER induced islet cell death in rat and human islets. Culture of beta-TC3 cells with recombinant PANDER had a dose-dependent inhibitory effect on cell viability. This effect was also time-dependent. PANDER caused apoptosis of beta-cells as assessed by electron microscopy, annexin V fluorescent staining, and flow-cytometric terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay. PANDER did not affect cytosolic Ca(2+) levels or nitric oxide levels. However, PANDER activated caspase-3. Hence, PANDER may have a role in the process of pancreatic beta-cell apoptosis.

  19. Effects of low intensity laser acupoint irradiation on inhibiting islet beta-cell apoptosis in rats with type 2 diabetes

    NASA Astrophysics Data System (ADS)

    Xiong, Guoxin; Xiong, Leilei; Li, Xinzhong

    2016-09-01

    To investigate the effects of low intensity semiconductor laser acupoint irradiation on inhibiting islet beta-cell apoptosis in rats with type 2 diabetes, a method using a high-fat diet and low-dose intraperitoneal injections of streptozotocin established a type 2 diabetes mellitus rat model. Model rats were randomly divided into a laser acupoint irradiation group, rosiglitazone control group, and placebo group; each group had 10 rats. In addition, 10 normal male rats were selected for the normal control group. The Housanli, Neiting and Yishu acupoints of the rats in the laser acupoint irradiation group were irradiated with a 10 mW semiconductor laser; each point was irradiated for 15 min, once every 2 d over 28 d, for a total of 14 episodes of irradiation. The rosiglitazone group rats were given rosiglitazone (0.2 mg kg-1) intragastrically; the placebo group rats were given 0.9% brine (0.2 mg kg-1) intragastrically, once daily, for four consecutive weeks. The change of fasting blood glucose was determined before and after each treatment. The islet beta-cell apoptosis was determined. The islet beta-cell apoptosis rates of the laser acupoint irradiation group and the rosiglitazone group were significantly lower than the rate of the placebo group. Even though the rate was lower in the laser acupoint irradiation group than in the rosiglitazone group, there was no significant difference between them. It is shown that acupoint irradiation with a semiconductor laser can effectively inhibit islet beta-cell apoptosis in rats with type 2 diabetes.

  20. Effects of low intensity laser acupoint irradiation on inhibiting islet beta-cell apoptosis in rats with type 2 diabetes

    NASA Astrophysics Data System (ADS)

    Xiong, Guoxin; Xiong, Leilei; Li, Xinzhong

    2016-09-01

    To investigate the effects of low intensity semiconductor laser acupoint irradiation on inhibiting islet beta-cell apoptosis in rats with type 2 diabetes, a method using a high-fat diet and low-dose intraperitoneal injections of streptozotocin established a type 2 diabetes mellitus rat model. Model rats were randomly divided into a laser acupoint irradiation group, rosiglitazone control group, and placebo group; each group had 10 rats. In addition, 10 normal male rats were selected for the normal control group. The Housanli, Neiting and Yishu acupoints of the rats in the laser acupoint irradiation group were irradiated with a 10 mW semiconductor laser; each point was irradiated for 15 min, once every 2 d over 28 d, for a total of 14 episodes of irradiation. The rosiglitazone group rats were given rosiglitazone (0.2 mg kg‑1) intragastrically; the placebo group rats were given 0.9% brine (0.2 mg kg‑1) intragastrically, once daily, for four consecutive weeks. The change of fasting blood glucose was determined before and after each treatment. The islet beta-cell apoptosis was determined. The islet beta-cell apoptosis rates of the laser acupoint irradiation group and the rosiglitazone group were significantly lower than the rate of the placebo group. Even though the rate was lower in the laser acupoint irradiation group than in the rosiglitazone group, there was no significant difference between them. It is shown that acupoint irradiation with a semiconductor laser can effectively inhibit islet beta-cell apoptosis in rats with type 2 diabetes.

  1. Free fatty acids and cytokines induce pancreatic beta-cell apoptosis by different mechanisms: role of nuclear factor-kappaB and endoplasmic reticulum stress.

    PubMed

    Kharroubi, Ilham; Ladrière, Laurence; Cardozo, Alessandra K; Dogusan, Zeynep; Cnop, Miriam; Eizirik, Décio L

    2004-11-01

    Apoptosis is probably the main form of beta-cell death in both type 1 diabetes mellitus (T1DM) and T2DM. In T1DM, cytokines contribute to beta-cell destruction through nuclear factor-kappaB (NF-kappaB) activation. Previous studies suggested that in T2DM high glucose and free fatty acids (FFAs) are beta-cell toxic also via NF-kappaB activation. The aims of this study were to clarify whether common mechanisms are involved in FFA- and cytokine-induced beta-cell apoptosis and determine whether TNFalpha, an adipocyte-derived cytokine, potentiates FFA toxicity through enhanced NF-kappaB activation. Apoptosis was induced in insulinoma (INS)-1E cells, rat islets, and fluorescence-activated cell sorting-purified beta-cells by oleate, palmitate, and/or cytokines (IL-1beta, interferon-gamma, TNFalpha). Palmitate and IL-1beta induced a similar percentage of apoptosis in INS-1E cells, whereas oleate was less toxic. TNFalpha did not potentiate FFA toxicity in primary beta-cells. The NF-kappaB-dependent genes inducible nitric oxide synthase and monocyte chemoattractant protein-1 were induced by IL-1beta but not by FFAs. Cytokines activated NF-kappaB in INS-1E and beta-cells, but FFAs did not. Moreover, FFAs did not enhance NF-kappaB activation by TNFalpha. Palmitate and oleate induced C/EBP homologous protein, activating transcription factor-4, and immunoglobulin heavy chain binding protein mRNAs, X-box binding protein-1 alternative splicing, and activation of the activating transcription factor-6 promoter in INS-1E cells, suggesting that FFAs trigger an endoplasmic reticulum (ER) stress response. We conclude that apoptosis is the main mode of FFA- and cytokine-induced beta-cell death but the mechanisms involved are different. Whereas cytokines induce NF-kappaB activation and ER stress (secondary to nitric oxide formation), FFAs activate an ER stress response via an NF-kappaB- and nitric oxide-independent mechanism. Our results argue against a unifying hypothesis for the

  2. Maternal antioxidants prevent beta cell apoptosis and promote formation of dual hormone-expressing endocrine cells in male offspring following fetal and neonatal nicotine exposure

    PubMed Central

    BRUIN, Jennifer E; WOYNILLOWICZ, Amanda K; HETTINGA, Bart P; TARNOPOLSKY, Mark A; MORRISON, Katherine M; GERSTEIN, Hertzel C; HOLLOWAY, Alison C

    2013-01-01

    Aim Fetal and neonatal nicotine exposure causes beta cell oxidative stress and apoptosis in neonates, leading to adult-onset dysglycemia. The goal of this study was to determine whether an antioxidant intervention could prevent nicotine-induced beta cell loss. Methods Nulliparous female Wistar rats received daily subcutaneous injections of either saline or nicotine bitartrate (1.0 mg/kg/d) for 2 weeks prior to mating until weaning. Nicotine-exposed dams received either normal chow or diet containing antioxidants (1000 IU/kg vitamin E, 0.25% w/w coenzyme Q10 and 0.1% w/w alpha-lipoic acid) during mating, pregnancy and lactation; saline-exposed dams received normal chow. Pancreas tissue was collected from male offspring at 3 weeks of age to measure beta cell fraction, apoptosis, proliferation and the presence of cells co-expressing insulin and glucagon. Results The birth weight of the offspring born to nicotine-exposed dams receiving dietary antioxidants was significantly reduced. Most interestingly, the antioxidant intervention to nicotine-exposed dams prevented the beta cell loss and apoptosis observed in nicotine exposed male offspring whose mothers did not receive antioxidants. Male pups born to nicotine-treated mothers receiving antioxidants also had a trend towards increased beta cell proliferation and a significant increase in islets containing insulin/glucagon bi-hormonal cells relative to the other two treatment groups. Conclusion This study demonstrates that exposure to maternal antioxidants protects beta cells from the damaging effects of nicotine thus preserving beta cell mass. PMID:22385833

  3. Exendin-4 Protects Mitochondria from Reactive Oxygen Species Induced Apoptosis in Pancreatic Beta Cells

    PubMed Central

    Li, Zhen; Zhou, Zhiguang; Huang, Gan; Hu, Fang; Xiang, Yufei; He, Lining

    2013-01-01

    Objective Mitochondrial oxidative stress is the basis for pancreatic β-cell apoptosis and a common pathway for numerous types of damage, including glucotoxicity and lipotoxicity. We cultivated mice pancreatic β-cell tumor Min6 cell lines in vitro and observed pancreatic β-cell apoptosis and changes in mitochondrial function before and after the addition of Exendin-4. Based on these observations, we discuss the protective role of Exendin-4 against mitochondrial oxidative damage and its relationship with Ca2+-independent phospholipase A2. Methods We established a pancreatic β-cell oxidative stress damage model using Min6 cell lines cultured in vitro with tert-buty1 hydroperoxide and hydrogen peroxide. We then added Exendin-4 to observe changes in the rate of cell apoptosis (Annexin-V-FITC-PI staining flow cytometry and DNA ladder). We detected the activity of the caspase 3 and 8 apoptotic factors, measured the mitochondrial membrane potential losses and reactive oxygen species production levels, and detected the expression of cytochrome c and Smac/DLAMO in the cytosol and mitochondria, mitochondrial Ca2-independent phospholipase A2 and Ca2+-independent phospholipase A2 mRNA. Results The time-concentration curve showed that different percentages of apoptosis occurred at different time-concentrations in tert-buty1 hydroperoxide- and hydrogen peroxide-induced Min6 cells. Incubation with 100 µmol/l of Exendin-4 for 48 hours reduced the Min6 cell apoptosis rate (p<0.05). The mitochondrial membrane potential loss and total reactive oxygen species levels decreased (p<0.05), and the release of cytochrome c and Smac/DLAMO from the mitochondria was reduced. The study also showed that Ca2+-independent phospholipase A2 activity was positively related to Exendin-4 activity. Conclusion Exendin-4 reduces Min6 cell oxidative damage and the cell apoptosis rate, which may be related to Ca2-independent phospholipase A2. PMID:24204601

  4. JNK1 Protects against Glucolipotoxicity-Mediated Beta-Cell Apoptosis

    PubMed Central

    Prause, Michala; Christensen, Dan Ploug; Billestrup, Nils; Mandrup-Poulsen, Thomas

    2014-01-01

    Pancreatic β-cell dysfunction is central to type 2 diabetes pathogenesis. Prolonged elevated levels of circulating free-fatty acids and hyperglycemia, also termed glucolipotoxicity, mediate β-cell dysfunction and apoptosis associated with increased c-Jun N-terminal Kinase (JNK) activity. Endoplasmic reticulum (ER) and oxidative stress are elicited by palmitate and high glucose concentrations further potentiating JNK activity. Our aim was to determine the role of the JNK subtypes JNK1, JNK2 and JNK3 in palmitate and high glucose-induced β-cell apoptosis. We established insulin-producing INS1 cell lines stably expressing JNK subtype specific shRNAs to understand the differential roles of the individual JNK isoforms. JNK activity was increased after 3 h of palmitate and high glucose exposure associated with increased expression of ER and mitochondrial stress markers. JNK1 shRNA expressing INS1 cells showed increased apoptosis and cleaved caspase 9 and 3 compared to non-sense shRNA expressing control INS1 cells when exposed to palmitate and high glucose associated with increased CHOP expression, ROS formation and Puma mRNA expression. JNK2 shRNA expressing INS1 cells did not affect palmitate and high glucose induced apoptosis or ER stress markers, but increased Puma mRNA expression compared to non-sense shRNA expressing INS1 cells. Finally, JNK3 shRNA expressing INS1 cells did not induce apoptosis compared to non-sense shRNA expressing INS1 cells when exposed to palmitate and high glucose but showed increased caspase 9 and 3 cleavage associated with increased DP5 and Puma mRNA expression. These data suggest that JNK1 protects against palmitate and high glucose-induced β-cell apoptosis associated with reduced ER and mitochondrial stress. PMID:24475223

  5. JunB Inhibits ER Stress and Apoptosis in Pancreatic Beta Cells

    PubMed Central

    Gurzov, Esteban N.; Ortis, Fernanda; Bakiri, Latifa; Wagner, Erwin F.; Eizirik, Decio L.

    2008-01-01

    Cytokines contribute to pancreatic β-cell apoptosis in type 1 diabetes (T1D) by modulation of β-cell gene expression networks. The transcription factor Activator Protein-1 (AP-1) is a key regulator of inflammation and apoptosis. We presently evaluated the function of the AP-1 subunit JunB in cytokine-mediated β-cell dysfunction and death. The cytokines IL-1β+IFN-γ induced an early and transitory upregulation of JunB by NF-κB activation. Knockdown of JunB by RNA interference increased cytokine-mediated expression of inducible nitric oxide synthase (iNOS) and endoplasmic reticulum (ER) stress markers, leading to increased apoptosis in an insulin-producing cell line (INS-1E) and in purified rat primary β-cells. JunB knockdown β-cells and junB−/− fibroblasts were also more sensitive to the chemical ER stressor cyclopiazonic acid (CPA). Conversely, adenoviral-mediated overexpression of JunB diminished iNOS and ER markers expression and protected β-cells from cytokine-induced cell death. These findings demonstrate a novel and unexpected role for JunB as a regulator of defense mechanisms against cytokine- and ER stress-mediated apoptosis. PMID:18716665

  6. Activation of PPAR{delta} up-regulates fatty acid oxidation and energy uncoupling genes of mitochondria and reduces palmitate-induced apoptosis in pancreatic {beta}-cells

    SciTech Connect

    Wan, Jun; Jiang, Li; Lue, Qingguo; Ke, Linqiu; Li, Xiaoyu; Tong, Nanwei

    2010-01-15

    Recent evidence indicates that decreased oxidative capacity, lipotoxicity, and mitochondrial aberrations contribute to the development of insulin resistance and type 2 diabetes. The goal of this study was to investigate the effects of peroxisome proliferator-activated receptor {delta} (PPAR{delta}) activation on lipid oxidation, mitochondrial function, and insulin secretion in pancreatic {beta}-cells. After HIT-T15 cells (a {beta}-cell line) were exposed to high concentrations of palmitate and GW501516 (GW; a selective agonist of PPAR{delta}), we found that administration of GW increased the expression of PPAR{delta} mRNA. GW-induced activation of PPAR{delta} up-regulated carnitine palmitoyltransferase 1 (CPT1), long-chain acyl-CoA dehydrogenase (LCAD), pyruvate dehydrogenase kinase 4 (PDK4), and uncoupling protein 2 (UCP2); alleviated mitochondrial swelling; attenuated apoptosis; and reduced basal insulin secretion induced by increased palmitate in HIT cells. These results suggest that activation of PPAR{delta} plays an important role in protecting pancreatic {beta}-cells against aberrations caused by lipotoxicity in metabolic syndrome and diabetes.

  7. PED/PEA-15 Inhibits Hydrogen Peroxide-Induced Apoptosis in Ins-1E Pancreatic Beta-Cells via PLD-1

    PubMed Central

    Raciti, Gregory Alexander; Zatterale, Federica; Nigro, Cecilia; Mirra, Paola; Falco, Roberta; Ulianich, Luca; Di Jeso, Bruno; Formisano, Pietro; Miele, Claudia; Beguinot, Francesco

    2014-01-01

    The small scaffold protein PED/PEA-15 is involved in several different physiologic and pathologic processes, such as cell proliferation and survival, diabetes and cancer. PED/PEA-15 exerts an anti-apoptotic function due to its ability to interfere with both extrinsic and intrinsic apoptotic pathways in different cell types. Recent evidence shows that mice overexpressing PED/PEA-15 present larger pancreatic islets and increased beta-cells mass. In the present work we investigated PED/PEA-15 role in hydrogen peroxide-induced apoptosis in Ins-1E beta-cells. In pancreatic islets isolated from TgPED/PEA-15 mice hydrogen peroxide-induced DNA fragmentation was lower compared to WT islets. TUNEL analysis showed that PED/PEA-15 overexpression increases the viability of Ins-1E beta-cells and enhances their resistance to apoptosis induced by hydrogen peroxide exposure. The activity of caspase-3 and the cleavage of PARP-1 were markedly reduced in Ins-1E cells overexpressing PED/PEA-15 (Ins-1EPED/PEA-15). In parallel, we observed a decrease of the mRNA levels of pro-apoptotic genes Bcl-xS and Bad. In contrast, the expression of the anti-apoptotic gene Bcl-xL was enhanced. Accordingly, DNA fragmentation was higher in control cells compared to Ins-1EPED/PEA-15 cells. Interestingly, the preincubation with propranolol, an inhibitor of the pathway of PLD-1, a known interactor of PED/PEA-15, responsible for its deleterious effects on glucose tolerance, abolishes the antiapoptotic effects of PED/PEA-15 overexpression in Ins-1E beta-cells. The same results have been obtained by inhibiting PED/PEA-15 interaction with PLD-1 in Ins-1EPED/PEA-15. These results show that PED/PEA-15 overexpression is sufficient to block hydrogen peroxide-induced apoptosis in Ins-1E cells through a PLD-1 mediated mechanism. PMID:25489735

  8. High glucose induces suppression of insulin signalling and apoptosis via upregulation of endogenous IL-1beta and suppressor of cytokine signalling-1 in mouse pancreatic beta cells.

    PubMed

    Venieratos, Panagiotis D; Drossopoulou, Garyfalia I; Kapodistria, Katerina D; Tsilibary, Effie C; Kitsiou, Paraskevi V

    2010-05-01

    Chronic hyperglycemia and inflammatory cytokines disrupt and/or attenuate signal transduction pathways that promote normal beta-cell survival, leading to the destruction of endocrine pancreas in type 2 diabetes. There is convincing evidence that autocrine insulin signalling exerts protective anti-apoptotic effects on beta cells. Suppressors of cytokine signalling (SOCS) were induced by several cytokines and inhibit insulin-initiated signal transduction. The aim of this study was to investigate whether high glucose can influence endogenous interleukin-1beta (IL-1beta) and SOCS expression thus affecting insulin signalling and survival in insulin-producing mouse pancreatic beta cells (betaTC-6). Results showed that prolonged exposure of betaTC-6 cells to increased glucose concentrations resulted in significant inhibition of insulin-induced tyrosine phosphorylation of the insulin receptor (IR), and insulin receptor substrate-2 (IRS-2) as well as PI3-kinase activation. These changes were accompanied by impaired activation of the anti-apoptotic signalling protein Akt and annulment of Akt-mediated suppression of the Forkhead family of transcription factors (FoxO) activation. Glucose-induced attenuation of IRS-2/Akt-mediated signalling was associated with increased IL-1beta expression. Enhanced endogenous IL-1beta specifically induced mRNA and protein expression of SOCS-1 in betaTC-6 cells. Inhibition of SOCS-1 expression by SOCS-1-specific small interfering RNA restored IRS-2/PI3K-mediated Akt phosphorylation suppressed by high glucose. The upregulation of endogenous cytokine signalling and FoxO activation were accompanied by enhanced caspase-3 activation and increased susceptibility of cells to apoptosis. These results indicated that glucose-induced endogenous IL-1beta expression increased betaTC-6 cells apoptosis by inhibiting, at least in part, IRS-2/Akt-mediated signalling through SOCS-1 upregulation.

  9. Expression of calbindin-D(28k) in a pancreatic islet beta-cell line protects against cytokine-induced apoptosis and necrosis.

    PubMed

    Rabinovitch, A; Suarez-Pinzon, W L; Sooy, K; Strynadka, K; Christakos, S

    2001-08-01

    Cytokines produced by immune system cells that infiltrate pancreatic islets are candidate mediators of islet beta-cell destruction in autoimmune (type 1) diabetes mellitus. Because the calcium binding protein, calbindin-D(28k), can prevent apoptotic cell death in different cell types, we investigated the possibility that calbindin-D(28k) may prevent cytokine-mediated islet beta-cell destruction. Using the expression vector BSRalpha, rat calbindin-D(28k) was stably expressed in the pancreatic islet beta-cell line, betaTC-3. Calbindin-D(28k) expression resulted in increased cell survival in the presence of the cytotoxic combination of the cytokines IL-1beta (30 U/ml), TNFalpha (10(3) U/ml), and interferon gamma (10(3) U/ml). The greatest protection was observed in the betaTC-3 cell clone expressing the highest concentration of calbindin-D(28k). Apoptotic cell death was detected by annexin V staining and by the TdT-mediated dUTP-X nick end labeling assay in vector-transfected betaTC-3 cells incubated with cytokines (14-15% apoptotic cells). The number of apoptotic cells was significantly decreased in calbindin-D(28k)-overexpressing betaTC-3 cells incubated with cytokines (5-6% apoptotic cells). To address the mechanism of the antiapoptotic effects of calbindin, studies were done to examine whether calbindin inhibits free radical formation. The stimulatory effects of the cytokines on lipid hydroperoxide, nitric oxide, and peroxynitrite production were significantly decreased in the calbindin-D(28k)-expressing betaTC-3 cells. Our findings indicate that calbindin-D(28k), by inhibiting free radical formation, can protect against cytokine-mediated apoptosis and destruction of beta-cells. These findings suggest that calbindin-D(28k) may be an important regulator of cell death that can protect pancreatic islet beta-cells from autoimmune destruction in type 1 diabetes.

  10. Deletion of Apoptosis Signal-Regulating Kinase 1 (ASK1) Protects Pancreatic Beta-Cells from Stress-Induced Death but Not from Glucose Homeostasis Alterations under Pro-Inflammatory Conditions

    PubMed Central

    Pepin, Emilie; Higa, Arisa; Schuster-Klein, Carole; Bernard, Catherine; Sulpice, Thierry; Guardiola, Beatrice; Chevet, Eric; Alquier, Thierry

    2014-01-01

    Background Type 2 diabetes is characterized by pancreatic beta-cell dysfunction and is associated with low-grade inflammation. Recent observations suggest that apoptosis signal-regulating kinase 1 (ASK1) is involved in beta-cell death in response to different stressors. In this study, we tested whether ASK1 deficiency protects beta-cells from glucolipotoxic conditions and cytokines treatment or from glucose homeostasis alteration induced by endotoxemia. Methodology/Principal Findings Insulin secretion was neither affected upon shRNA-mediated downregulation of ASK1 in MIN6 cells nor in islets from ASK1-deficient mice. ASK1 silencing in MIN6 cells and deletion in islets did not prevent the deleterious effect of glucolipotoxic conditions or cytokines on insulin secretion. However, it protected MIN6 cells from death induced by ER stress or palmitate and islets from short term caspase activation in response to cytokines. Moreover, endotoxemia induced by LPS infusion increased insulin secretion during hyperglycemic clamps but the response was similar in wild-type and ASK1-deficient mice. Finally, insulin sensitivity in the presence of LPS was not affected by ASK1-deficiency. Conclusions/Significance Our study demonstrates that ASK1 is not involved in beta-cell function and dysfunction but controls stress-induced beta-cell death. PMID:25383781

  11. Expansion of beta-cell mass in response to pregnancy.

    PubMed

    Rieck, Sebastian; Kaestner, Klaus H

    2010-03-01

    Inadequate beta-cell mass can lead to insulin insufficiency and diabetes. During times of prolonged metabolic demand for insulin, the endocrine pancreas can respond by increasing beta-cell mass, both by increasing cell size and by changing the balance between beta-cell proliferation and apoptosis. In this paper, we review recent advances in our understanding of the mechanisms that control the adaptive expansion of beta-cell mass, focusing on the islet's response to pregnancy, a physiological state of insulin resistance. Functional characterization of factors controlling both beta-cell proliferation and survival might not only lead to the development of successful therapeutic strategies to enhance the response of the beta-cell to increased metabolic loads, but also improve islet transplantation regimens.

  12. D-saccharic acid-1,4-lactone ameliorates alloxan-induced diabetes mellitus and oxidative stress in rats through inhibiting pancreatic beta-cells from apoptosis via mitochondrial dependent pathway

    SciTech Connect

    Bhattacharya, Semantee; Manna, Prasenjit; Sil, Parames C.

    2011-12-15

    Oxidative stress plays a vital role in diabetic complications. To suppress the oxidative stress mediated damage in diabetic pathophysiology, a special focus has been given on naturally occurring antioxidants present in normal diet. D-saccharic acid 1,4-lactone (DSL), a derivative of D-glucaric acid, is present in many dietary plants and is known for its detoxifying and antioxidant properties. The aim of the present study was to evaluate the beneficial role of DSL against alloxan (ALX) induced diabetes in the pancreas tissue of Swiss albino rats. A dose-dependent study for DSL (20-120 mg/kg body weight) was carried out to find the effective dose of the compound in ALX-induced diabetic rats. ALX exposure elevated the blood glucose, glycosylated Hb, decreased the plasma insulin and disturbed the intra-cellular antioxidant machineries whereas oral administration of DSL at a dose of 80 mg/kg body weight restored these alterations close to normal. Investigating the mechanism of the protective activity of DSL we observed that it prevented the pancreatic {beta}-cell apoptosis via mitochondria-dependent pathway. Results showed decreased mitochondrial membrane potential, enhanced cytochrome c release in the cytosol and reciprocal regulation of Bcl-2 family proteins in the diabetic rats. These events were also found to be associated with increased level of Apaf-1, caspase 9, and caspase 3 that ultimately led to pancreatic {beta}-cell apoptosis. DSL treatment, however, counteracted these changes. In conclusion, DSL possesses the capability of ameliorating the oxidative stress in ALX-induced diabetes and thus could be a promising approach in lessening diabetic complications. Highlights: Black-Right-Pointing-Pointer Oxidative stress is suggested as a key event in the pathogenesis of diabetes. Black-Right-Pointing-Pointer D-saccharic acid 1,4-lactone (DSL) reduces the alloxan-induced diabetes mellitus. Black-Right-Pointing-Pointer DSL normalizes cellular antioxidant machineries

  13. Lithospermic acid B protects beta-cells from cytokine-induced apoptosis by alleviating apoptotic pathways and activating anti-apoptotic pathways of Nrf2-HO-1 and Sirt1

    SciTech Connect

    Lee, Byung-Wan; Chun, Sung Wan; Kim, Soo Hyun; Lee, Yongho; Kang, Eun Seok; Cha, Bong-Soo; Lee, Hyun Chul

    2011-04-01

    Lithospermic acid B (LAB) has been reported to protect OLETF rats, an established type 2 diabetic animal model, from the development of diabetes-related vascular complications. We investigated whether magnesium lithospermate B (LAB) has a protective role under cytokine-induced apoptosis in INS-1 cells in vitro and whether it slows the development of diabetes in OLETF rats in vivo. Pretreatment with 50 {mu}M LAB significantly reduced the 1000 U/mL INF-{gamma} and 100 U/mL IL-1{beta}-induced INS-1 cell death. LAB significantly alleviated cytokine-induced phosphorylations of p38 and JNK in accordance with a decrease in cleaved caspase-3 activity in beta-cells. LAB also protected against the cytokine-induced caspase-3 apoptotic pathway via significant activation of Nrf2-HO (heme-oxigenase)-1 and Sirt1 expression. OLETF rats treated with 40 mg/kg/day LAB showed a significant improvement in glucose tolerance compared to untreated OLETF control rats in vivo. Our results suggest that the cytoprotective effects of LAB on pancreatic {beta}-cells are related with both alleviating apoptotic pathways and activating anti-apoptotic pathways of Nrf2-HO-1 and Sirt1.

  14. Interleukin-1beta induced vascular permeability is dependent on induction of endothelial tissue factor (TF) activity.

    PubMed

    Puhlmann, Markus; Weinreich, David M; Farma, Jeffrey M; Carroll, Nancy M; Turner, Ewa M; Alexander, H Richard

    2005-09-30

    IL-1beta is a pleotropic cytokine that may mediate increased procoagulant activity and permeability in endothelial tissue during inflammatory conditions. The procoagulant effects of IL-1beta are mediated through induction of tissue factor (TF) but its alterations on vascular permeability are not well characterized. We found that IL-1beta induced a rapid and dose-dependent increase in TF activity in human umbilical vein endothelial cells (ECs) under routine culture conditions. However, IL-1beta caused a rapid and marked increase in permeability across confluent EC monolayers using a two-compartment in vitro model only in the presence of factor VIII-deficient plasma that was completely abrogated by neutralizing anti-TF antibody pre-treatment. In vitro permeability was associated with loss of EC surface expression of VE-cadherin and contraction of F-actin cytoskeletal elements that resulted in EC intercellular gap formation. These data demonstrate that IL-1beta induces marked changes in permeability across activated endothelium via a TF dependent mechanism and suggest that modulation of TF activity may represent a strategy to treat various acute and chronic inflammatory conditions mediated by this cytokine.

  15. Glucolipotoxicity of the Pancreatic Beta Cell

    PubMed Central

    Poitout, Vincent; Amyot, Julie; Semache, Meriem; Zarrouki, Bader; Hagman, Derek; Fontés, Ghislaine

    2009-01-01

    Summary The concept of glucolipotoxicity refers to the combined, deleterious effects of elevated glucose and fatty acid levels on pancreatic beta-cell function and survival. Significant progress has been made in recent years towards a better understanding of the cellular and molecular basis of glucolipotoxicity in the beta cell. The permissive effect of elevated glucose on the detrimental actions of fatty acids stems from the influence of glucose on intracellular fatty-acid metabolism, promoting the synthesis of cellular lipids. The combination of excessive levels of fatty acids and glucose therefore leads to decreased insulin secretion, impaired insulin gene expression, and beta-cell death by apoptosis, all of which probably have distinct underlying mechanisms. Recent studies from our laboratory have identified several pathways implicated in fatty-acid inhibition of insulin gene expression, including the extracellular-regulated kinase (ERK1/2) pathway; the metabolic sensor Per-Arnt-Sim kinase (PASK); and the ATF6 branch of the unfolded protein response. We have also confirmed in vivo in rats that the decrease in insulin gene expression is an early defect which precedes any detectable abnormality in insulin secretion. While the role of glucolipotoxicity in humans is still debated, the inhibitory effects of chronically elevated fatty acid levels has been clearly demonstrated in several studies, at least in individuals genetically predisposed to developing type 2 diabetes. It is therefore likely that glucolipotoxicity contributes to beta-cell failure in type 2 diabetes as well as to the decline in beta-cell function observed after the onset of the disease. PMID:19715772

  16. Unraveling the contribution of pancreatic beta-cell suicide in autoimmune type 1 diabetes✩

    PubMed Central

    Jaberi-Douraki, Majid; Schnell, Santiago; Pietropaolo, Massimo; Khadra, Anmar

    2014-01-01

    In type 1 diabetes, an autoimmune disease mediated by autoreactive T-cells that attack insulin-secreting pancreatic beta-cells, it has been suggested that disease progression may additionally require protective mechanisms in the target tissue to impede such auto-destructive mechanisms. We hypothesize that the autoimmune attack against beta-cells causes endoplasmic reticulum stress by forcing the remaining beta-cells to synthesize and secrete defective insulin. To rescue beta-cell from the endoplasmic reticulum stress, beta-cells activate the unfolded protein response to restore protein homeostasis and normal insulin synthesis. Here we investigate the compensatory role of unfolded protein response by developing a multi-state model of type 1 diabetes that takes into account beta-cell destruction caused by pathogenic autoreactive T-cells and apoptosis triggered by endoplasmic reticulum stress. We discuss the mechanism of unfolded protein response activation and how it counters beta-cell extinction caused by an autoimmune attack and/or irreversible damage by endoplasmic reticulum stress. Our results reveal important insights about the balance between beta-cell destruction by autoimmune attack (beta-cell homicide) and beta-cell apoptosis by endoplasmic reticulum stress (beta-cell suicide). It also provides an explanation as to why the unfolded protein response may not be a successful therapeutic target to treat type 1 diabetes. PMID:24831415

  17. The replication of beta cells in normal physiology, in disease and for therapy.

    PubMed

    Butler, Peter C; Meier, Juris J; Butler, Alexandra E; Bhushan, Anil

    2007-11-01

    Replication of beta cells is an important source of beta-cell expansion in early childhood. The recent linkage of type 2 diabetes with several transcription factors involved in cell cycle regulation implies that growth of the beta-cell mass in early childhood might be an important determinant of risk for type 2 diabetes. Under some circumstances, including obesity and pregnancy, the beta-cell mass is adaptively increased in adult humans. The mechanisms by which this adaptive growth occurs and the relative contributions of beta-cell replication or of mechanisms independent of beta-cell replication are unknown. Also, although there is interest in the potential for beta-cell regeneration as a therapeutic approach in both type 1 and 2 diabetes, little is yet known about the potential sources of new beta cells in adult humans. In common with other cell types, replicating beta cells have an increased vulnerability to apoptosis, which is likely to limit the therapeutic value of inducing beta-cell replication in the proapoptotic environment of type 1 and 2 diabetes unless applied in conjunction with a strategy to suppress increased apoptosis.

  18. Regulation of pancreatic islet beta-cell mass by growth factor and hormone signaling.

    PubMed

    Huang, Yao; Chang, Yongchang

    2014-01-01

    Dysfunction and destruction of pancreatic islet beta cells is a hallmark of diabetes. Better understanding of cellular signals in beta cells will allow development of therapeutic strategies for diabetes, such as preservation and expansion of beta-cell mass and improvement of beta-cell function. During the past several decades, the number of studies analyzing the molecular mechanisms, including growth factor/hormone signaling pathways that impact islet beta-cell mass and function, has increased exponentially. Notably, somatolactogenic hormones including growth hormone (GH), prolactin (PRL), and insulin-like growth factor-1 (IGF-1) and their receptors (GHR, PRLR, and IGF-1R) are critically involved in beta-cell growth, survival, differentiation, and insulin secretion. In this chapter, we focus more narrowly on GH, PRL, and IGF-1 signaling, and GH-IGF-1 cross talk. We also discuss how these signaling aspects contribute to the regulation of beta-cell proliferation and apoptosis. In particular, our novel findings of GH-induced formation of GHR-JAK2-IGF-1R protein complex and synergistic effects of GH and IGF-1 on beta-cell signaling, proliferation, and antiapoptosis lead to a new concept that IGF-1R may serve as a proximal component of GH/GHR signaling.

  19. Protective effect of niacinamide on interleukin-1beta-induced annulus fibrosus type II collagen degeneration in vitro.

    PubMed

    Duan, Deyu; Yang, Shuhua; Shao, Zengwu; Wang, Hong; Xiong, Xiaoqian

    2007-02-01

    The protective effect of niacinamide on interleukin-1beta (IL-1beta)-induced annulus fibrosus (AF) type II collagen degeneration in vitro and the mechanism were investigated. Chiba's intervertebral disc (IVD) culture models in rabbits were established and 48 IVDs from 12 adult Japanese white rabbits were randomly divided into 4 groups: normal control group, niacinamide-treated group, type II collagen degneration group (IL-1beta) and treatment group (niacinamide+IL-1beta). After culture for one week, AFs were collected for inducible nitric oxide synthase (iNOS), cysteine containing aspartate specific protease-3 (Caspase-3) and type II collagen immunohistochemical examination, and type II collagen reverse transcription polymerase chain reaction (RT-PCR). The results showed that rate of iNOS positive staining AF cells in the 4 groups was 17.6%, 10.9%, 73.9% and 19.3% respectively. The positive rate in treatment group was significantly lower than in the type II collagen degeneration group (P<0.01). Rate of Caspase-3 positive staining AF cells in the 4 groups was 3.4%, 4.2%, 17.6% and 10.3% respectively. The positive rate in treatment group was lower than in the type II collagen degeneration group (P<0.01). Type II collagen staining demonstrated that lamellar structure and continuity of collagen in treatment group was better reversed than in the degeneration group. RT-PCR revealed that the expression of type II collagen in treatment group was significantly stronger than that in type II collagen degeneration group (P<0.01). It was concluded that niacinamide could effectively inhibit IL-1beta stimulated increase of iNOS and Caspase-3 in AF, and alleviate IL-1beta-caused destruction and synthesis inhibition of type II collagen. Niacinamide is of potential for clinical treatment of IVD degeneration.

  20. Inhibition of interleukin-1 beta-induced pyresis in the rabbit by peptide 204-212 of lipocortin 5.

    PubMed

    Palmi, M; Frosini, M; Sgaragli, G P; Becherucci, C; Perretti, M; Parente, L

    1995-07-25

    The intracerebroventricular administration of interleukin-1 beta (12.5 ng/kg) in rabbits caused a prompt rise of prostaglandin E2 concentration (+ 632.6 +/- 243.9%) in the cerebrospinal fluid followed by hyperthermia (+ 1.61 +/- 0.14 delta degrees C). The intracerebroventricular administration of an anti-inflammatory nonapeptide (amino acids 204-212, SHLRKVFDK) derived from lipocortin 5, thereafter referred to as lipocortin 5-(204-212)-peptide, inhibited in a significant manner both the increase in cerebrospinal fluid [prostaglandin E2] and the febrile response induced by the cytokine. This inhibitory effect is probably due to interference by the peptide with phospholipase A2 activity. A control peptide (FKRVHDLKS) formed by the same amino acids in a randomly shuffled sequence had no effect. These results show that, in addition to the anti-inflammatory effect previously reported, the peptide 204-212 of lipocortin 5 possesses, like glucocorticoids, anti-pyretic activity. The research on lipocortin-derived peptides may lead to the development of novel anti-inflammatory and anti-pyretic compounds. PMID:8566125

  1. Balancing needs and means: the dilemma of the beta-cell in the modern world.

    PubMed

    Leibowitz, G; Kaiser, N; Cerasi, E

    2009-11-01

    The insulin resistance of type 2 diabetes mellitus (T2DM), although important for its pathophysiology, is not sufficient to establish the disease unless major deficiency of beta-cell function coexists. This is demonstrated by the fact that near-physiological administration of insulin (CSII) achieved excellent blood glucose control with doses similar to those used in insulin-deficient type 1 diabetics. The normal beta-cell adapts well to the demands of insulin resistance. Also in hyperglycaemic states some degree of adaptation does exist and helps limit the severity of disease. We demonstrate here that the mammalian target of rapamycin (mTOR) system might play an important role in this adaptation, because blocking mTORC1 (complex 1) by rapamycin in the nutritional diabetes model Psammomys obesus caused severe impairment of beta-cell function, increased beta-cell apoptosis and progression of diabetes. On the other hand, under exposure to high glucose and FFA (gluco-lipotoxicity), blocking mTORC1 in vitro reduced endoplasmic reticulum (ER) stress and beta-cell death. Thus, according to the conditions of stress, mTOR may have beneficial or deleterious effects on the beta-cell. beta-Cell function in man can be reduced without T2DM/impaired glucose tolerance (IGT). Prospective studies have shown subjects with reduced insulin response to present, several decades later, an increased incidence of IGT/T2DM. From these and other studies we conclude that T2DM develops on the grounds of beta-cells whose adaptation capacity to increased nutrient intake and/or insulin resistance is in the lower end of the normal variation. Inborn and acquired factors that limit beta-cell function are diabetogenic only in a nutritional/metabolic environment that requires high functional capabilities from the beta-cell.

  2. Regulation of pancreatic beta-cell mass.

    PubMed

    Bouwens, Luc; Rooman, Ilse

    2005-10-01

    Beta-cell mass regulation represents a critical issue for understanding diabetes, a disease characterized by a near-absolute (type 1) or relative (type 2) deficiency in the number of pancreatic beta cells. The number of islet beta cells present at birth is mainly generated by the proliferation and differentiation of pancreatic progenitor cells, a process called neogenesis. Shortly after birth, beta-cell neogenesis stops and a small proportion of cycling beta cells can still expand the cell number to compensate for increased insulin demands, albeit at a slow rate. The low capacity for self-replication in the adult is too limited to result in a significant regeneration following extensive tissue injury. Likewise, chronically increased metabolic demands can lead to beta-cell failure to compensate. Neogenesis from progenitor cells inside or outside islets represents a more potent mechanism leading to robust expansion of the beta-cell mass, but it may require external stimuli. For therapeutic purposes, advantage could be taken from the surprising differentiation plasticity of adult pancreatic cells and possibly also from stem cells. Recent studies have demonstrated that it is feasible to regenerate and expand the beta-cell mass by the application of hormones and growth factors like glucagon-like peptide-1, gastrin, epidermal growth factor, and others. Treatment with these external stimuli can restore a functional beta-cell mass in diabetic animals, but further studies are required before it can be applied to humans. PMID:16183912

  3. Osteocalcin protects pancreatic beta cell function and survival under high glucose conditions

    SciTech Connect

    Kover, Karen; Yan, Yun; Tong, Pei Ying; Watkins, Dara; Li, Xiaoyu; Tasch, James; Hager, Melissa; Clements, Mark; Moore, Wayne V.

    2015-06-19

    Diabetes is characterized by progressive beta cell dysfunction and loss due in part to oxidative stress that occurs from gluco/lipotoxicity. Treatments that directly protect beta cell function and survival in the diabetic milieu are of particular interest. A growing body of evidence suggests that osteocalcin, an abundant non-collagenous protein of bone, supports beta cell function and proliferation. Based on previous gene expression data by microarray, we hypothesized that osteocalcin protects beta cells from glucose-induced oxidative stress. To test our hypothesis we cultured isolated rat islets and INS-1E cells in the presence of normal, high, or high glucose ± osteocalcin for up to 72 h. Oxidative stress and viability/mitochondrial function were measured by H{sub 2}O{sub 2} assay and Alamar Blue assay, respectively. Caspase 3/7 activity was also measured as a marker of apoptosis. A functional test, glucose stimulated insulin release, was conducted and expression of genes/protein was measured by qRT-PCR/western blot/ELISA. Osteocalcin treatment significantly reduced high glucose-induced H{sub 2}O{sub 2} levels while maintaining viability/mitochondrial function. Osteocalcin also significantly improved glucose stimulated insulin secretion and insulin content in rat islets after 48 h of high glucose exposure compared to untreated islets. As expected sustained high glucose down-regulated gene/protein expression of INS1 and BCL2 while increasing TXNIP expression. Interestingly, osteocalcin treatment reversed the effects of high glucose on gene/protein expression. We conclude that osteocalcin can protect beta cells from the negative effects of glucose-induced oxidative stress, in part, by reducing TXNIP expression, thereby preserving beta cell function and survival. - Highlights: • Osteocalcin reduces glucose-induced oxidative stress in beta cells. • Osteocalcin preserves beta cell function and survival under stress conditions. • Osteocalcin reduces glucose

  4. A synopsis of factors regulating beta cell development and beta cell mass.

    PubMed

    Prasadan, Krishna; Shiota, Chiyo; Xiangwei, Xiao; Ricks, David; Fusco, Joseph; Gittes, George

    2016-10-01

    The insulin-secreting beta cells in the endocrine pancreas regulate blood glucose levels, and loss of functional beta cells leads to insulin deficiency, hyperglycemia (high blood glucose) and diabetes mellitus. Current treatment strategies for type-1 (autoimmune) diabetes are islet transplantation, which has significant risks and limitations, or normalization of blood glucose with insulin injections, which is clearly not ideal. The type-1 patients can lack insulin counter-regulatory mechanism; therefore, hypoglycemia is a potential risk. Hence, a cell-based therapy offers a better alternative for the treatment of diabetes. Past research was focused on attempting to generate replacement beta cells from stem cells; however, recently there has been an increasing interest in identifying mechanisms that will lead to the conversion of pre-existing differentiated endocrine cells into beta cells. The goal of this review is to provide an overview of several of the key factors that regulate new beta cell formation (neogenesis) and beta cell proliferation. PMID:27105622

  5. Translation attenuation through eIF2α phosphorylation prevents oxidative stress and maintains the differentiated state in beta cells

    PubMed Central

    Back, Sung Hoon; Scheuner, Donalyn; Han, JaeSeok; Song, Benbo; Ribick, Mark; Wang, Junying; Gildersleeve, Robert D.; Pennathur, Subramaniam; Kaufman, Randal J.

    2009-01-01

    SUMMARY Accumulation of unfolded protein within the endoplasmic reticulum (ER) lumen attenuates mRNA translation through activation of the protein kinase PERK and subsequent phosphorylation of eukaryotic initiation factor 2 on Ser51 of the alpha subunit (eIF2α). Genetic disruption of the PERK/eIF2α pathway in humans and mice produces severe pancreatic beta cell deficiency and post-natal lethality. To elucidate the role of eIF2α phosphorylation in beta cells, we have rescued the lethality of homozygous eIF2α Ser51Ala mice by expression of a loxP-flanked wild-type eIF2α transgene. Beta cell-specific transgene deletion to prevent eIF2α phosphorylation caused a severe diabetic phenotype due to heightened, unregulated proinsulin translation, defective intracellular trafficking of secretory and plasma membrane proteins, increased oxidative damage, reduced expression of stress response and beta cell-specific genes, and apoptosis. However, glucose intolerance and beta cell death in these mice were attenuated by antioxidant treatment. We conclude that phosphorylation of eIF2α coordinately attenuates mRNA translation, prevents oxidative stress, and optimizes ER protein folding to support insulin production in the beta cell. These findings that show increased proinsulin synthesis causes oxidative stress leading to beta cell failure may reflect events in the beta cell loss associated with insulin resistance in type 2 diabetes. PMID:19583950

  6. Toward beta cell replacement for diabetes.

    PubMed

    Johannesson, Bjarki; Sui, Lina; Freytes, Donald O; Creusot, Remi J; Egli, Dieter

    2015-04-01

    The discovery of insulin more than 90 years ago introduced a life-saving treatment for patients with type 1 diabetes, and since then, significant progress has been made in clinical care for all forms of diabetes. However, no method of insulin delivery matches the ability of the human pancreas to reliably and automatically maintain glucose levels within a tight range. Transplantation of human islets or of an intact pancreas can in principle cure diabetes, but this approach is generally reserved for cases with simultaneous transplantation of a kidney, where immunosuppression is already a requirement. Recent advances in cell reprogramming and beta cell differentiation now allow the generation of personalized stem cells, providing an unlimited source of beta cells for research and for developing autologous cell therapies. In this review, we will discuss the utility of stem cell-derived beta cells to investigate the mechanisms of beta cell failure in diabetes, and the challenges to develop beta cell replacement therapies. These challenges include appropriate quality controls of the cells being used, the ability to generate beta cell grafts of stable cellular composition, and in the case of type 1 diabetes, protecting implanted cells from autoimmune destruction without compromising other aspects of the immune system or the functionality of the graft. Such novel treatments will need to match or exceed the relative safety and efficacy of available care for diabetes.

  7. Transcription factors involved in glucose-stimulated insulin secretion of pancreatic beta cells

    SciTech Connect

    Shao, Shiying; Fang, Zhong; Yu, Xuefeng; Zhang, Muxun

    2009-07-10

    GSIS, the most important function of pancreatic beta cell, is essential for maintaining the glucose homeostasis. Transcription factors are known to control different biological processes such as differentiation, proliferation and apoptosis. In pancreas, some transcription factors are involved in regulating the function of beta cells. In this review, the role of these transcription factors including Pdx-1, FoxO1, SREBP-1c, and MafA in GSIS is highlighted. The related molecular mechanisms are analyzed as well. Furthermore, the association between the role of transcription factors in GSIS and the development of T2DM is discussed.

  8. On the origin of the beta cell.

    PubMed

    Oliver-Krasinski, Jennifer M; Stoffers, Doris A

    2008-08-01

    The major forms of diabetes are characterized by pancreatic islet beta-cell dysfunction and decreased beta-cell numbers, raising hope for cell replacement therapy. Although human islet transplantation is a cell-based therapy under clinical investigation for the treatment of type 1 diabetes, the limited availability of human cadaveric islets for transplantation will preclude its widespread therapeutic application. The result has been an intense focus on the development of alternate sources of beta cells, such as through the guided differentiation of stem or precursor cell populations or the transdifferentiation of more plentiful mature cell populations. Realizing the potential for cell-based therapies, however, requires a thorough understanding of pancreas development and beta-cell formation. Pancreas development is coordinated by a complex interplay of signaling pathways and transcription factors that determine early pancreatic specification as well as the later differentiation of exocrine and endocrine lineages. This review describes the current knowledge of these factors as they relate specifically to the emergence of endocrine beta cells from pancreatic endoderm. Current therapeutic efforts to generate insulin-producing beta-like cells from embryonic stem cells have already capitalized on recent advances in our understanding of the embryonic signals and transcription factors that dictate lineage specification and will most certainly be further enhanced by a continuing emphasis on the identification of novel factors and regulatory relationships. PMID:18676806

  9. Role of Bioactive Food Components in Diabetes Prevention: Effects on Beta-Cell Function and Preservation

    PubMed Central

    Oh, Yoon Sin; Jun, Hee-Sook

    2014-01-01

    Bioactive compounds found in fruits and vegetables can have anti-oxidant, anti-inflammatory, and anti-carcinogenic effects and can be protective against various diseases and metabolic disorders. These beneficial effects make them good candidates for the development of new functional foods with potential protective and preventive properties for type 1 and type 2 diabetes. This review summarizes the most relevant results concerning the effects of various bioactive compounds such as flavonoids, vitamins, and carotenoids on several aspects of beta-cell functionality. Studies using animal models with induced diabetes and diabetic patients support the hypothesis that bioactive compounds could ameliorate diabetic phenotypes. Published data suggest that there might be direct effects of bioactive compounds on enhancing insulin secretion and preventing beta-cell apoptosis, and some compounds might modulate beta-cell proliferation. Further research is needed to establish any clinical effects of these compounds. PMID:25092987

  10. Plant-Derived Compounds Targeting Pancreatic Beta Cells for the Treatment of Diabetes

    PubMed Central

    Oh, Yoon Sin

    2015-01-01

    Diabetes is a global health problem and a national economic burden. Although several antidiabetic drugs are available, the need for novel therapeutic agents with improved efficacy and few side effects remains. Drugs derived from natural compounds are more attractive than synthetic drugs because of their diversity and minimal side effects. This review summarizes the most relevant effects of various plant-derived natural compounds on the functionality of pancreatic beta cells. Published data suggest that natural compounds directly enhance insulin secretion, prevent pancreatic beta cell apoptosis, and modulate pancreatic beta cell differentiation and proliferation. It is essential to continuously investigate natural compounds as sources of novel pharmaceuticals. Therefore, more studies into these compounds' mechanisms of action are warranted for their development as potential anti-diabetics. PMID:26587047

  11. ER stress and the decline and fall of pancreatic beta cells in type 1 diabetes

    PubMed Central

    Brozzi, Flora

    2016-01-01

    Components of the unfolded protein response (UPR) modulate beta cell inflammation and death in early type 1 diabetes (T1D). The UPR is a mechanism by which cells react to the accumulation of misfolded proteins in the endoplasmic reticulum (ER). It aims to restore cellular homeostasis, but in case of chronic or overwhelming ER stress the persistent activation of the UPR triggers apoptosis, contributing to the loss of beta cells in both T1D and type 2 diabetes. It remains to be determined how and why the transition from ‘physiological’ to ‘pathological’ UPR takes place. A key component of the UPR is the ER transmembrane protein IRE1α (inositol-requiring enzyme 1α). IRE1α activity is modulated by both intra-ER signals and by the formation of protein complexes at its cytosolic domain. The amplitude and duration of IRE1α signaling is critical for the transition between the adaptive and cell death programs, with particular relevance for the activation of the pro-apoptotic c-Jun N-terminal kinase (JNK) in beta cells. In the present review we discuss the available information on IRE1α-regulating proteins in beta cells and their downstream targets, and the important differences observed between cytokine-induced UPR in human and rodent beta cells. PMID:26899404

  12. Human Fucci Pancreatic Beta Cell Lines: New Tools to Study Beta Cell Cycle and Terminal Differentiation

    PubMed Central

    Carlier, Géraldine; Maugein, Alicia; Cordier, Corinne; Pechberty, Séverine; Garfa-Traoré, Meriem; Martin, Patrick; Scharfmann, Raphaël; Albagli, Olivier

    2014-01-01

    Regulation of cell cycle in beta cells is poorly understood, especially in humans. We exploited here the recently described human pancreatic beta cell line EndoC-βH2 to set up experimental systems for cell cycle studies. We derived 2 populations from EndoC-βH2 cells that stably harbor the 2 genes encoding the Fucci fluorescent indicators of cell cycle, either from two vectors, or from a unique bicistronic vector. In proliferating non-synchronized cells, the 2 Fucci indicators revealed cells in the expected phases of cell cycle, with orange and green cells being in G1 and S/G2/M cells, respectively, and allowed the sorting of cells in different substeps of G1. The Fucci indicators also faithfully red out alterations in human beta cell proliferative activity since a mitogen-rich medium decreased the proportion of orange cells and inflated the green population, while reciprocal changes were observed when cells were induced to cease proliferation and increased expression of some beta cell genes. In the last situation, acquisition of a more differentiated beta cell phenotype correlates with an increased intensity in orange fluorescence. Hence Fucci beta cell lines provide new tools to address important questions regarding human beta cell cycle and differentiation. PMID:25259951

  13. Beta cell mass in diabetes: a realistic therapeutic target?

    PubMed

    Meier, J J

    2008-05-01

    Beta cell deficiency underlies both type 1 and type 2 diabetes, and restoration or replacement of beta cell function is therefore the logical long-term solution to therapy. This review sets out to describe the defects in beta cell mass and function in both forms of diabetes, summarises current understanding of the underlying causes of beta cell death, and the methodological limitations of determining beta cell mass in vivo. Finally, the potential effects of current and future treatment regimens on beta cell mass and turnover are considered.

  14. When BAD is good for beta cells.

    PubMed

    Philipson, Louis H; Roe, Michael W

    2008-04-01

    BAD, a proapoptotic member of the Bcl-2 family of proteins, is regulated by phosphorylation. A recent study (Danial et al., 2008) suggests a phosphorylation-state-dependent bifunctional role of BAD in the regulation of glucose-stimulated insulin secretion and beta cell mass. PMID:18396130

  15. The effect of TRAIL molecule on cell viability in in vitro beta cell culture.

    PubMed

    Tekmen, I; Ozyurt, D; Pekçetin, C; Buldan, Z

    2007-06-01

    Insulin-dependent diabetes mellitus (IDDM) is an organ-specific autoimmune disorder triggered by autoreactive T cells directed to pancreas beta-cell antigens. In this disorder, more than 90% of beta cells are destroyed. Cell death may be mediated via soluble or membrane-bound cell death ligands. One of these ligands may be tumour necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), a member of the TNF-alpha superfamily. In the present study, we examined whether TRAIL had cytotoxic effects on adult rat pancreas beta cell cultures and INS1-E rat insulinoma cell line cultures or not. In this study, cell destruction models were built with TRAIL concentrations of 10, 100 and 1000 ng. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test was used for evaluating cell viability. It was detected that cell cultures with TRAIL added showed no differences statistically when compared with control cultures containing no toxic additions. These results showed that TRAIL did not have significant cytotoxic effects on pancreas beta cell culture and INS-1E rat insulinoma cell line cultures. Detection of the expression of TRAIL receptors and natural apoptosis inhibitor proteins will be favourable to investigate the resistance mechanisms to TRAIL-induced cell death in this cell culture system.

  16. Early peroxisome proliferator-activated receptor gamma regulated genes involved in expansion of pancreatic beta cell mass

    PubMed Central

    2011-01-01

    Background The progression towards type 2 diabetes depends on the allostatic response of pancreatic beta cells to synthesise and secrete enough insulin to compensate for insulin resistance. The endocrine pancreas is a plastic tissue able to expand or regress in response to the requirements imposed by physiological and pathophysiological states associated to insulin resistance such as pregnancy, obesity or ageing, but the mechanisms mediating beta cell mass expansion in these scenarios are not well defined. We have recently shown that ob/ob mice with genetic ablation of PPARγ2, a mouse model known as the POKO mouse failed to expand its beta cell mass. This phenotype contrasted with the appropriate expansion of the beta cell mass observed in their obese littermate ob/ob mice. Thus, comparison of these models islets particularly at early ages could provide some new insights on early PPARγ dependent transcriptional responses involved in the process of beta cell mass expansion Results Here we have investigated PPARγ dependent transcriptional responses occurring during the early stages of beta cell adaptation to insulin resistance in wild type, ob/ob, PPARγ2 KO and POKO mice. We have identified genes known to regulate both the rate of proliferation and the survival signals of beta cells. Moreover we have also identified new pathways induced in ob/ob islets that remained unchanged in POKO islets, suggesting an important role for PPARγ in maintenance/activation of mechanisms essential for the continued function of the beta cell. Conclusions Our data suggest that the expansion of beta cell mass observed in ob/ob islets is associated with the activation of an immune response that fails to occur in POKO islets. We have also indentified other PPARγ dependent differentially regulated pathways including cholesterol biosynthesis, apoptosis through TGF-β signaling and decreased oxidative phosphorylation. PMID:22208362

  17. Effects of exendin-4 on glucose tolerance, insulin secretion, and beta-cell proliferation depend on treatment dose, treatment duration and meal contents

    SciTech Connect

    Arakawa, Masayuki; Ebato, Chie; Mita, Tomoya; Hirose, Takahisa; Kawamori, Ryuzo; Fujitani, Yoshio; Watada, Hirotaka

    2009-12-18

    Beta-cell proliferation is regulated by various metabolic demands including peripheral insulin resistance, obesity, and hyperglycemia. In addition to enhancement of glucose-induced insulin secretion, agonists for glucagon-like peptide-1 receptor (GLP-1R) stimulate proliferation and inhibit apoptosis of beta-cells, thereby probably preserve beta-cell mass. To evaluate the beta-cell preserving actions of GLP-1R agonists, we assessed the acute and chronic effects of exendin-4 on beta-cell proliferation, mass and glucose tolerance in C57BL/6J mice under various conditions. Short-term administration of high-dose exendin-4 transiently stimulated beta-cell proliferation. Comparative transcriptomic analysis showed upregulation of IGF-1 receptor and its downstream effectors in islets. Treatment of mice with exendin-4 daily for 4 weeks (long-term administration) and feeding high-fat diet resulted in significant inhibition of weight gain and improvement of glucose tolerance with reduced insulin secretion and beta-cell mass. These findings suggest that long-term GLP-1 treatment results in insulin sensitization of peripheral organs, rather than enhancement of beta-cell proliferation and function, particularly when animals are fed high-fat diet. Thus, the effects of exendin-4 on glucose tolerance, insulin secretion, and beta-cell proliferation largely depend on treatment dose, duration of treatment and meal contents. While GLP-1 enhances proliferation of beta-cells in some diabetic mice models, our results suggest that GLP-1 stimulates beta-cell growth only when expansion of beta-cell mass is required to meet metabolic demands.

  18. DJ-1 Protects Pancreatic Beta Cells from Cytokine- and Streptozotocin-Mediated Cell Death.

    PubMed

    Jain, Deepak; Weber, Gesine; Eberhard, Daniel; Mehana, Amir E; Eglinger, Jan; Welters, Alena; Bartosinska, Barbara; Jeruschke, Kay; Weiss, Jürgen; Päth, Günter; Ariga, Hiroyoshi; Seufert, Jochen; Lammert, Eckhard

    2015-01-01

    A hallmark feature of type 1 and type 2 diabetes mellitus is the progressive dysfunction and loss of insulin-producing pancreatic beta cells, and inflammatory cytokines are known to trigger beta cell death. Here we asked whether the anti-oxidant protein DJ-1 encoded by the Parkinson's disease gene PARK7 protects islet cells from cytokine- and streptozotocin-mediated cell death. Wild type and DJ-1 knockout mice (KO) were treated with multiple low doses of streptozotocin (MLDS) to induce inflammatory beta cell stress and cell death. Subsequently, glucose tolerance tests were performed, and plasma insulin as well as fasting and random blood glucose concentrations were monitored. Mitochondrial morphology and number of insulin granules were quantified in beta cells. Moreover, islet cell damage was determined in vitro after streptozotocin and cytokine treatment of isolated wild type and DJ-1 KO islets using calcein AM/ethidium homodimer-1 staining and TUNEL staining. Compared to wild type mice, DJ-1 KO mice became diabetic following MLDS treatment. Insulin concentrations were substantially reduced, and fasting blood glucose concentrations were significantly higher in MLDS-treated DJ-1 KO mice compared to equally treated wild type mice. Rates of beta cell apoptosis upon MLDS treatment were twofold higher in DJ-1 KO mice compared to wild type mice, and in vitro inflammatory cytokines led to twice as much beta cell death in pancreatic islets from DJ-1 KO mice versus those of wild type mice. In conclusion, this study identified the anti-oxidant protein DJ-1 as being capable of protecting pancreatic islet cells from cell death induced by an inflammatory and cytotoxic setting. PMID:26422139

  19. Gallic acid protects RINm5F beta-cells from glucolipotoxicity by its antiapoptotic and insulin-secretagogue actions.

    PubMed

    Sameermahmood, Zaheer; Raji, Lenin; Saravanan, Thangavel; Vaidya, Ashok; Mohan, Viswanathan; Balasubramanyam, Muthuswamy

    2010-01-01

    Gallic acid is claimed to possess antioxidant, antiinflammatory and cytoprotective effects. Since pancreatic islets from Type 2 diabetic patients have functional defects, it was hypothesized that glucolipotoxicity might induce apoptosis in beta-cells and gallic acid could offer protection. To test this, RINm5F beta-cells were exposed to high glucose (25 microM) or palmitate (500 microM) or a combination of both for 24 h in the presence and absence of gallic acid. Cells subjected to glucolipotoxicity in the absence and presence of gallic acid were assessed for DNA damage by comet assay. Apoptosis was inferred by caspase-3 protein expression and caspase-3 activity and changes in Bcl-2 mRNA. RT-PCR was used to analyse PDX-1, insulin and UCP-2 mRNA expression in RINm5F beta-cells and insulin levels were quantified from the cell culture supernatant. NFkappaB signal was studied by EMSA, immunofluorescence and Western blot analysis. While RINm5F beta-cells subjected to glucolipotoxicity exhibited increased DNA damage, apoptotic markers and NFkappaB signals, all these apoptotic perturbations were resisted by gallic acid. Gallic acid dose-dependently increased insulin secretion in RINm5F beta-cells and upregulated mRNA of PDX-1 and insulin. It is suggested that the insulin-secretagogue and transcriptional regulatory action of gallic acid is a newly identified mechanism in our study.

  20. MicroRNAs as regulators of beta-cell function and dysfunction.

    PubMed

    Osmai, Mirwais; Osmai, Yama; Bang-Berthelsen, Claus H; Pallesen, Emil M H; Vestergaard, Anna L; Novotny, Guy W; Pociot, Flemming; Mandrup-Poulsen, Thomas

    2016-05-01

    In the last decade, there has been an explosion in both the number of and knowledge about miRNAs associated with both type 1 and type 2 diabetes. Even though we are presently in the initial stages of understanding how this novel class of posttranscriptional regulators are involved in diabetes, recent studies have demonstrated that miRNAs are important regulators of the islet transcriptome, controlling apoptosis, differentiation and proliferation, as well as regulating unique islet and beta-cell functions and pathways such as insulin expression, processing and secretion. Furthermore, a large number of miRNAs have been linked to diabetogenic processes induced by elevated levels of glucose, free fatty acids and inflammatory cytokines. Thus, miRNAs are novel therapeutic targets with the potential of protecting the beta-cell, and there is proof of principle that miRNA antagonists, so-called antagomirs, are effective in vivo for other disorders. miRNAs are exported out of cells in exosomes, raising the intriguing possibility of cell-to-cell communication between distant tissues via miRNAs and that miRNAs can be used as biomarkers of beta-cell function, mass and survival. The purpose of this review is to provide a status on how miRNAs control beta-cell function and viability in health and disease. PMID:26418758

  1. Insulin receptor alternative splicing is regulated by insulin signaling and modulates beta cell survival

    PubMed Central

    Malakar, Pushkar; Chartarifsky, Lital; Hija, Ayat; Leibowitz, Gil; Glaser, Benjamin; Dor, Yuval; Karni, Rotem

    2016-01-01

    Type 2 Diabetes (T2DM) affects more than 300 million people worldwide. One of the hallmarks of T2DM is peripheral insulin resistance, in part due to unproductive insulin signaling through the insulin receptor. The insulin receptor (INSR) exists as two isoforms, INSR-A and INSR-B, which results from skipping or inclusion of exon 11 respectively. What determines the relative abundance of the different insulin receptor splice variants is unknown. Moreover, it is not yet clear what the physiological roles of each of the isoforms are in normal and diseased beta cells. In this study, we show that insulin induces INSR exon 11 inclusion in pancreatic beta cells in both human and mouse. This occurs through activation of the Ras-MAPK/ERK signaling pathway and up-regulation of the splicing factor SRSF1. Induction of exon 11 skipping by a splice-site competitive antisense oligonucleotide inhibited the MAPK-ERK signaling pathway downstream of the insulin receptor, sensitizing the pancreatic β-cell line MIN6 to stress-induced apoptosis and lipotoxicity. These results assign to insulin a regulatory role in INSR alternative splicing through the Ras-MAPK/ERK signaling pathway. We suggest that in beta cells, INSR-B has a protective role, while INSR-A expression sensitizes beta cells to programmed cell death. PMID:27526875

  2. In vivo regeneration of insulin-producing beta-cells.

    PubMed

    Jun, Hee-Sook

    2010-01-01

    Type 1 and type 2 diabetes mellitus are considered to be caused by defective control of blood glucose resulting from a reduced beta-cell mass. Thus, the restoration of a functional beta-cell mass by replacing the damaged beta-cells or stimulating beta-cell regeneration is a logical approach for the treatment of diabetes. Strategies for increasing the beta-cell mass include stimulating beta-cell replication and differentiation and inhibiting beta-cell death. Treatment with various growth factors such as GLP-1, BTC, HGF, and EGF and forced expression of beta-cell transcription factors such as Pdx-1, NeuroD, and MafA resulted in the regeneration of beta-cells in vivo. Another approach is the administration of stem/progenitor cells, which can differentiate into insulin-producing cells. However, there are no satisfactory methods yet for clinical application. Understanding the mechanisms of the regenerative process of pancreatic beta-cells will pave the way for the development of regenerative medicine for treatment of diabetes. PMID:20217517

  3. Beta cell device using icosahedral boride compounds

    DOEpatents

    Aselage, Terrence L.; Emin, David

    2002-01-01

    A beta cell for converting beta-particle energies into electrical energy having a semiconductor junction that incorporates an icosahedral boride compound selected from B.sub.12 As.sub.2, B.sub.12 P.sub.2, elemental boron having an .alpha.-rhombohedral structure, elemental boron having a .beta.-rhombohedral structure, and boron carbides of the chemical formula B.sub.12-x C.sub.3-x, where 0.15

  4. Novel inhibitors of macrophage migration inhibitory factor prevent cytokine-induced beta cell death.

    PubMed

    Vujicic, Milica; Nikolic, Ivana; Krajnovic, Tamara; Cheng, Kai-Fan; VanPatten, Sonya; He, Mingzhu; Stosic-Grujicic, Stanislava; Stojanovic, Ivana; Al-Abed, Yousef; Saksida, Tamara

    2014-10-01

    Macrophage migration inhibitory factor is a multifunctional cytokine involved in the regulation of immune processes and also in apoptosis induction. Elevated MIF expression is detrimental for insulin-producing beta cells and MIF inhibition protected beta cells from several cytotoxic insults such as inflammatory cytokines, high fatty acids or high glucose concentrations. Therefore, the aim of this study was to investigate two newly synthesized small molecule MIF inhibitors (K664-1 and K647-1) and to compare them with previously established effects of the prototypical MIF inhibitor, ISO-1. Our results indicate that K664-1 and K647-1 are 160- and 40-fold more effective in inhibition of MIF׳s tautomerase activity than ISO-1. Also, new inhibitors confer beta cell protection from cytokine-triggered apoptosis at significantly lower concentrations than ISO-1. Although all three MIF inhibitors inhibit caspase 3 activity, K664-1 and K647-1 suppress pro-apoptotic BAX protein expression and up-regulate anti-apoptotic Bcl-2 mRNA. Finally, all three MIF inhibitors operate through blockade of nitric oxide production stimulated by cytokines. In conclusion, two novel MIF inhibitors are more potent than ISO-1 and operate through inhibition of the mitochondria-related apoptotic pathway. We propose that these compounds represent a unique class of anti-MIF antagonists that should be further tested for therapeutic use.

  5. Chlamydia pneumoniae Promotes Dysfunction of Pancreatic Beta Cells

    PubMed Central

    Rodriguez, Annette R.; Plascencia-Villa, Germán; Witt, Colleen M.; Yu, Jieh-Juen; José-Yacamán, Miguel; Chambers, James P.; Perry, George; Guentzel, M. Neal; Arulanandam, Bernard P.

    2015-01-01

    The human pathogen Chlamydia pneumoniae has been implicated in chronic inflammatory diseases including type 2 diabetes. Therefore, we designed a study to evaluate pancreatic beta cells and mast cells during chlamydial infection. Our study revealed that C. pneumoniae infected mast cells significantly (p< 0.005) decreased beta cell ATP and insulin production, in contrast to uninfected mast cells co-cultured with beta cells. Infected mast cells exhibited pyknotic nuclei and active caspase-3 and caspase-1 expression. Additionally, ex vivo analyses of tissues collected from C. pneumoniae infected mice showed increased interleukin-1β production in splenocytes and pancreatic tissues as was observed with in vitro mast cell-beta cell co-cultures during C. pneumoniae infection. Notably, infected mast cells promoted beta cell destruction. Our findings reveal the negative effect of C. pneumoniae on mast cells, and the consequential impact on pancreatic beta cell function and viability. PMID:25863744

  6. Prostaglandin E2 represses interleukin 1 beta-induced inflammatory mediator output from pregnant human myometrial cells through the EP2 and EP4 receptors.

    PubMed

    Mosher, Andrea A; Rainey, Kelly J; Giembycz, Mark A; Wood, Stephen; Slater, Donna M

    2012-07-01

    Inflammatory mediators, including prostaglandins, cytokines, and chemokines, are strongly implicated in the mechanism of human labor, though their precise roles remain unknown. Here we demonstrate that interleukin 1 beta (IL-1beta) significantly increased the expression and release of interleukin-8 (CXCL8), monocyte chemotactic protein-1 (CCL2), and granulocyte macrophage colony-stimulating factor (CSF2) by primary human myometrial cells. However, this effect was repressed by prostaglandin E(2) (PGE(2)). As PGE(2) can activate four distinct PGE(2) receptors (EP(1), EP(2), EP(3), and EP(4)) to elicit various responses, we sought to define the EP receptor(s) responsible for this repression. Using selective EP receptor agonists and a selective EP(4) antagonist, we show that PGE(2) mediates the repression of IL-1beta-induced release of CXCL8, CCL2, and CSF2 via activation of the EP(2) and EP(4) receptors. The use of siRNA gene-specific knockdown further confirmed a role for both receptors. Real-time RT-PCR demonstrated that EP(2) was the most highly expressed of all four EP receptors at the mRNA level in human myometrial cells, and immunocytochemistry showed that EP(2) protein is abundantly present throughout the cells. Interestingly, PGE(2) does not appear to reduce mRNA expression of CXCL8, CCL2, and CSF2. Our results demonstrate that PGE(2) can elicit anti-inflammatory responses via activation of the EP(2) and EP(4) receptors in lower segment term pregnant human myometrial cells. Further elucidation of the EP receptor-mediated signaling pathways in the pregnant human uterus may be beneficial for optimizing the maintenance of pregnancy, induction of labor or indeed treatment of preterm labor.

  7. Bone morphogenetic protein-7 inhibits constitutive and interleukin-1 beta-induced monocyte chemoattractant protein-1 expression in human mesangial cells: role for JNK/AP-1 pathway.

    PubMed

    Lee, Myung-Ja; Yang, Chul Woo; Jin, Dong Chan; Chang, Yoon Sik; Bang, Byung Kee; Kim, Yong-Soo

    2003-03-01

    Bone morphogenetic protein-7 (BMP-7), which belongs to the TGF-beta superfamily, has been shown to reduce macrophage infiltration and tissue injury in animal models of inflammatory renal disease. To explore the mechanism involved in the anti-inflammatory effect, we investigated the effect of BMP-7 on monocyte chemoattractant protein-1 (MCP-1) expression in cultured human mesangial cells. BMP- 7 significantly inhibited constitutive and IL-1 beta-induced MCP-1 protein production and MCP-1 mRNA expression by mesangial cells in a time- and concentration-dependent manner. BMP-7 also inhibited IL-1 beta-induced monocyte chemotactic activity released from the mesangial cells. We examined the role of transcription factors NF-kappa B and AP-1 in BMP-7 inhibition of IL-1 beta-induced MCP-1 expression. IL-1 beta increased NF-kappa B and AP-1 activity and both transcription factors mediated IL-1 beta-induced MCP-1 expression in mesangial cells. BMP-7 inhibited IL-1 beta-induced AP-1 activity in a concentration-dependent manner. In contrast, IL-1 beta-induced NF-kappa B activity and I kappa B alpha degradation were not affected by BMP-7. Furthermore, IL-1 beta-induced phosphorylation of c-Jun N-terminal kinase was inhibited by BMP-7. These data suggest that BMP-7 inhibits constitutive and IL-1 beta-induced MCP-1 expression in human mesangial cells partly by inhibiting c-Jun N-terminal kinase activity and subsequent AP-1 activity, and provide new insight into the therapeutic potential of BMP-7 in the inflammatory renal diseases.

  8. Interleukin-1 beta-induced up-regulation of opioid receptors in the untreated and morphine-desensitized U87 MG human astrocytoma cells

    PubMed Central

    2012-01-01

    Background Interleukin-1beta (IL-1β) is a pro-inflammatory cytokine that can be produced in the central nervous system during inflammatory conditions. We have previously shown that IL-1β expression is altered in the rat brain during a morphine tolerant state, indicating that this cytokine may serve as a convergent point between the immune challenge and opiate mediated biological pathways. We hypothesized that IL-1β up-regulates opioid receptors in human astrocytes in both untreated and morphine-desensitized states. Methods To test this hypothesis, we compared the basal expression of the mu (MOR), delta (DOR), and kappa (KOR) opioid receptors in the human U87 MG astrocytic cell line to SH-SY5Y neuronal and HL-60 immune cells using absolute quantitative real time RT-PCR (AQ-rt-RT-PCR). To demonstrate that IL-1β induced up-regulation of the MOR, DOR and KOR, U87 MG cells (2 x 105 cells/well) were treated with IL-1β (20 ng/mL or 40 ng/mL), followed by co-treatment with interleukin-1 receptor antagonist protein (IL-1RAP) (400 ng/mL or 400 ng/mL). The above experiment was repeated in the cells desensitized with morphine, where U87 MG cells were pre-treated with 100 nM morphine. The functionality of the MOR in U87 MG cells was then demonstrated using morphine inhibition of forksolin-induced intracellular cAMP, as determined by radioimmunoassay. Results U87 MG cells treated with IL-1β for 12 h showed a significant up-regulation of MOR and KOR. DOR expression was also elevated, although not significantly. Treatment with IL-1β also showed a significant up-regulation of the MOR in U87 MG cells desensitized with morphine. Co-treatment with IL-1β and interleukin-1 receptor antagonist protein (IL-1RAP) resulted in a significant decrease in IL-1β-mediated MOR up-regulation. Conclusion Our results indicate that the pro-inflammatory cytokine, IL-1β, affects opiate-dependent pathways by up-regulating the expression of the MOR in both untreated and morphine-desensitized U87 MG. PMID:23164507

  9. Interleukin-1 beta-induced disruption of the retinal vascular barrier of the central nervous system is mediated through leukocyte recruitment and histamine.

    PubMed Central

    Bamforth, S. D.; Lightman, S. L.; Greenwood, J.

    1997-01-01

    The vascular barriers of the central nervous system form a selective cellular interface between the blood and the neural parenchyma and restrict the transfer of both molecules and hematogenous cells. During immune-mediated diseases, leukocyte infiltration becomes dramatically up-regulated and the permeability of these barriers increases, leading to edema formation. The etiology of this damage remains largely unresolved although inflammatory cytokines have been implicated in the process. The effect of the proinflammatory cytokine interleukin (IL)-1 beta on the integrity of the rat blood-retinal barrier (BRB) was investigated up to 14 days after an intravitreal injection. The permeability of the BRB was evaluated quantitatively using the low molecular weight tracer [14C]mannitol. After IL-1 beta administration, a biphasic opening of the BRB to [14C]mannitol was recorded, peaking at 4 to 8 hours and 24 to 48 hours post-injection (PI). The early disruption coincided with the appearance of both polymorphonuclear and mononuclear leukocytes within the retina. By 12 hours PI, BRB permeability had returned to control values despite a continued increase in the number of infiltrating leukocytes. The second, more pronounced increase in barrier permeability detected at 24 to 48 hours PI corresponded with maximal leukocyte infiltration. Barrier dysfunction had resolved by 72 hours, and by 7 days the leukocyte infiltrate had disappeared. The IL-1 beta-induced increase in permeability could be completely abrogated at 4 and 24 hours PI by treating the animals with the histamine H2-receptor antagonist ranitidine, which also reduced leukocyte infiltration by 47.2%. The ability of histamine to disrupt the BRB was demonstrated by intravitreal and intravascular administration, which caused a rapid and significant increase in BRB permeability. Treatment of the animals with the cyclo-oxygenase inhibitor indomethacin had no effect on IL-1 beta-induced disruption of the BRB at 4 hours PI, but by 24 hours PI a significant reduction in permeability was observed that coincided with a 75.2% reduction in the leukocyte infiltrate. The depletion of circulating leukocytes to < 2% of control levels reduced the retinal leukocyte recruitment induced by IL-1 beta by 73.0% and decreased BRB permeability at both 4 and 24 hours after IL-1 beta injection. These data demonstrate that intravitreal IL-1 beta in the rat induces a biphasic opening of the BRB that appears to be mediated through recruited leukocytes and release of the vasoactive amine histamine. Images Figure 1 PMID:9006348

  10. Glucolipotoxicity age-dependently impairs beta cell function in rats despite a marked increase in beta cell mass

    PubMed Central

    Fontés, G.; Zarrouki, B.; Hagman, D. K.; Latour, M. G.; Semache, M.; Roskens, V.; Moore, P. C.; Prentki, M.; Rhodes, C. J.; Jetton, T. L.

    2010-01-01

    Aims/hypothesis Prolonged exposure of pancreatic beta cells to excessive levels of glucose and fatty acids, referred to as glucolipotoxicity, is postulated to contribute to impaired glucose homeostasis in patients with type 2 diabetes. However, the relative contribution of defective beta cell function vs diminished beta cell mass under glucolipotoxic conditions in vivo remains a subject of debate. We therefore sought to determine whether glucolipotoxicity in rats is due to impaired beta cell function and/or reduced beta cell mass, and whether older animals are more susceptible to glucolipotoxic condition. Methods Wistar rats (2 and 6 months old) received a 72 h infusion of glucose + intravenous fat emulsion or saline control. In vivo insulin secretion and sensitivity were assessed by hyperglycaemic clamps. Ex vivo insulin secretion, insulin biosynthesis and gene expression were measured in isolated islets. Beta cell mass and proliferation were examined by immunohistochemistry. Results A 72 h infusion of glucose + intravenous fat emulsion in 2-month-old Wistar rats did not affect insulin sensitivity, insulin secretion or beta cell mass. In 6-month-old rats by contrast it led to insulin resistance and reduced insulin secretion in vivo, despite an increase in beta cell mass and proliferation. This was associated with: (1) diminished glucose-stimulated second-phase insulin secretion and proinsulin biosynthesis; (2) lower insulin content; and (3) reduced expression of beta cell genes in isolated islets. Conclusions/interpretation In this in vivo model, glucolipotoxicity is characterised by an age-dependent impairment of glucose-regulated beta cell function despite a marked increase in beta cell mass. PMID:20628728

  11. Effect of Exendin-4 on Autophagy Clearance in Beta Cell of Rats with Tacrolimus-induced Diabetes Mellitus

    PubMed Central

    Lim, Sun Woo; Jin, Long; Jin, Jian; Yang, Chul Woo

    2016-01-01

    Growing evidence suggests that GLP-1 protects beta cells against various cellular injuries by modulating autophagy. In this study, we examined whether exendin-4 (Ex-4), a GLP-1 analog, had preventive effects on tacrolimus (Tac)-induced beta cell injury by improving autophagy clearance. Rats with Tac-induced diabetes mellitus exhibited increased autophagy-associated protein expression, light chain 3B levels, and autophagic vacuole numbers in pancreatic beta cells. Additionally, Tac increased autophagy in a dose- and time-dependent manner in vitro, and inhibition of autophagosome using 3-methyladenine reduced Tac-induced islet cell injury by decreasing reactive oxygen species production and apoptosis. Ex-4 treatment decreased Tac-induced hyperglycaemia, oxidative stress, and apoptosis, accompanied by decreased autophagy-associated protein expression and autophagosome numbers. In vivo and in vitro studies showed that Tac treatment impaired lysosomal function and autophagosome-lysosome fusion; these processes were improve by Ex-4 treatment. Moreover, addition of bafilomycin A1, an inhibitor of lysosomal function, abolished the protective effects of Ex-4. Our findings reveal that Tac-induced diabetes mellitus was a state of excessive burden of autophagosomes and impairment of autophagy clearance and that Ex-4 protected against Tac-induced pancreatic islet injury by reducing the burden of autophagosomes via activation of autophagosome clearance. Thus, Ex-4 had therapeutic effects on Tac-induced pancreatic beta cell injury. PMID:27436514

  12. Adenoviral infection or deferoxamine? Two approaches to overexpress VEGF in beta-cell lines.

    PubMed

    Langlois, Allan; Bietiger, William; Sencier, Marie-Christine; Maillard, Elisa; Pinget, Michel; Kessler, Laurence; Sigrist, Severine

    2009-07-01

    Rapid and adequate revascularization of transplanted islets is important for their survival and function during transplantation. Vascular endothelial growth factor (VEGF) could play a critical role with respect to islet revascularization. The aim of this study was to compare two strategies that are used to overexpress VEGF in beta-cells: (1) gene therapy through adenoviral infection and (2) a pharmacological approach using deferoxamine (DFO). beta-Cell lines from rat insulinoma (RINm5F) were either infected using an adenovirus encoding the gene of human VEGF 165 or incubated with DFO. One day after treatment, the viability of RINm5F cells was preserved with 10 micromol/L of DFO (103.95 +/- 5.66% toward control; n = 4). In addition, adenoviral infection maintained the viability of cells for all the concentrations used. In both treatments, overexpression of VEGF was in a comparable level. Finally, the ratio of Bax/Bcl-2 indicated that the apoptosis increased in infected beta-cells whereas treatment with DFO seems to be antiapoptotic. Our results suggest that the use of DFO could be a realistic approach to improve the vascularization of islets during transplantation. PMID:19527112

  13. Uncovering Factors Related to Pancreatic Beta-Cell Function

    PubMed Central

    Curran, Aoife M.; Ryan, Miriam F.; Drummond, Elaine; Gibney, Eileen R.; Gibney, Michael J.; Roche, Helen M.; Brennan, Lorraine

    2016-01-01

    Aim The incidence of type 2 diabetes has increased rapidly on a global scale. Beta-cell dysfunction contributes to the overall pathogenesis of type 2 diabetes. However, factors contributing to beta-cell function are not clear. The aims of this study were (i) to identify factors related to pancreatic beta-cell function and (ii) to perform mechanistic studies in vitro. Methods Three specific measures of beta-cell function were assessed for 110 participants who completed an oral glucose tolerance test as part of the Metabolic Challenge Study. Anthropometric and biochemical parameters were assessed as potential modulators of beta-cell function. Subsequent in vitro experiments were performed using the BRIN-BD11 pancreatic beta-cell line. Validation of findings were performed in a second human cohort. Results Waist-to-hip ratio was the strongest anthropometric modulator of beta-cell function, with beta-coefficients of -0.33 (p = 0.001) and -0.30 (p = 0.002) for beta-cell function/homeostatic model assessment of insulin resistance (HOMA-IR), and disposition index respectively. Additionally, the resistin-to-adiponectin ratio (RA index) emerged as being strongly associated with beta-cell function, with beta-coefficients of -0.24 (p = 0.038) and -0.25 (p = 0.028) for beta-cell function/HOMA-IR, and disposition index respectively. Similar results were obtained using a third measure for beta-cell function. In vitro experiments revealed that the RA index was a potent regulator of acute insulin secretion where a high RA index (20ng ml-1 resistin, 5nmol l-1 g-adiponectin) significantly decreased insulin secretion whereas a low RA index (10ng ml-1 resistin, 10nmol l-1 g-adiponectin) significantly increased insulin secretion. The RA index was successfully validated in a second human cohort with beta-coefficients of -0.40 (p = 0.006) and -0.38 (p = 0.008) for beta-cell function/ HOMA-IR, and disposition index respectively. Conclusions Waist-to-hip ratio and RA index were identified

  14. Sources of beta cells inside the pancreas.

    PubMed

    De Groef, Sofie; Staels, Willem; Van Gassen, Naomi; Lemper, Marie; Yuchi, Yixing; Sojoodi, Mozhdeh; Bussche, Leen; Heremans, Yves; Leuckx, Gunter; De Leu, Nico; Van de Casteele, Mark; Baeyens, Luc; Heimberg, Harry

    2016-09-01

    The generation of beta(-like) cells to compensate for their absolute or relative shortage in type 1 and type 2 diabetes is an obvious therapeutic strategy. Patients first received grafts of donor islet cells over 25 years ago, but this procedure has not become routine in clinical practice because of a donor cell shortage and (auto)immune problems. Transplantation of differentiated embryonic and induced pluripotent stem cells may overcome some but not all the current limitations. Reprogramming exocrine cells towards functional beta(-like) cells would offer an alternative abundant and autologous source of beta(-like) cells. This review focuses on work by our research group towards achieving such a source of cells. It summarises a presentation given at the 'Can we make a better beta cell?' symposium at the 2015 annual meeting of the EASD. It is accompanied by two other reviews on topics from this symposium (by Amin Ardestani and Kathrin Maedler, DOI: 10.1007/s00125-016-3892-9 , and by Heiko Lickert and colleagues, DOI: 10.1007/s00125-016-3949-9 ) and a commentary by the Session Chair, Shanta Persaud (DOI: 10.1007/s00125-016-3870-2 ). PMID:27053238

  15. Structure-function studies of PANDER, an islet specific cytokine inducing cell death of insulin-secreting beta cells.

    PubMed

    Yang, Jichun; Gao, Zhiyong; Robert, Claudia E; Burkhardt, Brant R; Gaweska, Helena; Wagner, Amary; Wu, Jianmei; Greene, Scott R; Young, Robert A; Wolf, Bryan A

    2005-08-30

    PANDER (pancreatic derived factor, FAM3B) is a novel cytokine, present in insulin secretory granules, that induces apoptosis of alpha and beta cells of mouse, rat, and human islets in a dose- and time-dependent manner, and may be implicated in diabetes. PANDER has the predicted secondary structure of 4 alpha-helical bundles with an up-up-down-down topology, and two disulfide bonds. Eleven mutated PANDERs were constructed and expressed in beta-TC3 cells to identify the essential region of PANDER involved in beta-cell death. Beta-cell function was assessed by assays of cell viability and insulin secretion. Based on quantitative real-time RT-PCR all mutant PANDERs had similar mRNA expression levels in beta-TC3 cells. Immunoblotting showed that ten of eleven mutant PANDER proteins were synthesized and detected in beta-TC3 cells. A mutant PANDER with no signal peptide, however, was not expressed. Truncation of helix D alone caused a 40-50% decrease in PANDER's activity, while truncation of both helices C and D resulted in a 75% loss of activity. In contrast, truncation of the N-terminus of PANDER (helix A, the loop between helices A and B, and the first two cysteines) had no effect on PANDER-induced beta-cell death. The third and fourth cysteines of PANDER, C91 and C229, were shown to form one disulfide bond and be functionally important. Finally, the region between Cys91 and Phe152 constitutes the active part of PANDER, based on the demonstration that mutants with truncation of helix B or C caused decreased beta-cell death and did not inhibit insulin secretion, as compared to wild-type PANDER. Hence, helices B and C and the second disulfide bond of PANDER are essential for PANDER-induced beta-cell death.

  16. Crucial role of PDX-1 in pancreas development, beta-cell differentiation, and induction of surrogate beta-cells.

    PubMed

    Kaneto, Hideaki; Miyatsuka, Takeshi; Shiraiwa, Toshihiko; Yamamoto, Kaoru; Kato, Ken; Fujitani, Yoshio; Matsuoka, Taka-aki

    2007-01-01

    Pancreatic and duodenal homeobox factor-1 (PDX-1) plays a crucial role in pancreas development, beta-cell differentiation, and maintaining mature beta-cell function. At an early stage of embryonic development, PDX-1 is initially expressed in the gut region when the foregut endoderm becomes committed to common pancreatic precursor cells. During pancreas development, PDX-1 expression is maintained in precursor cells, and later it becomes restricted to beta-cells. In mature beta-cells, PDX-1 transactivates the insulin gene and other genes involved in glucose sensing and metabolism, such as GLUT2 and glucokinase. MafA is a recently isolated beta-cell-specific transcription factor which functions as a potent activator of insulin gene transcription. During pancreas development, MafA expression is first detected at the beginning of the principal phase of insulin-producing cell production. Furthermore, these transcription factors play a crucial role in inducing surrogate beta-cells from non-beta-cells and thus could be therapeutic targets for diabetes.

  17. Serotonin competence of mouse beta cells during pregnancy.

    PubMed

    Goyvaerts, Lotte; Schraenen, Anica; Schuit, Frans

    2016-07-01

    Pregnancy is a key mammalian reproductive event in which growth and differentiation of the fetus imposes extra metabolic and hormonal demands on the mother. Its successful outcome depends on major changes in maternal blood circulation, metabolism and endocrine function. One example is the endocrine pancreas, where beta cells undergo a number of changes in pregnancy that result in enhanced functional beta cell mass in order to compensate for the rising metabolic needs for maternal insulin. During the last 5 years, a series of studies have increased our understanding of the molecular events involved in this functional adaptation. In the mouse, a prominent functional change during pregnancy is the capacity of some beta cells to produce serotonin. In this review we will discuss the mechanism and potential effects of pregnancy-related serotonin production in beta cells, considering functional consequences at the local intra-islet and systemic level. PMID:27056372

  18. Insights from lncRNAs Profiling of MIN6 Beta Cells Undergoing Inflammation

    PubMed Central

    Sun, Chuntao; Xue, Lihua; Zhu, Ziyang; Zhang, Fan; Yang, Ruixue; Yuan, Xuewen

    2016-01-01

    Type 1 diabetes mellitus (T1DM) is an organ-specific autoimmune disease characterized by chronic and progressive apoptotic destruction of pancreatic beta cells. During the initial phases of T1DM, cytokines and other inflammatory mediators released by immune cells progressively infiltrate islet cells, induce alterations in gene expression, provoke functional impairment, and ultimately lead to apoptosis. Long noncoding RNAs (lncRNAs) are a new important class of pervasive genes that have a variety of biological functions and play key roles in many diseases. However, whether they have a function in cytokine-induced beta cell apoptosis is still uncertain. In this study, lncRNA microarray technology was used to identify the differently expressed lncRNAs and mRNAs in MIN6 cells exposed to proinflammatory cytokines. Four hundred forty-four upregulated and 279 downregulated lncRNAs were detected with a set filter fold-change ≧2.0. To elucidate the potential functions of these lncRNAs, Gene Ontology (GO) and pathway analyses were used to evaluate the potential functions of differentially expressed lncRNAs. Additionally, a lncRNA-mRNA coexpression network was constructed to predict the interactions between the most strikingly regulated lncRNAs and mRNAs. This study may be utilized as a background or reference resource for future functional studies on lncRNAs related to the diagnosis and development of new therapies for T1DM.

  19. Insights from lncRNAs Profiling of MIN6 Beta Cells Undergoing Inflammation

    PubMed Central

    Sun, Chuntao; Xue, Lihua; Zhu, Ziyang; Zhang, Fan; Yang, Ruixue; Yuan, Xuewen

    2016-01-01

    Type 1 diabetes mellitus (T1DM) is an organ-specific autoimmune disease characterized by chronic and progressive apoptotic destruction of pancreatic beta cells. During the initial phases of T1DM, cytokines and other inflammatory mediators released by immune cells progressively infiltrate islet cells, induce alterations in gene expression, provoke functional impairment, and ultimately lead to apoptosis. Long noncoding RNAs (lncRNAs) are a new important class of pervasive genes that have a variety of biological functions and play key roles in many diseases. However, whether they have a function in cytokine-induced beta cell apoptosis is still uncertain. In this study, lncRNA microarray technology was used to identify the differently expressed lncRNAs and mRNAs in MIN6 cells exposed to proinflammatory cytokines. Four hundred forty-four upregulated and 279 downregulated lncRNAs were detected with a set filter fold-change ≧2.0. To elucidate the potential functions of these lncRNAs, Gene Ontology (GO) and pathway analyses were used to evaluate the potential functions of differentially expressed lncRNAs. Additionally, a lncRNA-mRNA coexpression network was constructed to predict the interactions between the most strikingly regulated lncRNAs and mRNAs. This study may be utilized as a background or reference resource for future functional studies on lncRNAs related to the diagnosis and development of new therapies for T1DM. PMID:27698546

  20. Pancreatic beta cells express a diverse set of homeobox genes.

    PubMed Central

    Rudnick, A; Ling, T Y; Odagiri, H; Rutter, W J; German, M S

    1994-01-01

    Homeobox genes, which are found in all eukaryotic organisms, encode transcriptional regulators involved in cell-type differentiation and development. Several homeobox genes encoding homeodomain proteins that bind and activate the insulin gene promoter have been described. In an attempt to identify additional beta-cell homeodomain proteins, we designed primers based on the sequences of beta-cell homeobox genes cdx3 and lmx1 and the Drosophila homeodomain protein Antennapedia and used these primers to amplify inserts by PCR from an insulinoma cDNA library. The resulting amplification products include sequences encoding 10 distinct homeodomain proteins; 3 of these proteins have not been described previously. In addition, an insert was obtained encoding a splice variant of engrailed-2, a homeodomain protein previously identified in the central nervous system. Northern analysis revealed a distinct pattern of expression for each homeobox gene. Interestingly, the PCR-derived clones do not represent a complete sampling of the beta-cell library because no inserts encoding cdx3 or lmx1 protein were obtained. Beta cells probably express additional homeobox genes. The abundance and diversity of homeodomain proteins found in beta cells illustrate the remarkable complexity and redundancy of the machinery controlling beta-cell development and differentiation. Images PMID:7991607

  1. New insights into fatty acid modulation of pancreatic beta-cell function.

    PubMed

    Haber, Esther P; Procópio, Joaquim; Carvalho, Carla R O; Carpinelli, Angelo R; Newsholme, Philip; Curi, Rui

    2006-01-01

    Insulin resistance states as found in type 2 diabetes and obesity are frequently associated with hyperlipidemia. Both stimulatory and detrimental effects of free fatty acids (FFA) on pancreatic beta cells have long been recognized. Acute exposure of the pancreatic beta cell to both high glucose concentrations and saturated FFA results in a substantial increase of insulin release, whereas a chronic exposure results in desensitization and suppression of secretion. Reduction of plasma FFA levels in fasted rats or humans severely impairs glucose-induced insulin release but palmitate can augment insulin release in the presence of nonstimulatory concentrations of glucose. These results imply that changes in physiological plasma levels of FFA are important for regulation of beta-cell function. Although it is widely accepted that fatty acid (FA) metabolism (notably FA synthesis and/or formation of LC-acyl-CoA) is necessary for stimulation of insulin secretion, the key regulatory molecular mechanisms controlling the interplay between glucose and fatty acid metabolism and thus insulin secretion are not well understood but are now described in detail in this review. Indeed the correct control of switching between FA synthesis or oxidation may have critical implications for beta-cell function and integrity both in vivo and in vitro. LC-acyl-CoA (formed from either endogenously synthesized or exogenous FA) controls several aspects of beta-cell function including activation of certain types of PKC, modulation of ion channels, protein acylation, ceramide- and/or NO-mediated apoptosis, and binding to and activating nuclear transcriptional factors. The present review also describes the possible effects of FAs on insulin signaling. We have previously reported that acute exposure of islets to palmitate up-regulates some key components of the intracellular insulin signaling pathway in pancreatic islets. Another aspect considered in this review is the potential source of fatty acids

  2. Detailed transcriptome atlas of the pancreatic beta cell

    PubMed Central

    Kutlu, Burak; Burdick, David; Baxter, David; Rasschaert, Joanne; Flamez, Daisy; Eizirik, Decio L; Welsh, Nils; Goodman, Nathan; Hood, Leroy

    2009-01-01

    Background Gene expression patterns provide a detailed view of cellular functions. Comparison of profiles in disease vs normal conditions provides insights into the processes underlying disease progression. However, availability and integration of public gene expression datasets remains a major challenge. The aim of the present study was to explore the transcriptome of pancreatic islets and, based on this information, to prepare a comprehensive and open access inventory of insulin-producing beta cell gene expression, the Beta Cell Gene Atlas (BCGA). Methods We performed Massively Parallel Signature Sequencing (MPSS) analysis of human pancreatic islet samples and microarray analyses of purified rat beta cells, alpha cells and INS-1 cells, and compared the information with available array data in the literature. Results MPSS analysis detected around 7600 mRNA transcripts, of which around a third were of low abundance. We identified 2000 and 1400 transcripts that are enriched/depleted in beta cells compared to alpha cells and INS-1 cells, respectively. Microarray analysis identified around 200 transcription factors that are differentially expressed in either beta or alpha cells. We reanalyzed publicly available gene expression data and integrated these results with the new data from this study to build the BCGA. The BCGA contains basal (untreated conditions) gene expression level estimates in beta cells as well as in different cell types in human, rat and mouse pancreas. Hierarchical clustering of expression profile estimates classify cell types based on species while beta cells were clustered together. Conclusion Our gene atlas is a valuable source for detailed information on the gene expression distribution in beta cells and pancreatic islets along with insulin producing cell lines. The BCGA tool, as well as the data and code used to generate the Atlas are available at the T1Dbase website (T1DBase.org). PMID:19146692

  3. Islet Stellate Cells Isolated from Fibrotic Islet of Goto-Kakizaki Rats Affect Biological Behavior of Beta-Cell.

    PubMed

    Li, Feng-Fei; Chen, Bi-Jun; Li, Wei; Li, Ling; Zha, Min; Zhou, S; Bachem, M G; Sun, Zi-Lin

    2016-01-01

    We previously isolated islet stellate cells (ISCs) from healthy Wistar rat islets. In the present study, we isolated "already primed by diabetic environment" ISCs from islets of Goto-Kakizaki rats, determined the gene profile of these cells, and assessed the effects of these ISCs on beta-cell function and survival. We detected gene expression of ISCs by digital gene expression. INS-1 cell proliferation, apoptosis, and insulin production were measured after being treated with ISCs supernatant (SN). We observed the similar expression pattern of ISCs and PSCs, but 1067 differentially expressed genes. Insulin production in INS-1 cells cultured with ISC-SN was significantly reduced. The 5-ethynyl-2'-deoxyuridine-positive INS-1 cells treated with ISC-SN were decreased. Propidium iodide- (PI-) positive INS-1 cells were 2.6-fold higher than those in control groups. Caspase-3 activity was increased. In conclusion, ISCs presented in fibrotic islet of GK rats might be special PSCs, which impaired beta-cell function and proliferation and increased beta-cell apoptosis.

  4. Islet Stellate Cells Isolated from Fibrotic Islet of Goto-Kakizaki Rats Affect Biological Behavior of Beta-Cell

    PubMed Central

    Li, Feng-Fei; Chen, Bi-Jun; Li, Wei; Li, Ling; Zha, Min; Zhou, S.; Bachem, M. G.; Sun, Zi-Lin

    2016-01-01

    We previously isolated islet stellate cells (ISCs) from healthy Wistar rat islets. In the present study, we isolated “already primed by diabetic environment” ISCs from islets of Goto-Kakizaki rats, determined the gene profile of these cells, and assessed the effects of these ISCs on beta-cell function and survival. We detected gene expression of ISCs by digital gene expression. INS-1 cell proliferation, apoptosis, and insulin production were measured after being treated with ISCs supernatant (SN). We observed the similar expression pattern of ISCs and PSCs, but 1067 differentially expressed genes. Insulin production in INS-1 cells cultured with ISC-SN was significantly reduced. The 5-ethynyl-2′-deoxyuridine-positive INS-1 cells treated with ISC-SN were decreased. Propidium iodide- (PI-) positive INS-1 cells were 2.6-fold higher than those in control groups. Caspase-3 activity was increased. In conclusion, ISCs presented in fibrotic islet of GK rats might be special PSCs, which impaired beta-cell function and proliferation and increased beta-cell apoptosis. PMID:26697502

  5. Trefoil factor 3 stimulates human and rodent pancreatic islet beta-cell replication with retention of function.

    PubMed

    Fueger, Patrick T; Schisler, Jonathan C; Lu, Danhong; Babu, Daniella A; Mirmira, Raghavendra G; Newgard, Christopher B; Hohmeier, Hans E

    2008-05-01

    Both major forms of diabetes involve a decline in beta-cell mass, mediated by autoimmune destruction of insulin-producing cells in type 1 diabetes and by increased rates of apoptosis secondary to metabolic stress in type 2 diabetes. Methods for controlled expansion of beta-cell mass are currently not available but would have great potential utility for treatment of these diseases. In the current study, we demonstrate that overexpression of trefoil factor 3 (TFF3) in rat pancreatic islets results in a 4- to 5-fold increase in [(3)H]thymidine incorporation, with full retention of glucose-stimulated insulin secretion. This increase was almost exclusively due to stimulation of beta-cell replication, as demonstrated by studies of bromodeoxyuridine incorporation and co-immunofluorescence analysis with anti-bromodeoxyuridine and antiinsulin or antiglucagon antibodies. The proliferative effect of TFF3 required the presence of serum or 0.5 ng/ml epidermal growth factor. The ability of TFF3 overexpression to stimulate proliferation of rat islets in serum was abolished by the addition of epidermal growth factor receptor antagonist AG1478. Furthermore, TFF3-induced increases in [3H]thymidine incorporation in rat islets cultured in serum was blocked by overexpression of a dominant-negative Akt protein or treatment with triciribine, an Akt inhibitor. Finally, overexpression of TFF3 also caused a doubling of [3H]thymidine incorporation in human islets. In summary, our findings reveal a novel TFF3-mediated pathway for stimulation of beta-cell replication that could ultimately be exploited for expansion or preservation of islet beta-cell mass.

  6. Expression of adiponectin receptors in pancreatic beta cells.

    PubMed

    Kharroubi, Ilham; Rasschaert, Joanne; Eizirik, Décio L; Cnop, Miriam

    2003-12-26

    Pancreatic beta cell dysfunction is an early and crucial pathogenic factor in the development of type 2 diabetes. Free fatty acids (FFA) and adipokines released from adipose tissues lead to both the development of insulin resistance and beta cell dysfunction. Adiponectin is a novel adipokine with antidiabetic properties. Its circulating concentrations are reduced in subjects with increased visceral adiposity, insulin resistance, or type 2 diabetes. Very recently, the cloning of two adiponectin receptors AdipoR1 and AdipoR2 was reported. AdipoR1 is abundantly expressed in muscle, while AdipoR2 is predominantly expressed in liver. Here we report the marked expression of mRNAs for the adiponectin receptors AdipoR1 and AdipoR2 in human and rat pancreatic beta cells, at levels similar to liver and greater than muscle. Adiponectin receptor expression is increased by beta cell exposure to the unsaturated FFA oleate, and treatment of insulin-producing cells with globular adiponectin induces lipoprotein lipase expression. Regulated adiponectin receptor expression on pancreatic beta cells might be a novel mechanism modulating the effects of circulating adiponectin. PMID:14651988

  7. Pancreatic beta-cells expressing GLP-1 are resistant to the toxic effects of immunosuppressive drugs.

    PubMed

    D'Amico, Eugenio; Hui, Hongxiang; Khoury, Nasif; Di Mario, Umberto; Perfetti, Riccardo

    2005-04-01

    Glucose intolerance is often observed after pancreatic islet cell transplantation. The administration of immunosuppressive agents (ISD), necessary to avoid tissue rejection, is in part responsible for hyperglycemia. To investigate whether mouse insulinoma (MIN6) cells transfected with the glucagon like peptide-1 (GLP-1) fragment of the proglucagon gene (RIP/GLP-1 MIN6 cells) are resistant to the toxicity derived from the administration of ISD. RIP/GLP-1 MIN6 cells, as well as parental MIN6 cells, were exposed to a cocktail of ISD. The secretion of insulin and the expression of apoptosis-related proteins were investigated by RIA and western blot analysis. Cell apoptosis was quantified by FACS analysis. Finally, to study whether the antiapoptotic action of GLP-1 was a function of its effect on insulin secretion, or rather it was a direct effect of GLP-1, cells were cultured with or without diazoxide or exendin-9. GLP-1 improved the functional activity and the viability of cells exposed to ISD. The insulin secretion of RIP/GLP-1 MIN6 cells after exposure to ISD was preserved. The expression of GLP-1 by beta-cells reduced the number of apoptotic cells and increased the expression of the antiapoptotic protein Bcl-2. GLP-1 also decreased the abundance of the proapoptotic markers PARP-p85 and Smac/Diablo. Treatment of cells with the diazoxide did not abolish the protective advantage that cells transfected with GLP-1 had; conversely the exposure of cells to exendin-9 was associated with a restored susceptibility to apoptosis. This report demonstrates that GLP-1 is capable of preserving beta-cell function and protecting cells from apoptotic cell death.

  8. Mechanisms of Beta Cell Dysfunction Associated With Viral Infection.

    PubMed

    Petzold, Antje; Solimena, Michele; Knoch, Klaus-Peter

    2015-10-01

    Type 1 diabetes (T1D) results from genetic predisposition and environmental factors leading to the autoimmune destruction of pancreatic beta cells. Recently, a rapid increase in the incidence of childhood T1D has been observed worldwide; this is too fast to be explained by genetic factors alone, pointing to the spreading of environmental factors linked to the disease. Enteroviruses (EVs) are perhaps the most investigated environmental agents in relationship to the pathogenesis of T1D. While several studies point to the likelihood of such correlation, epidemiological evidence in its support is inconclusive or in some instances even against it. Hence, it is still unknown if and how EVs are involved in the development of T1D. Here we review recent findings concerning the biology of EV in beta cells and the potential implications of this knowledge for the understanding of beta cell dysfunction and autoimmune destruction in T1D.

  9. Islet distribution of Peptide YY and its regulatory role in primary mouse islets and immortalised rodent and human beta-cell function and survival.

    PubMed

    Khan, Dawood; Vasu, Srividya; Moffett, R Charlotte; Irwin, Nigel; Flatt, Peter R

    2016-11-15

    Recent evidence suggests that the classic gut peptide, Peptide YY (PYY), could play a fundamental role in endocrine pancreatic function. In the present study expression of PYY and its NPY receptors on mouse islets and immortalised rodent and human beta-cells was examined together with the effects of both major circulating forms of PYY, namely PYY(1-36) and PYY(3-36), on beta-cell function, murine islet adaptions to insulin deficiency/resistance, as well as direct effects on cultured beta-cell proliferation and apoptosis. In vivo administration of PYY(3-36), but not PYY(1-36), markedly (p < 0.05) decreased food intake in overnight fasted mice. Neither form of PYY affected glucose disposal or insulin secretion following an i.p. glucose challenge. However, in vitro, PYY(1-36) and PYY(3-36) inhibited (p < 0.05 to p < 0.001) glucose, alanine and GLP-1 stimulated insulin secretion from immortalised rodent and human beta-cells, as well as isolated mouse islets, by impeding alterations in membrane potential, [Ca(2+)]i and elevations of cAMP. Mice treated with multiple low dose streptozotocin presented with severe (p < 0.01) loss of beta-cell mass accompanied by notable increases (p < 0.001) in alpha and PP cell numbers. In contrast, hydrocortisone-induced insulin resistance increased islet number (p < 0.01) and beta-cell mass (p < 0.001). PYY expression was consistently observed in alpha-, PP- and delta-, but not beta-cells. Streptozotocin decreased islet PYY co-localisation with PP (p < 0.05) and somatostatin (p < 0.001), whilst hydrocortisone increased PYY co-localisation with glucagon (p < 0.05) in mice. More detailed in vitro investigations revealed that both forms of PYY augmented (p < 0.05 to p < 0.01) immortalised human and rodent beta-cell proliferation and protected against streptozotocin-induced cytotoxicity, to a similar or superior extent as the well characterised beta-cell proliferative and anti-apoptotic agent GLP-1. Taken together

  10. On the coherent behavior of pancreatic beta cell clusters

    NASA Astrophysics Data System (ADS)

    Loppini, Alessandro; Capolupo, Antonio; Cherubini, Christian; Gizzi, Alessio; Bertolaso, Marta; Filippi, Simonetta; Vitiello, Giuseppe

    2014-09-01

    Beta cells in pancreas represent an example of coupled biological oscillators which via communication pathways, are able to synchronize their electrical activity, giving rise to pulsatile insulin release. In this work we numerically analyze scale free self-similarity features of membrane voltage signal power density spectrum, through a stochastic dynamical model for beta cells in the islets of Langerhans fine tuned on mouse experimental data. Adopting the algebraic approach of coherent state formalism, we show how coherent molecular domains can arise from proper functional conditions leading to a parallelism with “phase transition” phenomena of field theory.

  11. Tumor necrosis factor-alpha-induced changes in insulin-producing beta-cells.

    PubMed

    Parkash, Jai; Chaudhry, Muhammad A; Rhoten, William B

    2005-10-01

    The migration of macrophages and lymphocytes that produce cytokines such as tumor necrosis factor-alpha (TNF-alpha) causes beta-cell death, leading to type 1 diabetes. Similarly, in type 2 diabetes, the adipocyte-derived cytokines including TNF-alpha are elevated in the circulation, causing inflammation and insulin resistance. Thus, the studies described in this article using TNF-alpha are relevant to furthering our understanding of the pathogenesis of diabetes mellitus. We used RINr1046-38 (RIN) insulin-producing beta-cells, which constitutively express calbindin-D(28k), to characterize the effect of TNF-alpha on apoptosis, replication, insulin release, and gene and protein expression. Western blots of TNF-alpha-treated RIN cells revealed a decrease in calbindin-D(28k). By ELISA, TNF-alpha-treated beta-cells had 47% less calbindin-D(28k) than controls. In association with the decline in calbindin-D(28k), TNF-alpha treatment of RIN cells led to a 73% greater increase in changes in intracellular calcium concentration (Delta[Ca(2+)](i)) in TNF-alpha-treated cells as compared to that in control RIN cells upon treatment with 50 mM KCl; caused a greater increase in the [Ca(2+)](i) following the addition of 5.5 microM ionomycin; increased by more than threefold the apoptotic rate, expressed as the percentage of TUNEL-positive nuclei to total nuclei; decreased the rate of cell replication by 36%; and increased and decreased selectively the expression of specific genes as determined by microarray analysis. The subcellular localizations of Bcl-2, an antiapoptotic protein, and Bax, a proapoptotic protein, within RIN cells were altered with TNF-alpha treatment such that the two were colocalized with mitochondria in the perinuclear region. We conclude that the proapoptotic action of TNF-alpha on beta-cells is manifested via decreased expression of calbindin-D(28k) and is mediated at least in part by [Ca(2+)](i). PMID:16114068

  12. Beta-cell Assembly for the Quad Gas Sampling Detector

    SciTech Connect

    Cooper, Matthew W.; Bowyer, Ted W.; McIntyre, Justin I.; Hayes, James C.; Heimbigner, Tom R.; Ripplinger, Michael D.; Thompson, Robert C.

    2008-05-05

    The beta-cells used in the beta-gamma detector have taken time to develop and to standardize the assembly of them. In making the assembly routine it is important to have step by step assembly instructions as well as a list of potential problems and their solutions. This document attempts to accomplish these goals.

  13. Regulation of. beta. -cell glucose transporter gene expression

    SciTech Connect

    Chen, Ling; Alam, Tausif; Johnson, J.H.; Unger, R.H. Department of Veterans Affairs Medical Center, Dallas, TX ); Hughes, S.; Newgard, C.B. )

    1990-06-01

    It has been postulated that a glucose transporter of {beta} cells (GLUT-2) may be important in glucose-stimulated insulin secretion. To determine whether this transporter is constitutively expressed or regulated, the authors subjected conscious unrestrained Wistar rats to perturbations in glucose homeostasis and quantitated {beta}-cell GLUT-2 mRNA by in situ hybridization. After 3 hr of hypoglycemia, GLUT-2 and proinsulin mRNA signal densities were reduced by 25% of the level in control rats. After 4 days, GLUT-2 and proinsulin mRNA densities were reduced by 85% and 65%, respectively. After 12 days of hypoglycemia, the K{sub m} for 3-O-methyl-D-glucose transport in isolated rat islets, normally 18-20 mM, was 2.5 mM. This provides functional evidence of a profound reduction of high K{sub m} glucose transporter in {beta} cells. In contrast, GLUT-2 was only slightly reduced by hypoglycemia in liver. To determine the effect of prolonged hyperglycemia, they also infused animals with 50% (wt/vol) glucose for 5 days. Hyperglycemic clamping increased GLUT-2 mRNA by 46% whereas proinsulin mRNA doubled. They conclude that GLUT-2 expression in {beta} cells, but not liver, is subject to regulation by certain perturbations in blood glucose homeostasis.

  14. Loss of TRB3 alters dynamics of MLK3-JNK signaling and inhibits cytokine-activated pancreatic beta cell death.

    PubMed

    Humphrey, Rohan K; Ray, Anamika; Gonuguntla, Sumati; Hao, Ergeng; Jhala, Ulupi S

    2014-10-24

    Disabling cellular defense mechanisms is essential for induction of apoptosis. We have previously shown that cytokine-mediated activation of the MAP3K MLK3 stabilizes TRB3 protein levels to inhibit AKT and compromise beta cell survival. Here, we show that genetic deletion of TRB3 results in basal activation of AKT, preserves mitochondrial integrity, and confers resistance against cytokine-induced pancreatic beta cell death. Mechanistically, we find that TRB3 stabilizes MLK3, most likely by suppressing AKT-directed phosphorylation, ubiquitination, and proteasomal degradation of MLK3. Accordingly, TRB3(-/-) islets show a decrease in both the amplitude and duration of cytokine-stimulated MLK3 induction and JNK activation. It is well known that JNK signaling is facilitated by a feed forward loop of sequential kinase phosphorylation and is reinforced by a mutual stabilization of the module components. The failure of TRB3(-/-) islets to mount an optimal JNK activation response, coupled with the ability of TRB3 to engage and maintain steady state levels of MLK3, recasts TRB3 as an integral functional component of the JNK module in pancreatic beta cells. PMID:25204656

  15. IDO-Expressing Fibroblasts Protect Islet Beta Cells From Immunological Attack and Reverse Hyperglycemia in Non-Obese Diabetic Mice.

    PubMed

    Zhang, Yun; Jalili, Reza B; Kilani, Ruhangiz T; Elizei, Sanam Salimi; Farrokhi, Ali; Khosravi-Maharlooei, Mohsen; Warnock, Garth L; Ao, Ziliang; Marzban, Lucy; Ghahary, Aziz

    2016-09-01

    Indoleamine 2,3-dioxygenase (IDO) induces immunological tolerance in physiological and pathological conditions. Therefore, we used dermal fibroblasts with stable IDO expression as a cell therapy to: (i) Investigate the factors determining the efficacy of this cell therapy for autoimmune diabetes in non-obese diabetic (NOD) mice; (ii) Scrutinize the potential immunological mechanisms. Newly diabetic NOD mice were randomly injected with either 10 × 10(6) (10M) or 15 × 10(6) (15M) IDO-expressing dermal fibroblasts. Blood glucose levels (BGLs), body weight, plasma kynurenine levels, insulitis severity, islet beta cell function, autoreactive CD8(+) T cells, Th17 cells and regulatory T cells (Tregs) were then investigated in these mice. IL-1β and cleaved caspase-3 levels were assessed in islets co-cultured with IDO-expressing fibroblasts. BGLs in 83% mice treated with 15M IDO-expressing fibroblasts recovered to normal up to 120 days. However, only 17% mice treated with 10M IDO-expressing cells were reversed to normoglycemia. A 15M IDO-expressing fibroblasts significantly reduced infiltrated immune cells in islets and recovered the functionality of remaining islet beta cells in NOD mice. Additionally, they successfully inhibited autoreactive CD8(+) T cells and Th17 cells as well as increased Tregs in different organs of NOD mice. Islet beta cells co-cultured with IDO-expressing fibroblasts had reduced IL-1β levels and cell apoptosis. Both cell number and IDO enzymatic activity contributes to the efficiency of IDO cell therapy. Optimized IDO-expressing fibroblasts successfully reverse the progression of diabetes in NOD mice through induction of Tregs as well as inhibition of beta cell specific autoreactive CD8(+) T cells and Th17 cells. J. Cell. Physiol. 231: 1964-1973, 2016. © 2016 Wiley Periodicals, Inc.

  16. The role of autophagy in pancreatic beta-cell and diabetes.

    PubMed

    Fujitani, Yoshio; Kawamori, Ryuzo; Watada, Hirotaka

    2009-02-01

    Pancreatic beta-cells play a key role in glucose homeostasis in mammals. Although large-scale protein synthesis and degradation occur in pancreatic beta-cells, the mechanism underlying dynamic protein turnover in beta-cells remains largely unknown. We found low-level constitutive autophagy in beta-cells of C57BL/6 mice fed a standard diet; however, autophagy was markedly upregulated in mice fed a high-fat diet. beta-cells of diabetic db/db mice contained large numbers of autophagosomes, compared with nondiabetic db/misty controls. The functional importance of autophagy was analyzed using beta-cell-specific Atg7 knockout mice. Autophagy-deficient mice showed degeneration of beta-cells and impaired glucose tolerance with reduced insulin secretion. While a high-fat diet stimulated beta-cell autophagy in control mice, it induced a profound deterioration of glucose intolerance in beta-cell autophagy-deficient mutants, partly because of the lack of a compensatory increase in beta-cell mass. These results suggest that the degradation of unnecessary cellular components by autophagy is essential for maintenance of the architecture and function of beta-cells. Autophagy also serves as a crucial element of stress responses to protect beta-cells under insulin-resistant states. Impairment of autophagic machinery could thus predispose individuals to type 2 diabetes. PMID:19158492

  17. A Figure-of-Merit for Beta Cell Detector Characterization

    SciTech Connect

    Foxe, Michael P.; Miller, Brian W.; Suarez, Rey; Hayes, James C.

    2015-09-02

    In order to decrease the minimum detectable activities (MDAs) of beta-gamma radioxenon detectors, it is important to increase the ability to resolve the individual isotopes. One proposed method for doing this is to increase the energy resolution of the beta cell through the use of silicon detectors. While silicon detectors can improve the energy resolution, it is accompanied with a decrease in detection efficiency compared to plastic scintillator beta cells. Due to the uncertainty on the impact of the competing variables, we have developed a figure-of-merit (FOM) capable of determining the impact of detector parameters on the MDAs. By utilizing the FOM to analyze different detectors, we are able to directly compare current and future detectors and estimate their impact on the radioxenon MDAs.

  18. Circadian Transcription from Beta Cell Function to Diabetes Pathophysiology.

    PubMed

    Perelis, Mark; Ramsey, Kathryn Moynihan; Marcheva, Biliana; Bass, Joseph

    2016-08-01

    The mammalian circadian clock plays a central role in the temporal coordination of physiology across the 24-h light-dark cycle. A major function of the clock is to maintain energy constancy in anticipation of alternating periods of fasting and feeding that correspond with sleep and wakefulness. While it has long been recognized that humans exhibit robust variation in glucose tolerance and insulin sensitivity across the sleep-wake cycle, experimental genetic analysis has now revealed that the clock transcription cycle plays an essential role in insulin secretion and metabolic function within pancreatic beta cells. This review addresses how studies of the beta cell clock may elucidate the etiology of subtypes of diabetes associated with circadian and sleep cycle disruption, in addition to more general forms of the disease. PMID:27440914

  19. Disruption of Tsc2 in pancreatic beta cells induces beta cell mass expansion and improved glucose tolerance in a TORC1-dependent manner.

    PubMed

    Rachdi, Latif; Balcazar, Norman; Osorio-Duque, Fernando; Elghazi, Lynda; Weiss, Aaron; Gould, Aaron; Chang-Chen, Karen J; Gambello, Michael J; Bernal-Mizrachi, Ernesto

    2008-07-01

    Regulation of pancreatic beta cell mass and function is a major determinant for the development of diabetes. Growth factors and nutrients are important regulators of beta cell mass and function. The signaling pathways by which these growth signals modulate these processes have not been completely elucidated. Tsc2 is an attractive candidate to modulate these processes, because it is a converging point for growth factor and nutrient signals. In these experiments, we generated mice with conditional deletion of Tsc2 in beta cells (betaTsc2(-/-)). These mice exhibited decreased glucose levels and hyperinsulinemia in the fasting and fed state. Improved glucose tolerance in these mice was observed as early as 4 weeks of age and was still present in 52-week-old mice. Deletion of Tsc2 in beta cells induced expansion of beta cell mass by increased proliferation and cell size. Rapamycin treatment reversed the metabolic changes in betaTsc2(-/-) mice by induction of insulin resistance and reduction of beta cell mass. The reduction of beta cell mass in betaTsc2(-/-) mice by inhibition of the mTOR/Raptor (TORC1) complex with rapamycin treatment suggests that TORC1 mediates proliferative and growth signals induced by deletion of Tsc2 in beta cells. These studies uncover a critical role for the Tsc2/mTOR pathway in regulation of beta cell mass and carbohydrate metabolism in vivo.

  20. Reduction in placental growth factor impaired gestational beta-cell proliferation through crosstalk between beta-cells and islet endothelial cells

    PubMed Central

    Xu, Xiaosheng; Shen, Jian

    2016-01-01

    Reduced placental growth factor (PLGF) during pregnancy is known to be a reason for developing preeclampsia (PE) and gestational diabetes mellitus (GDM), but the underlying mechanisms remain unclear. Recently, it has been shown that reduced PLGF may induce GDM through suppressing beta-cell mass growth in a PI3k/Akt signalling-dependent manner. Here, we dissected the interaction between beta-cells and islet endothelial cells in this model. We analysed proliferation of beta-cells and islet endothelial cells at different time points of gestation in mice. We cultured mouse islet endothelial cells (MS1), with or without PLGF. We cultured primary mouse beta-cells in conditioned media from PLGF-treated MS1. We cultured MS1 cells in conditioned media from proliferating beta-cells that were activated with conditioned media from PLGF-treated MS1 cells. We analysed cell proliferation by BrdU incorporation. We analysed cell growth by a MTT assay. We found that during mouse gestation, the increases in cell proliferation occurred earlier in beta-cells than in islet endothelial cells. In vitro, PLGF itself failed to induce proliferation of MS1 cells. However, conditioned media from the PLGF-treated MS1 cells induced beta-cell proliferation, resulting in increases in beta-cell number. Moreover, proliferation of MS1 cells significantly increased when MS1 cells were cultured in conditioned media from proliferating beta-cells activated with conditioned media from PLGF-treated MS1 cells. Thus, our data suggest that gestational PLGF may stimulate islet endothelial cells to release growth factors to promote beta-cell proliferation, and proliferating beta-cells in turn release endothelial cell growth factor to increase proliferation of endothelial cells. PE-associated reduction in PLGF impairs these processes to result in islet growth impairment, and subsequently the onset of GDM. PMID:27725870

  1. PDX-1 and MafA play a crucial role in pancreatic beta-cell differentiation and maintenance of mature beta-cell function.

    PubMed

    Kaneto, Hideaki; Miyatsuka, Takeshi; Kawamori, Dan; Yamamoto, Kaoru; Kato, Ken; Shiraiwa, Toshihiko; Katakami, Naoto; Yamasaki, Yoshimitsu; Matsuhisa, Munehide; Matsuoka, Taka-Aki

    2008-05-01

    Pancreatic and duodenal homeobox factor-1 (PDX-1) plays a crucial role in pancreas development, beta-cell differentiation, and maintenance of mature beta-cell function. PDX-1 expression is maintained in pancreatic precursor cells during pancreas development but becomes restricted to beta-cells in mature pancreas. In mature beta-cells, PDX-1 transactivates the insulin and other genes involved in glucose sensing and metabolism such as GLUT2 and glucokinase. MafA is a recently isolated beta-cell-specific transcription factor which functions as a potent activator of insulin gene transcription. Furthermore, these transcription factors play an important role in induction of insulin-producing cells in various non-beta-cells and thus could be therapeutic targets for diabetes. On the other hand, under diabetic conditions, expression and/or activities of PDX-1 and MafA in beta-cells are reduced, which leads to suppression of insulin biosynthesis and secretion. It is likely that alteration of such transcription factors explains, at least in part, the molecular mechanism for beta-cell glucose toxicity found in diabetes.

  2. Regulating the beta cell mass as a strategy for type-2 diabetes treatment.

    PubMed

    Song, Imane; Muller, Christo; Louw, Johan; Bouwens, Luc

    2015-01-01

    The incidence of type 2 diabetes (T2D) increases dramatically worldwide and has created an enormous health care burden. Obesity, dyslipidemia and insulin resistance are major risk factors for the development of T2D, but the major factor leading to the disease is failure of the insulin-producing beta cell mass to compensate for increasing insulin demands of the body. Progression of the disease further diminishes the beta cell mass as a result of lipotoxicity and glucotoxicity for which beta cells are particularly sensitive. Hence, treatment aiming to prevent beta cell loss or increase the number of beta cells could inhibit diabetes progression or lead to restoration of normal metabolism. Whereas current and new antidiabetic drugs are mainly targeting insulin secretion and action or glucose uptake, newer interventions must be found that prevent beta cell loss or increase beta cell number. The targets for this are beta cell proliferation, neogenesis and survival. This review examines major evidence from animal experiments suggesting that it is feasible to regulate the beta cell mass by bioactive compounds like growth factors, cytokines, hormones, phytochemicals and small molecules. Often the mode of action remains unclear due to inadequate methods to assess the effects of the compounds on the beta cell dynamics. Furthermore, a major challenge is to identify compounds with sufficient specificity in order to avoid unwanted effects on other cell types. Provided such safety issues can be solved, this may provide a curative approach for diabetes treatment.

  3. Role of MicroRNAs in Islet Beta-Cell Compensation and Failure during Diabetes

    PubMed Central

    Plaisance, Valérie; Waeber, Gérard

    2014-01-01

    Pancreatic beta-cell function and mass are markedly adaptive to compensate for the changes in insulin requirement observed during several situations such as pregnancy, obesity, glucocorticoids excess, or administration. This requires a beta-cell compensation which is achieved through a gain of beta-cell mass and function. Elucidating the physiological mechanisms that promote functional beta-cell mass expansion and that protect cells against death, is a key therapeutic target for diabetes. In this respect, several recent studies have emphasized the instrumental role of microRNAs in the control of beta-cell function. MicroRNAs are negative regulators of gene expression, and are pivotal for the control of beta-cell proliferation, function, and survival. On the one hand, changes in specific microRNA levels have been associated with beta-cell compensation and are triggered by hormones or bioactive peptides that promote beta-cell survival and function. Conversely, modifications in the expression of other specific microRNAs contribute to beta-cell dysfunction and death elicited by diabetogenic factors including, cytokines, chronic hyperlipidemia, hyperglycemia, and oxidized LDL. This review underlines the importance of targeting the microRNA network for future innovative therapies aiming at preventing the beta-cell decline in diabetes. PMID:24734255

  4. Parathyroid Hormone-Related Peptide (1-36) Enhances Beta Cell Regeneration and Increases Beta Cell Mass in a Mouse Model of Partial Pancreatectomy

    PubMed Central

    Mozar, Anaïs; Lin, Hugo; Williams, Katoura; Chin, Connie; Li, Rosemary; Kondegowda, Nagesha Guthalu; Stewart, Andrew F.; Garcia-Ocaña, Adolfo; Vasavada, Rupangi Chhaya

    2016-01-01

    Aims/Hypothesis Finding ways to stimulate the regeneration of endogenous pancreatic beta cells is an important goal in the treatment of diabetes. Parathyroid hormone-related protein (PTHrP), the full-length (1–139) and amino-terminal (1–36) peptides, enhance beta cell function, proliferation, and survival. Therefore, we hypothesize that PTHrP(1–36) has the potential to regenerate endogenous beta cells. Methods The partial pancreatectomy (PPx) mouse model of beta cell injury was used to test this hypothesis. Male Balb/c mice underwent either sham-operation or PPx, and were subsequently injected with PTHrP(1–36) (160μg/kg) or vehicle (veh), for 7, 30, or 90 days. The four groups of mice, sham-veh, sham-PTHrP, PPx-veh, and PPx-PTHrP were assessed for PTHrP and receptor expression, and glucose and beta cell homeostasis. Results PTHrP-receptor, but not the ligand, was significantly up-regulated in islets from mice that underwent PPx compared to sham-operated mice. This suggests that exogenous PTHrP could further enhance beta cell regeneration after PPx. PTHrP did not significantly affect body weight, blood glucose, plasma insulin, or insulin sensitivity, in either sham or PPx mice. Glucose tolerance improved in the PPx-PTHrP versus PPx-veh mice only in the early stages of treatment. As hypothesized, there was a significant increase in beta cell proliferation in PPx-PTHrP mice at days 7 and 30; however, this was normalized by day 90, compared to PPx-veh mice. Enhanced beta cell proliferation translated to a marked increase in beta cell mass at day 90, in PPx-PTHrP versus PPx-veh mice. Conclusions PTHrP(1–36) significantly enhances beta cell regeneration through increased beta cell proliferation and beta cell mass after PPx. Future studies will determine the potential of PTHrP to enhance functional beta cell mass in the setting of diabetes. PMID:27391423

  5. Cdk5 inhibitory peptide (CIP) inhibits Cdk5/p25 activity induced by high glucose in pancreatic beta cells and recovers insulin secretion from p25 damage.

    PubMed

    Zheng, Ya-Li; Li, Congyu; Hu, Ya-Fang; Cao, Li; Wang, Hui; Li, Bo; Lu, Xiao-Hua; Bao, Li; Luo, Hong-Yan; Shukla, Varsha; Amin, Niranjana D; Pant, Harish C

    2013-01-01

    Cdk5/p25 hyperactivity has been demonstrated to lead to neuron apoptosis and degenerations. Chronic exposure to high glucose (HG) results in hyperactivity of Cdk5 and reduced insulin secretion. Here, we set out to determine whether abnormal upregulation of Cdk5/p25 activity may be induced in a pancreatic beta cell line, Min6 cells. We first confirmed that p25 were induced in overexpressed p35 cells treated with HG and increased time course dependence. Next, we showed that no p25 was detected under short time HG stimulation (4-12 hrs), however was detectable in the long exposure in HG cells (24 hrs and 48 hrs). Cdk5 activity in the above cells was much higher than low glucose treated cells and resulted in more than 50% inhibition of insulin secretion. We confirmed these results by overexpression of p25 in Min6 cells. As in cortical neurons, CIP, a small peptide, inhibited Cdk5/p25 activity and restored insulin secretion. The same results were detected in co-infection of dominant negative Cdk5 (DNCdk5) with p25. CIP also reduced beta cells apoptosis induced by Cdk5/p25. These studies indicate that Cdk5/p25 hyperactivation deregulates insulin secretion and induces cell death in pancreatic beta cells and suggests that CIP may serve as a therapeutic agent for type 2 diabetes. PMID:24039692

  6. Treatment with CNX-011-67, a novel GPR40 agonist, delays onset and progression of diabetes and improves beta cell preservation and function in male ZDF rats

    PubMed Central

    2013-01-01

    Background The role of G protein-coupled receptor (GPR40), which is highly expressed in pancreatic beta cells, has been studied extensively in the amelioration of beta cell dysfunction in T2D using rat and mouse islets, beta cell lines and in animal models of diabetes. But its potential as a therapeutic target has not been fully explored. This aim of the study is to evaluate the therapeutic potential of CNX-011-67, a highly selective, potent and orally bioavailable GPR40 agonist, in controlling diabetes and other metabolic parameters. Methods Seven week old male ZDF rats were treated with either vehicle or CNX-011-67, 5 mg/kg twice daily, for seven weeks. The animals were subjected to oral glucose tolerance and insulin tolerance tests. Plasma glucose, insulin, triglyceride, HbA1c, fructosamine and free fatty acids were measured at selected time points. Pancreas from control and treated animals were subjected to insulin and pancreatic and duodenal homeobox 1 (PDX1) immunohistochemistry and were also evaluated by electron microscopy. Also the potential impact of CNX-011-67 on islet insulin secretion, content, ATP levels and markers of both glucose oxidation, beta cell health in rat islets under chronic glucolipotoxic conditions was evaluated. Results Treatment of male ZDF rats with CNX-011-67 for 7 weeks significantly enhanced insulin secretion in response to oral glucose load, delayed the onset of fasting hyperglycemia by 3 weeks, reduced nonfasting glucose excursions, fasting free fatty acids and triglyceride levels. A significant increase in PDX1 expression and insulin content and reduction in plasma fructosamine, HOMA-IR, and beta cell apoptosis were observed. CNX-011-67 improves glucose mediated insulin secretion, insulin gene transcription and islet insulin content in cultured rat islets under chronic glucolipotoxic condition. Also enhanced glucose oxidation in the form of increased islet ATP content and overall improvement in beta cell health in the form of

  7. Whole organism high content screening identifies stimulators of pancreatic beta-cell proliferation.

    PubMed

    Tsuji, Naoki; Ninov, Nikolay; Delawary, Mina; Osman, Sahar; Roh, Alex S; Gut, Philipp; Stainier, Didier Y R

    2014-01-01

    Inducing beta-cell mass expansion in diabetic patients with the aim to restore glucose homeostasis is a promising therapeutic strategy. Although several in vitro studies have been carried out to identify modulators of beta-cell mass expansion, restoring endogenous beta-cell mass in vivo has yet to be achieved. To identify potential stimulators of beta-cell replication in vivo, we established transgenic zebrafish lines that monitor and allow the quantification of cell proliferation by using the fluorescent ubiquitylation-based cell cycle indicator (FUCCI) technology. Using these new reagents, we performed an unbiased chemical screen, and identified 20 small molecules that markedly increased beta-cell proliferation in vivo. Importantly, these structurally distinct molecules, which include clinically-approved drugs, modulate three specific signaling pathways: serotonin, retinoic acid and glucocorticoids, showing the high sensitivity and robustness of our screen. Notably, two drug classes, retinoic acid and glucocorticoids, also promoted beta-cell regeneration after beta-cell ablation. Thus, this study establishes a proof of principle for a high-throughput small molecule-screen for beta-cell proliferation in vivo, and identified compounds that stimulate beta-cell proliferation and regeneration.

  8. Nkx6.1 is essential for maintaining the functional state of pancreatic beta cells.

    PubMed

    Taylor, Brandon L; Liu, Fen-Fen; Sander, Maike

    2013-09-26

    Recently, loss of beta-cell-specific traits has been proposed as an early cause of beta cell failure in diabetes. However, the molecular mechanisms that underlie the loss of beta cell features remain unclear. Here, we identify an Nkx6.1-controlled gene regulatory network as essential for maintaining the functional and molecular traits of mature beta cells. Conditional Nkx6.1 inactivation in adult mice caused rapid-onset diabetes and hypoinsulinemia. Genome-wide analysis of Nkx6.1-regulated genes and functional assays further revealed a critical role for Nkx6.1 in the control of insulin biosynthesis, insulin secretion, and beta cell proliferation. Over time, Nkx6.1-deficient beta cells acquired molecular characteristics of delta cells, revealing a molecular link between impaired beta cell functional properties and loss of cell identity. Given that Nkx6.1 levels are reduced in human type 2 diabetic beta cells, our study lends support to the concept that loss of beta cell features could contribute to the pathogenesis of diabetes. PMID:24035389

  9. Maturation of stem cell-derived beta-cells guided by the expression of urocortin 3.

    PubMed

    van der Meulen, Talitha; Huising, Mark O

    2014-01-01

    Type 1 diabetes (T1D) is a devastating disease precipitated by an autoimmune response directed at the insulin-producing beta-cells of the pancreas for which no cure exists. Stem cell-derived beta-cells show great promise for a cure as they have the potential to supply unlimited numbers of cells that could be derived from a patient's own cells, thus eliminating the need for immunosuppression. Current in vitro protocols for the differentiation of stem cell-derived beta-cells can successfully generate pancreatic endoderm cells. In diabetic rodents, such cells can differentiate further along the beta-cell lineage until they are eventually capable of restoring normoglycemia. While these observations demonstrate that stem cell-derived pancreatic endoderm has the potential to differentiate into mature, glucose-responsive beta-cells, the signals that direct differentiation and maturation from pancreatic endoderm onwards remain poorly understood. In this review, we analyze the sequence of events that culminates in the formation of beta-cells during embryonic development. and summarize how current protocols to generate beta-cells have sought to capitalize on this ontogenic template. We place particular emphasis on the current challenges and opportunities which occur in the later stages of beta-cell differentiation and maturation of transplantable stem cell-derived beta-cells. Another focus is on the question how the use of recently identified maturation markers such as urocortin 3 can be instrumental in guiding these efforts. PMID:25148370

  10. Quantitative-Proteomic Comparison of Alpha and Beta Cells to Uncover Novel Targets for Lineage Reprogramming

    PubMed Central

    Mertins, Philipp; Udeshi, Namrata D.; Dančík, Vlado; Fomina-Yadlin, Dina; Kubicek, Stefan; Clemons, Paul A.; Schreiber, Stuart L.; Carr, Steven A.; Wagner, Bridget K.

    2014-01-01

    Type-1 diabetes (T1D) is an autoimmune disease in which insulin-secreting pancreatic beta cells are destroyed by the immune system. An emerging strategy to regenerate beta-cell mass is through transdifferentiation of pancreatic alpha cells to beta cells. We previously reported two small molecules, BRD7389 and GW8510, that induce insulin expression in a mouse alpha cell line and provide a glimpse into potential intermediate cell states in beta-cell reprogramming from alpha cells. These small-molecule studies suggested that inhibition of kinases in particular may induce the expression of several beta-cell markers in alpha cells. To identify potential lineage reprogramming protein targets, we compared the transcriptome, proteome, and phosphoproteome of alpha cells, beta cells, and compound-treated alpha cells. Our phosphoproteomic analysis indicated that two kinases, BRSK1 and CAMKK2, exhibit decreased phosphorylation in beta cells compared to alpha cells, and in compound-treated alpha cells compared to DMSO-treated alpha cells. Knock-down of these kinases in alpha cells resulted in expression of key beta-cell markers. These results provide evidence that perturbation of the kinome may be important for lineage reprogramming of alpha cells to beta cells. PMID:24759943

  11. Inorganic mercury causes pancreatic beta-cell death via the oxidative stress-induced apoptotic and necrotic pathways

    SciTech Connect

    Chen Yawen; Huang Chunfa; Yang Chingyao; Yen Chengchieh; Tsai Kehsung; Liu Shinghwa

    2010-03-15

    Mercury is a well-known highly toxic metal. In this study, we characterize and investigate the cytotoxicity and its possible mechanisms of inorganic mercury in pancreatic beta-cells. Mercury chloride (HgCl{sub 2}) dose-dependently decreased the function of insulin secretion and cell viability in pancreatic beta-cell-derived HIT-T15 cells and isolated mouse pancreatic islets. HgCl{sub 2} significantly increased ROS formation in HIT-T15 cells. Antioxidant N-acetylcysteine effectively reversed HgCl{sub 2}-induced insulin secretion dysfunction in HIT-T15 cells and isolated mouse pancreatic islets. Moreover, HgCl{sub 2} increased sub-G1 hypodiploids and annexin-V binding in HIT-T15 cells, indicating that HgCl{sub 2} possessed ability in apoptosis induction. HgCl{sub 2} also displayed several features of mitochondria-dependent apoptotic signals including disruption of the mitochondrial membrane potential, increase of mitochondrial cytochrome c release and activations of poly (ADP-ribose) polymerase (PARP) and caspase 3. Exposure of HIT-T15 cells to HgCl{sub 2} could significantly increase both apoptotic and necrotic cell populations by acridine orange/ethidium bromide dual staining. Meanwhile, HgCl{sub 2} could also trigger the depletion of intracellular ATP levels and increase the LDH release from HIT-T15 cells. These HgCl{sub 2}-induced cell death-related signals could be significantly reversed by N-acetylcysteine. The intracellular mercury levels were markedly elevated in HgCl{sub 2}-treated HIT-T15 cells. Taken together, these results suggest that HgCl{sub 2}-induced oxidative stress causes pancreatic beta-cell dysfunction and cytotoxicity involved the co-existence of apoptotic and necrotic cell death.

  12. In-vitro differentiation of pancreatic beta-cells.

    PubMed

    Soria, B

    2001-10-01

    Stem cell biology is a new field that holds promise for in-vitro mass production of pancreatic beta-cells, which are responsible for insulin synthesis, storage, and release. Lack or defect of insulin produces diabetes mellitus, a devastating disease suffered by 150 million people in the world. Transplantation of insulin-producing cells could be a cure for type 1 and some cases of type 2 diabetes, however this procedure is limited by the scarcity of material. Obtaining pancreatic beta-cells from embryonic stem cells would overcome this problem. We have derived insulin-producing cells from mouse embryonic stem cells by a 3-step in-vitro differentiation method consisting of directed differentiation, cell-lineage selection, and maturation. These insulin-producing cells normalize blood glucose when transplanted into streptozotocin-diabetic mice. Strategies to increase islet precursor cells from embryonic stem cells include the expression of relevant transcription factors (Pdx1, Ngn3, Isl-1, etc), together with the use of extracellular factors. Once a high enough proportion of islet precursors has been obtained there is a need for cell-lineage selection in order to purify the desired cell population. For this purpose, we designed a cell-trapping method based on a chimeric gene that fuses the human insulin gene regulatory region with the structural gene that confers resistance to neomycin. When incorporated into embryonic stem cells, this fusion gene will generate neomycin resistance in those cells that initiate the synthesis of insulin. Not only embryonic, but also adult stem cells are potential sources for insulin-containing cells. Duct cells from the adult pancreas are committed to differentiate into the four islet cell types; other possibilities may include nestin-positive cells from islets and adult pluripotent stem cells from other origins. Whilst the former are committed to be islet cells but have a reduced capacity to expand, the latter are more pluripotent and

  13. UPR in palmitate-treated pancreatic beta-cells is not affected by altering oxidation of the fatty acid

    PubMed Central

    2011-01-01

    Background Elevated levels of lipids are detrimental for beta-cell function and mass. One of the mechanisms of how fatty acids induce apoptosis is development of the unfolded protein response (UPR). It is still far from understood how fatty acids activate the UPR, however. Methods We examined how palmitate-induced activation of the UPR was affected by altering the metabolism of the fatty acid in insulin-secreting INS-1E and MIN6 cell lines and intact human islets. To increase oxidation, we used low glucose (5.5 mM) or AICAR; and to reduce oxidation, we used high glucose (25 mM) or etomoxir. UPR was measured after 3, 24 and 48 hours of palmitate treatment. Results Modulation of palmitate oxidation by either glucose or the pharmacological agents did not affect palmitate-induced UPR activation. Conclusion Our finding suggests that other factors than oxidation of palmitate play a role in the activation of UPR in fatty acid-treated beta-cells. PMID:21978671

  14. Gene Expression Profiles of Beta-Cell Enriched Tissue Obtained by Laser Capture Microdissection from Subjects with Type 2 Diabetes

    PubMed Central

    Marselli, Lorella; Thorne, Jeffrey; Dahiya, Sonika; Sgroi, Dennis C.; Sharma, Arun; Bonner-Weir, Susan; Marchetti, Piero; Weir, Gordon C.

    2010-01-01

    Background Changes in gene expression in pancreatic beta-cells from type 2 diabetes (T2D) should provide insights into their abnormal insulin secretion and turnover. Methodology/Principal Findings Frozen sections were obtained from cadaver pancreases of 10 control and 10 T2D human subjects. Beta-cell enriched samples were obtained by laser capture microdissection (LCM). RNA was extracted, amplified and subjected to microarray analysis. Further analysis was performed with DNA-Chip Analyzer (dChip) and Gene Set Enrichment Analysis (GSEA) software. There were changes in expression of genes linked to glucotoxicity. Evidence of oxidative stress was provided by upregulation of several metallothionein genes. There were few changes in the major genes associated with cell cycle, apoptosis or endoplasmic reticulum stress. There was differential expression of genes associated with pancreatic regeneration, most notably upregulation of members of the regenerating islet gene (REG) family and metalloproteinase 7 (MMP7). Some of the genes found in GWAS studies to be related to T2D were also found to be differentially expressed. IGF2BP2, TSPAN8, and HNF1B (TCF2) were upregulated while JAZF1 and SLC30A8 were downregulated. Conclusions/Significance This study made possible by LCM has identified many novel changes in gene expression that enhance understanding of the pathogenesis of T2D. PMID:20644627

  15. Oxidative Stress Type Influences the Properties of Antioxidants Containing Polyphenols in RINm5F Beta Cells

    PubMed Central

    Auberval, Nathalie; Dal, Stéphanie; Bietiger, William; Seyfritz, Elodie; Peluso, Jean; Muller, Christian; Zhao, Minjie; Marchioni, Eric; Pinget, Michel; Jeandidier, Nathalie; Maillard, Elisa; Schini-Kerth, Valérie; Sigrist, Séverine

    2015-01-01

    The in vitro methods currently used to screen bioactive compounds focus on the use of a single model of oxidative stress. However, this simplistic view may lead to conflicting results. The aim of this study was to evaluate the antioxidant properties of two natural extracts (a mix of red wine polyphenols (RWPs) and epigallocatechin gallate (EGCG)) with three models of oxidative stress induced with hydrogen peroxide (H2O2), a mixture of hypoxanthine and xanthine oxidase (HX/XO), or streptozotocin (STZ) in RINm5F beta cells. We employed multiple approaches to validate their potential as therapeutic treatment options, including cell viability, reactive oxygen species production, and antioxidant enzymes expression. All three oxidative stresses induced a decrease in cell viability and an increase in apoptosis, whereas the level of ROS production was variable depending on the type of stress. The highest level of ROS was found for the HX/XO-induced stress, an increase that was reflected by higher expression antioxidant enzymes. Further, both antioxidant compounds presented beneficial effects during oxidative stress, but EGCG appeared to be a more efficient antioxidant. These data indicate that the efficiency of natural antioxidants is dependent on both the nature of the compound and the type of oxidative stress generated. PMID:26508986

  16. Beta-cell deterioration during diabetes: what's in the gun?

    PubMed

    Robertson, R Paul

    2009-10-01

    Clinical management of patients with type 2 diabetes (T2D) successfully prevents extreme hyperglycemia but does not precisely control glucose levels throughout the day. The pathogenesis of T2D is akin to a double-barrel shotgun. The first trigger causes an explosion that sets genetic expression of the disease in motion; the second trigger discharges a host of environmental factors that worsen its clinical course. Candidate shells include glucolipotoxicity, cytokines, oxidative and endoplasmic reticulum stress and insulin resistance. This review considers how each candidate adversely impacts beta-cell function to create the downward spiral of glycemic control. Their roles in pathogenesis raise possibilities for new drug therapies designed to protect against adverse effects of residual hyperglycemia in patients treated with conventional drugs.

  17. Nanomaterial Solutions for the Protection of Insulin Producing Beta Cells

    NASA Astrophysics Data System (ADS)

    Atchison, Nicole Ann

    Islet transplantation is a promising treatment for type 1 diabetes. However, even with the many successes, islet transplantation has yet to reach its full potential. Limited islet sources, loss of cell viability during isolation and culture, and post-transplant graft loss are a few of the issues preventing extensive use of islet transplantation. The application of biomaterial systems to alleviate some of the stresses affecting islet viability has led to improvements in isolation and transplantation outcomes, but problems persist. In this work we approach two distinct issues affecting islet viability; ischemic conditions and immunological attack post-transplant. Ischemic conditions have been linked to a loss of islet graft function and occur during organ preservation, islet isolation and culture, and after islets are transplanted. We show that liposomal delivery of adenosine triphosphate (ATP) to beta cells can limit cell death and loss of function in ischemic conditions. We demonstrate that by functionalizing liposomes with the fibronectin-mimetic peptide PR_b, delivery of liposomes to porcine islets and rat beta cells is increased compared to nontargeted controls. Additionally, liposomes are shown to protect by providing both ATP and lipids to the ischemic cells. The delivery of ATP was investigated here but application of PR_b functionalized liposomes could be extended to other interesting cargos as well. The second area of investigation involves encapsulation of islets with silica nanoparticles to create a permselective barrier. Silica nanoparticles are an interesting material for encapsulation given their ability to be fine-tuned and further functionalized. We demonstrate that size-tunable, fluorescent silica nanoparticles can be assembled layer-by-layer on the surface of cells and that silica nanoparticle encapsulated islets are able to secrete insulin in response to a glucose challenge.

  18. Glucose activates prenyltransferases in pancreatic islet beta-cells.

    PubMed

    Goalstone, Marc; Kamath, Vasudeva; Kowluru, Anjaneyulu

    2010-01-01

    A growing body of evidence implicates small G-proteins [e.g., Cdc42 and Rac1] in glucose-stimulated insulin secretion [GSIS] in the islet beta-cell. These signaling proteins undergo post-translational modifications [e.g., prenylation] at their C-terminal cysteine residue and appear to be essential for the transport and fusion of insulin-containing secretory granules with the plasma membrane and the exocytotic secretion of insulin. However, potential regulation of the prenylating enzymes by physiological insulin secretogues [e.g., glucose] has not been investigated thus far. Herein, we report immunological localization, sub-cellular distribution and regulation of farnesyltransferases [FTases] and geranylgeranyltransferase [GGTase] by glucose in insulin-secreting INS 832/13 beta-cells and normal rat islets. Our findings suggest that an insulinotropic concentration of glucose [20mM] markedly stimulated the expression of the alpha-subunits of FTase/GGTase-1, but not the beta-subunits of FTase or GGTase-1 without significantly affecting the predominantly cytosolic distribution of these holoenzymes in INS 832/13 cells and rodent islets. Under these conditions, glucose significantly stimulated [2.5- to 4.0-fold over basal] the activities of both FTase and GGTase-1 in both cell types. Together, these findings provide the first evidence to suggest that GSIS involves activation of the endogenous islet prenyltransferases by glucose, culminating in the activation of their respective G-protein substrates, which is necessary for cytoskeletal rearrangement, vesicular transport, fusion and secretion of insulin.

  19. Glucose activates prenyltransferases in pancreatic islet {beta}-cells

    SciTech Connect

    Goalstone, Marc; Kamath, Vasudeva; Kowluru, Anjaneyulu

    2010-01-01

    A growing body of evidence implicates small G-proteins [e.g., Cdc42 and Rac1] in glucose-stimulated insulin secretion [GSIS] in the islet {beta}-cell. These signaling proteins undergo post-translational modifications [e.g., prenylation] at their C-terminal cysteine residue and appear to be essential for the transport and fusion of insulin-containing secretory granules with the plasma membrane and the exocytotic secretion of insulin. However, potential regulation of the prenylating enzymes by physiological insulin secretogues [e.g., glucose] has not been investigated thus far. Herein, we report immunological localization, sub-cellular distribution and regulation of farnesyltransferases [FTases] and geranylgeranyltransferase [GGTase] by glucose in insulin-secreting INS 832/13 {beta}-cells and normal rat islets. Our findings suggest that an insulinotropic concentration of glucose [20 mM] markedly stimulated the expression of the {alpha}-subunits of FTase/GGTase-1, but not the {beta}-subunits of FTase or GGTase-1 without significantly affecting the predominantly cytosolic distribution of these holoenzymes in INS 832/13 cells and rodent islets. Under these conditions, glucose significantly stimulated [2.5- to 4.0-fold over basal] the activities of both FTase and GGTase-1 in both cell types. Together, these findings provide the first evidence to suggest that GSIS involves activation of the endogenous islet prenyltransferases by glucose, culminating in the activation of their respective G-protein substrates, which is necessary for cytoskeletal rearrangement, vesicular transport, fusion and secretion of insulin.

  20. Specialized Hub Beta Cells Trade Maximal Insulin Production for Perfect Timing.

    PubMed

    Kolic, Jelena; Johnson, James D

    2016-09-13

    The pulsatility of insulin release is disturbed early in type 2 diabetes, but it is not clear whether specialized pacemaker cells drive islet oscillations. In this issue of Cell Metabolism, Johnston et al. (2016) show that specialized hubs, identified as 1%-10% of beta cells with more active mitochondria and less insulin, synchronize beta cell oscillations.

  1. Quantification of the relationship between glycemia and beta-cell mass adaptation in vivo.

    PubMed

    Atkinson, Laura L; Topp, Brian G; Au, Jenny; Vinerian, Horatiu V; Dhatt, Narinder; Finegood, Diane T

    2009-08-01

    Beta-cell mass dynamics play an important role in the adaptation to obesity, as well as in the pathogenesis of type 2 diabetes. Here we used a 24-hour modified hyperglycemic clamp protocol to investigate the effect of increasing glucose concentrations (15, 20, 25, or 35 mmol/L) on beta-cell mass and rates of beta-cell replication, death, and neogenesis in 6-week-old Sprague Dawley rats (n = 40). During the first 4 h of glucose infusion, plasma insulin levels rose to an approximate steady state in each group, but by the end of 24 h, there was no difference in insulin levels between any of the groups. There was also no difference in beta-cell mass between groups. Mean beta-cell replication rates displayed a linear relationship to mean plasma glucose levels in all hyperglycemic animals (r(2) = 0.98, p < 0.05). Relative to the uninfused basal control animals, replication rates were significantly reduced in the 15 mmol/L glucose group. The percentage of TUNEL-positive beta-cells was not different between groups. There was also no significant difference in markers of neogenesis. Thus, these data demonstrate that hyperglycemia for 24 h had no effect on beta-cell mass, death, or neogenesis in 6-week-old Sprague Dawley rats. We demonstrate a linear relationship, however, between hyperglycemia and beta-cell replication rates in vivo.

  2. Specialized Hub Beta Cells Trade Maximal Insulin Production for Perfect Timing.

    PubMed

    Kolic, Jelena; Johnson, James D

    2016-09-13

    The pulsatility of insulin release is disturbed early in type 2 diabetes, but it is not clear whether specialized pacemaker cells drive islet oscillations. In this issue of Cell Metabolism, Johnston et al. (2016) show that specialized hubs, identified as 1%-10% of beta cells with more active mitochondria and less insulin, synchronize beta cell oscillations. PMID:27626196

  3. Marked over expression of uncoupling protein-2 in beta cells exerts minor effects on mitochondrial metabolism

    SciTech Connect

    Hals, Ingrid K.; Ogata, Hirotaka; Pettersen, Elin; Ma, Zuheng; Bjoerklund, Anneli; Skorpen, Frank; Egeberg, Kjartan Wollo; Grill, Valdemar

    2012-06-29

    Highlights: Black-Right-Pointing-Pointer The impact of UCP-2 over expression on mitochondrial function is controversial. Black-Right-Pointing-Pointer We tested mitochondrial functions at defined levels of overexpression. Black-Right-Pointing-Pointer We find minor increases of fatty acid oxidation and uncoupling. Black-Right-Pointing-Pointer Effects were seen only at high level (fourfold) of over expression. Black-Right-Pointing-Pointer Hence it is doubtful whether these effects are of importance in diabetes. -- Abstract: Evidence is conflicting as to the impact of elevated levels of uncoupling protein-2 (UCP-2) on insulin-producing beta cells. Here we investigated effects of a fourfold induction of UCP-2 protein primarily on mitochondrial parameters and tested for replication of positive findings at a lower level of induction. We transfected INS-1 cells to obtain a tet-on inducible cell line. A 48 h exposure to 1 {mu}g/ml of doxycycline (dox) induced UCP-2 fourfold (424 {+-} 113%, mean {+-} SEM) and 0.1 {mu}g/ml twofold (178 {+-} 29%, n = 3). Fourfold induced cells displayed normal viability (MTT, apoptosis), normal cellular insulin contents and, glucose-induced insulin secretion (+27 {+-} 11%) as well as D-[U-{sup 14}C]-glucose oxidation (+5 {+-} 9% at 11 mM glucose). Oxidation of [1-{sup 14}C]-oleate was increased from 4088 to 5797 fmol/{mu}g prot/2 h at 3.3 mM glucose, p < 0.03. Oxidation of L-[{sup 14}C(U)]-glutamine was unaffected. Induction of UCP-2 did not significantly affect measures of mitochondrial membrane potential (Rhodamine 123) or mitochondrial mass (Mitotracker Green) and did not affect ATP levels. Oligomycin-inhibited oxygen consumption (a measure of mitochondrial uncoupling) was marginally increased, the effect being significant in comparison with dox-only treated cells, p < 0.05. Oxygen radicals, assessed by dichlorofluorescin diacetate, were decreased by 30%, p < 0.025. Testing for the lower level of UCP-2 induction did not reproduce any of the

  4. Beta-cell function and mass in type 2 diabetes.

    PubMed

    Larsen, Marianne O

    2009-08-01

    The aim of the work described here was to improve our understanding of beta-cell function (BCF) and beta-cell mass (BCM) and their relationship in vivo using the minipig as a model for some of the aspects of human type 2 diabetes (T2DM). More specifically, the aim was to evaluate the following questions: How is BCF, especially high frequency pulsatile insulin secretion, affected by a primary reduction in BCM or by primary obesity or a combination of the two in the minipig? Can evaluation of BCF in vivo be used as a surrogate measure to predict BCM in minipigs over a range of BCM and body weight? We first developed a minipig model of reduced BCM and mild diabetes using administration of a combination of streptozotocin (STZ) and nicotinamide (NIA) as a tool to study effects of a primary reduction of BCM on BCF. The model was characterized using a mixed-meal oral glucose tolerance test and intravenous stimulation with glucose and arginine as well as by histology of the pancreas after euthanasia. It was shown that stable, moderate diabetes can be induced and that the model is characterized by fasting and postprandial hyperglycemia, reduced insulin secretion and reduced BCM. Several defects in insulin secretion are well documented in human T2DM; however, the role in the pathogenesis and the possible clinical relevance of high frequency (rapid) pulsatile insulin secretion is still debated. We therefore investigated this phenomenon in normal minipigs and found easily detectable pulses in peripheral vein plasma samples that were shown to be correlated with pulses found in portal vein plasma. Furthermore, the rapid kinetics of insulin in the minipig strongly facilitates pulse detection. These characteristics make the minipig particularly suitable for studying the occurrence of disturbed pulsatility in relation to T2DM. Disturbances of rapid pulsatile insulin secretion have been reported to be a very early event in the development of T2DM and include disorderliness of pulses

  5. Beta-cell function and mass in type 2 diabetes.

    PubMed

    Larsen, Marianne O

    2009-08-01

    The aim of the work described here was to improve our understanding of beta-cell function (BCF) and beta-cell mass (BCM) and their relationship in vivo using the minipig as a model for some of the aspects of human type 2 diabetes (T2DM). More specifically, the aim was to evaluate the following questions: How is BCF, especially high frequency pulsatile insulin secretion, affected by a primary reduction in BCM or by primary obesity or a combination of the two in the minipig? Can evaluation of BCF in vivo be used as a surrogate measure to predict BCM in minipigs over a range of BCM and body weight? We first developed a minipig model of reduced BCM and mild diabetes using administration of a combination of streptozotocin (STZ) and nicotinamide (NIA) as a tool to study effects of a primary reduction of BCM on BCF. The model was characterized using a mixed-meal oral glucose tolerance test and intravenous stimulation with glucose and arginine as well as by histology of the pancreas after euthanasia. It was shown that stable, moderate diabetes can be induced and that the model is characterized by fasting and postprandial hyperglycemia, reduced insulin secretion and reduced BCM. Several defects in insulin secretion are well documented in human T2DM; however, the role in the pathogenesis and the possible clinical relevance of high frequency (rapid) pulsatile insulin secretion is still debated. We therefore investigated this phenomenon in normal minipigs and found easily detectable pulses in peripheral vein plasma samples that were shown to be correlated with pulses found in portal vein plasma. Furthermore, the rapid kinetics of insulin in the minipig strongly facilitates pulse detection. These characteristics make the minipig particularly suitable for studying the occurrence of disturbed pulsatility in relation to T2DM. Disturbances of rapid pulsatile insulin secretion have been reported to be a very early event in the development of T2DM and include disorderliness of pulses

  6. Beta Cell Mass Restoration in Alloxan-Diabetic Mice Treated with EGF and Gastrin.

    PubMed

    Song, Imane; Patel, Oelfah; Himpe, Eddy; Muller, Christo J F; Bouwens, Luc

    2015-01-01

    One week of treatment with EGF and gastrin (EGF/G) was shown to restore normoglycemia and to induce islet regeneration in mice treated with the diabetogenic agent alloxan. The mechanisms underlying this regeneration are not fully understood. We performed genetic lineage tracing experiments to evaluate the contribution of beta cell neogenesis in this model. One day after alloxan administration, mice received EGF/G treatment for one week. The treatment could not prevent the initial alloxan-induced beta cell mass destruction, however it did reverse glycemia to control levels within one day, suggesting improved peripheral glucose uptake. In vitro experiments with C2C12 cell line showed that EGF could stimulate glucose uptake with an efficacy comparable to that of insulin. Subsequently, EGF/G treatment stimulated a 3-fold increase in beta cell mass, which was partially driven by neogenesis and beta cell proliferation as assessed by beta cell lineage tracing and BrdU-labeling experiments, respectively. Acinar cell lineage tracing failed to show an important contribution of acinar cells to the newly formed beta cells. No appearance of transitional cells co-expressing insulin and glucagon, a hallmark for alpha-to-beta cell conversion, was found, suggesting that alpha cells did not significantly contribute to the regeneration. An important fraction of the beta cells significantly lost insulin positivity after alloxan administration, which was restored to normal after one week of EGF/G treatment. Alloxan-only mice showed more pronounced beta cell neogenesis and proliferation, even though beta cell mass remained significantly depleted, suggesting ongoing beta cell death in that group. After one week, macrophage infiltration was significantly reduced in EGF/G-treated group compared to the alloxan-only group. Our results suggest that EGF/G-induced beta cell regeneration in alloxan-diabetic mice is driven by beta cell neogenesis, proliferation and recovery of insulin. The

  7. Bmal1 and Beta cell clock are required for adaptation to circadian disruption, and their loss of function leads to oxidative stress-induced Beta cell failure in mice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Circadian disruption has deleterious effects on metabolism. Global deletion of Bmal1, a core clock gene, results in Beta cell dysfunction and diabetes. However, it is unknown if this is due to loss of cell-autonomous function of Bmal1 in Beta cells. To address this, we generated mice with Beta cell ...

  8. Stem cells to pancreatic beta-cells: new sources for diabetes cell therapy.

    PubMed

    Guo, Tingxia; Hebrok, Matthias

    2009-05-01

    The number of patients worldwide suffering from the chronic disease diabetes mellitus is growing at an alarming rate. Insulin-secreting beta-cells in the islet of Langerhans are damaged to different extents in diabetic patients, either through an autoimmune reaction present in type 1 diabetic patients or through inherent changes within beta-cells that affect their function in patients suffering from type 2 diabetes. Cell replacement strategies via islet transplantation offer potential therapeutic options for diabetic patients. However, the discrepancy between the limited number of donor islets and the high number of patients who could benefit from such a treatment reflects the dire need for renewable sources of high-quality beta-cells. Human embryonic stem cells (hESCs) are capable of self-renewal and can differentiate into components of all three germ layers, including all pancreatic lineages. The ability to differentiate hESCs into beta-cells highlights a promising strategy to meet the shortage of beta-cells. Here, we review the different approaches that have been used to direct differentiation of hESCs into pancreatic and beta-cells. We will focus on recent progress in the understanding of signaling pathways and transcription factors during embryonic pancreas development and how this knowledge has helped to improve the methodology for high-efficiency beta-cell differentiation in vitro.

  9. Effects of ORP150 on appearance and function of pancreatic beta cells following acute necrotizing pancreatitis.

    PubMed

    Deng, Wen-Hong; Chen, Chen; Wang, Wei-Xing; Yu, Jia; Li, Jin-You; Liu, Lei

    2011-06-15

    Pancreatic beta cells produce and release insulin, which decreases the blood glucose level. Endoplasmic reticulum stress caused pancreatic beta cell dysfunction and death in acute necrotizing pancreatitis (ANP). The 150kD oxygen-regulated protein (ORP150) took part in the process of endoplasmic reticulum stress. This study investigated the effect of ORP150 on appearance and function of pancreatic beta cells in ANP. Acute necrotizing pancreatitis relied on retrograde infusion of 5% sodium taurocholate into the bile-pancreatic duct. The severity of ANP was estimated by serum amylase, secretory phospholipase A(2,) and pancreatic histopathology. The changes in appearance and function of pancreatic beta cells were detected by light and electron microscopy and the levels of serum glucose, insulin, and C-peptide. ORP150 expression was studied using western blot and immunohistochemisty assay. The expression of ORP150 mainly appeared on pancreatic beta cells and decreased gradually during the pathogenesis of ANP. The results of light and electron microscopy indicated pancreatic beta cell dysfunction and death, concomitant with elevation of serum glucose, insulin, and C-peptide in ANP. These results imply a probable role of ORP150 in the changes in appearance and function of pancreatic beta cells following acute necrotizing pancreatitis, through the pathway of endoplasmic reticulum stress.

  10. Secondary prevention of type 1 diabetes mellitus: stopping immune destruction and promoting beta-cell regeneration.

    PubMed

    Couri, C E B; Foss, M C; Voltarelli, J C

    2006-10-01

    Type 1 diabetes mellitus results from a cell-mediated autoimmune attack against pancreatic beta-cells. Traditional treatments involve numerous daily insulin dosages/injections and rigorous glucose control. Many efforts toward the identification of beta-cell precursors have been made not only with the aim of understanding the physiology of islet regeneration, but also as an alternative way to produce beta-cells to be used in protocols of islet transplantation. In this review, we summarize the most recent studies related to precursor cells implicated in the regeneration process. These include embryonic stem cells, pancreas-derived multipotent precursors, pancreatic ductal cells, hematopoietic stem cells, mesenchymal stem cells, hepatic oval cells, and mature beta-cells. There is controversial evidence of the potential of these cell sources to regenerate beta-cell mass in diabetic patients. However, clinical trials using embryonic stem cells, umbilical cord blood or adult bone marrow stem cells are under way. The results of various immunosuppressive regimens aiming at blocking autoimmunity against pancreatic beta-cells and promoting beta-cell preservation are also analyzed. Most of these regimens provide transient and partial effect on insulin requirements, but new regimens are beginning to be tested. Our own clinical trial combines a high dose immunosuppression with mobilized peripheral blood hematopoietic stem cell transplantation in early-onset type 1 diabetes mellitus.

  11. Cocoa phenolic extract protects pancreatic beta cells against oxidative stress.

    PubMed

    Martín, María Angeles; Ramos, Sonia; Cordero-Herrero, Isabel; Bravo, Laura; Goya, Luis

    2013-08-01

    Diabetes mellitus is associated with reductions in glutathione, supporting the critical role of oxidative stress in its pathogenesis. Antioxidant food components such as flavonoids have a protective role against oxidative stress-induced degenerative and age-related diseases. Flavonoids constitute an important part of the human diet; they can be found in most plant foods, including green tea, grapes or cocoa and possess multiple biological activities. This study investigates the chemo-protective effect of a cocoa phenolic extract (CPE) containing mainly flavonoids against oxidative stress induced by tert-butylhydroperoxide (t-BOOH) on Ins-1E pancreatic beta cells. Cell viability and oxidative status were evaluated. Ins-1E cells treatment with 5-20 μg/mL CPE for 20 h evoked no cell damage and did not alter ROS production. Addition of 50 μM t-BOOH for 2 h increased ROS and carbonyl groups content and decreased reduced glutathione level. Pre-treatment of cells with CPE significantly prevented the t-BOOH-induced ROS and carbonyl groups and returned antioxidant defences to adequate levels. Thus, Ins-1E cells treated with CPE showed a remarkable recovery of cell viability damaged by t-BOOH, indicating that integrity of surviving machineries in the CPE-treated cells was notably protected against the oxidative insult. PMID:23912326

  12. Cocoa Phenolic Extract Protects Pancreatic Beta Cells against Oxidative Stress

    PubMed Central

    Martín, María Ángeles; Ramos, Sonia; Cordero-Herrero, Isabel; Bravo, Laura; Goya, Luis

    2013-01-01

    Diabetes mellitus is associated with reductions in glutathione, supporting the critical role of oxidative stress in its pathogenesis. Antioxidant food components such as flavonoids have a protective role against oxidative stress-induced degenerative and age-related diseases. Flavonoids constitute an important part of the human diet; they can be found in most plant foods, including green tea, grapes or cocoa and possess multiple biological activities. This study investigates the chemo-protective effect of a cocoa phenolic extract (CPE) containing mainly flavonoids against oxidative stress induced by tert-butylhydroperoxide (t-BOOH) on Ins-1E pancreatic beta cells. Cell viability and oxidative status were evaluated. Ins-1E cells treatment with 5–20 μg/mL CPE for 20 h evoked no cell damage and did not alter ROS production. Addition of 50 μM t-BOOH for 2 h increased ROS and carbonyl groups content and decreased reduced glutathione level. Pre-treatment of cells with CPE significantly prevented the t-BOOH-induced ROS and carbonyl groups and returned antioxidant defences to adequate levels. Thus, Ins-1E cells treated with CPE showed a remarkable recovery of cell viability damaged by t-BOOH, indicating that integrity of surviving machineries in the CPE-treated cells was notably protected against the oxidative insult. PMID:23912326

  13. Reduced expression of the liver/beta-cell glucose transporter isoform in glucose-insensitive pancreatic beta cells of diabetic rats.

    PubMed Central

    Thorens, B; Weir, G C; Leahy, J L; Lodish, H F; Bonner-Weir, S

    1990-01-01

    Rats injected with a single dose of streptozocin at 2 days of age develop non-insulin-dependent diabetes 6 weeks later. The pancreatic beta islet cells of these diabetic rats display a loss of glucose-induced insulin secretion while maintaining sensitivity to other secretagogues such as arginine. We analyzed the level of expression of the liver/beta-cell glucose transporter isoform in diabetic islets by immunofluorescence staining of pancreas sections and by Western blotting of islet lysates. Islets from diabetic animals have a reduced expression of this beta-cell-specific glucose transporter isoform and the extent of reduction is correlated with the severity of hyperglycemia. In contrast, expression of this transporter isoform in liver is minimally modified by the diabetes. Thus a decreased expression of the liver/beta-cell glucose transporter isoform in beta cells is associated with the impaired glucose sensing characteristic of diabetic islets; our data suggest that this glucose transporter may be part of the beta-cell glucose sensor. Images PMID:2204056

  14. Reduced Expression of the Liver/Beta-Cell Glucose Transporter Isoform in Glucose-Insensitive Pancreatic Beta Cells of Diabetic Rats

    NASA Astrophysics Data System (ADS)

    Thorens, Bernard; Weir, Gordon C.; Leahy, John L.; Lodish, Harvey F.; Bonner-Weir, Susan

    1990-09-01

    Rats injected with a single dose of streptozocin at 2 days of age develop non-insulin-dependent diabetes 6 weeks later. The pancreatic beta islet cells of these diabetic rats display a loss of glucose-induced insulin secretion while maintaining sensitivity to other secretagogues such as arginine. We analyzed the level of expression of the liver/beta-cell glucose transporter isoform in diabetic islets by immunofluorescence staining of pancreas sections and by Western blotting of islet lysates. Islets from diabetic animals have a reduced expression of this beta-cell-specific glucose transporter isoform and the extent of reduction is correlated with the severity of hyperglycemia. In contrast, expression of this transporter isoform in liver is minimally modified by the diabetes. Thus a decreased expression of the liver/beta-cell glucose transporter isoform in beta cells is associated with the impaired glucose sensing characteristic of diabetic islets; our data suggest that this glucose transporter may be part of the beta-cell glucose sensor.

  15. Beta-cell gene expression and functional characterisation of the human insulinoma cell line CM.

    PubMed

    Baroni, M G; Cavallo, M G; Mark, M; Monetini, L; Stoehrer, B; Pozzilli, P

    1999-04-01

    Animal insulinoma cell lines are widely used to study physiological and pathophysiological mechanisms involved in glucose metabolism and to establish in vitro models for studies on beta-cells. In contrast, human insulinoma cell lines are rarely used because of difficulties in obtaining and culturing them for long periods. The aim of our study was to investigate, under different experimental conditions, the capacity of the human insulinoma cell line CM to retain beta-cell function, particularly the expression of constitutive beta-cell genes (insulin, the glucose transporters GLUT1 and GLUT2, glucokinase), intracellular and secreted insulin, beta-cell granules, and cAMP content. Results showed that CM cells from an early-passage express specific beta-cell genes in response to glucose stimulation, in particular the insulin and GLUT genes. Such capacity is lost at later passages when cells are cultured at standard glucose concentrations. However, if cultured at lower glucose concentration (0.8 mM) for a longer time, CM cells re-acquire the capacity to respond to glucose stimulation, as shown by the increased expression of beta-cell genes (insulin, GLUT2, glucokinase). Nonetheless, insulin secretion could not be restored under such experimental conditions despite the presence of intracellular insulin, although cAMP response to a potent activator of adenylate cyclase, forskolin, was present indicating a viable system. In conclusion, these data show that the human insulinoma cell line CM, at both early-passage and late-passage, posseses a functional glucose-signalling pathway and insulin mRNA expression similar to normal beta-cells, representing, therefore, a good model for studies concerning the signalling and expression of beta-cells. Furthermore, we have previously shown that it is also a good model for immunological studies. In this respect it is important to note that the CM cell line is one of the very few existing human beta-cell lines in long-term culture.

  16. Beta-Cell ARNT Is Required for Normal Glucose Tolerance in Murine Pregnancy

    PubMed Central

    Lau, Sue Mei; Cha, Kuan Minn; Karunatillake, Ayesha; Stokes, Rebecca A.; Cheng, Kim; McLean, Mark; Cheung, N. W.; Gonzalez, Frank J.; Gunton, Jenny E.

    2013-01-01

    Aims Insulin secretion increases in normal pregnancy to meet increasing demands. Inability to increase beta-cell function results in gestational diabetes mellitus (GDM). We have previously shown that the expression of the transcription factor ARNT (Aryl-hydrocarbon Receptor Nuclear Translocator) is reduced in the islets of humans with type 2 diabetes. Mice with a beta-cell specific deletion of ARNT (β-ARNT mice) have impaired glucose tolerance secondary to defective insulin secretion. We hypothesised that ARNT is required to increase beta-cell function during pregnancy, and that β-ARNT mice would be unable to compensate for the beta-cell stress of pregnancy. The aims of this study were to investigate the mechanisms of ARNT regulation of beta-cell function and glucose tolerance in pregnancy. Methods β-ARNT females were mated with floxed control (FC) males and FC females with β-ARNT males. Results During pregnancy, β-ARNT mice had a marked deterioration in glucose tolerance secondary to defective insulin secretion. There was impaired beta-cell proliferation in late pregnancy, associated with decreased protein and mRNA levels of the islet cell-cycle regulator cyclinD2. There was also reduced expression of Irs2 and G6PI. In contrast, in control mice, pregnancy was associated with a 2.1-fold increase in ARNT protein and a 1.6-fold increase in cyclinD2 protein, and with increased beta-cell proliferation. Conclusions Islet ARNT increases in normal murine pregnancy and beta-cell ARNT is required for cyclinD2 induction and increased beta-cell proliferation in pregnancy. PMID:24204824

  17. Transplantation of mesenchymal stem cells recruits trophic macrophages to induce pancreatic beta cell regeneration in diabetic mice.

    PubMed

    Cao, Xiaocang; Han, Zhi-Bo; Zhao, Hui; Liu, Qiang

    2014-08-01

    Alleviation of hyperglycemia in chemical-induced diabetic mice has been reported after bone marrow transplantation. Nevertheless, the underlying mechanism remains elusive. In the present study, we transplanted genetically labeled primary mouse mesenchymal stem cells into the pancreas of the streptozotocin-treated hyperglycemic isogeneic mice, resulting in a decrease in blood glucose due to a recovery in beta cell mass. Further analysis revealed that the increase in beta cell mass was predominantly attributable to beta cell replication. The grafted mesenchymal stem cells did not transdifferentiate into beta cells themselves but recruited and polarized macrophages in a Stromal cell-derived factor 1-dependent manner, which in turn promoted beta cell replication. Our finding thus suggests that transplantation of autogenic mesenchymal stem cells may increase functional beta cell mass by boosting beta cell replication in diabetes. PMID:24915493

  18. Maturation of adult beta-cells revealed using a Pdx1/insulin dual-reporter lentivirus.

    PubMed

    Szabat, Marta; Luciani, Dan S; Piret, James M; Johnson, James D

    2009-04-01

    The enigmatic process of beta-cell maturation has significant implications for diabetes pathogenesis, and potential diabetes therapies. This study examined the dynamics and heterogeneity of insulin and pancreatic duodenal homeobox (Pdx)-1 gene expression in adult beta-cells. Insulin and Pdx1 expression were monitored in human and mouse islet cells and MIN6 cells using a Pdx1-monomeric red fluorescent protein/insulin-enhanced green fluorescent protein dual-reporter lentivirus. The majority of fluorescent cells were highly positive for both Pdx1 and insulin. Cells expressing Pdx1 but little or no insulin (Pdx1(+)/Ins(low)) comprised 15-25% of the total population. Time-lapse imaging demonstrated that Pdx1(+)/Ins(low) primary beta-cells and MIN6 cells could convert to Pdx1(+)/Ins(+) cells without cell division. Genes involved in the mature beta-cell phenotype (Glut2, MafA) were expressed at higher levels in Pdx1(+)/Ins(+) cells relative to Pdx1(+)/Ins(low) cells. Conversely, genes implicated in early beta-cell development (MafB, Nkx2.2) were enriched in Pdx1(+)/Ins(low) cells. Sorted Pdx1(+)/Ins(low) MIN6 cells had a higher replication rate and secreted less insulin relative to double-positive cells. Long-term phenotype tracking of Pdx1(+)/Ins(low) cells showed two groups, one that matured into Pdx1(+)/Ins(+) cells and one that remained immature. These results demonstrate that adult beta-cells pass through distinct maturation states, which is consistent with previously observed heterogeneity in insulin and Pdx1 expression in adult beta-cells. At a given time, a proportion of adult beta-cells share similar characteristics to functionally immature embryonic beta-cell progenitors. The maturation of adult beta-cells recapitulates development in that Pdx1 expression precedes the robust expression of insulin and other mature beta-cell genes. These results have implications for harnessing the maturation process for therapeutic purposes. PMID:19095744

  19. Bmal1 is required for beta cell compensatory expansion, survival and metabolic adaptation to diet-induced obesity in mice

    PubMed Central

    Rakshit, Kuntol; Hsu, Tu Wen

    2016-01-01

    Aims/hypothesis Obesity and consequent insulin resistance are known risk factors for type 2 diabetes. A compensatory increase in beta cell function and mass in response to insulin resistance permits maintenance of normal glucose homeostasis, whereas failure to do so results in beta cell failure and type 2 diabetes. Recent evidence suggests that the circadian system is essential for proper metabolic control and regulation of beta cell function. We set out to address the hypothesis that the beta cell circadian clock is essential for the appropriate functional and morphological beta cell response to insulin resistance. Methods We employed conditional deletion of the Bmal1 (also known as Arntl) gene (encoding a key circadian clock transcription factor) in beta cells using the tamoxifen-inducible CreERT recombination system. Upon adulthood, Bmal1 deletion in beta cells was achieved and mice were exposed to either chow or high fat diet (HFD). Changes in diurnal glycaemia, glucose tolerance and insulin secretion were longitudinally monitored in vivo and islet morphology and turnover assessed by immunofluorescence. Isolated islet experiments in vitro were performed to delineate changes in beta cell function and transcriptional regulation of cell proliferation. Results Adult Bmal1 deletion in beta cells resulted in failed metabolic adaptation to HFD characterised by fasting and diurnal hyperglycaemia, glucose intolerance and loss of glucose-stimulated insulin secretion. Importantly, HFD-induced beta cell expansion was absent following beta cell Bmal1 deletion indicating impaired beta cell proliferative and regenerative potential, which was confirmed by assessment of transcriptional profiles in isolated islets. Conclusion/interpretation Results of the study suggest that the beta cell circadian clock is a novel regulator of compensatory beta cell expansion and function in response to increased insulin demand associated with diet-induced obesity. PMID:26762333

  20. Regulation of Pancreatic Beta Cell Stimulus-Secretion Coupling by microRNAs

    PubMed Central

    Esguerra, Jonathan L. S.; Mollet, Inês G.; Salunkhe, Vishal A.; Wendt, Anna; Eliasson, Lena

    2014-01-01

    Increased blood glucose after a meal is countered by the subsequent increased release of the hypoglycemic hormone insulin from the pancreatic beta cells. The cascade of molecular events encompassing the initial sensing and transport of glucose into the beta cell, culminating with the exocytosis of the insulin large dense core granules (LDCVs) is termed “stimulus-secretion coupling.” Impairment in any of the relevant processes leads to insufficient insulin release, which contributes to the development of type 2 diabetes (T2D). The fate of the beta cell, when exposed to environmental triggers of the disease, is determined by the possibility to adapt to the new situation by regulation of gene expression. As established factors of post-transcriptional regulation, microRNAs (miRNAs) are well-recognized mediators of beta cell plasticity and adaptation. Here, we put focus on the importance of comprehending the transcriptional regulation of miRNAs, and how miRNAs are implicated in stimulus-secretion coupling, specifically those influencing the late stages of insulin secretion. We suggest that efficient beta cell adaptation requires an optimal balance between transcriptional regulation of miRNAs themselves, and miRNA-dependent gene regulation. The increased knowledge of the beta cell transcriptional network inclusive of non-coding RNAs such as miRNAs is essential in identifying novel targets for the treatment of T2D. PMID:25383562

  1. Acute overexpression of lactate dehydrogenase-A perturbs beta-cell mitochondrial metabolism and insulin secretion.

    PubMed

    Ainscow, E K; Zhao, C; Rutter, G A

    2000-07-01

    Islet beta-cells express low levels of lactate dehydrogenase and have high glycerol phosphate dehydrogenase activity. To determine whether this configuration favors oxidative glucose metabolism via mitochondria in the beta-cell and is important for beta-cell metabolic signal transduction, we have determined the effects on glucose metabolism and insulin secretion of acute overexpression of the skeletal muscle isoform of lactate dehydrogenase (LDH)-A. Monitored in single MIN6 beta-cells, LDH hyperexpression (achieved by intranuclear cDNA microinjection or adenoviral infection) diminished the response to glucose of both phases of increases in mitochondrial NAD(P)H, as well as increases in mitochondrial membrane potential, cytosolic free ATP, and cystolic free Ca2+. These effects were observed at all glucose concentrations, but were most pronounced at submaximal glucose levels. Correspondingly, adenoviral vector-mediated LDH-A overexpression reduced insulin secretion stimulated by 11 mmol/l glucose and the subsequent response to stimulation with 30 mmol/l glucose, but it was without significant effect when the concentration of glucose was raised acutely from 3 to 30 mmol/l. Thus, overexpression of LDH activity interferes with normal glucose metabolism and insulin secretion in the islet beta-cell type, and it may therefore be directly responsible for insulin secretory defects in some forms of type 2 diabetes. The results also reinforce the view that glucose-derived pyruvate metabolism in the mitochondrion is critical for glucose-stimulated insulin secretion in the beta-cell.

  2. The electrophysiology of the beta-cell based on single transmembrane protein characteristics.

    PubMed

    Meyer-Hermann, Michael E

    2007-10-15

    The electrophysiology of beta-cells is at the origin of insulin secretion. beta-Cells exhibit a complex behavior upon stimulation with glucose including repeated bursts and continuous spiking. Mathematical modeling is most suitable to improve knowledge about the function of various transmembrane currents provided the model is based on reliable data. This is the first attempt to build a mathematical model for the beta-cell electrophysiology in a bottom-up approach that relies on single protein conductance data. The results of previous whole-cell-based models are reconsidered. The full simulation including all prominent transmembrane proteins in beta-cells is used to provide a functional interpretation of their role in beta-cell bursting and an updated vantage point of beta-cell electrophysiology. As a result of a number of in silico knock-out and block experiments the novel model makes some unexpected predictions: single-channel conductance data imply that large-conductance calcium-gated potassium currents acquire the potential of driving oscillations at supralarge glucose levels. A more complex burst interruption model is presented. It also turns out that, depending on the species, sodium currents may be more relevant than considered so far. Experiments are proposed to verify these predictions. PMID:17573431

  3. Effects of amino acids on membrane potential and 86Rb+ fluxes in pancreatic beta-cells

    SciTech Connect

    Henquin, J.C.; Meissner, H.P.

    1981-03-01

    The membrane potential of beta-cells was studied with microelectrodes in mouse islets and their potassium permeability was evaluated by measuring 86Rb+ fluxes in rat islets. In the absence of glucose, L-leucine, its metabolite ketoisocaproate, and its nonmetabolized analogue 2-aminonorbornane-2-carboxylic acid (BCH) depolarized beta-cells and triggered bursts of electrical activity like glucose. The effect of leucine was weak, but was potentiated by a low concentration of glucose or by theophylline; the effect of ketoisocaproate was stronger and faster than that of an equimolar concentration of glucose. Arginine alone produced only a fast depolarization of beta-cells, insufficient to trigger electrical activity. Leucine and arginine potentiated the activity induced by glucose. In a glucose-free medium, alanine only slightly depolarized beta cells, whereas isoleucine and phenylalanie had no effect. Leucine, ketoisocaproate, and BCH reversibly decreased 86Rb+ efflux from islets perifused in the absence of glucose and increased 86Rb+ uptake. By contrast, both in the absence or presence of glucose, arginine increased 86Rb+ efflux and decreased 86Rb+ uptake. It is proposed that leucine, ketoisocaproate, and BCH, as glucose, deplolarize beta-cells by decreasing their potassium permeability, whereas arginine acts differently. The appearance of bursts of electrical activity with secretagogues unrelated to glucose suggests that they reflect an intrinsic property of the beta-cell membrane.

  4. The effect of smoking cessation pharmacotherapies on pancreatic beta cell function

    SciTech Connect

    Woynillowicz, Amanda K.; Raha, Sandeep; Nicholson, Catherine J.; Holloway, Alison C.

    2012-11-15

    The goal of our study was to evaluate whether drugs currently used for smoking cessation (i.e., nicotine replacement therapy, varenicline [a partial agonist at nicotinic acetylcholine receptors (nAChR)] and bupropion [which acts in part as a nAChR antagonist]) can affect beta cell function and determine the mechanism(s) of this effect. INS-1E cells, a rat beta cell line, were treated with nicotine, varenicline and bupropion to determine their effects on beta cell function, mitochondrial electron transport chain enzyme activity and cellular/oxidative stress. Treatment of INS-1E cells with equimolar concentrations (1 μM) of three test compounds resulted in an ablation of normal glucose-stimulated insulin secretion by the cells. This disruption of normal beta cell function was associated with mitochondrial dysfunction since all three compounds tested significantly decreased the activity of mitochondrial electron transport chain enzyme activity. These results raise the possibility that the currently available smoking cessation pharmacotherapies may also have adverse effects on beta cell function and thus glycemic control in vivo. Therefore whether or not the use of nicotine replacement therapy, varenicline and bupropion can cause endocrine changes which are consistent with impaired pancreatic function warrants further investigation. -- Highlights: ► Smoking cessation drugs have the potential to disrupt beta cell function in vitro. ► The effects of nicotine, varenicline and bupropion are similar. ► The impaired beta cell function is mediated by mitochondrial dysfunction. ► If similar effects are seen in vivo, these drugs may increase the risk of diabetes.

  5. BMP4-BMPR1A signaling in beta cells is required for and augments glucose-stimulated insulin secretion.

    PubMed

    Goulley, Joan; Dahl, Ulf; Baeza, Nathalie; Mishina, Yuji; Edlund, Helena

    2007-03-01

    Impaired glucose-stimulated insulin secretion (GSIS) and perturbed proinsulin processing are hallmarks of beta cell dysfunction in type 2 diabetes. Signals that can preserve and/or enhance beta cell function are therefore of great therapeutic interest. Here we show that bone morphogenetic protein 4 (Bmp4) and its high-affinity receptor, Bmpr1a, are expressed in beta cells. Mice with attenuated BMPR1A signaling in beta cells show decreased expression of key genes involved in insulin gene expression, proinsulin processing, glucose sensing, secretion stimulus coupling, incretin signaling, and insulin exocytosis and develop diabetes due to impaired insulin secretion. We also show that transgenic expression of Bmp4 in beta cells enhances GSIS and glucose clearance and that systemic administration of BMP4 protein to adult mice significantly stimulates GSIS and ameliorates glucose tolerance in a mouse model of glucose intolerance. Thus, BMP4-BMPR1A signaling in beta cells plays a key role in GSIS.

  6. ROS signaling, oxidative stress and Nrf2 in pancreatic beta-cell function

    SciTech Connect

    Pi Jingbo; Zhang Qiang; Fu Jingqi; Woods, Courtney G.; Hou Yongyong; Corkey, Barbara E.; Collins, Sheila; Andersen, Melvin E.

    2010-04-01

    This review focuses on the emerging evidence that reactive oxygen species (ROS) derived from glucose metabolism, such as H{sub 2}O{sub 2}, act as metabolic signaling molecules for glucose-stimulated insulin secretion (GSIS) in pancreatic beta-cells. Particular emphasis is placed on the potential inhibitory role of endogenous antioxidants, which rise in response to oxidative stress, in glucose-triggered ROS and GSIS. We propose that cellular adaptive response to oxidative stress challenge, such as nuclear factor E2-related factor 2 (Nrf2)-mediated antioxidant induction, plays paradoxical roles in pancreatic beta-cell function. On the one hand, induction of antioxidant enzymes protects beta-cells from oxidative damage and possible cell death, thus minimizing oxidative damage-related impairment of insulin secretion. On the other hand, the induction of antioxidant enzymes by Nrf2 activation blunts glucose-triggered ROS signaling, thus resulting in reduced GSIS. These two premises are potentially relevant to impairment of beta-cells occurring in the late and early stage of Type 2 diabetes, respectively. In addition, we summarized our recent findings that persistent oxidative stress due to absence of uncoupling protein 2 activates cellular adaptive response which is associated with impaired pancreatic beta-cell function.

  7. 4-Phenylbutyric Acid Attenuates Pancreatic Beta-Cell Injury in Rats with Experimental Severe Acute Pancreatitis

    PubMed Central

    Guo, Wen-yi; Zhao, Liang; Xiang, Ming-wei; Mei, Fang-chao; Abliz, Ablikim; Hu, Peng; Deng, Wen-hong; Yu, Jia

    2016-01-01

    Endoplasmic reticulum (ER) stress is a particular process with an imbalance of homeostasis, which plays an important role in pancreatitis, but little is known about how ER stress is implicated in severe acute pancreatitis (SAP) induced pancreatic beta-cell injury. To investigate the effect of 4-phenylbutyric acid (4-PBA) on the beta-cell injury following SAP and the underlying mechanism, twenty-four Sprague-Dawley rats were randomly divided into sham-operation (SO) group, SAP model group, and 4-PBA treatment group. SAP model was induced by infusion of 5% sodium taurocholate into the biliopancreatic duct. 4-PBA or normal saline was injected intraperitoneally for 3 days in respective group before successful modeling. Results showed that 4-PBA attenuated the following: (1) pancreas and islet pathological injuries, (2) serum TNF-α and IL-1β, (3) serum insulin and glucose, (4) beta-cell ultrastructural changes, (5) ER stress markers (BiP, ORP150, and CHOP), Caspase-3, and insulin expression in islet. These results suggested that 4-PBA mitigates pancreatic beta-cell injury and endocrine disorder in SAP, presumably because of its role in inhibiting excessive endoplasmic reticulum stress. This may serve as a new therapeutic target for reducing pancreatic beta-cell injury and endocrine disorder in SAP upon 4-PBA treatment.

  8. Islet beta-cell secretion determines glucagon release from neighbouring alpha-cells.

    PubMed

    Ishihara, Hisamitsu; Maechler, Pierre; Gjinovci, Asllan; Herrera, Pedro-Luis; Wollheim, Claes B

    2003-04-01

    Homeostasis of blood glucose is maintained by hormone secretion from the pancreatic islets of Langerhans. Glucose stimulates insulin secretion from beta-cells but suppresses the release of glucagon, a hormone that raises blood glucose, from alpha-cells. The mechanism by which nutrients stimulate insulin secretion has been studied extensively: ATP has been identified as the main messenger and the ATP-sensitive potassium channel as an essential transducer in this process. By contrast, much less is known about the mechanisms by which nutrients modulate glucagon secretion. Here we use conventional pancreas perfusion and a transcriptional targeting strategy to analyse cell-type-specific signal transduction and the relationship between islet alpha- and beta-cells. We find that pyruvate, a glycolytic intermediate and principal substrate of mitochondria, stimulates glucagon secretion. Our analyses indicate that, although alpha-cells, like beta-cells, possess the inherent capacity to respond to nutrients, secretion from alpha-cells is normally suppressed by the simultaneous activation of beta-cells. Zinc released from beta-cells may be implicated in this suppression. Our results define the fundamental mechanisms of differential responses to identical stimuli between cells in a microorgan.

  9. Alpha cells secrete acetylcholine as a non-neuronal paracrine signal priming human beta cell function

    PubMed Central

    Rodriguez-Diaz, Rayner; Dando, Robin; Jacques-Silva, M. Caroline; Fachado, Alberto; Molina, Judith; Abdulreda, Midhat; Ricordi, Camillo; Roper, Stephen D.; Berggren, Per-Olof; Caicedo, Alejandro

    2011-01-01

    Acetylcholine is a neurotransmitter that plays a major role in the function of the insulin secreting pancreatic beta cell1,2. Parasympathetic innervation of the endocrine pancreas, the islets of Langerhans, has been shown to provide cholinergic input to the beta cell in several species1,3,4, but the role of autonomic innervation in human beta cell function is at present unclear. Here we show that, in contrast to mouse islets, cholinergic innervation of human islets is sparse. Instead, we find that the alpha cells of the human islet provide paracrine cholinergic input to surrounding endocrine cells. Human alpha cells express the vesicular acetylcholine transporter and release acetylcholine when stimulated with kainate or a lowering in glucose concentration. Acetylcholine secretion by alpha cells in turn sensitizes the beta cell response to increases in glucose concentration. Our results demonstrate that in human islets acetylcholine is a paracrine signal that primes the beta cell to respond optimally to subsequent increases in glucose concentration. We anticipate these results to revise models about neural input and cholinergic signaling in the endocrine pancreas. Cholinergic signaling within the islet represents a potential therapeutic target in diabetes5, highlighting the relevance of this advance to future drug development. PMID:21685896

  10. 4-Phenylbutyric Acid Attenuates Pancreatic Beta-Cell Injury in Rats with Experimental Severe Acute Pancreatitis

    PubMed Central

    Guo, Wen-yi; Zhao, Liang; Xiang, Ming-wei; Mei, Fang-chao; Abliz, Ablikim; Hu, Peng; Deng, Wen-hong; Yu, Jia

    2016-01-01

    Endoplasmic reticulum (ER) stress is a particular process with an imbalance of homeostasis, which plays an important role in pancreatitis, but little is known about how ER stress is implicated in severe acute pancreatitis (SAP) induced pancreatic beta-cell injury. To investigate the effect of 4-phenylbutyric acid (4-PBA) on the beta-cell injury following SAP and the underlying mechanism, twenty-four Sprague-Dawley rats were randomly divided into sham-operation (SO) group, SAP model group, and 4-PBA treatment group. SAP model was induced by infusion of 5% sodium taurocholate into the biliopancreatic duct. 4-PBA or normal saline was injected intraperitoneally for 3 days in respective group before successful modeling. Results showed that 4-PBA attenuated the following: (1) pancreas and islet pathological injuries, (2) serum TNF-α and IL-1β, (3) serum insulin and glucose, (4) beta-cell ultrastructural changes, (5) ER stress markers (BiP, ORP150, and CHOP), Caspase-3, and insulin expression in islet. These results suggested that 4-PBA mitigates pancreatic beta-cell injury and endocrine disorder in SAP, presumably because of its role in inhibiting excessive endoplasmic reticulum stress. This may serve as a new therapeutic target for reducing pancreatic beta-cell injury and endocrine disorder in SAP upon 4-PBA treatment. PMID:27656209

  11. New therapies aimed at the preservation or restoration of beta cell function in type 1 diabetes.

    PubMed

    Keymeulen, B

    2006-01-01

    Type 1 diabetes is caused by an immune-mediated destruction of the insulin-secreting beta cells in the pancreas. The disease can become clinically apparent at any age. At diagnosis, there is invariably some residual beta cell function and more so in adults than in children. Recent studies--including one conducted mainly in Belgium--have provided proof of principle that short-term anti-T-cell antibody treatment is able to preserve residual beta cell function for at least 18 months. The resultant stabilizing effect on metabolic control is expected to delay or limit chronic complications in these patients. With a similar goal in mind, nonuremic C-peptide negative patients are offered beta cell transplantation. The outcome of these implants looks promising but their final applicability hinges on finding ways to induce immune tolerance to the donor beta cells. A widespread application, however, will only occur if the shortage of viable human donor cells can be overcome. Both xenotransplantation and stem cell therapy provide possible strategies to solve this problem and represent areas of intense investigation. The ultimate goal is prevention of clinical disease. Studies by the Belgian Diabetes Registry and others in first degree family members of type 1 diabetic patients have refined the identification of individuals at very high risk of hyperglycaemia so that new immunological treatments can be tested in the prediabetic phase. PMID:17240745

  12. 4-Phenylbutyric Acid Attenuates Pancreatic Beta-Cell Injury in Rats with Experimental Severe Acute Pancreatitis.

    PubMed

    Hong, Yu-Pu; Guo, Wen-Yi; Wang, Wei-Xing; Zhao, Liang; Xiang, Ming-Wei; Mei, Fang-Chao; Abliz, Ablikim; Hu, Peng; Deng, Wen-Hong; Yu, Jia

    2016-01-01

    Endoplasmic reticulum (ER) stress is a particular process with an imbalance of homeostasis, which plays an important role in pancreatitis, but little is known about how ER stress is implicated in severe acute pancreatitis (SAP) induced pancreatic beta-cell injury. To investigate the effect of 4-phenylbutyric acid (4-PBA) on the beta-cell injury following SAP and the underlying mechanism, twenty-four Sprague-Dawley rats were randomly divided into sham-operation (SO) group, SAP model group, and 4-PBA treatment group. SAP model was induced by infusion of 5% sodium taurocholate into the biliopancreatic duct. 4-PBA or normal saline was injected intraperitoneally for 3 days in respective group before successful modeling. Results showed that 4-PBA attenuated the following: (1) pancreas and islet pathological injuries, (2) serum TNF-α and IL-1β, (3) serum insulin and glucose, (4) beta-cell ultrastructural changes, (5) ER stress markers (BiP, ORP150, and CHOP), Caspase-3, and insulin expression in islet. These results suggested that 4-PBA mitigates pancreatic beta-cell injury and endocrine disorder in SAP, presumably because of its role in inhibiting excessive endoplasmic reticulum stress. This may serve as a new therapeutic target for reducing pancreatic beta-cell injury and endocrine disorder in SAP upon 4-PBA treatment. PMID:27656209

  13. High fat programming of beta cell compensation, exhaustion, death and dysfunction.

    PubMed

    Cerf, Marlon E

    2015-03-01

    Programming refers to events during critical developmental windows that shape progeny health outcomes. Fetal programming refers to the effects of intrauterine (in utero) events. Lactational programming refers to the effects of events during suckling (weaning). Developmental programming refers to the effects of events during both fetal and lactational life. Postnatal programming refers to the effects of events either from birth (lactational life) to adolescence or from weaning (end of lactation) to adolescence. Islets are most plastic during the early life course; hence programming during fetal and lactational life is most potent. High fat (HF) programming is the maintenance on a HF diet (HFD) during critical developmental life stages that alters progeny metabolism and physiology. HF programming induces variable diabetogenic phenotypes dependent on the timing and duration of the dietary insult. Maternal obesity reinforces HF programming effects in progeny. HF programming, through acute hyperglycemia, initiates beta cell compensation. However, HF programming eventually leads to chronic hyperglycemia that triggers beta cell exhaustion, death and dysfunction. In HF programming, beta cell dysfunction often co-presents with insulin resistance. Balanced, healthy nutrition during developmental windows is critical for preserving beta cell structure and function. Thus early positive nutritional interventions that coincide with the development of beta cells may reduce the overwhelming burden of diabetes and metabolic disease.

  14. Nutrient regulation of insulin secretion and beta-cell functional integrity.

    PubMed

    Newsholme, Philip; Gaudel, Celine; McClenaghan, Neville H

    2010-01-01

    Pancreatic beta-cells are often referred to as "fuel sensors" as they continually monitor and respond to dietary nutrients, under the modulation of additional neurohormonal signals, in order to secrete insulin to best meet the needs of the organism. beta-cell nutrient sensing requires metabolic activation, resulting in production of stimulus-secretion coupling signals that promote insulin biosynthesis and release. The primary stimulus for insulin secretion is glucose, and islet beta-cells are particularly responsive to this important nutrient secretagogue, It is important to consider individual effects of different classes of nutrient or other physiological or pharmacological agents on metabolism and insulin secretion. However, given that beta-cells are continually exposed to a complex milieu of nutrients and other circulating factors, it is important to also acknowledge and examine the interplay between glucose metabolism and that of the two other primary nutrient classes, the amino acids and fatty acids. It is the mixed nutrient sensing and outputs of glucose, amino and fatty acid metabolism that generate the metabolic coupling factors (MCFs) involved in signaling for insulin exocytosis. Primary MCFs in the beta-cell include ATP, NADPH, glutamate, long chain acyl-CoA and diacylglycerol and are discussed in detail in this article.

  15. The responses of I beta cells to increases in the rate of lung inflation.

    PubMed

    Marino, P L; Davies, R O; Pack, A I

    1981-08-31

    The activity of inspiratory cells in the region of the nucleus of the tractus solitarius (NTS) was recorded extracellularly in paralyzed, artificially ventilated cats either during chloralose-urethane anesthesia or following midcollicular decerebration. Twenty-three of the 68 inspiratory cells recorded in the region of the NTS were classified as I beta cells on the basis of their response to withholding lung inflation. The dynamic sensitivity of I beta cells was determined by studying their response to increases in the rate of lung inflation at constant peak volume. The I beta cells in this study showed 3 distinct patterns of response to increases in the rate of inflation. Five cells showed no change in firing pattern (fixed firing pattern). Ten cells showed an increase in the rate of rise of cell activity but no change in peak frequency (low dynamic sensitivity). Eight cells showed increases in both the rate of rise of cell activity and peak frequency (high dynamic sensitivity). It was concluded that I beta cells are not a functionally homogeneous population, at least in terms of their dynamic sensitivity. Cells showing fixed firing patterns have the characteristics of off-switch neurons. Cells with low levels of dynamic sensitivity may receive afferents from pulmonary stretch receptors. Cells showing a high degree of dynamic sensitivity may receive afferents from rapidly adapting receptors. The fact that I beta cells are not a functionally homogeneous population may explain the many divergent observations reported from studies of these cells.

  16. Hormone-sensitive lipase, the rate-limiting enzyme in triglyceride hydrolysis, is expressed and active in beta-cells.

    PubMed

    Mulder, H; Holst, L S; Svensson, H; Degerman, E; Sundler, F; Ahrén, B; Rorsman, P; Holm, C

    1999-01-01

    Triglycerides in the beta-cell may be important for stimulus-secretion coupling, through provision of a lipid-derived signal, and for pathogenetic events in NIDDM, where lipids may adversely affect beta-cell function. In adipose tissues, hormone-sensitive lipase (HSL) is rate-limiting in triglyceride hydrolysis. Here, we investigated whether this enzyme is also expressed and active in beta-cells. Northern blot analysis and reverse transcription-polymerase chain reaction demonstrated that HSL is expressed in rat islets and in the clonal beta-cell lines INS-1, RINm5F, and HIT-T15. Western blot analysis identified HSL in mouse and rat islets and the clonal beta-cells. In mouse and rat, immunocytochemistry showed a predominant occurrence of HSL in beta-cells, with a presumed cytoplasmic localization. Lipase activity in homogenates of the rodent islets and clonal beta-cells constituted 2.1 +/- 0.6% of that in adipocytes; this activity was immunoinhibited by use of antibodies to HSL. The established HSL expression and activity in beta-cells offer a mechanism whereby lipids are mobilized from intracellular stores. Because HSL in adipocytes is activated by cAMP-dependent protein kinase (PKA), PKA-regulated triglyceride hydrolysis in beta-cells may participate in the regulation of insulin secretion, possibly by providing a lipid-derived signal, e.g., long-chain acyl-CoA and diacylglycerol.

  17. Pluripotent stem cells, a potential source of beta-cells for diabetes therapy.

    PubMed

    Hansson, Mattias; Madsen, Ole D

    2010-04-01

    Although the reconstitution of a functional beta-cell mass by transplantation of isolated islets can restore euglycemia in the absence of insulin treatment, a shortage of donor material is one of the factors preventing the general use of cell replacement therapy for the treatment of type 1 diabetes mellitus (T1DM). Advances in the directed differentiation of pluripotent stem cells toward beta-cells via the stepwise recapitulation of embryonic development have generated proof of concept demonstrating that stem cells may be an appropriate source of cells for the generation of therapeutic beta-cells. However, progress toward a clinical application of this technology is slow and challenging. This review highlights some of the critical issues impeding the translation of stem cell-based diabetes therapies to the clinic. PMID:20336589

  18. Zip4 Mediated Zinc Influx Stimulates Insulin Secretion in Pancreatic Beta Cells

    PubMed Central

    Hardy, Alexandre B.; Prentice, Kacey J.; Froese, Sean; Liu, Ying; Andrews, Glen K.; Wheeler, Michael B.

    2015-01-01

    Zinc has an important role in normal pancreatic beta cell physiology as it regulates gene transcription, insulin crystallization and secretion, and cell survival. Nevertheless, little is known about how zinc is transported through the plasma membrane of beta cells and which of the class of zinc influx transporters (Zip) is involved. Zip4 was previously shown to be expressed in human and mouse beta cells; however, its function there is still unknown. Therefore, the aim of this study was to define the zinc transport role of Zip4 in beta cells. To investigate this, Zip4 was over-expressed in MIN6 beta cells using a pCMV6-Zip4GFP plasmid. Organelle staining combined with confocal microscopy showed that Zip4 exhibits a widespread localization in MIN6 cells. Time-lapse zinc imaging experiments showed that Zip4 increases cytoplasmic zinc levels. This resulted in increased granular zinc content and glucose-stimulated insulin secretion. Interestingly, it is unlikely that the increased glucose stimulated insulin secretion was triggered by a modulation of mitochondrial function, as mitochondrial membrane potential remained unchanged. To define the role of Zip4 in-vivo, we generated a beta cell-specific knockout mouse model (Zip4BKO). Deletion of the Zip4 gene was confirmed in Zip4BKO islets by PCR, RT-PCR, and immuno-histochemistry. Zip4BKO mice showed slightly improved glucose homeostasis but no change in insulin secretion during an oral glucose tolerance test. While Zip4 was not found to be essential for proper glucose homeostasis and insulin secretion in vivo in mice, this study also found that Zip4 mediates increases in cytoplasmic and granular zinc pools and stimulates glucose dependant insulin secretion in-vitro. PMID:25806541

  19. Precise expression of Fis1 is important for glucose responsiveness of beta cells.

    PubMed

    Schultz, Julia; Waterstradt, Rica; Kantowski, Tobias; Rickmann, Annekatrin; Reinhardt, Florian; Sharoyko, Vladimir; Mulder, Hindrik; Tiedge, Markus; Baltrusch, Simone

    2016-07-01

    Mitochondrial network functionality is vital for glucose-stimulated insulin secretion in pancreatic beta cells. Altered mitochondrial dynamics in pancreatic beta cells are thought to trigger the development of type 2 diabetes mellitus. Fission protein 1 (Fis1) might be a key player in this process. Thus, the aim of this study was to investigate mitochondrial morphology in dependence of beta cell function, after knockdown and overexpression of Fis1. We demonstrate that glucose-unresponsive cells with impaired glucose-stimulated insulin secretion (INS1-832/2) showed decreased mitochondrial dynamics compared with glucose-responsive cells (INS1-832/13). Accordingly, mitochondrial morphology visualised using MitoTracker staining differed between the two cell lines. INS1-832/2 cells formed elongated and clustered mitochondria, whereas INS1-832/13 cells showed a homogenous mitochondrial network. Fis1 overexpression using lentiviral transduction significantly improved glucose-stimulated insulin secretion and mitochondrial network homogeneity in glucose-unresponsive cells. Conversely, Fis1 downregulation by shRNA, both in primary mouse beta cells and glucose-responsive INS1-832/13 cells, caused unresponsiveness and significantly greater numbers of elongated mitochondria. Overexpression of FIS1 in primary mouse beta cells indicated an upper limit at which higher FIS1 expression reduced glucose-stimulated insulin secretion. Thus, FIS1 was overexpressed stepwise up to a high concentration in RINm5F cells using the RheoSwitch system. Moderate FIS1 expression improved glucose-stimulated insulin secretion, whereas high expression resulted in loss of glucose responsiveness and in mitochondrial artificial loop structures and clustering. Our data confirm that FIS1 is a key regulator in pancreatic beta cells, because both glucose-stimulated insulin secretion and mitochondrial dynamics were clearly adapted to precise expression levels of this fission protein. PMID:27179109

  20. Glucose sensing in the pancreatic beta cell: a computational systems analysis

    PubMed Central

    2010-01-01

    Background Pancreatic beta-cells respond to rising blood glucose by increasing oxidative metabolism, leading to an increased ATP/ADP ratio in the cytoplasm. This leads to a closure of KATP channels, depolarization of the plasma membrane, influx of calcium and the eventual secretion of insulin. Such mechanism suggests that beta-cell metabolism should have a functional regulation specific to secretion, as opposed to coupling to contraction. The goal of this work is to uncover contributions of the cytoplasmic and mitochondrial processes in this secretory coupling mechanism using mathematical modeling in a systems biology approach. Methods We describe a mathematical model of beta-cell sensitivity to glucose. The cytoplasmic part of the model includes equations describing glucokinase, glycolysis, pyruvate reduction, NADH and ATP production and consumption. The mitochondrial part begins with production of NADH, which is regulated by pyruvate dehydrogenase. NADH is used in the electron transport chain to establish a proton motive force, driving the F1F0 ATPase. Redox shuttles and mitochondrial Ca2+ handling were also modeled. Results The model correctly predicts changes in the ATP/ADP ratio, Ca2+ and other metabolic parameters in response to changes in substrate delivery at steady-state and during cytoplasmic Ca2+ oscillations. Our analysis of the model simulations suggests that the mitochondrial membrane potential should be relatively lower in beta cells compared with other cell types to permit precise mitochondrial regulation of the cytoplasmic ATP/ADP ratio. This key difference may follow from a relative reduction in respiratory activity. The model demonstrates how activity of lactate dehydrogenase, uncoupling proteins and the redox shuttles can regulate beta-cell function in concert; that independent oscillations of cytoplasmic Ca2+ can lead to slow coupled metabolic oscillations; and that the relatively low production rate of reactive oxygen species in beta-cells

  1. Efficient (18)F-Labeling of Synthetic Exendin-4 Analogues for Imaging Beta Cells.

    PubMed

    Keliher, Edmund J; Reiner, Thomas; Thurber, Greg M; Upadhyay, Rabi; Weissleder, Ralph

    2012-08-01

    A number of exendin derivatives have been developed to target glucagon-like peptide 1 (GLP-1) receptors on beta cells in vivo. Modifications of exendin analogues have been shown to have significant effects on pharmacokinetics and, as such, have been used to develop a variety of therapeutic compounds. Here, we show that an exendin-4, modified at position 12 with a cysteine conjugated to a tetrazine, can be labeled with (18)F-trans-cyclooctene and converted into a PET imaging agent at high yields and with good selectivity. The agent accumulates in beta cells in vivo and has sufficiently high accumulation in mouse models of insulinomas to enable in vivo imaging.

  2. Endoplasmic Reticulum Stress in Beta Cells and Development of Diabetes

    PubMed Central

    Fonseca, Sonya G.; Burcin, Mark; Gromada, Jesper; Urano, Fumihiko

    2009-01-01

    The endoplasmic reticulum (ER) is a cellular compartment responsible for multiple important cellular functions including the biosynthesis and folding of newly synthesized proteins destined for secretion, such as insulin. A myriad of pathological and physiological factors perturb ER function and cause dysregulation of ER homeostasis, leading to ER stress. ER stress elicits a signaling cascade to mitigate stress, the Unfolded Protein Response (UPR). As long as the UPR can relieve stress, cells can produce the proper amount of proteins and maintain ER homeostasis. If the UPR, however, fails to maintain ER homeostasis, cells will undergo apoptosis. Activation of the UPR is critical to the survival of insulin-producing pancreatic β-cells with high secretory protein production. Any disruption of ER homeostasis in β-cells can lead to cell death and contribute to the pathogenesis of diabetes. There are several models of ER stress-mediated diabetes. In this review, we outline the underlying molecular mechanisms of ER stress-mediated β-cell dysfunction and death during the progression of diabetes. PMID:19665428

  3. Coating nanofiber scaffolds with beta cell membrane to promote cell proliferation and function.

    PubMed

    Chen, Wansong; Zhang, Qiangzhe; Luk, Brian T; Fang, Ronnie H; Liu, Younian; Gao, Weiwei; Zhang, Liangfang

    2016-05-21

    The cell membrane cloaking technique has emerged as an intriguing strategy in nanomaterial functionalization. Coating synthetic nanostructures with natural cell membranes bestows the nanostructures with unique cell surface antigens and functions. Previous studies have focused primarily on development of cell membrane-coated spherical nanoparticles and the uses thereof. Herein, we attempt to extend the cell membrane cloaking technique to nanofibers, a class of functional nanomaterials that are drastically different from nanoparticles in terms of dimensional and mechanophysical characteristics. Using pancreatic beta cells as a model cell line, we demonstrate successful preparation of cell membrane-coated nanofibers and validate that the modified nanofibers possess an antigenic exterior closely resembling that of the source beta cells. When such nanofiber scaffolds are used to culture beta cells, both cell proliferation rate and function are significantly enhanced. Specifically, glucose-dependent insulin secretion from the cells is increased by near five-fold compared with the same beta cells cultured in regular, unmodified nanofiber scaffolds. Overall, coating cell membranes onto nanofibers could add another dimension of flexibility and controllability in harnessing cell membrane functions and offer new opportunities for innovative applications. PMID:27139582

  4. Beta cell function and BMI in ethnically diverse children with newly diagnosed autoimmune type 1 diabetes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of our study was to examine the relationship between BMI and beta-cell function at diagnosis of autoimmune type 1 diabetes (T1D) in a large group of ethnically diverse children. Cross-sectional analysis of 524 children (60.8% White, 19.5% Hispanic, 14.5% African-American, 5.2% other n...

  5. Merocytic dendritic cells break T cell tolerance to beta cell antigens in NOD mouse diabetes1

    PubMed Central

    Katz, Jonathan D; Ondr, Jennifer K; Opoka, Robert J; Garcia, Zacharias; Janssen, Edith M

    2010-01-01

    In type 1 diabetes (T1D), the breach of central and peripheral tolerance results in autoreactive T cells destroying insulin-producing, pancreatic beta cells. Herein, we identify a critical sub-population of dendritic cells responsible for mediating both the cross-presentation of islet antigen to CD8+ T cells and the direct presentation of beta cell antigen to CD4+ T cells. These cells, termed merocytic dendritic cells (mcDC), are more numerous in nonobese diabetic (NOD) mouse, and when antigen-loaded rescue CD8+ T cells from peripheral anergy and deletion, while stimulating islet-reactive CD4+ T cells. When purified from the pancreatic lymph nodes of overtly diabetic NOD mice, mcDC break peripheral T cell tolerance to beta cells in vivo and induce rapid onset T1D in young NOD mouse. Thus, the mcDC subset appears to represent the long-sought APC responsible for breaking peripheral tolerance to beta cell antigen in vivo. PMID:20644171

  6. Pancreatic beta cells are highly susceptible to oxidative and ER stresses during the development of diabetes.

    PubMed

    Gorasia, Dhana G; Dudek, Nadine L; Veith, Paul D; Shankar, Renu; Safavi-Hemami, Helena; Williamson, Nicholas A; Reynolds, Eric C; Hubbard, Michael J; Purcell, Anthony W

    2015-02-01

    The complex interplay of many cell types and the temporal heterogeneity of pancreatic islet composition obscure the direct role of resident alpha and beta cells in the development of Type 1 diabetes. Therefore, in addition to studying islets isolated from non-obese diabetic mice, we analyzed homogeneous cell populations of murine alpha (αTC-1) and beta (NIT-1) cell lines to understand the role and differential survival of these two predominant islet cell populations. A total of 56 proteins in NIT-1 cells and 50 in αTC-1 cells were differentially expressed when exposed to proinflammatory cytokines. The major difference in the protein expression between cytokine-treated NIT-1 and αTC-1 cells was free radical scavenging enzymes. A similar observation was made in cytokine-treated whole islets, where a comprehensive analysis of subcellular fractions revealed that 438 unique proteins were differentially expressed under inflammatory conditions. Our data indicate that beta cells are relatively susceptible to ER and oxidative stress and reveal key pathways that are dysregulated in beta cells during cytokine exposure. Additionally, in the islets, inflammation also leads to enhanced antigen presentation, which completes a three-way insult on beta cells, rendering them targets of infiltrating T lymphocytes.

  7. A red-shifted photochromic sulfonylurea for the remote control of pancreatic beta cell function.

    PubMed

    Broichhagen, J; Frank, J A; Johnston, N R; Mitchell, R K; Šmid, K; Marchetti, P; Bugliani, M; Rutter, G A; Trauner, D; Hodson, D J

    2015-04-01

    Azobenzene photoresponsive elements can be installed on sulfonylureas, yielding optical control over pancreatic beta cell function and insulin release. An obstacle to such photopharmacological approaches remains the use of ultraviolet-blue illumination. Herein, we synthesize and test a novel yellow light-activated sulfonylurea based on a heterocyclic azobenzene bearing a push-pull system. PMID:25744824

  8. Nuclear SREBP-1a causes loss of pancreatic {beta}-cells and impaired insulin secretion

    SciTech Connect

    Iwasaki, Yuko; Iwasaki, Hitoshi; Yatoh, Shigeru; Ishikawa, Mayumi; Kato, Toyonori; Matsuzaka, Takashi; Nakagawa, Yoshimi; Yahagi, Naoya; Kobayashi, Kazuto; Takahashi, Akimitsu; Suzuki, Hiroaki; Yamada, Nobuhiro; Shimano, Hitoshi

    2009-01-16

    Transgenic mice expressing nuclear sterol regulatory element-binding protein-1a under the control of the insulin promoter were generated to determine the role of SREBP-1a in pancreatic {beta}-cells. Only low expressors could be established, which exhibited mild hyperglycemia, impaired glucose tolerance, and reduced plasma insulin levels compared to C57BL/6 controls. The islets isolated from the transgenic mice were fewer and smaller, and had decreased insulin content and unaltered glucagon staining. Both glucose- and potassium-stimulated insulin secretions were decreased. The transgenic islets consistently expressed genes for fatty acids and cholesterol synthesis, resulting in accumulation of triglycerides but not cholesterol. PDX-1, {beta}{epsilon}{tau}{alpha}2, MafA, and IRS-2 were suppressed, partially explaining the loss and dysfunction of {beta}-cell mass. The transgenic mice on a high fat/high sucrose diet still exhibited impaired insulin secretion and continuous {beta}-cell growth defect. Therefore, nuclear SREBP-1a, even at a low level, strongly disrupts {beta}-cell mass and function.

  9. Nitric oxide stimulates insulin gene transcription in pancreatic {beta}-cells

    SciTech Connect

    Campbell, S.C. . E-mail: s.c.campbell@ncl.ac.uk; Richardson, H.; Ferris, W.F.; Butler, C.S.; Macfarlane, W.M.

    2007-02-23

    Recent studies have identified a positive role for nitric oxide (NO) in the regulation of pancreatic {beta}-cell function. The aim of this study was to determine the effects of short-term exposure to NO on {beta}-cell gene expression and the activity of the transcription factor PDX-1. NO stimulated the activity of the insulin gene promoter in Min6 {beta}-cells and endogenous insulin mRNA levels in both Min6 and isolated islets of Langerhans. Addition of wortmannin prior to NO stimulation blocked the observed increases in insulin gene promoter activity. Although NO addition stimulated the phosphorylation of p38, inhibition by SB203580 did not block the effect of NO on the insulin gene promoter. NO addition also stimulated both the nuclear accumulation and the DNA binding activity of PDX-1. This study has shown that over 24 h, NO stimulates insulin gene expression, PI-3-kinase activity and the activity of the critical {beta}-cell transcription factor PDX-1.

  10. Measuring beta-cell function relative to insulin sensitivity in youth: Does the hyperglycemic clamp suffice?

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To compare beta-cell function relative to insulin sensitivity, disposition index (DI), calculated from two clamps (2cDI, insulin sensitivity from the hyperinsulinemic-euglycemic clamp and first-phase insulin from the hyperglycemic clamp) with the DI calculated from the hyperglycemic clamp alone (hcD...

  11. Generating pancreatic beta-cells from embryonic stem cells by manipulating signaling pathways.

    PubMed

    Champeris Tsaniras, Spyridon; Jones, Peter M

    2010-07-01

    Type 1 diabetes results from an insufficiency of insulin production as a result of autoimmune destruction of the insulin-secreting pancreatic beta-cells. It can be treated by transplantation of islets of Langerhans from human donors, but widespread application of this therapy is restricted by the scarcity of donor tissue. Generation of functional beta-cells from embryonic stem (ES) cells in vitro could provide a source of an alternative graft material. Several ES cell differentiation protocols have reported the production of insulin-producing cells by mimicking the in vivo developmental stages of pancreatic organogenesis in which cells are transitioned through mesendoderm, definitive endoderm, foregut endoderm, pancreatic endoderm, and the endocrine precursor stage, until mature beta-cells are obtained. These studies provide proof of concept that recapitulating pancreatic development in vitro offers a useful strategy for generating beta-cells, but current differentiation protocols employ a bewildering variety of growth factors, mitogens, and pharmacological agents. In this review, we will attempt to clarify the functions of these agents in in vitro differentiation strategies by focusing on the intracellular signaling pathways through which they operate - phosphatidylinositol 3-kinase, transforming growth factor beta, Wnt/beta-catenin, Hedgehog, and Notch. PMID:20385725

  12. Metabolomic profiling of amino acids and beta-cell function relative to insulin sensitivity in youth

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In longitudinal studies of adults, elevated amino acid (AA) concentrations predicted future type 2 diabetes mellitus (T2DM). The aim of the present investigation was to examine whether increased plasma AA concentrations are associated with impaired beta-cell function relative to insulin sensitivity ...

  13. Ataxin-10 interacts with O-GlcNAc transferase OGT in pancreatic {beta} cells

    SciTech Connect

    Andrali, Sreenath S.; Maerz, Pia; Oezcan, Sabire . E-mail: sozcan@uky.edu

    2005-11-11

    Several nuclear and cytoplasmic proteins in metazoans are modified by O-linked N-acetylglucosamine (O-GlcNAc). This modification is dynamic and reversible similar to phosphorylation and is catalyzed by the O-linked GlcNAc transferase (OGT). Hyperglycemia has been shown to increase O-GlcNAc levels in pancreatic {beta} cells, which appears to interfere with {beta}-cell function. To obtain a better understanding of the role of O-linked GlcNAc modification in {beta} cells, we have isolated OGT interacting proteins from a cDNA library made from the mouse insulinoma MIN6 cell line. We describe here the identification of Ataxin-10, encoded by the SCA10 (spinocerebellar ataxia type 10) gene as an OGT interacting protein. Mutations in the SCA10 gene cause progressive cerebellar ataxias and seizures. We demonstrate that SCA10 interacts with OGT in vivo and is modified by O-linked glycosylation in MIN6 cells, suggesting a novel role for the Ataxin-10 protein in pancreatic {beta} cells.

  14. Coating nanofiber scaffolds with beta cell membrane to promote cell proliferation and function

    NASA Astrophysics Data System (ADS)

    Chen, Wansong; Zhang, Qiangzhe; Luk, Brian T.; Fang, Ronnie H.; Liu, Younian; Gao, Weiwei; Zhang, Liangfang

    2016-05-01

    The cell membrane cloaking technique has emerged as an intriguing strategy in nanomaterial functionalization. Coating synthetic nanostructures with natural cell membranes bestows the nanostructures with unique cell surface antigens and functions. Previous studies have focused primarily on development of cell membrane-coated spherical nanoparticles and the uses thereof. Herein, we attempt to extend the cell membrane cloaking technique to nanofibers, a class of functional nanomaterials that are drastically different from nanoparticles in terms of dimensional and mechanophysical characteristics. Using pancreatic beta cells as a model cell line, we demonstrate successful preparation of cell membrane-coated nanofibers and validate that the modified nanofibers possess an antigenic exterior closely resembling that of the source beta cells. When such nanofiber scaffolds are used to culture beta cells, both cell proliferation rate and function are significantly enhanced. Specifically, glucose-dependent insulin secretion from the cells is increased by near five-fold compared with the same beta cells cultured in regular, unmodified nanofiber scaffolds. Overall, coating cell membranes onto nanofibers could add another dimension of flexibility and controllability in harnessing cell membrane functions and offer new opportunities for innovative applications.

  15. Beta-cell metabolic alterations under chronic nutrient overload in rat and human islets

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The aim of this study was to assess multifactorial Beta-cell responses to metabolic perturbations in primary rat and human islets. Treatment of dispersed rat islet cells with elevated glucose and free fatty acids (FFAs, oleate:palmitate = 1:1 v/v) resulted in increases in the size and the number of ...

  16. [New aspects of pancreatic beta cell functions and their possible therapeutic applications].

    PubMed

    Tiedge, M

    2006-12-01

    Using the metabolic stimulus-secretion coupling of pancreatic beta cells as an example, this review illustrates how new strategies in the treatment of type 2 diabetes mellitus can be developed from the results of basic research. Metabolic stimulus-secretion coupling presupposes the metabolizing of those stimuli of insulin secretion that have the properties of nutritional substances. Changes in the ATP/ADP ratio within the beta cells will then trigger the release of insulin granules from them. Glucokinase, a glucose phosphorylating enzyme, functions as a metabolic glucose sensor, which couples changes in physiological glucose concentration in the pancreatic beta cells and in the liver to the intermediary metabolism, i.e. glycolysis, the citrate cycle and respiratory-chain phosphorylation. In this way insulin secretion and hepatic metabolism are positively influenced. Several pharmaceutical companies (Roche, Merck, Astra-Zeneca, Lilly) have recently developed first examples of glucokinase-activating compounds and demonstrated in animal models their efficacy in the treatment of type 2 diabetes mellitus. These glucokinase activators prevent glucokinase from changing into a catalytically inactive structure. They also increase glucose affinity of the enzyme and stabilize a catalytically active form of glucokinase proteins. In this way glucokinase activators increase glucose-induced insulin secretion and inhibit hepatic glucogenesis. Glucokinase activators are an interesting innovation in the future treatment of type 2 diabetes, because their action on beta cells and the liver is caused by changes in blood glucose concentration.

  17. Selective deletion of Pten in pancreatic beta cells leads to increased islet mass and resistance to STZ-induced diabetes.

    PubMed

    Stiles, Bangyan L; Kuralwalla-Martinez, Christine; Guo, Wei; Gregorian, Caroline; Wang, Ying; Tian, Jide; Magnuson, Mark A; Wu, Hong

    2006-04-01

    Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) is a lipid phosphatase. PTEN inhibits the action of phosphatidylinositol-3-kinase and reduces the levels of phosphatidylinositol triphosphate, a crucial second messenger for cell proliferation and survival, as well as insulin signaling. In this study, we deleted Pten specifically in the insulin producing beta cells during murine pancreatic development. Pten deletion leads to increased cell proliferation and decreased cell death, without significant alteration of beta-cell differentiation. Consequently, the mutant pancreas generates more and larger islets, with a significant increase in total beta-cell mass. PTEN loss also protects animals from developing streptozotocin-induced diabetes. Our data demonstrate that PTEN loss in beta cells is not tumorigenic but beneficial. This suggests that modulating the PTEN-controlled signaling pathway is a potential approach for beta-cell protection and regeneration therapies. PMID:16537919

  18. Selective deletion of Pten in pancreatic beta cells leads to increased islet mass and resistance to STZ-induced diabetes.

    PubMed

    Stiles, Bangyan L; Kuralwalla-Martinez, Christine; Guo, Wei; Gregorian, Caroline; Wang, Ying; Tian, Jide; Magnuson, Mark A; Wu, Hong

    2006-04-01

    Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) is a lipid phosphatase. PTEN inhibits the action of phosphatidylinositol-3-kinase and reduces the levels of phosphatidylinositol triphosphate, a crucial second messenger for cell proliferation and survival, as well as insulin signaling. In this study, we deleted Pten specifically in the insulin producing beta cells during murine pancreatic development. Pten deletion leads to increased cell proliferation and decreased cell death, without significant alteration of beta-cell differentiation. Consequently, the mutant pancreas generates more and larger islets, with a significant increase in total beta-cell mass. PTEN loss also protects animals from developing streptozotocin-induced diabetes. Our data demonstrate that PTEN loss in beta cells is not tumorigenic but beneficial. This suggests that modulating the PTEN-controlled signaling pathway is a potential approach for beta-cell protection and regeneration therapies.

  19. Conversion of embryonic stem cells into pancreatic beta-cell surrogates guided by ontogeny.

    PubMed

    Lees, Justin G; Tuch, Bernard E

    2006-05-01

    Cellular therapies to treat Type 1 diabetes are being devised and the use of human embryonic stem cells (hESCs) offers a solution to the issue of supply, because hESCs can be maintained in a pluripotent state indefinitely. Furthermore, hESCs have advantages in terms of their plasticity and reduced immunogenicity. Several strategies that have so far been investigated indicate that hESCs are capable of differentiating into insulin producing beta-cell surrogates. However the efficiency of the differentiation procedures used is generally quite low and the cell populations derived are often highly heterogenous. A strategy that appears to have long term potential is to design differentiation procedures based on the ontogeny of the beta-cell. The focus of this strategy is to replicate signaling processes that are known to be involved in the maturation of a beta-cell. The earliest pancreatic progenitors found in the developing vertebrate fetus are produced via a process known as gastrulation and form part of the definitive endoderm germ layer. hESCs have recently been differentiated into definitive endoderm with high efficiency using a differentiation procedure that mimics the signaling that occurs during gastrulation and the formation of the definitive endoderm. Subsequent events during pancreas development involve a section of the definitive endoderm forming into pancreatic epithelium, which then branches into the pancreatic mesenchyme to form islet clusters of endocrine cells. A proportion of the endocrine precursor cells within islets develop into insulin producing beta-cells. The challenge currently is to design hESC differentiation procedures that mimic the combined events of these stages of beta-cell development.

  20. Therapies aimed at preservation or restoration of beta cell function in type 1 diabetes.

    PubMed

    Keymeulen, B

    2008-01-01

    Type 1 diabetes is caused by an immune mediated destruction of the insulin-secreting beta cells in the pancreas. The disease can become clinically apparent at any age. At clinical diagnosis, there is invariably some residual beta cell function. Recent studies--including one mainly conducted in Belgium--have provided proof of principle that short-term humanized anti-T-cell antibody treatment is able to preserve residual beta cell function for at least 18 months in adult type 1 diabetic patients with a recent clinical onset of disease. The effect of anti-T-cell antibody treatment is more pronounced among patients with initial higher residual beta-cell function. The resultant stabilizing effect on metabolic control is expected to delay chronic complications and avoid hypoglycemia in these patients. With a similar goal in mind, non-uremic C-peptide negative type 1 diabetic patients are offered beta cell transplantation. During the last years the one year survival of these grafts under immune suppression with Anti-Thymocyte-Globulin, tacrolimus and mycophenolate mofetil exceeds 80% with virtually no cases of primary non-function. Widespread application will however only occur if ways are found to induce operational graft tolerance and the shortage of viable human donor cells can be overcome. Both islet xenotransplantation and stem cell therapy provide possible strategies to solve this problem and represent areas of intense investigation. The ultimate goal is prevention of clinical disease. Studies by the Belgian Diabetes Registry and others in first degree family members of type 1 diabetic patients have refined identification of individuals at very high risk of hyperglycemia so that new immunological treatments can be tested in the prediabetic phase. PMID:18630722

  1. Fibroblast growth factor receptor-1 signaling in pancreatic islet beta-cells is modulated by the extracellular matrix.

    PubMed

    Kilkenny, Dawn M; Rocheleau, Jonathan V

    2008-01-01

    Maintenance of pancreatic beta-cell mass depends on extracellular stimuli that promote survival and proliferation. In the islet, these stimuli come from the beta-cell microenvironment and include extracellular matrix deposited by associated vascular endothelial cells. Fibroblast growth factor receptor-1 (FGFR1) has recently been implicated as a signaling pathway that is important for normal beta-cell function. We would like to understand how extracellular matrix and FGFR1 signaling interact to promote beta-cell survival and proliferation. To examine beta-cell-specific receptor responses, we created lentiviral vectors with rat insulin promoter-driven expression of Venus fluorescent protein-tagged full-length (R1betav) and kinase-deficient (KDR1betav) FGFR1. Significant FGF-1-dependent activation of ERK1/2 was observed in betaTC3 cells, dispersed beta-cells, and beta-cells in intact islets. This response was enhanced by R1betav expression and reduced by KDR1betav expression. Plating-dispersed beta-cells on collagen type IV resulted in enhanced expression of endogenous FGFR1 that was associated with sustained activation of ERK1/2. Conversely, plating cells on laminin reduced expression of FGFR1, and this reduction was associated with transient activation of ERK1/2. Addition of neutralizing antibodies to inhibit beta-cell attachment to laminin via alpha(6)-integrin increased high-affinity FGF-1-binding at the plasma membrane and resulted in sustained ERK1/2 activity similar to cells plated on collagen type IV. These data show that the FGF-stimulated beta-cell response is negatively affected by alpha(6)-integrin binding to laminin and suggest regulation associated with vascular endothelial cell remodeling. PMID:17916654

  2. Minireview: beta-cell replacement therapy for diabetes in the 21st century: manipulation of cell fate by directed differentiation.

    PubMed

    Yechoor, Vijay; Chan, Lawrence

    2010-08-01

    Pancreatic beta-cell failure underlies type 1 diabetes; it also contributes in an essential way to type 2 diabetes. beta-Cell replacement is an important component of any cure for diabetes. The current options of islet and pancreas transplantation are not satisfactory as definitive forms of therapy. Here, we review strategies for induced de novo pancreatic beta-cell formation, which depend on the targeted differentiation of cells into pancreatic beta-cells. With this objective in mind, one can manipulate the fate of three different types of cells: 1) from terminally differentiated cells, e.g. exocrine pancreatic cells, into beta-cells; 2) from multipotent adult stem cells, e.g. hepatic oval cells, into pancreatic islets; and 3) from pluripotent stem cells, e.g. embryonic stem cells and induced pluripotent stem cells, into beta-cells. We will examine the pros and cons of each strategy as well as the hurdles that must be overcome before these approaches to generate new beta-cells will be ready for clinical application. PMID:20219891

  3. Minireview: beta-cell replacement therapy for diabetes in the 21st century: manipulation of cell fate by directed differentiation.

    PubMed

    Yechoor, Vijay; Chan, Lawrence

    2010-08-01

    Pancreatic beta-cell failure underlies type 1 diabetes; it also contributes in an essential way to type 2 diabetes. beta-Cell replacement is an important component of any cure for diabetes. The current options of islet and pancreas transplantation are not satisfactory as definitive forms of therapy. Here, we review strategies for induced de novo pancreatic beta-cell formation, which depend on the targeted differentiation of cells into pancreatic beta-cells. With this objective in mind, one can manipulate the fate of three different types of cells: 1) from terminally differentiated cells, e.g. exocrine pancreatic cells, into beta-cells; 2) from multipotent adult stem cells, e.g. hepatic oval cells, into pancreatic islets; and 3) from pluripotent stem cells, e.g. embryonic stem cells and induced pluripotent stem cells, into beta-cells. We will examine the pros and cons of each strategy as well as the hurdles that must be overcome before these approaches to generate new beta-cells will be ready for clinical application.

  4. Ketosis-onset diabetes in Tunisian adults: immunological markers and beta-cell function.

    PubMed

    Harzallah, F; Ben Brahim, A; Laadhar, L; Feki, M; Zitouni, M; Makni, S; Kaabachi, N; Slimane, H

    2010-01-01

    The aim of this study in Tunisia was to classify ketosis-onset diabetes in adult patients. All patients aged > 30 years without known diabetes, presenting with ketosis and admitted to our department were studied. Patients with secondary or gestational diabetes and those on corticoid therapy or with coinciding infection were excluded. The data included clinical characteristics, immunological markers and beta-cell function. Of the 63 patients, islet-cell antibodies were present in 27.0%, glutamic acid decarboxylase antibodies in 25.4% and thyrosin phosphatase antibodies in 19.0%. Beta-cell functional reserve was preserved in 54.0%. Our results confirm that patients with ketosis-onset diabetes mellitus in adulthood are a heterogeneous group. PMID:20214161

  5. Efficient (18)F-Labeling of Synthetic Exendin-4 Analogues for Imaging Beta Cells.

    PubMed

    Keliher, Edmund J; Reiner, Thomas; Thurber, Greg M; Upadhyay, Rabi; Weissleder, Ralph

    2012-08-01

    A number of exendin derivatives have been developed to target glucagon-like peptide 1 (GLP-1) receptors on beta cells in vivo. Modifications of exendin analogues have been shown to have significant effects on pharmacokinetics and, as such, have been used to develop a variety of therapeutic compounds. Here, we show that an exendin-4, modified at position 12 with a cysteine conjugated to a tetrazine, can be labeled with (18)F-trans-cyclooctene and converted into a PET imaging agent at high yields and with good selectivity. The agent accumulates in beta cells in vivo and has sufficiently high accumulation in mouse models of insulinomas to enable in vivo imaging. PMID:23997998

  6. Efficient 18F-Labeling of Synthetic Exendin-4 Analogues for Imaging Beta Cells

    PubMed Central

    Keliher, Edmund J; Reiner, Thomas; Thurber, Greg M; Upadhyay, Rabi; Weissleder, Ralph

    2012-01-01

    A number of exendin derivatives have been developed to target glucagon-like peptide 1 (GLP-1) receptors on beta cells in vivo. Modifications of exendin analogues have been shown to have significant effects on pharmacokinetics and, as such, have been used to develop a variety of therapeutic compounds. Here, we show that an exendin-4, modified at position 12 with a cysteine conjugated to a tetrazine, can be labeled with 18F-trans-cyclooctene and converted into a PET imaging agent at high yields and with good selectivity. The agent accumulates in beta cells in vivo and has sufficiently high accumulation in mouse models of insulinomas to enable in vivo imaging. PMID:23997998

  7. Rapid regulation of pancreatic alpha- and beta- cell signalling systems by estrogens.

    PubMed

    Ripoll, Cristina; Ropero, Ana B; Alonso-Magdalena, Paloma; Quesada, Ivan; Fuentes, Esther; Nadal, Angel

    2008-03-01

    Rapid estrogen actions are triggered after estrogens are bound to a variety of proteins in organelles other than the nucleus. Those include classic estrogen receptors ERalpha and ERbeta, novel membrane proteins that behave as estrogen receptors such as GPR30, ion channels, and other ligand receptors. In pancreatic alpha and beta-cells, estrogens binding to a non-classical membrane estrogen receptors at physiological concentrations regulate ion channels and [Ca(2+)](i) signals, provoking important physiological responses. In beta-cells, 17beta-estradiol regulates K(ATP) channel activity and glucose-induced [Ca(2+)](i) oscillations, eliciting changes in insulin release and the activation of Ca(2+)-dependent transcription factors. In alpha-cells, 17beta-estradiol abolishes low glucose-induced [Ca(2+)](i) oscillations.

  8. Glucose-modulated Tyrosine Nitration in Beta-Cells: Targets and Consequences

    PubMed Central

    Koeck, Thomas; Corbett, John A.; Crabb, John W.; Stuehr, Dennis J.; Aulak, Kulwant S.

    2009-01-01

    Hyperglycemia, key factor of the pre-diabetic and diabetic pathology, is associated with cellular oxidative stress that promotes oxidative protein modifications. We report that protein nitration is responsive to changes in glucose concentrations in islets of Langerhans and insulinoma beta-cells. Alterations in the extent of tyrosine nitration as well as the cellular nitroproteome profile correlated tightly with changing glucose concentrations. The target proteins we identified function in protein folding, energy metabolism, antioxidant capacity, and membrane permeability. Nitration of heat shock protein 60 in vitro was found to decrease its ATP hydrolysis and interaction with proinsulin, suggesting a mechanism by which protein nitration could diminish insulin secretion. This was supported by our finding of a decrease in stimulated insulin secretion following glycolytic stress in cultured cells. Our results reveal that protein tyrosine nitration may be a previously unrecognized factor in beta-cell dysfunction and the pathogenesis of diabetes. PMID:19402213

  9. Immune responses to an encapsulated allogeneic islet {beta}-cell line in diabetic NOD mice

    SciTech Connect

    Black, Sasha P. . E-mail: Sasha.Black@ca.crl.com; Constantinidis, Ioannis; Cui, Hong; Tucker-Burden, Carol; Weber, Collin J.; Safley, Susan A.

    2006-02-03

    Our goal is to develop effective islet grafts for treating type 1 diabetes. Since human islets are scarce, we evaluated the efficacy of a microencapsulated insulin-secreting conditionally transformed allogeneic {beta}-cell line ({beta}TC-tet) in non-obese diabetic mice treated with tetracycline to inhibit cell growth. Relatively low serum levels of tetracycline controlled proliferation of {beta}TC-tet cells without inhibiting effective control of hyperglycemia in recipients. There was no significant host cellular reaction to the allografts or host cell adherence to microcapsules, and host cytokine levels were similar to those of sham-operated controls. We conclude that encapsulated allogeneic {beta}-cell lines may be clinically relevant, because they effectively restore euglycemia and do not elicit a strong cellular immune response following transplantation. To our knowledge, this is First extensive characterization of the kinetics of host cellular and cytokine responses to an encapsulated islet cell line in an animal model of type 1 diabetes.

  10. LGR5 and Nanog identify stem cell signature of pancreas beta cells which initiate pancreatic cancer.

    PubMed

    Amsterdam, Abraham; Raanan, Calanit; Schreiber, Letizia; Polin, Nava; Givol, David

    2013-04-01

    Pancreas cancer, is the fourth leading cause of cancer death but its cell of origin is controversial. We compared the localization of stem cells in normal and cancerous pancreas using antibodies to the stem cell markers Nanog and LGR5. Here we show, for the first time, that LGR5 is expressed in normal pancreas, exclusively in the islets of Langerhans and it is co-localized, surprisingly, with Nanog and insulin in clusters of beta cells. In cancerous pancreas Nanog and LGR5 are expressed in the remaining islets and in all ductal cancer cells. We observed insulin staining among the ductal cancer cells, but not in metastases. This indicates that the islet's beta cells, expressing LGR5 and Nanog markers are the initiating cells of pancreas cancer, which migrated from the islets to form the ductal cancerous tissue, probably after mutation and de-differentiation. This discovery may facilitate treatment of this devastating cancer.

  11. ER stress in pancreatic beta cells: the thin red line between adaptation and failure.

    PubMed

    Eizirik, Decio L; Cnop, Miriam

    2010-01-01

    Secretory cells, such as pancreatic beta cells, face the challenge of increasing protein synthesis severalfold during acute or chronic stimulation. This poses a burden on the endoplasmic reticulum (ER), the organelle where proinsulin synthesis and folding takes place. Thus, beta cells use various adaptive mechanisms to adjust the functional capacity of the ER to the prevailing demand. These check-and-balance mechanisms are collectively known as the unfolded protein response (UPR). It remains unclear how UPR signaling is ultimately regulated and what delineates the boundaries between a physiological and a pathological response. New discoveries point to the divergent effects of acute and chronic metabolic fluxes and chemical ER stressors on the formation of complexes among UPR transducers, scaffold proteins, and phosphatases. These and other findings provide a first glimpse on how different signals trigger diverging UPR outcomes. PMID:20179270

  12. Insulin granule trafficking in beta-cells: mathematical model of glucose-induced insulin secretion.

    PubMed

    Bertuzzi, Alessandro; Salinari, Serenella; Mingrone, Geltrude

    2007-07-01

    A mathematical model that represents the dynamics of intracellular insulin granules in beta-cells is proposed. Granule translocation and exocytosis are controlled by signals assumed to be essentially related to ATP-to-ADP ratio and cytosolic Ca(2+) concentration. The model provides an interpretation of the roles of the triggering and amplifying pathways of glucose-stimulated insulin secretion. Values of most of the model parameters were inferred from available experimental data. The numerical simulations represent a variety of experimental conditions, such as the stimulation by high K(+) and by different time courses of extracellular glucose, and the predicted responses agree with published experimental data. Model capacity to represent data measured in a hyperglycemic clamp was also tested. Model parameter changes that may reflect alterations of beta-cell function present in type 2 diabetes are investigated, and the action of pharmacological agents that bind to sulfonylurea receptors is simulated.

  13. Glucocorticoids Inhibit Basal and Hormone-Induced Serotonin Synthesis in Pancreatic Beta Cells.

    PubMed

    Hasni Ebou, Moina; Singh-Estivalet, Amrit; Launay, Jean-Marie; Callebert, Jacques; Tronche, François; Ferré, Pascal; Gautier, Jean-François; Guillemain, Ghislaine; Bréant, Bernadette; Blondeau, Bertrand; Riveline, Jean-Pierre

    2016-01-01

    Diabetes is a major complication of chronic Glucocorticoids (GCs) treatment. GCs induce insulin resistance and also inhibit insulin secretion from pancreatic beta cells. Yet, a full understanding of this negative regulation remains to be deciphered. In the present study, we investigated whether GCs could inhibit serotonin synthesis in beta cell since this neurotransmitter has been shown to be involved in the regulation of insulin secretion. To this aim, serotonin synthesis was evaluated in vitro after treatment with GCs of either islets from CD1 mice or MIN6 cells, a beta-cell line. We also explored the effect of GCs on the stimulation of serotonin synthesis by several hormones such as prolactin and GLP 1. We finally studied this regulation in islet in two in vivo models: mice treated with GCs and with liraglutide, a GLP1 analog, and mice deleted for the glucocorticoid receptor in the pancreas. We showed in isolated islets and MIN6 cells that GCs decreased expression and activity of the two key enzymes of serotonin synthesis, Tryptophan Hydroxylase 1 (Tph1) and 2 (Tph2), leading to reduced serotonin contents. GCs also blocked the induction of serotonin synthesis by prolactin or by a previously unknown serotonin activator, the GLP-1 analog exendin-4. In vivo, activation of the Glucagon-like-Peptide-1 receptor with liraglutide during 4 weeks increased islet serotonin contents and GCs treatment prevented this increase. Finally, islets from mice deleted for the GR in the pancreas displayed an increased expression of Tph1 and Tph2 and a strong increased serotonin content per islet. In conclusion, our results demonstrate an original inhibition of serotonin synthesis by GCs, both in basal condition and after stimulation by prolactin or activators of the GLP-1 receptor. This regulation may contribute to the deleterious effects of GCs on beta cells.

  14. Age-related differences in the pancreatic beta-cell response to hyperglycemia after eccentric exercise.

    PubMed

    Krishnan, R K; Hernandez, J M; Williamson, D L; O'Gorman, D J; Evans, W J; Kirwan, J P

    1998-09-01

    Eccentric exercise (ECC) causes muscle damage, insulin resistance, and increased pancreatic beta-cell secretion in young individuals. However, the effects of age on the pancreatic beta-cell response to glucose after ECC are unknown. Hyperglycemic clamps (180 min, 10.0 mM) were performed on eight young (age 22 +/- 1 yr) and eight older (age 66 +/- 2 yr) healthy sedentary males without exercise (CONT) and 48 h after ECC. ECC increased (P < 0.02) muscle soreness ratings and plasma creatine kinase concentrations in both groups. Insulin and C-peptide secretions were similar between young and older subjects during CONT clamps. ECC increased (P < 0.05) first-phase (0-10 min) C-peptide area under the curve in young (4.2 +/- 0.4 vs. 3.7 +/- 0.6 nM . min; ECC vs. CONT, respectively) but not in older subjects (3.2 +/- 0.7 vs. 3.5 +/- 0.7 nM . min; ECC vs. CONT), with significant group differences (P < 0.02). Indeed, ECC repressed (P < 0.05) first-phase peak C-peptide concentrations in older subjects (0. 93 +/- 0.16 vs. 1.12 +/- 0.11 nM; ECC vs. CONT). Moreover, first-phase C-peptide-to-insulin molar ratios suggest age-related differences (P < 0.05) in insulin/C-peptide clearance after ECC. Furthermore, the observed C-peptide response after ECC was related to abdominal adiposity [r = -0.62, P < 0.02, and r = -0.66, P < 0. 006, for first and second (10-180 min) phases, respectively]. In conclusion, older individuals did not exhibit the compensatory increase in beta-cell secretion observed among young individuals after ECC. Thus, with increasing age, the pancreatic beta-cell may be less responsive to the physiological stress associated with ECC. PMID:9725813

  15. Low Adiponectin Concentration in Pregnancy Predicts Postpartum Insulin Resistance, Beta-cell Dysfunction, and Fasting Glycaemia

    PubMed Central

    Retnakaran, R; Qi, Y; Connelly, PW; Sermer, M; Hanley, AJ; Zinman, B

    2010-01-01

    Aims/Hypothesis The postpartum following gestational diabetes (GDM) is characterized by subtle metabolic defects, including beta-cell dysfunction that is believed to mediate the increased future risk of type 2 diabetes in this patient population. Recently, low circulating levels of adiponectin and increased leptin and C-reactive protein (CRP) have emerged as novel diabetic risk factors, although their relevance to GDM and subsequent diabetes has not been characterized. Thus, we sought to determine whether adiponectin, leptin and CRP in pregnancy relate to the postpartum metabolic defects linking GDM with type 2 diabetes. Methods 487 women underwent metabolic characterization, including oral glucose tolerance test (OGTT), in pregnancy and at 3-months postpartum. Based on the antepartum OGTT, there were 137 women with GDM, 91 with gestational impaired glucose tolerance, and 259 with normal glucose tolerance. Results Adiponectin levels were lowest (p<0.0001) and CRP levels highest (p=0.0008) in women with GDM. Leptin did not differ between the glucose tolerance groups (p=0.4483). Adiponectin (r=0.41,p<0.0001), leptin (r=−0.36,p<0.0001) and CRP (r=−0.30,p<0.0001) in pregnancy were all associated with postpartum insulin sensitivity (ISOGTT). Intriguingly, adiponectin was also related to postpartum beta-cell function (insulinogenic index/HOMA-IR) (r=0.16,p=0.0009). Indeed, on multiple linear regression analyses, adiponectin in pregnancy independently predicted both postpartum insulin sensitivity (t=3.97,p<0.0001) and beta-cell function (t=2.37,p=0.0181), even after adjustment for GDM. Furthermore, adiponectin emerged as a significant negative independent determinant of postpartum fasting glucose (t=−3.01,p=0.0027). Conclusions Hypoadiponectinemia in pregnancy predicts postpartum insulin resistance, beta-cell dysfunction, and fasting glycaemia, and hence may be relevant to the pathophysiology relating GDM with type 2 diabetes. PMID:19937225

  16. Sodium arsenite impairs insulin secretion and transcription in pancreatic {beta}-cells

    SciTech Connect

    Diaz-Villasenor, Andrea; Sanchez-Soto, M. Carmen; Cebrian, Mariano E.; Ostrosky-Wegman, Patricia; Hiriart, Marcia . E-mail: mhiriart@ifc.unam.mx

    2006-07-01

    Human studies have shown that chronic inorganic arsenic (iAs) exposure is associated with a high prevalence and incidence of type 2 diabetes. However, the mechanism(s) underlying this effect are not well understood, and practically, there is no information available on the effects of arsenic on pancreatic {beta}-cells functions. Thus, since insulin secreted by the pancreas plays a crucial role in maintaining glucose homeostasis, our aim was to determine if sodium arsenite impairs insulin secretion and mRNA expression in single adult rat pancreatic {beta}-cells. Cells were treated with 0.5, 1, 2, 5 and 10 {mu}M sodium arsenite and incubated for 72 and 144 h. The highest dose tested (10 {mu}M) decreased {beta}-cell viability, by 33% and 83%, respectively. Insulin secretion and mRNA expression were evaluated in the presence of 1 and 5 {mu}M sodium arsenite. Basal insulin secretion, in 5.6 mM glucose, was not significantly affected by 1 or 5 {mu}M treatment for 72 h, but basal secretion was reduced when cells were exposed to 5 {mu}M sodium arsenite for 144 h. On the other hand, insulin secretion in response to 15.6 mM glucose decreased with sodium arsenite in a dose-dependent manner in such a way that cells were no longer able to distinguish between different glucose concentrations. We also showed a significant decrease in insulin mRNA expression of cells exposed to 5 {mu}M sodium arsenite during 72 h. Our data suggest that arsenic may contribute to the development of diabetes mellitus by impairing pancreatic {beta}-cell functions, particularly insulin synthesis and secretion.

  17. Glucocorticoids Inhibit Basal and Hormone-Induced Serotonin Synthesis in Pancreatic Beta Cells

    PubMed Central

    Hasni Ebou, Moina; Singh-Estivalet, Amrit; Launay, Jean-Marie; Callebert, Jacques; Tronche, François; Ferré, Pascal; Gautier, Jean-François; Guillemain, Ghislaine; Bréant, Bernadette

    2016-01-01

    Diabetes is a major complication of chronic Glucocorticoids (GCs) treatment. GCs induce insulin resistance and also inhibit insulin secretion from pancreatic beta cells. Yet, a full understanding of this negative regulation remains to be deciphered. In the present study, we investigated whether GCs could inhibit serotonin synthesis in beta cell since this neurotransmitter has been shown to be involved in the regulation of insulin secretion. To this aim, serotonin synthesis was evaluated in vitro after treatment with GCs of either islets from CD1 mice or MIN6 cells, a beta-cell line. We also explored the effect of GCs on the stimulation of serotonin synthesis by several hormones such as prolactin and GLP 1. We finally studied this regulation in islet in two in vivo models: mice treated with GCs and with liraglutide, a GLP1 analog, and mice deleted for the glucocorticoid receptor in the pancreas. We showed in isolated islets and MIN6 cells that GCs decreased expression and activity of the two key enzymes of serotonin synthesis, Tryptophan Hydroxylase 1 (Tph1) and 2 (Tph2), leading to reduced serotonin contents. GCs also blocked the induction of serotonin synthesis by prolactin or by a previously unknown serotonin activator, the GLP-1 analog exendin-4. In vivo, activation of the Glucagon-like-Peptide-1 receptor with liraglutide during 4 weeks increased islet serotonin contents and GCs treatment prevented this increase. Finally, islets from mice deleted for the GR in the pancreas displayed an increased expression of Tph1 and Tph2 and a strong increased serotonin content per islet. In conclusion, our results demonstrate an original inhibition of serotonin synthesis by GCs, both in basal condition and after stimulation by prolactin or activators of the GLP-1 receptor. This regulation may contribute to the deleterious effects of GCs on beta cells. PMID:26901633

  18. Vanadyl Sulfate Treatment Stimulates Proliferation and Regeneration of Beta Cells in Pancreatic Islets

    PubMed Central

    Missaoui, Samira; Ben Rhouma, Khémais; Yacoubi, Mohamed-Tahar; Sakly, Mohsen; Tebourbi, Olfa

    2014-01-01

    We examined the effects of vanadium sulfate (VOSO4) treatment at 5 and 10 mg/kg for 30 days on endocrine pancreas activity and histology in nondiabetic and STZ-induced diabetic rats. In diabetic group, blood glucose levels significantly increased while insulinemia level markedly decreased. At the end of treatment, VOSO4 at a dose of 10 mg/Kg normalized blood glucose level in diabetic group, restored insulinemia, and significantly improved insulin sensitivity. VOSO4 also increased in a dose-dependent manner the number of insulin immunopositive beta cells in pancreatic islets of nondiabetic rats. Furthermore, in the STZ-diabetic group, the decrease in the number of insulin immunopositive beta cells was corrected to reach the control level mainly with the higher dose of vanadium. Therefore, VOSO4 treatment normalized plasma glucose and insulin levels and improved insulin sensitivity in STZ-experimental diabetes and induced beta cells proliferation and/or regeneration in normal or diabetic rats. PMID:25215302

  19. The Microtubule-Associated Protein Tau and Its Relevance for Pancreatic Beta Cells.

    PubMed

    Maj, Magdalena; Hoermann, Gregor; Rasul, Sazan; Base, Wolfgang; Wagner, Ludwig; Attems, Johannes

    2016-01-01

    Structural and biochemical alterations of the microtubule-associated protein tau (MAPT) are associated with degenerative disorders referred to as tauopathies. We have previously shown that MAPT is present in human islets of Langerhans, human insulinomas, and pancreatic beta-cell line models, with biophysical similarities to the pathological MAPT in the brain. Here, we further studied MAPT in pancreatic endocrine tissue to better understand the mechanisms that lead to functional dysregulation of pancreatic beta cells. We found upregulation of MAPT protein expression in human insulinomas when compared to human pancreatic islets of Langerhans and an imbalance between MAPT isoforms in insulinomas tissue. We cloned one 3-repeat domain MAPT and transduced this into a beta-cell derived rodent cell line Rin-5F. Proliferation experiments showed higher growth rates and metabolic activities of cells overexpressing MAPT protein. We observed that a MAPT overexpressing cell line demonstrates altered insulin transcription, translation, and insulin secretion rates. We found the relative insulin secretion rates were significantly decreased in a MAPT overexpressing cell line and these findings could be confirmed using partial MAPT knock-down cell lines. Our findings support that MAPT may play an important role in insulin granule trafficking and indicate the importance of balanced MAPT phosphorylation and dephosphorylation for adequate insulin release.

  20. Vanadyl sulfate treatment stimulates proliferation and regeneration of beta cells in pancreatic islets.

    PubMed

    Missaoui, Samira; Ben Rhouma, Khémais; Yacoubi, Mohamed-Tahar; Sakly, Mohsen; Tebourbi, Olfa

    2014-01-01

    We examined the effects of vanadium sulfate (VOSO4) treatment at 5 and 10 mg/kg for 30 days on endocrine pancreas activity and histology in nondiabetic and STZ-induced diabetic rats. In diabetic group, blood glucose levels significantly increased while insulinemia level markedly decreased. At the end of treatment, VOSO4 at a dose of 10 mg/Kg normalized blood glucose level in diabetic group, restored insulinemia, and significantly improved insulin sensitivity. VOSO4 also increased in a dose-dependent manner the number of insulin immunopositive beta cells in pancreatic islets of nondiabetic rats. Furthermore, in the STZ-diabetic group, the decrease in the number of insulin immunopositive beta cells was corrected to reach the control level mainly with the higher dose of vanadium. Therefore, VOSO4 treatment normalized plasma glucose and insulin levels and improved insulin sensitivity in STZ-experimental diabetes and induced beta cells proliferation and/or regeneration in normal or diabetic rats.

  1. GLP-1 receptor antagonist as a potential probe for pancreatic {beta}-cell imaging

    SciTech Connect

    Mukai, Eri; Toyoda, Kentaro; Kimura, Hiroyuki; Kawashima, Hidekazu; Fujimoto, Hiroyuki; Ueda, Masashi; Temma, Takashi; Hirao, Konomu; Nagakawa, Kenji; Saji, Hideo; Inagaki, Nobuya

    2009-11-20

    We examined exendin(9-39), an antagonist of glucagon-like peptide-1 (GLP-1) receptor (GLP-1R), as a potential probe for imaging of pancreatic {beta}-cells. To evaluate in vitro receptor specificity, binding assay was performed using dispersed mouse islet cells. Binding assay showed competitive inhibition of [{sup 125}I]BH-exendin(9-39) binding by non-radioactive exendin(9-39). To assess in vivo selectivity, the biodistribution was evaluated by intravenous administration of [{sup 125}I]BH-exendin(9-39) to mice. Radioactivity of harvested pancreas reached highest levels at 60 and 120 min among organs examined except lung. Pre-administration of excess non-radioactive exendin(9-39) remarkably and specifically blocked the radioactivity of pancreas. After [{sup 125}I]BH-exendin(9-39) injection into transgenic mice with pancreatic {beta}-cells expressing GFP, fluorescent and radioactive signals of sections of pancreas were evaluated with an image analyzer. Imaging analysis showed that the fluorescent GFP signals and the radioactive signals were correspondingly located. Thus, the GLP-1R antagonist exendin(9-39) may serve as a useful probe for pancreatic {beta}-cell imaging.

  2. Calcium co-regulates oxidative metabolism and ATP synthase-dependent respiration in pancreatic beta cells.

    PubMed

    De Marchi, Umberto; Thevenet, Jonathan; Hermant, Aurelie; Dioum, Elhadji; Wiederkehr, Andreas

    2014-03-28

    Mitochondrial energy metabolism is essential for glucose-induced calcium signaling and, therefore, insulin granule exocytosis in pancreatic beta cells. Calcium signals are sensed by mitochondria acting in concert with mitochondrial substrates for the full activation of the organelle. Here we have studied glucose-induced calcium signaling and energy metabolism in INS-1E insulinoma cells and human islet beta cells. In insulin secreting cells a surprisingly large fraction of total respiration under resting conditions is ATP synthase-independent. We observe that ATP synthase-dependent respiration is markedly increased after glucose stimulation. Glucose also causes a very rapid elevation of oxidative metabolism as was followed by NAD(P)H autofluorescence. However, neither the rate of the glucose-induced increase nor the new steady-state NAD(P)H levels are significantly affected by calcium. Our findings challenge the current view, which has focused mainly on calcium-sensitive dehydrogenases as the target for the activation of mitochondrial energy metabolism. We propose a model of tight calcium-dependent regulation of oxidative metabolism and ATP synthase-dependent respiration in beta cell mitochondria. Coordinated activation of matrix dehydrogenases and respiratory chain activity by calcium allows the respiratory rate to change severalfold with only small or no alterations of the NAD(P)H/NAD(P)(+) ratio.

  3. The Microtubule-Associated Protein Tau and Its Relevance for Pancreatic Beta Cells

    PubMed Central

    Maj, Magdalena; Hoermann, Gregor; Rasul, Sazan; Base, Wolfgang; Wagner, Ludwig; Attems, Johannes

    2016-01-01

    Structural and biochemical alterations of the microtubule-associated protein tau (MAPT) are associated with degenerative disorders referred to as tauopathies. We have previously shown that MAPT is present in human islets of Langerhans, human insulinomas, and pancreatic beta-cell line models, with biophysical similarities to the pathological MAPT in the brain. Here, we further studied MAPT in pancreatic endocrine tissue to better understand the mechanisms that lead to functional dysregulation of pancreatic beta cells. We found upregulation of MAPT protein expression in human insulinomas when compared to human pancreatic islets of Langerhans and an imbalance between MAPT isoforms in insulinomas tissue. We cloned one 3-repeat domain MAPT and transduced this into a beta-cell derived rodent cell line Rin-5F. Proliferation experiments showed higher growth rates and metabolic activities of cells overexpressing MAPT protein. We observed that a MAPT overexpressing cell line demonstrates altered insulin transcription, translation, and insulin secretion rates. We found the relative insulin secretion rates were significantly decreased in a MAPT overexpressing cell line and these findings could be confirmed using partial MAPT knock-down cell lines. Our findings support that MAPT may play an important role in insulin granule trafficking and indicate the importance of balanced MAPT phosphorylation and dephosphorylation for adequate insulin release. PMID:26824039

  4. Matrix metalloproteinase-9 is essential for physiological Beta cell function and islet vascularization in adult mice.

    PubMed

    Christoffersson, Gustaf; Waldén, Tomas; Sandberg, Monica; Opdenakker, Ghislain; Carlsson, Per-Ola; Phillipson, Mia

    2015-04-01

    The availability of paracrine factors in the islets of Langerhans, and the constitution of the beta cell basement membrane can both be affected by proteolytic enzymes. This study aimed to investigate the effects of the extracellular matrix-degrading enzyme gelatinase B/matrix metalloproteinase-9 (Mmp-9) on islet function in mice. Islet function of Mmp9-deficient (Mmp9(-/-)) mice and their wild-type littermates was evaluated both in vivo and in vitro. The pancreata of Mmp9(-/-) mice did not differ from wild type in islet mass or distribution. However, Mmp9(-/-) mice had an impaired response to a glucose load in vivo, with lower serum insulin levels. The glucose-stimulated insulin secretion was reduced also in vitro in isolated Mmp9(-/-) islets. The vascular density of Mmp9(-/-) islets was lower, and the capillaries had fewer fenestrations, whereas the islet blood flow was threefold higher. These alterations could partly be explained by compensatory changes in the expression of matrix-related proteins. This in-depth investigation of the effects of the loss of MMP-9 function on pancreatic islets uncovers a deteriorated beta cell function that is primarily due to a shift in the beta cell phenotype, but also due to islet vascular aberrations. This likely reflects the importance of a normal islet matrix turnover exerted by MMP-9, and concomitant release of paracrine factors sequestered on the matrix.

  5. Early-life origins of type 2 diabetes: fetal programming of the beta-cell mass.

    PubMed

    Portha, Bernard; Chavey, Audrey; Movassat, Jamileh

    2011-01-01

    A substantial body of evidence suggests that an abnormal intrauterine milieu elicited by maternal metabolic disturbances as diverse as undernutrition, placental insufficiency, diabetes or obesity, may program susceptibility in the fetus to later develop chronic degenerative diseases, such as obesity, hypertension, cardiovascular diseases and diabetes. This paper examines the developmental programming of glucose intolerance/diabetes by disturbed intrauterine metabolic condition experimentally obtained in various rodent models of maternal protein restriction, caloric restriction, overnutrition or diabetes, with a focus on the alteration of the developing beta-cell mass. In most of the cases, whatever the type of initial maternal metabolic stress, the beta-cell adaptive growth which normally occurs during gestation, does not take place in the pregnant offspring and this results in the development of gestational diabetes. Therefore gestational diabetes turns to be the ultimate insult targeting the offspring beta-cell mass and propagates diabetes risk to the next generation again. The aetiology and the transmission of spontaneous diabetes as encountered in the GK/Par rat model of type 2 diabetes, are discussed in such a perspective. This review also discusses the non-genomic mechanisms involved in the installation of the programmed effect as well as in its intergenerational transmission.

  6. Impact of fetal and neonatal environment on beta cell function and development of diabetes.

    PubMed

    Nielsen, Jens H; Haase, Tobias N; Jaksch, Caroline; Nalla, Amarnadh; Søstrup, Birgitte; Nalla, Anjana A; Larsen, Louise; Rasmussen, Morten; Dalgaard, Louise T; Gaarn, Louise W; Thams, Peter; Kofod, Hans; Billestrup, Nils

    2014-11-01

    The global epidemic of diabetes is a serious threat against health and healthcare expenses. Although genetics is important it does not explain the dramatic increase in incidence, which must involve environmental factors. Two decades ago the concept of the thrifty phenotype was introduced, stating that the intrauterine environment during pregnancy has an impact on the gene expression that may persist until adulthood and cause metabolic diseases like obesity and type 2 diabetes. As the pancreatic beta cells are crucial in the regulation of metabolism this article will describe the influence of normal pregnancy on the beta cells in both the mother and the fetus and how various conditions like diabetes, obesity, overnutrition and undernutrition during and after pregnancy may influence the ability of the offspring to adapt to changes in insulin demand later in life. The influence of environmental factors including nutrients and gut microbiota on appetite regulation, mitochondrial activity and the immune system that may affect beta cell growth and function directly and indirectly is discussed. The possible role of epigenetic changes in the transgenerational transmission of the adverse programming may be the most threatening aspect with regard to the global diabetes epidemics. Finally, some suggestions for intervention are presented.

  7. Islet Autoimmunity Identifies a Unique Pattern of Impaired Pancreatic Beta-Cell Function, Markedly Reduced Pancreatic Beta Cell Mass and Insulin Resistance in Clinically Diagnosed Type 2 Diabetes

    PubMed Central

    Subauste, Angela; Gianani, Roberto; Chang, Annette M.; Plunkett, Cynthia; Pietropaolo, Susan L.; Zhang, Ying-Jian; Barinas-Mitchell, Emma; Kuller, Lewis H.; Galecki, Andrzej; Halter, Jeffrey B.; Pietropaolo, Massimo

    2014-01-01

    There is a paucity of literature describing metabolic and histological data in adult-onset autoimmune diabetes. This subgroup of diabetes mellitus affects at least 5% of clinically diagnosed type 2 diabetic patients (T2DM) and it is termed Latent Autoimmune Diabetes in Adults (LADA). We evaluated indexes of insulin secretion, metabolic assessment, and pancreatic pathology in clinically diagnosed T2DM patients with and without the presence of humoral islet autoimmunity (Ab). A total of 18 patients with at least 5-year duration of clinically diagnosed T2DM were evaluated in this study. In those subjects we assessed acute insulin responses to arginine, a glucose clamp study, whole-body fat mass and fat-free mass. We have also analyzed the pancreatic pathology of 15 T2DM and 43 control cadaveric donors, using pancreatic tissue obtained from all the T2DM organ donors available from the nPOD network through December 31, 2013. The presence of islet Ab correlated with severely impaired β-cell function as demonstrated by remarkably low acute insulin response to arginine (AIR) when compared to that of the Ab negative group. Glucose clamp studies indicated that both Ab positive and Ab negative patients exhibited peripheral insulin resistance in a similar fashion. Pathology data from T2DM donors with Ab or the autoimmune diabetes associated DR3/DR4 allelic class II combination showed reduction in beta cell mass as well as presence of autoimmune-associated pattern A pathology in subjects with either islet autoantibodies or the DR3/DR4 genotype. In conclusion, we provide compelling evidence indicating that islet Ab positive long-term T2DM patients exhibit profound impairment of insulin secretion as well as reduced beta cell mass seemingly determined by an immune-mediated injury of pancreatic β-cells. Deciphering the mechanisms underlying beta cell destruction in this subset of diabetic patients may lead to the development of novel immunologic therapies aimed at halting the

  8. TIRF imaging of docking and fusion of single insulin granule motion in primary rat pancreatic beta-cells: different behaviour of granule motion between normal and Goto-Kakizaki diabetic rat beta-cells.

    PubMed

    Ohara-Imaizumi, Mica; Nishiwaki, Chiyono; Kikuta, Toshiteru; Nagai, Shintaro; Nakamichi, Yoko; Nagamatsu, Shinya

    2004-07-01

    We imaged and analysed the motion of single insulin secretory granules near the plasma membrane in live pancreatic beta-cells, from normal and diabetic Goto-Kakizaki (GK) rats, using total internal reflection fluorescence microscopy (TIRFM). In normal rat primary beta-cells, the granules that were fusing during the first phase originate from previously docked granules, and those during the second phase originate from 'newcomers'. In diabetic GK rat beta-cells, the number of fusion events from previously docked granules were markedly reduced, and, in contrast, the fusion from newcomers was still preserved. The dynamic change in the number of docked insulin granules showed that, in GK rat beta-cells, the total number of docked insulin granules was markedly decreased to 35% of the initial number after glucose stimulation. Immunohistochemistry with anti-insulin antibody observed by TIRFM showed that GK rat beta-cells had a marked decline of endogenous insulin granules docked to the plasma membrane. Thus our results indicate that the decreased number of docked insulin granules accounts for the impaired insulin release during the first phase of insulin release in diabetic GK rat beta-cells.

  9. A high-throughput chemical screen reveals that harmine-mediated inhibition of DYRK1A increases human pancreatic beta cell replication.

    PubMed

    Wang, Peng; Alvarez-Perez, Juan-Carlos; Felsenfeld, Dan P; Liu, Hongtao; Sivendran, Sharmila; Bender, Aaron; Kumar, Anil; Sanchez, Roberto; Scott, Donald K; Garcia-Ocaña, Adolfo; Stewart, Andrew F

    2015-04-01

    Types 1 and 2 diabetes affect some 380 million people worldwide. Both ultimately result from a deficiency of functional pancreatic insulin-producing beta cells. Beta cells proliferate in humans during a brief temporal window beginning around the time of birth, with a peak percentage (∼2%) engaged in the cell cycle in the first year of life. In embryonic life and after early childhood, beta cell replication is barely detectable. Whereas beta cell expansion seems an obvious therapeutic approach to beta cell deficiency, adult human beta cells have proven recalcitrant to such efforts. Hence, there remains an urgent need for antidiabetic therapeutic agents that can induce regeneration and expansion of adult human beta cells in vivo or ex vivo. Here, using a high-throughput small-molecule screen (HTS), we find that analogs of the small molecule harmine function as a new class of human beta cell mitogenic compounds. We also define dual-specificity tyrosine-regulated kinase-1a (DYRK1A) as the likely target of harmine and the nuclear factors of activated T cells (NFAT) family of transcription factors as likely mediators of human beta cell proliferation and differentiation. Using three different mouse and human islet in vivo-based models, we show that harmine is able to induce beta cell proliferation, increase islet mass and improve glycemic control. These observations suggest that harmine analogs may have unique therapeutic promise for human diabetes therapy. Enhancing the potency and beta cell specificity of these compounds are important future challenges. PMID:25751815

  10. Factors Associated with Beta-Cell Dysfunction in Type 2 Diabetes: The BETADECLINE Study

    PubMed Central

    Russo, Giuseppina T.; Giorda, Carlo Bruno; Cercone, Stefania; Nicolucci, Antonio; Cucinotta, Domenico

    2014-01-01

    Aims Beta-cell dysfunction is an early event in the natural history of type 2 diabetes. However, its progression is variable and potentially influenced by several clinical factors. We report the baseline data of the BetaDecline study, an Italian prospective multicenter study on clinical predictors of beta-cell dysfunction in type 2 diabetes. Materials and Methods Clinical, lifestyle, and laboratory data, including circulating levels of inflammatory markers and non-esterified fatty acids, were collected in 507 type 2 diabetic outpatients on stable treatment with oral hypoglycemic drugs or diet for more than 1 year. Beta-cell dysfunction was evaluated by calculating the proinsulin/insulin ratio (P/I). Results At baseline, the subjects in the upper PI/I ratio quartile were more likely to be men and receiving secretagogue drugs; they also showed a borderline longer diabetes duration (P = 0.06) and higher serum levels of glycated hemoglobin (HbA1c), fasting blood glucose, and triglycerides. An inverse trend across all PI/I quartiles was noted for BMI and serum levels of total cholesterol (T-C), LDL-C, HDL-C and C reactive protein (CRP), and with homeostatic model assessment (HOMA-B) and HOMA of insulin resistance (HOMA-IR) values (P<0.05 for all). At multivariate analysis, the risk of having a P/I ratio in the upper quartile was higher in the subjects on secretagogue drugs (odds ratio [OR] 4.2; 95% confidence interval [CI], 2.6–6.9) and in the males (OR 1.8; 95% CI, 1.1–2.9). Conclusions In the BetaDecline study population, baseline higher PI/I values, a marker of beta-cell dysfunction, were more frequent in men and in patients on secretagogues drugs. Follow-up of this cohort will allow the identification of clinical predictors of beta-cell failure in type 2 diabetic outpatients. PMID:25347846

  11. Modulation of the bursting properties of single mouse pancreatic beta-cells by artificial conductances.

    PubMed Central

    Kinard, T A; de Vries, G; Sherman, A; Satin, L S

    1999-01-01

    Glucose triggers bursting activity in pancreatic islets, which mediates the Ca2+ uptake that triggers insulin secretion. Aside from the channel mechanism responsible for bursting, which remains unsettled, it is not clear whether bursting is an endogenous property of individual beta-cells or requires an electrically coupled islet. While many workers report stochastic firing or quasibursting in single cells, a few reports describe single-cell bursts much longer (minutes) than those of islets (15-60 s). We studied the behavior of single cells systematically to help resolve this issue. Perforated patch recordings were made from single mouse beta-cells or hamster insulinoma tumor cells in current clamp at 30-35 degrees C, using standard K+-rich pipette solution and external solutions containing 11.1 mM glucose. Dynamic clamp was used to apply artificial KATP and Ca2+ channel conductances to cells in current clamp to assess the role of Ca2+ and KATP channels in single cell firing. The electrical activity we observed in mouse beta-cells was heterogeneous, with three basic patterns encountered: 1) repetitive fast spiking; 2) fast spikes superimposed on brief (<5 s) plateaus; or 3) periodic plateaus of longer duration (10-20 s) with small spikes. Pattern 2 was most similar to islet bursting but was significantly faster. Burst plateaus lasting on the order of minutes were only observed when recordings were made from cell clusters. Adding gCa to cells increased the depolarizing drive of bursting and lengthened the plateaus, whereas adding gKATP hyperpolarized the cells and lengthened the silent phases. Adding gCa and gKATP together did not cancel out their individual effects but could induce robust bursts that resembled those of islets, and with increased period. These added currents had no slow components, indicating that the mechanisms of physiological bursting are likely to be endogenous to single beta-cells. It is unlikely that the fast bursting (class 2) was due to

  12. Generation of Beta Cells from Human Pluripotent Stem Cells: Potential for Regenerative Medicine

    PubMed Central

    Nostro, Maria Cristina; Keller, Gordon

    2015-01-01

    The loss of beta cells in Type I Diabetes ultimately leads to insulin dependence and major complications that are difficult to manage by insulin injections. Given the complications associated with long-term administration of insulin, cell-replacement therapy is now under consideration as an alternative treatment that may someday provide a cure for this disease. Over the past 10 years, islet transplantation trials have demonstrated that it is possible to replenish beta cell function in Type I Diabetes patients and, at least temporarily, eliminate their dependency on insulin. While not yet optimal, the success of these trials has provided proof-of-principle that cell replacement therapy is a viable option for treating diabetes. Limited access to donor islets has launched a search for alternative source of beta cells for cell therapy purposes and focused the efforts of many investigators on the challenge of deriving such cells from human embryonic and induced pluripotent stem cells. Over the past five years, significant advances have been made in understanding the signaling pathways that control lineage development from hPSCs and as a consequence, it is now possible to routinely generate human insulin producing cells from both hESCs and hiPSCs. While these achievements are impressive, significant challenges do still exist, as the majority of insulin producing cells generated under these conditions are polyhormonal and non functional, likely reflecting the emergence of the polyhormonal population that is known to arise in the early embryo during the phase of pancreatic development known as the ‘first transition’. Functional beta cells, which arise during the second phase or transition of pancreatic development have been generated from hPSCs, however they are detected only following transplantation of progenitor stage cells into immunocompromised mice. With this success, our challenge now is to define the pathways that control the development and maturation of this

  13. Evidence for glucagon-like peptide-1 receptor signaling to activate ATP-sensitive potassium channels in pancreatic beta cells.

    PubMed

    Kwon, Hye-Jung; Park, Hyun-Sun; Park, Sung-Hee; Park, Jae-Hyung; Shin, Su-Kyung; Song, Seung Eun; Hwang, Meeyul; Cho, Ho-Chan; Song, Dae-Kyu

    2016-01-01

    Glucagon-like peptide-1 (GLP-1) is a gut peptide that promotes insulin release from pancreatic beta cells. GLP-1 has been shown to confer glucose-insensitive beta cells with glucose sensitivity by modulation of the activity of the ATP-sensitive potassium (KATP) channel. The channel closing effect of GLP-1, interacting with corresponding G-protein-coupled receptors, has been well established; however, to our knowledge, no study has shown whether GLP-1 directly induces activation of beta-cell KATP channels. Here, we aimed to evaluate whether the activation of beta-cell KATP channels by GLP-1 exists and affects intracellular Ca(2+) levels ([Ca(2+)]i). KATP channel activity was measured in isolated rat pancreatic beta cells by whole-cell perforated patch-clamp recordings with a diazoxide-containing pipette solution. Changes in [Ca(2+)]i and the subcellular localization of KATP channels were observed using the calcium-sensitive dye fura-4/AM and anti-Kir6.2 antibodies in INS-1 beta cells, respectively. To eliminate the well-known inhibitory effects of GLP-1 on KATP channel activity, channels were fully inhibited by pretreatment with methyl pyruvate and epigallocatechin-3-gallate. In the pretreated beta cells, GLP-1 and exendin-4 promptly activated the channels, reducing [Ca(2+)]i. The phosphoinositide 3-kinase (PI3K) inhibitor LY294002 blocked the effects of GLP-1 on channel activity. Moreover, phosphatidylinositol-3,4,5-trisphosphate mimicked the effects of GLP-1. These results suggested that beta-cell GLP-1 receptor signaling involved activation of KATP channels via a PI3K-dependent pathway. This alternative mechanism of GLP-1 function may act as a negative feedback pathway, modulating the glucose-dependent GLP-1 inhibition on KATP channel activity. PMID:26655814

  14. The quest to make fully functional human pancreatic beta cells from embryonic stem cells: climbing a mountain in the clouds.

    PubMed

    Johnson, James D

    2016-10-01

    The production of fully functional insulin-secreting cells to treat diabetes is a major goal of regenerative medicine. In this article, I review progress towards this goal over the last 15 years from the perspective of a beta cell biologist. I describe the current state-of-the-art, and speculate on the general approaches that will be required to identify and achieve our ultimate goal of producing functional beta cells. The need for deeper phenotyping of heterogeneous cultures of stem cell derived islet-like cells in parallel with a better understanding of the heterogeneity of the target cell type(s) is emphasised. This deep phenotyping should include high-throughput single-cell analysis, as well as comprehensive 'omics technologies to provide unbiased characterisation of cell products and human beta cells. There are justified calls for more detailed and well-powered studies of primary human pancreatic beta cell physiology, and I propose online databases of standardised human beta cell responses to physiological stimuli, including both functional and metabolomic/proteomic/transcriptomic profiles. With a concerted, community-wide effort, including both basic and applied scientists, beta cell replacement will become a clinical reality for patients with diabetes. PMID:27473069

  15. The quest to make fully functional human pancreatic beta cells from embryonic stem cells: climbing a mountain in the clouds.

    PubMed

    Johnson, James D

    2016-10-01

    The production of fully functional insulin-secreting cells to treat diabetes is a major goal of regenerative medicine. In this article, I review progress towards this goal over the last 15 years from the perspective of a beta cell biologist. I describe the current state-of-the-art, and speculate on the general approaches that will be required to identify and achieve our ultimate goal of producing functional beta cells. The need for deeper phenotyping of heterogeneous cultures of stem cell derived islet-like cells in parallel with a better understanding of the heterogeneity of the target cell type(s) is emphasised. This deep phenotyping should include high-throughput single-cell analysis, as well as comprehensive 'omics technologies to provide unbiased characterisation of cell products and human beta cells. There are justified calls for more detailed and well-powered studies of primary human pancreatic beta cell physiology, and I propose online databases of standardised human beta cell responses to physiological stimuli, including both functional and metabolomic/proteomic/transcriptomic profiles. With a concerted, community-wide effort, including both basic and applied scientists, beta cell replacement will become a clinical reality for patients with diabetes.

  16. Effect of Hypertriglyceridemia on Beta Cell Mass and Function in ApoC3 Transgenic Mice.

    PubMed

    Liu, Yun-Zi; Cheng, Xiaoyun; Zhang, Ting; Lee, Sojin; Yamauchi, Jun; Xiao, Xiangwei; Gittes, George; Qu, Shen; Jiang, Chun-Lei; Dong, H Henry

    2016-07-01

    Hypertriglyceridemia results from increased production and decreased clearance of triglyceride-rich very low-density lipoproteins, a pathological condition that accounts for heightened risk of ischemic vascular diseases in obesity and type 2 diabetes. Despite its intimate association with insulin resistance, whether hypertriglyceridemia constitutes an independent risk for beta cell dysfunction in diabetes is unknown. Answering this fundamental question is stymied by the fact that hypertriglyceridemia is intertwined with hyperglycemia and insulin resistance in obese and diabetic subjects. To circumvent this limitation, we took advantage of apolipoprotein C3 (ApoC3)-transgenic mice, a model with genetic predisposition to hypertriglyceridemia. We showed that ApoC3-transgenic mice, as opposed to age/sex-matched wild-type littermates, develop hypertriglyceridemia with concomitant elevations in plasma cholesterol and non-esterified fatty acid levels. Anti-insulin and anti-glucagon dual immunohistochemistry in combination with morphometric analysis revealed that ApoC3-transgenic and wild-type littermates had similar beta cell and alpha cell masses as well as islet size and architecture. These effects correlated with similar amplitudes of glucose-stimulated insulin secretion and similar degrees of postprandial glucose excursion in ApoC3-transgenic versus wild-type littermates. Oil Red O histology did not visualize lipid infiltration into islets, correlating with the lack of ectopic triglyceride and cholesterol depositions in the pancreata of ApoC3-transgenic versus wild-type littermates. ApoC3-transgenic mice, despite persistent hypertriglyceridemia, maintained euglycemia under both fed and fasting conditions without manifestation of insulin resistance and fasting hyperinsulinemia. Thus, hypertriglyceridemia per se is not an independent risk factor for beta cell dysfunction in ApoC3 transgenic mice. PMID:27226540

  17. Regulation of islet beta-cell pyruvate metabolism: interactions of prolactin, glucose, and dexamethasone.

    PubMed

    Arumugam, Ramamani; Horowitz, Eric; Noland, Robert C; Lu, Danhong; Fleenor, Donald; Freemark, Michael

    2010-07-01

    Prolactin (PRL) induces beta-cell proliferation and glucose-stimulated insulin secretion (GSIS) and counteracts the effects of glucocorticoids on insulin production. The mechanisms by which PRL up-regulates GSIS are unknown. We used rat islets and insulinoma (INS-1) cells to explore the interactions of PRL, glucose, and dexamethasone (DEX) in the regulation of beta-cell pyruvate carboxylase (PC), pyruvate dehydrogenase (PDH), and the pyruvate dehydrogenase kinases (PDKs), which catalyze the phosphorylation and inactivation of PDH. PRL increased GSIS by 37% (P < 0.001) in rat islets. Glucose at supraphysiological concentrations (11 mm) increased PC mRNA in islets; in contrast, PRL suppressed PC mRNA levels in islets and INS-1 cells, whereas DEX was without effect. Neither PRL nor DEX altered PC protein or activity levels. In INS-1 cells, PRL increased PDH activity 1.4- to 2-fold (P < 0.05-0.001) at glucose concentrations ranging from 2.5-11 mm. DEX reduced PDH activity; this effect was reversed by PRL. PDK1, -2, -3, and -4 mRNAs were detected in both islets and insulinoma cells, but the latter expressed trivial amounts of PDK4. PRL reduced PDK2 mRNA and protein levels in rat islets and INS-1 cells and PDK4 mRNA in islets; DEX increased PDK2 mRNA in islets and INS-1 cells; this effect was reversed by PRL. Our findings suggest that PRL induction of GSIS is mediated by increases in beta-cell PDH activity; this is facilitated by suppression of PDKs. PRL counteracts the effects of DEX on PDH and PDK expression, suggesting novel roles for the lactogens in the defense against diabetes. PMID:20484462

  18. Age-Related Impairment of Pancreatic Beta-Cell Function: Pathophysiological and Cellular Mechanisms

    PubMed Central

    De Tata, Vincenzo

    2014-01-01

    The incidence of type 2 diabetes significantly increases with age. The relevance of this association is dramatically magnified by the concomitant global aging of the population, but the underlying mechanisms remain to be fully elucidated. Here, some recent advances in this field are reviewed at the level of both the pathophysiology of glucose homeostasis and the cellular senescence of pancreatic islets. Overall, recent results highlight the crucial role of beta-cell dysfunction in the age-related impairment of pancreatic endocrine function and delineate the possibility of new original therapeutic interventions. PMID:25232350

  19. Suppression of Beta-cell secretion by somatostatin does not fully reverse the disproportionate proinsulinemia of type 2 diabetes.

    PubMed

    Røder, Michael E; Kahn, Steven E

    2004-12-01

    Disproportionate hyperproinsulinemia is a feature of beta-cell dysfunction in type 2 diabetes. It has been hypothesized that this abnormality represents an intrinsic abnormality of the beta-cell and/or may result from an increase in beta-cell secretory demand. To address this, six patients with type 2 diabetes and six age- and BMI-matched normal subjects received a combined 3-h insulin and somatostatin clamp to decrease beta-cell secretory demand. An arginine stimulation test was performed before and at the end of the clamp to measure beta-cell peptide release. In keeping with the reduction in secretory demand, C-peptide levels were suppressed by 60-80% during the clamp, as were proinsulin (PI) levels. The arginine-stimulated PI/C-peptide ratio decreased in the diabetic subjects from 4.4 +/- 1.5% before to 1.8 +/- 0.5% after the clamp (P < 0.01). This latter ratio was similar to that observed in the normal subjects before the somatostatin infusion (1.5 +/- 0.3%). In the normal subjects, after the clamp the PI/C-peptide ratio had decreased to 0.8 +/- 0.3% (P <0.01). Thus, the postclamp PI/C-peptide ratio in the subjects with type 2 diabetes was elevated compared with that in the normal subjects (P <0.05). Based on these observations, while relief of secretory demand on beta-cells by somatostatin decreases the disproportionate elevation in PI levels in patients with type 2 diabetes, the failure to normalize this measure suggests that an intrinsic abnormality of beta-cell function exists in subjects with type 2 diabetes that may be aggravated by increased secretory demand.

  20. Glucagon-like peptide-1 improves beta-cell antioxidant capacity via extracellular regulated kinases pathway and Nrf2 translocation.

    PubMed

    Fernández-Millán, E; Martín, M A; Goya, L; Lizárraga-Mollinedo, E; Escrivá, F; Ramos, S; Álvarez, C

    2016-06-01

    Oxidative stress plays an important role in the development of beta-cell dysfunction and insulin resistance, two major pathophysiological abnormalities of type 2 diabetes. Expression levels of antioxidant enzymes in beta cells are very low, rendering them more susceptible to damage caused by reactive oxygen species (ROS). Although the antioxidant effects of glucagon-like peptide-1 (GLP-1) and its analogs have been previously reported, the exact mechanisms involved are still unclear. In this study, we demonstrated that GLP-1 was able to effectively inhibit oxidative stress and cell death of INS-1E beta cells induced by the pro-oxidant tert-butyl hydroperoxide (tert-BOOH). Incubation with GLP-1 enhanced cellular levels of glutathione and the activity of its related enzymes, glutathione-peroxidase (GPx) and -reductase (GR) in beta cells. However, inhibition of ERK, but not of the PI3K/AKT pathway abolished, at least in part, the antioxidant effect of GLP-1. Moreover, ERK activation seems to be protein kinase A (PKA)-dependent because inhibition of PKA with H-89 was sufficient to block the GLP-1-derived protective effect on beta cells. GLP-1 likewise increased the synthesis of GR and favored the translocation of the nuclear transcription factor erythroid 2p45-related factor (Nrf2), a transcription factor implicated in the expression of several antioxidant/detoxificant enzymes. Glucose-stimulated insulin secretion was also preserved in beta-cells challenged with tert-BOOH but pre-treated with GLP-1, probably through the down-regulation of the mitochondrial uncoupling-protein2 (UCP2). Thus, our results provide additional mechanisms of action of GLP-1 to prevent oxidative damage in beta cells through the modulation of signaling pathways involved in antioxidant enzyme regulation. PMID:26968794

  1. Glucagon-like peptide-1 improves beta-cell antioxidant capacity via extracellular regulated kinases pathway and Nrf2 translocation.

    PubMed

    Fernández-Millán, E; Martín, M A; Goya, L; Lizárraga-Mollinedo, E; Escrivá, F; Ramos, S; Álvarez, C

    2016-06-01

    Oxidative stress plays an important role in the development of beta-cell dysfunction and insulin resistance, two major pathophysiological abnormalities of type 2 diabetes. Expression levels of antioxidant enzymes in beta cells are very low, rendering them more susceptible to damage caused by reactive oxygen species (ROS). Although the antioxidant effects of glucagon-like peptide-1 (GLP-1) and its analogs have been previously reported, the exact mechanisms involved are still unclear. In this study, we demonstrated that GLP-1 was able to effectively inhibit oxidative stress and cell death of INS-1E beta cells induced by the pro-oxidant tert-butyl hydroperoxide (tert-BOOH). Incubation with GLP-1 enhanced cellular levels of glutathione and the activity of its related enzymes, glutathione-peroxidase (GPx) and -reductase (GR) in beta cells. However, inhibition of ERK, but not of the PI3K/AKT pathway abolished, at least in part, the antioxidant effect of GLP-1. Moreover, ERK activation seems to be protein kinase A (PKA)-dependent because inhibition of PKA with H-89 was sufficient to block the GLP-1-derived protective effect on beta cells. GLP-1 likewise increased the synthesis of GR and favored the translocation of the nuclear transcription factor erythroid 2p45-related factor (Nrf2), a transcription factor implicated in the expression of several antioxidant/detoxificant enzymes. Glucose-stimulated insulin secretion was also preserved in beta-cells challenged with tert-BOOH but pre-treated with GLP-1, probably through the down-regulation of the mitochondrial uncoupling-protein2 (UCP2). Thus, our results provide additional mechanisms of action of GLP-1 to prevent oxidative damage in beta cells through the modulation of signaling pathways involved in antioxidant enzyme regulation.

  2. IFN-{gamma} sensitizes MIN6N8 insulinoma cells to TNF-{alpha}-induced apoptosis by inhibiting NF-{kappa}B-mediated XIAP upregulation

    SciTech Connect

    Kim, Hun Sik; Kim, Sunshin; Lee, Myung-Shik . E-mail: mslee@smc.samsung.co.kr

    2005-10-28

    Although X-linked inhibitor of apoptosis protein (XIAP) is an important intracellular suppressor of apoptosis in a variety of cell types, its role in cytokine-induced pancreatic {beta}-cell apoptosis remains unclear. Here, we found that: (i) XIAP level was inversely correlated with tumor necrosis factor (TNF)-{alpha}-induced apoptosis in MIN6N8 insulinoma cells; (ii) adenoviral XIAP overexpression abrogated the TNF-{alpha}-induced apoptosis through inhibition of caspase activity; (iii) downregulation of XIAP by antisense oligonucleotide or Smac peptide sensitized MIN6N8 cells to TNF-{alpha}-induced apoptosis; (iv) XIAP expression was induced by TNF-{alpha} through a nuclear factor-{kappa}B (NF-{kappa}B)-dependent pathway, and interferon (IFN)-{gamma} prevented such an induction in a manner independent of NF-{kappa}B, which presents a potential mechanism underlying cytotoxic IFN-{gamma}/TNF-{alpha} synergism. Taken together, our results suggest that XIAP is an important modulator of TNF-{alpha}-induced apoptosis of MIN6N8 cells, and XIAP regulation in pancreatic {beta}-cells might play an important role in pancreatic {beta}-cell apoptosis and in the pathogenesis of type 1 diabetes.

  3. [Analysis of the effect of diabetes type 2 duration on beta cell secretory function and insulin resistance].

    PubMed

    Popović, Ljiljana; Zamaklar, Miroslava; Lalić, Katarina; Vasović, Olga

    2006-01-01

    Diabetes type 2 is a chronic metabolic disorder. Pathogenesis of diabetes type 2 results from the impaired insulin secretion, impaired insulin action and increased endogenous glucose production. Diabetes evolves through several phases characterized by qualitative and quantitative changes of beta cell secretory function. The aim of our study was to analyze the impact of diabetes duration on beta cell secretory function and insulin resistance. The results indicated significant negative correlation of diabetes duration and fasting insulinemia, as well as beta cell secretory function assessed by HOMA beta index. Our study also found significant negative correlation of diabetes duration and insulin resistance assessed by HOMA IR index. Significant positive correlation was established between beta cell secretory capacity (fasting insulinemia and HOMA beta) and insulin resistance assessed by HOMA IR index, independently of diabetes duration. These results indicate that: beta cell secretory capacity, assessed by HOMA beta index, significantly decreases with diabetes duration. In parallel with decrease of fasting insulinemia, reduction of insulin resistance assessed by HOMA IR index was found as well.

  4. The protective activity of Urtica dioica leaves on blood glucose concentration and beta-cells in streptozotocin-diabetic rats.

    PubMed

    Golalipour, Mohammad Jafar; Khori, Vahid

    2007-04-15

    This study was done to determine the protective activity of the hydroalcholic extract of Urtica dioica leaves on Hyperglycemia and beta-cells in hyperglycemic rats. Thirty Wistar rats were allocated in groups of normal, Diabetic and treatment. Hyperglycemia in Rats induced by 80 mg kg(-1) streptozotocin. In treatment group, animals received hydroalcholic extract of Urtica dioica 100 mg kg(-1) day(-1) for five days, intraperitoneally and then hyperglycemia induced by streptozotocin. The blood glucose concentration was measured by using a Glucometer in 1st, 3rd and 5th weeks. In the end of 5th weeks the animals in each group were sacrificed by anesthesia and whole pancreas in three groups extracted and fixed in bouin's fluid and stained by chromealum hematoxiline-phloxine and beta cells were counted in three groups by Olympus microscope. Mean +/- SE of blood glucose concentrations in the end of fifth weeks were 99.4 +/-5.0, 454.7 +/- 34.5 and 303.6 +/- 100.6 in control, diabetic and treatment groups, respectively (p < 0.05). The percentages of beta-cells in control, diabetic and treatment groups were 73.6, 1.9 and 22.9%, respectively. The percentage of beta-cells in treatment group comparing with diabetic group was significant (p < 0.05). This study showed that the protective administration of hydroalcholic extract of Urtica dioica has hypoglycemic effect and protective activity of beta-cells of langerhans in hyperglycemic rats. PMID:19069917

  5. Bioluminescence imaging reveals dynamics of beta cell loss in the non-obese diabetic (NOD) mouse model.

    PubMed

    Virostko, John; Radhika, Armandla; Poffenberger, Greg; Dula, Adrienne N; Moore, Daniel J; Powers, Alvin C

    2013-01-01

    We generated a mouse model (MIP-Luc-VU-NOD) that enables non-invasive bioluminescence imaging (BLI) of beta cell loss during the progression of autoimmune diabetes and determined the relationship between BLI and disease progression. MIP-Luc-VU-NOD mice displayed insulitis and a decline in bioluminescence with age which correlated with beta cell mass, plasma insulin, and pancreatic insulin content. Bioluminescence declined gradually in female MIP-Luc-VU-NOD mice, reaching less than 50% of the initial BLI at 10 weeks of age, whereas hyperglycemia did not ensue until mice were at least 16 weeks old. Mice that did not become diabetic maintained insulin secretion and had less of a decline in bioluminescence than mice that became diabetic. Bioluminescence measurements predicted a decline in beta cell mass prior to the onset of hyperglycemia and tracked beta cell loss. This model should be useful for investigating the fundamental processes underlying autoimmune diabetes and developing new therapies targeting beta cell protection and regeneration.

  6. Alpha cells secrete acetylcholine as a non-neuronal paracrine signal priming beta cell function in humans.

    PubMed

    Rodriguez-Diaz, Rayner; Dando, Robin; Jacques-Silva, M Caroline; Fachado, Alberto; Molina, Judith; Abdulreda, Midhat H; Ricordi, Camillo; Roper, Stephen D; Berggren, Per-Olof; Caicedo, Alejandro

    2011-06-19

    Acetylcholine is a neurotransmitter that has a major role in the function of the insulin-secreting pancreatic beta cell. Parasympathetic innervation of the endocrine pancreas, the islets of Langerhans, has been shown to provide cholinergic input to the beta cell in several species, but the role of autonomic innervation in human beta cell function is at present unclear. Here we show that, in contrast to the case in mouse islets, cholinergic innervation of human islets is sparse. Instead, we find that the alpha cells of human islets provide paracrine cholinergic input to surrounding endocrine cells. Human alpha cells express the vesicular acetylcholine transporter and release acetylcholine when stimulated with kainate or a lowering in glucose concentration. Acetylcholine secretion by alpha cells in turn sensitizes the beta cell response to increases in glucose concentration. Our results demonstrate that in human islets acetylcholine is a paracrine signal that primes the beta cell to respond optimally to subsequent increases in glucose concentration. Cholinergic signaling within islets represents a potential therapeutic target in diabetes, highlighting the relevance of this advance to future drug development.

  7. Beta Cell Formation in vivo Through Cellular Networking, Integration and Processing (CNIP) in Wild Type Adult Mice.

    PubMed

    Doiron, Bruno; Hu, Wenchao; DeFronzo, Ralph A

    2016-01-01

    Insulin replacement therapy is essential in type 1 diabetic individuals and is required in ~40- 50% of type 2 diabetics during their lifetime. Prior attempts at beta cell regeneration have relied upon pancreatic injury to induce beta cell proliferation, dedifferentiation and activation of the embryonic pathway, or stem cell replacement. We report an alternative method to transform adult non-stem (somatic) cells into pancreatic beta cells. The Cellular Networking, Integration and Processing (CNIP) approach targets cellular mechanisms involved in pancreatic function in the organ's adult state and utilizes a synergistic mechanism that integrates three important levels of cellular regulation to induce beta cell formation: (i) glucose metabolism, (ii) membrane receptor function, and (iii) gene transcription. The aim of the present study was to induce pancreatic beta cell formation in vivo in adult animals without stem cells and without dedifferentiating cells to recapitulate the embryonic pathway as previously published (1-3). Our results employing CNIP demonstrate that: (i) insulin secreting cells can be generated in adult pancreatic tissue in vivo and circumvent the problem of generating endocrine (glucagon and somatostatin) cells that exert deleterious effects on glucose homeostasis, and (ii) longterm normalization of glucose tolerance and insulin secretion can be achieved in a wild type diabetic mouse model. The CNIP cocktail has the potential to be used as a preventative or therapeutic treatment or cure for both type 1 and type 2 diabetes. PMID:26696016

  8. Detection of C-Peptide in Urine as a Measure of Ongoing Beta Cell Function.

    PubMed

    McDonald, T J; Perry, M H

    2016-01-01

    C-peptide is a protein secreted by the pancreatic beta cells in equimolar quantities with insulin, following the cleavage of proinsulin into insulin. Measurement of C-peptide is used as a surrogate marker of endogenous insulin secretory capacity. Assessing C-peptide levels can be useful in classifying the subtype of diabetes as well as assessing potential treatment choices in the management of diabetes.Standard measures of C-peptide involve blood samples collected either fasted or, most often, after a fixed stimulus (such as oral glucose, mixed meal, or IV glucagon). Despite the established clinical utility of blood C-peptide measurement, its widespread use is limited. In many instances this is due to perceived practical restrictions associated with sample collection.Urine C-peptide measurement is an attractive noninvasive alternative to blood measures of beta-cell function. Urine C-peptide creatinine ratio measured in a single post stimulated sample has been shown to be a robust, reproducible measure of endogenous C-peptide which is stable for three days at room temperature when collected in boric acid. Modern high sensitivity immunoassay technologies have facilitated measurement of C-peptide down to single picomolar concentrations. PMID:27083170

  9. Autoregulation of free radicals via uncoupling protein control in pancreatic beta-cell mitochondria.

    PubMed

    Heuett, William J; Periwal, Vipul

    2010-01-20

    Pancreatic beta-cells sense the ambient blood-glucose concentration and secrete insulin to signal other tissues to take up glucose. Mitochondria play a key role in this response as they metabolize nutrients to produce ATP and reactive oxygen species (ROS), both of which are involved in insulin secretion signaling. Based on data available in the literature and previously developed mathematical models, we present a model of glucose-stimulated mitochondrial respiration, ATP synthesis, and ROS production and control in beta-cells. The model is consistent with a number of experimental observations reported in the literature. Most notably, it captures the nonlinear rise in the proton leak rate at high membrane potential and the increase in this leak due to uncoupling protein (UCP) activation by ROS. The functional forms used to model ROS production and UCP regulation yield insight into these mechanisms, as many details have not yet been unraveled in the experimental literature. We examine short- and long-term effects of UCP activation inhibition and changes in the mitochondrial density on mitochondrial responses to glucose. Results suggest increasing mitochondrial density while decreasing UCP activity may be an effective way to increase glucose-stimulated insulin secretion while decreasing oxidative stress.

  10. The pancreatic beta cell as a paradigm for advances in inositide research.

    PubMed

    Barker, Christopher J; Berggren, Per-Olof

    2012-09-01

    In a previous review for Advances in Enzyme Research (Berggren and Barker, 2008) we outlined the history of our involvement in discovering important roles for inositides in the insulin secreting pancreatic beta cell. In this current appraisal we bring the work up to date and project how we believe this field will continue to develop in the future. Recently, we have seen an important synergism between the growth in our understanding of inositide function and our knowledge of beta cell stimulus-secretion coupling in both physiological and pathophysiological contexts. Important advances have been made in three areas. 1. The classic regulation of cytoplasmic free Ca(2+) concentration [Ca(2+)](i) by Inositol 1,4,5-trisphosphate (Ins(1,4,5)P(3)) and its receptor, 2. A novel role of the inositol pyrophosphates, especially 5-diphosphoinositol pentakisphosphate (5-PP-InsP(5)), in exocytosis, and 3. The unique signaling roles of PI3K pathways instituted by the engagement of the insulin receptor in an autocrine, positive feed-back loop. We examine each of these in turn and close with an assessment of the likely future directions the research will take.

  11. Chemiluminescence Imaging of Superoxide Anion Detects Beta-Cell Function and Mass

    PubMed Central

    Bronsart, Laura L.; Stokes, Christian; Contag, Christopher H.

    2016-01-01

    Superoxide anion is produced during normal cellular respiration and plays key roles in cellular physiology with its dysregulation being associated with a variety of diseases. Superoxide anion is a short-lived molecule and, therefore, its homeostatic regulation and role in biology and disease requires dynamic quantification with fine temporal resolution. Here we validated coelenterazine as a reporter of intracellular superoxide anion concentration and used it as a dynamic measure both in vitro and in vivo. Chemiluminescence was dependent upon superoxide anion levels, including those produced during cellular respiration, and concentrations varied both kinetically and temporally in response to physiologically relevant fluctuations in glucose levels. In vivo imaging with coelenterazine revealed that beta cells of the pancreas have increased levels of superoxide anion, which acted as a measure of beta-cell function and mass and could predict the susceptibility of mice to diabetes mellitus. Glucose response and regulation are key elements of cellular physiology and organismal biology, and superoxide anion appears to play a fundamental and dynamic role in both of these processes. PMID:26752052

  12. Chemiluminescence Imaging of Superoxide Anion Detects Beta-Cell Function and Mass.

    PubMed

    Bronsart, Laura L; Stokes, Christian; Contag, Christopher H

    2016-01-01

    Superoxide anion is produced during normal cellular respiration and plays key roles in cellular physiology with its dysregulation being associated with a variety of diseases. Superoxide anion is a short-lived molecule and, therefore, its homeostatic regulation and role in biology and disease requires dynamic quantification with fine temporal resolution. Here we validated coelenterazine as a reporter of intracellular superoxide anion concentration and used it as a dynamic measure both in vitro and in vivo. Chemiluminescence was dependent upon superoxide anion levels, including those produced during cellular respiration, and concentrations varied both kinetically and temporally in response to physiologically relevant fluctuations in glucose levels. In vivo imaging with coelenterazine revealed that beta cells of the pancreas have increased levels of superoxide anion, which acted as a measure of beta-cell function and mass and could predict the susceptibility of mice to diabetes mellitus. Glucose response and regulation are key elements of cellular physiology and organismal biology, and superoxide anion appears to play a fundamental and dynamic role in both of these processes. PMID:26752052

  13. INS-gene mutations: From genetics and beta cell biology to clinical disease

    PubMed Central

    Liu, Ming; Sun, Jinhong; Cui, Jinqiu; Chen, Wei; Guo, Huan; Barbetti, Fabrizio; Arvan, Peter

    2015-01-01

    A growing list of insulin gene mutations causing a new form of monogenic diabetes has drawn increasing attention over the past seven years. The mutations have been identified in the untranslated regions of the insulin gene as well as the coding sequence of preproinsulin including within the signal peptide, insulin B-chain, C-peptide, insulin A-chain, and the proteolytic cleavage sites both for signal peptidase and the prohormone convertases. These mutations affect a variety of different steps of insulin biosynthesis in pancreatic beta cells. Importantly, although many of these mutations cause proinsulin misfolding with early onset autosomal dominant diabetes, some of the mutant alleles appear to engage different cellular and molecular mechanisms that underlie beta cell failure and diabetes. In this article, we review the most recent advances in the field and discuss challenges as well as potential strategies to prevent/delay the development and progression of autosomal dominant diabetes caused by INS-gene mutations. It is worth noting that although diabetes caused by INS gene mutations is rare, increasing evidence suggests that defects in the pathway of insulin biosynthesis may also be involved in the progression of more common types of diabetes. Collectively, the (pre)proinsulin mutants provide insightful molecular models to better understand the pathogenesis of all forms of diabetes in which preproinsulin processing defects, proinsulin misfolding, and ER stress are involved. PMID:25542748

  14. Beta cell antigens in type 1 diabetes: triggers in pathogenesis and therapeutic targets

    PubMed Central

    Mauvais, François-Xavier; Diana, Julien; van Endert, Peter

    2016-01-01

    Research focusing on type 1 diabetes (T1D) autoantigens aims to explore our understanding of these beta cell proteins in order to design assays for monitoring the pathogenic autoimmune response, as well as safe and efficient therapies preventing or stopping it. In this review, we will discuss progress made in the last 5 years with respect to mechanistic understanding, diagnostic monitoring, and therapeutic modulation of the autoantigen-specific cellular immune response in T1D. Some technical progress in monitoring tools has been made; however, the potential of recent technologies for highly multiplexed exploration of human cellular immune responses remains to be exploited in T1D research, as it may be the key to the identification of surrogate markers of disease progression that are still wanting. Detailed analysis of autoantigen recognition by T cells suggests an important role of non-conventional antigen presentation and processing in beta cell-directed autoimmunity, but the impact of this in human T1D has been little explored. Finally, therapeutic administration of autoantigens to T1D patients has produced disappointing results. The application of novel modes of autoantigen administration, careful translation of mechanistic understanding obtained in preclinical studies and in vitro with human cells, and combination therapies including CD3 antibodies may help to make autoantigen-based immunotherapy for T1D a success story in the future. PMID:27158463

  15. Beta cell antigens in type 1 diabetes: triggers in pathogenesis and therapeutic targets.

    PubMed

    Mauvais, François-Xavier; Diana, Julien; van Endert, Peter

    2016-01-01

    Research focusing on type 1 diabetes (T1D) autoantigens aims to explore our understanding of these beta cell proteins in order to design assays for monitoring the pathogenic autoimmune response, as well as safe and efficient therapies preventing or stopping it. In this review, we will discuss progress made in the last 5 years with respect to mechanistic understanding, diagnostic monitoring, and therapeutic modulation of the autoantigen-specific cellular immune response in T1D. Some technical progress in monitoring tools has been made; however, the potential of recent technologies for highly multiplexed exploration of human cellular immune responses remains to be exploited in T1D research, as it may be the key to the identification of surrogate markers of disease progression that are still wanting. Detailed analysis of autoantigen recognition by T cells suggests an important role of non-conventional antigen presentation and processing in beta cell-directed autoimmunity, but the impact of this in human T1D has been little explored. Finally, therapeutic administration of autoantigens to T1D patients has produced disappointing results. The application of novel modes of autoantigen administration, careful translation of mechanistic understanding obtained in preclinical studies and in vitro with human cells, and combination therapies including CD3 antibodies may help to make autoantigen-based immunotherapy for T1D a success story in the future. PMID:27158463

  16. Symmetric Fold/Super-Hopf Bursting, Chaos and Mixed-Mode Oscillations in Pernarowski Model of Pancreatic Beta-Cells

    NASA Astrophysics Data System (ADS)

    Fallah, Haniyeh

    Pancreatic beta-cells produce insulin to regularize the blood glucose level. Bursting is important in beta cells due to its relation to the release of insulin. Pernarowski model is a simple polynomial model of beta-cell activities indicating bursting oscillations in these cells. This paper presents bursting behaviors of symmetric type in this model. In addition, it is shown that the current system exhibits the phenomenon of period doubling cascades of canards which is a route to chaos. Canards are also observed symmetrically near folds of slow manifold which results in a chaotic transition between n and n + 1 spikes symmetric bursting. Furthermore, mixed-mode oscillations (MMOs) and combination of symmetric bursting together with MMOs are illustrated during the transition between symmetric bursting and continuous spiking.

  17. Phosphorylation events implicating p38 and PI3K mediate tungstate-effects in MIN6 beta cells

    SciTech Connect

    Piquer, Sandra; Gomis, Ramon . E-mail: rgomis@clinic.ub.es

    2007-06-29

    Oral administration of sodium tungstate is an effective treatment for diabetes in animal models. Several lines of evidence indicate the pancreatic beta cell as one of the targets of tungstate action. Here, we examined the molecular mechanism by which this compound exerts its effects on the beta cell line MIN6. Tungstate treatment induced phosphorylation and subsequent activation of p38 and PI3K which in turn are implicated in tungstate PDX-1 nuclear localization and activation. Although no effect was observed in glucose-induced insulin secretion we found that tungstate activates basal insulin release, a process driven, at least in part, by activation of p38. These results show a direct involvement of p38 and PI3K phosphorylation in the mechanism of action of tungstate in the beta cell.

  18. Hepatocyte nuclear factor 3beta is involved in pancreatic beta-cell-specific transcription of the pdx-1 gene.

    PubMed Central

    Wu, K L; Gannon, M; Peshavaria, M; Offield, M F; Henderson, E; Ray, M; Marks, A; Gamer, L W; Wright, C V; Stein, R

    1997-01-01

    The mammalian homeobox gene pdx-1 is expressed in pluripotent precursor cells in the dorsal and ventral pancreatic bud and duodenal endoderm, which will produce the pancreas and the rostral duodenum. In the adult, pdr-1 is expressed principally within insulin-secreting pancreatic islet beta cells and cells of the duodenal epithelium. Our objective in this study was to localize sequences within the mouse pdx-1 gene mediating selective expression within the islet. Studies of transgenic mice in which a genomic fragment of the mouse pdx-1 gene from kb -4.5 to +8.2 was used to drive a beta-galactosidase reporter showed that the control sequences sufficient for appropriate developmental and adult specific expression were contained within this region. Three nuclease-hypersensitive sites, located between bp -2560 and -1880 (site 1), bp -1330 and -800 (site 2), and bp -260 and +180 (site 3), were identified within the 5'-flanking region of the endogenous pdx-1 gene. Pancreatic beta-cell-specific expression was shown to be controlled by sequences within site 1 from an analysis of the expression pattern of various pdr-1-herpes simplex virus thymidine kinase promoter expression constructs in transfected beta-cell and non-beta-cell lines. Furthermore, we also established that this region was important in vivo by demonstrating that expression from a site 1-driven beta-galactosidase reporter construct was directed to islet beta-cells in transgenic mice. The activity of the site 1-driven constructs was reduced substantially in beta-cell lines by mutating a hepatocyte nuclear factor 3 (HNF3)-like site located between nucleotides -2007 and -1996. Gel shift analysis indicated that HNF3beta present in islet beta cells binds to this element. Immunohistochemical studies revealed that HNF3beta was present within the nuclei of almost all islet beta cells and subsets of pancreatic acinar cells. Together, these results suggest that HNF3beta, a key regulator of endodermal cell lineage

  19. Activation of transmembrane bile acid receptor TGR5 stimulates insulin secretion in pancreatic {beta} cells

    SciTech Connect

    Kumar, Divya P.; Rajagopal, Senthilkumar; Mahavadi, Sunila; Mirshahi, Faridoddin; Grider, John R.; Murthy, Karnam S.; Sanyal, Arun J.

    2012-10-26

    Highlights: Black-Right-Pointing-Pointer G protein coupled receptor TGR5 is expressed in mouse and human islets. Black-Right-Pointing-Pointer TGR5 is coupled to activation of Gs and Ca{sup 2+} release via cAMP/Epac/PLC-{epsilon} pathway. Black-Right-Pointing-Pointer Activation of TGR5 by bile salts and selective ligands causes insulin secretion. Black-Right-Pointing-Pointer TGR5 could be a potential therapeutic target to treat diabetes. -- Abstract: Bile acids act as signaling molecules and stimulate the G protein coupled receptor, TGR5, in addition to nuclear farnesoid X receptor to regulate lipid, glucose and energy metabolism. Bile acid induced activation of TGR5 in the enteroendocrine cells promotes glucagon like peptide-1 (GLP-1) release, which has insulinotropic effect in the pancreatic {beta} cells. In the present study, we have identified the expression of TGR5 in pancreatic {beta} cell line MIN6 and also in mouse and human pancreatic islets. TGR5 selective ligands, oleanolic acid (OA) and INT-777 selectively activated G{alpha}{sub s} and caused an increase in intracellular cAMP and Ca{sup 2+}. OA and INT-777 also increased phosphoinositide (PI) hydrolysis and the increase was blocked by NF449 (a selective G{alpha}{sub s} inhibitor) or (U73122) (PI hydrolysis inhibitor). OA, INT-777 and lithocholic acid increased insulin release in MIN6 and human islets and the increase was inhibited by treatment with NF449, (U73122) or BAPTA-AM (chelator of calcium), but not with myristoylated PKI (PKA inhibitor), suggesting that the release is dependent on G{sub s}/cAMP/Ca{sup 2+} pathway. 8-pCPT-2 Prime -O-Me-cAMP, a cAMP analog, which activates Epac, but not PKA also stimulated PI hydrolysis. In conclusion, our study demonstrates that the TGR5 expressed in the pancreatic {beta} cells regulates insulin secretion and highlights the importance of ongoing therapeutic strategies targeting TGR5 in the control of glucose homeostasis.

  20. LIGHT/IFN-γ triggers β cells apoptosis via NF-κB/Bcl2-dependent mitochondrial pathway.

    PubMed

    Zheng, Quan-You; Cao, Zhao-Hui; Hu, Xiao-Bo; Li, Gui-Qing; Dong, Shi-Fang; Xu, Gui-Lian; Zhang, Ke-Qin

    2016-10-01

    LIGHT recruits and activates naive T cells in the islets at the onset of diabetes. IFN-γ secreted by activated T lymphocytes is involved in beta cell apoptosis. However, whether LIGHT sensitizes IFNγ-induced beta cells destruction remains unclear. In this study, we used the murine beta cell line MIN6 and primary islet cells as models for investigating the underlying cellular mechanisms involved in LIGHT/IFNγ - induced pancreatic beta cell destruction. LIGHT and IFN-γ synergistically reduced MIN6 and primary islet cells viability; decreased cell viability was due to apoptosis, as demonstrated by a significant increase in Annexin V(+) cell percentage, detected by flow cytometry. In addition to marked increases in cytochrome c release and NF-κB activation, the combination of LIGHT and IFN-γ caused an obvious decrease in expression of the anti-apoptotic proteins Bcl-2 and Bcl-xL, but an increase in expression of the pro-apoptotic proteins Bak and Bax in MIN6 cells. Accordingly, LIGHT deficiency led to a decrease in NF-κB activation and Bak expression, and peri-insulitis in non-obese diabetes mice. Inhibition of NF-κB activation with the specific NF-κB inhibitor, PDTC (pyrrolidine dithiocarbamate), reversed Bcl-xL down-regulation and Bax up-regulation, and led to a significant increase in LIGHT- and IFN-γ-treated cell viability. Moreover, cleaved caspase-9, -3, and PARP (poly (ADP-ribose) polymerase) were observed after LIGHT and IFN-γ treatment. Pretreatment with caspase inhibitors remarkably attenuated LIGHT- and IFNγ-induced cell apoptosis. Taken together, our results indicate that LIGHT signalling pathway combined with IFN-γ induces beta cells apoptosis via an NF-κB/Bcl2-dependent mitochondrial pathway.

  1. Is Transforming Stem Cells to Pancreatic Beta Cells Still the Holy Grail for Type 2 Diabetes?

    PubMed

    Kahraman, Sevim; Okawa, Erin R; Kulkarni, Rohit N

    2016-08-01

    Diabetes is a progressive disease affecting millions of people worldwide. There are several medications and treatment options to improve the life quality of people with diabetes. One of the strategies for the treatment of diabetes could be the use of human pluripotent stem cells or induced pluripotent stem cells. The recent advances in differentiation of stem cells into insulin-secreting beta-like cells in vitro make the transplantation of the stem cell-derived beta-like cells an attractive approach for treatment of type 1 and type 2 diabetes. While stem cell-derived beta-like cells provide an unlimited cell source for beta cell replacement therapies, these cells can also be used as a platform for drug screening or modeling diseases.

  2. UCP-2 and UCP-3 Proteins Are Differentially Regulated in Pancreatic Beta-Cells

    PubMed Central

    Li, Yunfeng; Maedler, Kathrin; Shu, Luan; Haataja, Leena

    2008-01-01

    Background Increased uncoupling protein-2 (UCP-2) expression has been associated with impaired insulin secretion, whereas UCP-3 protein levels are decreased in the skeleton muscle of type-2 diabetic subjects. In the present studies we hypothesize an opposing effect of glucose on the regulation of UCP-2 and UCP-3 in pancreatic islets. Methodology Dominant negative UCP-2 and wild type UCP-3 adenoviruses were generated, and insulin release by transduced human islets was measured. UCP-2 and UCP-3 mRNA levels were determined using quantitative PCR. UCP-2 and UCP-3 protein expression was investigated in human islets cultured in the presence of different glucose concentrations. Human pancreatic sections were analyzed for subcellular localization of UCP-3 using immunohistochemistry. Principal Findings Dominant negative UCP-2 expression in human islets increased insulin secretion compared to control islets (p<0.05). UCP-3 mRNA is expressed in human islets, but the relative abundance of UCP-2 mRNA was 8.1-fold higher (p<0.05). Immunohistochemical analysis confirmed co-localization of UCP-3 protein with mitochondria in human beta-cells. UCP-2 protein expression in human islets was increased ∼2-fold after high glucose exposure, whereas UCP-3 protein expression was decreased by ∼40% (p<0.05). UCP-3 overexpression improved glucose-stimulated insulin secretion. Conclusions UCP-2 and UCP-3 may have distinct roles in regulating beta-cell function. Increased expression of UCP-2 and decreased expression of UCP-3 in humans with chronic hyperglycemia may contribute to impaired glucose-stimulated insulin secretion. These data imply that mechanisms that suppress UCP-2 or mechanisms that increase UCP-3 expression and/or function are potential therapeutic targets to offset defects of insulin secretion in humans with type-2 diabetes. PMID:18167556

  3. Molecular targets of a human HNF1 alpha mutation responsible for pancreatic beta-cell dysfunction.

    PubMed

    Wang, H; Antinozzi, P A; Hagenfeldt, K A; Maechler, P; Wollheim, C B

    2000-08-15

    The reverse tetracycline-dependent transactivator system was employed in insulinoma INS-1 cells to achieve controlled inducible expression of hepatocyte nuclear factor-1 alpha (HNF1 alpha)-P291fsinsC, the most common mutation associated with subtype 3 of maturity-onset diabetes of the young (MODY3). Nuclear localized HNF1 alpha-P291fsinsC protein exerts its dominant-negative effects by competing with endogenous HNF1 alpha for the cognate DNA-binding site. HNF1 alpha controls multiple genes implicated in pancreatic beta-cell function and notably in metabolism- secretion coupling. In addition to reduced expression of the genes encoding insulin, glucose transporter-2, L-pyruvate kinase, aldolase B and 3-hydroxy-3-methylglutaryl coenzyme A reductase, induction of HNF1 alpha-P291fsinsC also significantly inhibits expression of mitochondrial 2-oxoglutarate dehydrogenase (OGDH) E1 subunit mRNA and protein. OGDH enzyme activity and [(14)C]pyruvate oxidation were also reduced. In contrast, the mRNA and protein levels of mitochondrial uncoupling protein-2 were dramatically increased by HNF1 alpha-P291fsinsC induction. As predicted from this altered gene expression profile, HNF1 alpha-P291fsinsC also inhibits insulin secretory responses to glucose and leucine, correlated with impaired nutrient-evoked mitochondrial ATP production and mitochondrial membrane hyperpolarization. These unprecedented results suggest the molecular mechanism of HNF1 alpha-P291fsinsC causing beta-cell dysfunction. PMID:10944108

  4. Role of humoral beta-cell autoimmunity in type 1 diabetes.

    PubMed

    Knip, Mikael; Siljander, Heli; Ilonen, Jorma; Simell, Olli; Veijola, Riitta

    2016-07-01

    Islet cell antibodies (ICA) were found for the first time more than 40 yr ago in patients with autoimmune endocrine deficiencies, including type 1 diabetes (T1D). ICA detected by indirect immunofluorescence represent a heterogeneous group of autoantibodies targeting a series of biochemical autoantigens, such as the protein tyrosine phosphatase related islet antigen 2 (IA-2), the 65 kD isoform of glutamic acid decarboxylase (GA65), and zinc transporter 8 (ZnT8) as well as currently unidentified autoantigens. The general view is that the diabetes-associated autoantibodies are not directly involved in beta-cell destruction but function as biomarkers of an ongoing destructive process. The diabetes-associated autoantibodies remain the strongest predictive marker for future development of T1D. Positivity for multiple (≥2) autoantibodies is highly predictive of clinical disease both among first-degree relatives and in the general population. Autoantibody titers are highly variable during the preclinical phase, but in many cases the titers tend to decrease before diagnosis. The first signs of beta-cell autoimmunity may appear early during the first months of life. The majority of those individuals diagnosed with T1D before puberty seroconvert to autoantibody positivity before the age of 3 yr. The natural course and duration of preclinical diabetes vary substantially from one individual to another. The characteristics of the isotype-specific response during preclinical diabetes appear to be antigen-specific. Diabetes-associated autoantibodies may be useful surrogate markers of the subsequent development of T1D in primary prevention trials. T1D may occur, albeit rarely, in the absence of any signs of humoral autoimmunity at diagnosis. PMID:27411432

  5. Stimulus-secretion coupling in beta-cells: modulation by pH.

    PubMed

    Pace, C S; Tarvin, J T; Smith, J S

    1983-01-01

    We have examined the influence of changes in pH on the oscillatory pattern of electrical activity (EA) in the beta-cell by altering medium pH (pHo) and using permeable weak buffers to alter intracellular pH (pHi). A decrease in pH in the presence of glucose elicited depolarization to the active phase and constant spike activity, whereas an increase in pH elicited a decrease in spike activity or silent hyperpolarization. On inhibition of HCO3:Cl antiport by addition of DIDS (4,4'-diisothiocyano-2,2'-stilbene disulfonic acid), probenecid, or withdrawal of medium HCO-3, there was an increase in the duration of the active phase. A similar result was obtained on the inhibition of Na:H antiport by the addition of amiloride or the reduction of medium [Na+]. The influence of H+ and glucose has been proposed to decrease K+ permeability (PK). However, the influence of pH on 86Rb+ efflux was most effective at subthreshold or 4.2 mM glucose; only a moderate decrease in PK occurred at 8.3 mM glucose, and no effect was obtained at 16.7 mM glucose. Alteration of pHi, and not pHo, induces similar effects on glucose-induced electrical and secretory events. There is a clear dissociation between the influence of inhibitors of the Na:H and HCO3:Cl antiporters on the electrical and secretory events. DIDS and amiloride increased glucose-induced EA, but markedly inhibited the secretory response to glucose. It is evident that pH modulates the electrical events and cationic fluxes and ultimately influences the transduction of information to the mechanisms controlling the secretory process in the beta-cell. PMID:6295181

  6. Serpine1 Mediates Porphyromonas gingivalis Induced Insulin Secretion in the Pancreatic Beta Cell Line MIN6

    PubMed Central

    Bhat, Uppoor G.; Watanabe, Keiko

    2015-01-01

    Periodontitis is an inflammatory disease resulting in destruction of gingiva and alveolar bone caused by an exuberant host immunological response to periodontal pathogens. Results from a number of epidemiological studies indicate a close association between diabetes and periodontitis. Results from cross-sectional studies indicate that subjects with periodontitis have a higher odds ratio of developing insulin resistance (IR). However, the mechanisms by which periodontitis influences the development of diabetes are not known. Results from our previous studies using an animal model of periodontitis suggest that periodontitis accelerates the onset of hyperinsulinemia and IR. In addition, LPS from a periodontal pathogen, Porphyromonas gingivalis (Pg), stimulates Serpine1 expression in the pancreatic beta cell line MIN6. Based on these observations, we hypothesized that a periodontal pathogen induces hyperinsulinemia and Serpine1 may be involved in this process. To test this hypothesis, we co-incubated Pg with the pancreatic beta cell line MIN6 and measured the effect on insulin secretion by MIN6 cells. We further determined the involvement of Serpine1 in insulin secretion by downregulating Serpine1 expression. Our results indicated that Pg stimulated insulin secretion by approximately 3.0 fold under normoglycemic conditions. In a hyperglycemic state, Pg increased insulin secretion by 1.5 fold. Pg significantly upregulated expression of the Serpine1 gene and this was associated with increased secretion of insulin by MIN6 cells. However, cells with downregulated Serpine1 expression were resistant to Pg stimulated insulin secretion under normoglycemic conditions. We conclude that the periodontal pathogen, Pg, induced insulin secretion by MIN6 cells and this induction was, in part, Serpine1 dependent. Thus, Serpine1 may play a pivotal role in insulin secretion during the accelerated development of hyperinsulinemia and the resulting IR in the setting of periodontitis. PMID

  7. cAMP mediators of pulsatile insulin secretion from glucose-stimulated single beta-cells.

    PubMed

    Idevall-Hagren, Olof; Barg, Sebastian; Gylfe, Erik; Tengholm, Anders

    2010-07-23

    Pulsatile insulin release from glucose-stimulated beta-cells is driven by oscillations of the Ca(2+) and cAMP concentrations in the subplasma membrane space ([Ca(2+)](pm) and [cAMP](pm)). To clarify mechanisms by which cAMP regulates insulin secretion, we performed parallel evanescent wave fluorescence imaging of [cAMP](pm), [Ca(2+)](pm), and phosphatidylinositol 3,4,5-trisphosphate (PIP(3)) in the plasma membrane. This lipid is formed by autocrine insulin receptor activation and was used to monitor insulin release kinetics from single MIN6 beta-cells. Elevation of the glucose concentration from 3 to 11 mm induced, after a 2.7-min delay, coordinated oscillations of [Ca(2+)](pm), [cAMP](pm), and PIP(3). Inhibitors of protein kinase A (PKA) markedly diminished the PIP(3) response when applied before glucose stimulation, but did not affect already manifested PIP(3) oscillations. The reduced PIP(3) response could be attributed to accelerated depolarization causing early rise of [Ca(2+)](pm) that preceded the elevation of [cAMP](pm). However, the amplitude of the PIP(3) response after PKA inhibition was restored by a specific agonist to the cAMP-dependent guanine nucleotide exchange factor Epac. Suppression of cAMP formation with adenylyl cyclase inhibitors reduced already established PIP(3) oscillations in glucose-stimulated cells, and this effect was almost completely counteracted by the Epac agonist. In cells treated with small interfering RNA targeting Epac2, the amplitudes of the glucose-induced PIP(3) oscillations were reduced, and the Epac agonist was without effect. The data indicate that temporal coordination of the triggering [Ca(2+)](pm) and amplifying [cAMP](pm) signals is important for glucose-induced pulsatile insulin release. Although both PKA and Epac2 partake in initiating insulin secretion, the cAMP dependence of established pulsatility is mediated by Epac2.

  8. Species-specific vesicular monoamine transporter 2 (VMAT2) expression in mammalian pancreatic beta cells: implications for optimising radioligand-based human beta cell mass (BCM) imaging in animal models

    PubMed Central

    Hartwig, N. R.; Kalmbach, N.; Klietz, M.; Anlauf, M.; Eiden, L. E.; Weihe, E.

    2014-01-01

    Aims/hypothesis Imaging of beta cell mass (BCM) is a major challenge in diabetes research. The vesicular monoamine transporter 2 (VMAT2) is abundantly expressed in human beta cells. Radiolabelled analogues of tetrabenazine (TBZ; a low-molecular-weight, cell-permeant VMAT2-selective ligand) have been employed for pancreatic islet imaging in humans. Since reports on TBZ-based VMAT2 imaging in rodent pancreas have been fraught with confusion, we compared VMAT2 gene expression patterns in the mouse, rat, pig and human pancreas, to identify appropriate animal models with which to further validate and optimise TBZ imaging in humans. Methods We used a panel of highly sensitive VMAT2 antibodies developed against equivalently antigenic regions of the transporter from each species in combination with immunostaining for insulin and species-specific in situ hybridisation probes. Individual pancreatic islets were obtained by laser-capture microdissection and subjected to analysis of mRNA expression of VMAT2. Results The VMAT2 protein was not expressed in beta cells in the adult pancreas of common mouse or rat laboratory strains, in contrast to its expression in beta cells (but not other pancreatic endocrine cell types) in the pancreas of pigs and humans. VMAT2- and tyrosine hydroxylase co-positive (catecholaminergic) innervation was less abundant in humans than in rodents. VMAT2-positive mast cells were identified in the pancreas of all species. Conclusions/interpretation Primates and pigs are suitable models for TBZ imaging of beta cells. Rodents, because of a complete lack of VMAT2 expression in the endocrine pancreas, are a ‘null’ model for assessing interference with BCM measurements by VMAT2-positive mast cells and sympathetic innervation in the pancreas. PMID:23404442

  9. Expression of neurexin, neuroligin, and their cytoplasmic binding partners in the pancreatic beta-cells and the involvement of neuroligin in insulin secretion.

    PubMed

    Suckow, Arthur T; Comoletti, Davide; Waldrop, Megan A; Mosedale, Merrie; Egodage, Sonya; Taylor, Palmer; Chessler, Steven D

    2008-12-01

    The composition of the beta-cell exocytic machinery is very similar to that of neuronal synapses, and the developmental pathway of beta-cells and neurons substantially overlap. beta-Cells secrete gamma-aminobutyric acid and express proteins that, in the brain, are specific markers of inhibitory synapses. Recently, neuronal coculture experiments have identified three families of synaptic cell-surface molecules (neurexins, neuroligins, and SynCAM) that drive synapse formation in vitro and that control the differentiation of nascent synapses into either excitatory or inhibitory fully mature nerve terminals. The inhibitory synapse-like character of the beta-cells led us to hypothesize that members of these families of synapse-inducing adhesion molecules would be expressed in beta-cells and that the pattern of expression would resemble that associated with neuronal inhibitory synaptogenesis. Here, we describe beta-cell expression of the neuroligins, neurexins, and SynCAM, and show that neuroligin expression affects insulin secretion in INS-1 beta-cells and rat islet cells. Our findings demonstrate that neuroligins and neurexins are expressed outside the central nervous system and help confer an inhibitory synaptic-like phenotype onto the beta-cell surface. Analogous to their role in synaptic neurotransmission, neurexin-neuroligin interactions may play a role in the formation of the submembrane insulin secretory apparatus.

  10. Beta-cell function, incretin effect, and incretin hormones in obese youth along the span of glucose tolerance from normal to prediabetes to Type 2 diabetes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Using the hyperglycemic and euglycemic clamp, we demonstrated impaired Beta-cell function in obese youth with increasing dysglycemia. Herein we describe oral glucose tolerance test (OGTT)-modeled Beta-cell function and incretin effect in obese adolescents spanning the range of glucose tolerance. Bet...

  11. Endoplasmic Reticulum Stress Links Oxidative Stress to Impaired Pancreatic Beta-Cell Function Caused by Human Oxidized LDL.

    PubMed

    Plaisance, Valérie; Brajkovic, Saška; Tenenbaum, Mathie; Favre, Dimitri; Ezanno, Hélène; Bonnefond, Amélie; Bonner, Caroline; Gmyr, Valéry; Kerr-Conte, Julie; Gauthier, Benoit R; Widmann, Christian; Waeber, Gérard; Pattou, François; Froguel, Philippe; Abderrahmani, Amar

    2016-01-01

    Elevated plasma concentration of the pro-atherogenic oxidized low density lipoprotein cholesterol (LDL) triggers adverse effects in pancreatic beta-cells and is associated with type 2 diabetes. Here, we investigated whether the endoplasmic reticulum (ER) stress is a key player coupling oxidative stress to beta-cell dysfunction and death elicited by human oxidized LDL. We found that human oxidized LDL activates ER stress as evidenced by the activation of the inositol requiring 1α, and the elevated expression of both DDIT3 (also called CHOP) and DNAJC3 (also called P58IPK) ER stress markers in isolated human islets and the mouse insulin secreting MIN6 cells. Silencing of Chop and inhibition of ER stress markers by the chemical chaperone phenyl butyric acid (PBA) prevented cell death caused by oxidized LDL. Finally, we found that oxidative stress accounts for activation of ER stress markers induced by oxidized LDL. Induction of Chop/CHOP and p58IPK/P58IPK by oxidized LDL was mimicked by hydrogen peroxide and was blocked by co-treatment with the N-acetylcystein antioxidant. As a conclusion, the harmful effects of oxidized LDL in beta-cells requires ER stress activation in a manner that involves oxidative stress. This mechanism may account for impaired beta-cell function in diabetes and can be reversed by antioxidant treatment. PMID:27636901

  12. Beta Cells Secrete Significant and Regulated Levels of Insulin for Long Periods when Seeded onto Acellular Micro-Scaffolds.

    PubMed

    Sionov, Ronit Vogt; Finesilver, Gershon; Sapozhnikov, Lena; Soroker, Avigail; Zlotkin-Rivkin, Efrat; Saad, Yocheved; Kahana, Meygal; Bodaker, Matan; Alpert, Evgenia; Mitrani, Eduardo

    2015-11-01

    The aim of this work is to obtain significant and regulated insulin secretion from human beta cells ex vivo. Long-term culture of human pancreatic islets and attempts at expanding human islet cells normally result in loss of beta-cell phenotype. We propose that to obtain proper ex vivo beta cell function, there is a need to develop three-dimensional structures that mimic the natural islet tissue microenvironment. We here describe the preparation of endocrine micro-pancreata (EMPs) that are made up of acellular organ-derived micro-scaffolds seeded with human intact or enzymatically dissociated islets. We show that EMPs constructed by seeding whole islets, freshly enzymatically-dissociated islets or even dissociated islets grown first in standard monolayer cultures express high levels of key beta-cell specific genes and secrete quantities of insulin per cell similar to freshly isolated human islets in a glucose-regulated manner for more than 3 months in vitro. PMID:26416226

  13. Down-regulation of zinc transporter 8 (SLC30A8) in pancreatic beta-cells promotes cell survival

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The pancreatic islet contains high levels of zinc in granular vesicles of beta-cells where insulin is matured, crystallized, and stored before secretion. Zinc is an essential co-factor for insulin crystallization forming dense core in secretory granules. In insulin-containing secretory granules, zin...

  14. Endoplasmic Reticulum Stress Links Oxidative Stress to Impaired Pancreatic Beta-Cell Function Caused by Human Oxidized LDL

    PubMed Central

    Favre, Dimitri; Ezanno, Hélène; Bonnefond, Amélie; Bonner, Caroline; Gmyr, Valéry; Kerr-Conte, Julie; Gauthier, Benoit R.; Widmann, Christian; Waeber, Gérard; Pattou, François; Froguel, Philippe; Abderrahmani, Amar

    2016-01-01

    Elevated plasma concentration of the pro-atherogenic oxidized low density lipoprotein cholesterol (LDL) triggers adverse effects in pancreatic beta-cells and is associated with type 2 diabetes. Here, we investigated whether the endoplasmic reticulum (ER) stress is a key player coupling oxidative stress to beta-cell dysfunction and death elicited by human oxidized LDL. We found that human oxidized LDL activates ER stress as evidenced by the activation of the inositol requiring 1α, and the elevated expression of both DDIT3 (also called CHOP) and DNAJC3 (also called P58IPK) ER stress markers in isolated human islets and the mouse insulin secreting MIN6 cells. Silencing of Chop and inhibition of ER stress markers by the chemical chaperone phenyl butyric acid (PBA) prevented cell death caused by oxidized LDL. Finally, we found that oxidative stress accounts for activation of ER stress markers induced by oxidized LDL. Induction of Chop/CHOP and p58IPK/P58IPK by oxidized LDL was mimicked by hydrogen peroxide and was blocked by co-treatment with the N-acetylcystein antioxidant. As a conclusion, the harmful effects of oxidized LDL in beta-cells requires ER stress activation in a manner that involves oxidative stress. This mechanism may account for impaired beta-cell function in diabetes and can be reversed by antioxidant treatment. PMID:27636901

  15. Coordinated regulation by Shp2 tyrosine phosphatase of signaling events controlling insulin biosynthesis in pancreatic beta-cells.

    PubMed

    Zhang, Sharon S; Hao, Ergeng; Yu, Jianxiu; Liu, Wen; Wang, Jing; Levine, Fred; Feng, Gen-Sheng

    2009-05-01

    Intracellular signaling by which pancreatic beta-cells synthesize and secrete insulin in control of glucose homeostasis is not fully understood. Here we show that Shp2, a cytoplasmic tyrosine phosphatase possessing 2 SH2 domains, coordinates signaling events required for insulin biosynthesis in beta-cells. Mice with conditional ablation of the Shp2/Ptpn11 gene in the pancreas exhibited defective glucose-stimulated insulin secretion and impaired glucose tolerance. Consistently, siRNA-mediated Shp2-knockdown in rat insulinoma INS-1 832/13 cells resulted in decreased insulin production and secretion despite an increase in cellular ATP. Shp2 modulates the strength of signals flowing through Akt/FoxO1 and Erk pathways, culminating in control of Pdx1 expression and activity on Ins1 and Ins2 promoters, and forced Pdx1 expression rescued insulin production in Shp2-knockdown beta-cells. Therefore, Shp2 acts as a signal coordinator in beta-cells, orchestrating multiple pathways controlling insulin biosynthesis to maintain glucose homeostasis.

  16. Induction of beta-cell resistance to hypoxia and technologies for oxygen delivery to transplanted pancreatic islets.

    PubMed

    Lazard, Daniel; Vardi, Pnina; Bloch, Konstantin

    2012-09-01

    Hypoxia is believed to be a crucial factor involved in cell adaptation to environmental stress. Islet transplantation, especially with immunoisolated islets, interrupts vascular connections, resulting in the substantially decreased delivery of oxygen and nutrients to islet cells. Insulin-producing pancreatic beta cells are known to be highly susceptible to oxygen deficiency. Such susceptibility to hypoxia is believed to be one of the main causes of beta-cell death in the post-transplantation period. Different strategies have been developed for the protection of beta cells against hypoxic injury and for oxygen delivery to transplanted islets. The enhancement of beta-cell defense properties against hypoxia has been achieved using various techniques such as gene transfection, drug supplementation, co-culturing with stem cells and cell selection. Technologies for oxygen delivery to transplanted islets include local neovascularization of subcutaneous sites, electrochemical and photosynthetic oxygen generation, oxygen refuelling of bio-artificial pancreas and whole body oxygenation by using hyperbaric therapy. Progress in the field of oxygen technologies for islet transplantation requires a multidisciplinary approach to explore and optimize the interaction between components of the biological system and different technological processes. This review article focuses mainly on the recently developed strategies for oxygenation and protection from hypoxic injury - to achieve stable and long-term normoglycaemia in diabetic patients with transplanted pancreatic islets. PMID:22389124

  17. Proinsulin maturation disorder is a contributor to the defect of subsequent conversion to insulin in {beta}-cells

    SciTech Connect

    Wang, Jie; Osei, Kwame

    2011-07-22

    Highlights: {yields} Primary proinsulin maturation disorder is inherent in Ins2{sup +/Akita} islets/{beta}-cells. {yields} A consequence is the inefficient conversion of proinsulin to insulin. {yields} Post-translational defects occur as well in the involved PC1/3 and PC2 convertases. {yields} Proinsulin maturation chaos results in defects in the following conversion process. {yields} A link of the proinsulin maturation disorder and hyperproinsulinemia is suggested. -- Abstract: Disproportionate hyperproinsulinemia is an indicator of {beta}-cell dysfunction in diabetes and the basis underlying this abnormality remains obscure. Recently, we have found proinsulin is an aggregation-prone molecule inherent with a low relative folding rate and maintains a homeostatic balance of natively and plentiful non-natively folded states (i.e., proinsulin homeostasis, PIHO) in normal {beta}-cells as a result of the integration of maturation and disposal processes. PIHO is susceptible to environmental and genetic influences. Perturbation of PIHO produces a number of toxic consequences with known association to {beta}-cell failure in diabetes. To explore whether the perturbation of PIHO has a link to disproportionate hyperproinsulinemia, we investigated proinsulin conversion and the involved prohormone convertase 1/3 (PC1/3) and 2 (PC2) in mouse Ins2{sup +/Akita} islets/{beta}-cells that preserve a primary PIHO disorder due to a mutation (C96Y) in the insulin 2 (Ins2) gene. Our metabolic-labeling studies found an increased ratio of proinsulin to insulin in the cellular or released proteins of Ins2{sup +/Akita} islets. Histological, metabolic-labeling, and RT-PCR analyses revealed decreases of the PC1/3 and PC2 immunoreactivities in the {beta}-cells of Ins2{sup +/Akita} islets in spite of no declines of these two convertases at the transcriptional and translational levels. Immunoblot analyses in cloned Ins2{sup +/Akita} {beta}-cells further confirmed the increased ratio of proinsulin

  18. Correlation of insulin resistance, beta cell function and insulin sensitivity with serum sFas and sFasL in newly diagnosed type 2 diabetes.

    PubMed

    Kumar, Hemant; Mishra, Manish; Bajpai, Surabhi; Pokhria, Deepa; Arya, Awadhesh Kumar; Singh, Rakesh Kumar; Tripathi, Kamlakar

    2013-08-01

    Pancreatic beta cell dysfunction and reduced insulin sensitivity are fundamental factors associated with glucotoxicity, lipotoxicity and oxidative stress in type 2 diabetic patients (T2DM). Diabetic milieu can induce apoptosis in several types of cells. The aim of present study was to compare circulating soluble apoptotic markers (sFas and sFas-L) with HOMA-IR, HOMA-%S, HOMA-%B in the serum of newly diagnosed T2DM and healthy subjects. For this study, 94 T2DM and 60 healthy subjects were enroled and evaluated for various parameters. Biochemical quantifications were performed with Syncron CX5 auto-analyzer. The levels of serum sFas-L, TNF-α and IL-6 were estimated by flowcytometry. The fasting serum insulin and sFas quantified by ELISA. HOMA-IR, HOMA-%S and HOMA-%B were calculated with HOMA calculator v2.2.2. The levels of TC, TG, LDL-C, VLDL-C were augmented and HDL declined significantly (P < 0.001) in diabetics. The levels of serum insulin, TNF-α, IL-6, sFas, HOMA-IR were raised (P < 0.001) and sFas-L, HOMA-%S and HOMA-%B were decreased significantly (P < 0.001) in T2DM subjects than healthy. In diabetics, serum sFas was positively correlated with HOMA-IR (r = 0.720, P < 0.001) and negatively with HOMA-%B (r = -0.642, P < 0.001) significantly while serum sFasL was negatively correlated with HOMA-IR (r = -0.483, P < 0.001) and positively with HOMA-%B (r = 0.466, P < 0.001) significantly. Further, the multivariate stepwise regression analysis shows that HOMA-IR contributes significantly to the variance of sFas and sFasL. Our findings suggest that the pancreatic beta cell dysfunction along with increased insulin resistance appears to be associated with apoptotic markers. PMID:21695404

  19. PARP-1 and YY1 Are Important Novel Regulators of CXCL12 Gene Transcription in Rat Pancreatic Beta Cells

    PubMed Central

    Marković, Jelena; Grdović, Nevena; Dinić, Svetlana; Karan-Djurašević, Teodora; Uskoković, Aleksandra; Arambašić, Jelena; Mihailović, Mirjana; Pavlović, Sonja; Poznanović, Goran; Vidaković, Melita

    2013-01-01

    Despite significant progress, the molecular mechanisms responsible for pancreatic beta cell depletion and development of diabetes remain poorly defined. At present, there is no preventive measure against diabetes. The positive impact of CXCL12 expression on the pancreatic beta cell prosurvival phenotype initiated this study. Our aim was to provide novel insight into the regulation of rat CXCL12 gene (Cxcl12) transcription. The roles of poly(ADP-ribose) polymerase-1 (PARP-1) and transcription factor Yin Yang 1 (YY1) in Cxcl12 transcription were studied by examining their in vitro and in vivo binding affinities for the Cxcl12 promoter in a pancreatic beta cell line by the electrophoretic mobility shift assay and chromatin immunoprecipitation. The regulatory activities of PARP-1 and YY1 were assessed in transfection experiments using a reporter vector with a Cxcl12 promoter sequence driving luciferase gene expression. Experimental evidence for PARP-1 and YY1 revealed their trans-acting potential, wherein PARP-1 displayed an inhibitory, and YY1 a strong activating effect on Cxcl12 transcription. Streptozotocin (STZ)-induced general toxicity in pancreatic beta cells was followed by changes in Cxcl12 promoter regulation. PARP-1 binding to the Cxcl12 promoter during basal and in STZ-compromised conditions led us to conclude that PARP-1 regulates constitutive Cxcl12 expression. During the early stage of oxidative stress, YY1 exhibited less affinity toward the Cxcl12 promoter while PARP-1 displayed strong binding. These interactions were accompanied by Cxcl12 downregulation. In the later stages of oxidative stress and intensive pancreatic beta cell injury, YY1 was highly expressed and firmly bound to Cxcl12 promoter in contrast to PARP-1. These interactions resulted in higher Cxcl12 expression. The observed ability of PARP-1 to downregulate, and of YY1 to upregulate Cxcl12 promoter activity anticipates corresponding effects in the natural context where the functional

  20. Effect of prolonged exposure to sublethal concentrations of DDT and DDE on protein expression in human pancreatic beta cells.

    PubMed

    Pavlikova, Nela; Smetana, Pavel; Halada, Petr; Kovar, Jan

    2015-10-01

    Pollution of the environment represents one of less explored potential reasons for the worldwide epidemic of type 2 diabetes. One of the most prevalent organochlorine pollutants remains the pesticide DDT and its degradation product DDE. Despite some epidemiologic correlations between levels of DDT and DDE in human organism and the prevalence of diabetes, there is almost no information about the exact targets of these compounds inside pancreatic beta cells. To detect functional areas of pancreatic beta cells that could be affected by exposure to DDT and DDE, we analyzed changes in protein expression in the NES2Y human pancreatic beta cell line exposed to three sublethal concentrations (0.1 μM, 1 μM, 10 μM) of DDT and DDE for 1 month. Protein separation and identification was achieved using high-resolution 2D-electrophoresis, computer analysis and mass spectrometry. With these techniques, four proteins were found downregulated after exposure to 10 μM DDT: three cytoskeletal proteins (cytokeratin 8, cytokeratin 18 and actin) and one protein involved in glycolysis (alpha-enolase). Two proteins were downregulated after exposure to 10 μM DDE: cytokeratin 18 and heterogenous nuclear ribonucleoprotein H1 (HNRH1). These changes correlate with previously described effects of other stress conditions (e.g. exposure to palmitate, hyperglycemia, imidazoline derivative, and cytokines) on protein expression in pancreatic beta cells. We conclude that cytoskeletal proteins and their processing, glucose metabolism, and mRNA processing may represent targets affected by exposure to conditions hostile to pancreatic beta cells, including exposure to DDT and DDE.

  1. Transforming growth factor-beta/Smad3 signaling regulates insulin gene transcription and pancreatic islet beta-cell function.

    PubMed

    Lin, Huei-Min; Lee, Ji-Hyeon; Yadav, Hariom; Kamaraju, Anil K; Liu, Eric; Zhigang, Duan; Vieira, Anthony; Kim, Seong-Jin; Collins, Heather; Matschinsky, Franz; Harlan, David M; Roberts, Anita B; Rane, Sushil G

    2009-05-01

    Pancreatic islet beta-cell dysfunction is a signature feature of Type 2 diabetes pathogenesis. Consequently, knowledge of signals that regulate beta-cell function is of immense clinical relevance. Transforming growth factor (TGF)-beta signaling plays a critical role in pancreatic development although the role of this pathway in the adult pancreas is obscure. Here, we define an important role of the TGF-beta pathway in regulation of insulin gene transcription and beta-cell function. We identify insulin as a TGF-beta target gene and show that the TGF-beta signaling effector Smad3 occupies the insulin gene promoter and represses insulin gene transcription. In contrast, Smad3 small interfering RNAs relieve insulin transcriptional repression and enhance insulin levels. Transduction of adenoviral Smad3 into primary human and non-human primate islets suppresses insulin content, whereas, dominant-negative Smad3 enhances insulin levels. Consistent with this, Smad3-deficient mice exhibit moderate hyperinsulinemia and mild hypoglycemia. Moreover, Smad3 deficiency results in improved glucose tolerance and enhanced glucose-stimulated insulin secretion in vivo. In ex vivo perifusion assays, Smad3-deficient islets exhibit improved glucose-stimulated insulin release. Interestingly, Smad3-deficient islets harbor an activated insulin-receptor signaling pathway and TGF-beta signaling regulates expression of genes involved in beta-cell function. Together, these studies emphasize TGF-beta/Smad3 signaling as an important regulator of insulin gene transcription and beta-cell function and suggest that components of the TGF-beta signaling pathway may be dysregulated in diabetes.

  2. Adult Human Pancreatic Islet Beta-Cells Display Limited Turnover and Long Lifespan as Determined by In-Vivo Thymidine Analog Incorporation and Radiocarbon Dating

    SciTech Connect

    Perl, S; Kushner, J A; Buchholz, B A; Meeker, A K; Stein, G M; Hsieh, M; Kirby, M; Pechhold, S; Liu, E H; Harlan, D M; Tisdale, J F

    2010-03-15

    Diabetes mellitus results from an absolute or relative deficiency of insulin producing pancreatic beta-cells. The adult human beta-cell's turnover rate remains unknown. We employed novel techniques to examine adult human islet beta-cell turnover and longevity in vivo. Subjects enrolled in NIH clinical trials received thymidine analogues [iododeoxyuridine (IdU) or bromodeoxyuridine (BrdU)] 8-days to 4-years prior to death. Archival autopsy samples from ten patients (aged 17-74 years) were employed to assess beta-cell turnover by scoring nuclear analog labeling within insulin staining cells. Human adult beta-cell longevity was determined by estimating the cells genomic DNA integration of atmospheric carbon-14 ({sup 14}C). DNA was purified from pancreatic islets isolated from cadaveric donors; whole islet prep DNA was obtained from a 15 year old donor, and purified beta-cell DNA was obtained from two donors (age 48 and 80 years). {sup 14}C levels were then determined using accelerator mass spectrometry (AMS). Cellular 'birth date' was determined by comparing the subject's DNA {sup 14}C content relative to a well-established {sup 14}C atmospheric prevalence curve. In the two subjects less than age 20 years, 1-2% of the beta-cell nuclei co-stained for BrdU/IdU. No beta-cell nuclei co-stained in the eight patients more than 30 years old. Consistent with the BrdU/IdU turnover data, beta-cell DNA {sup 14}C content indicated the cells 'birth date' occurred within the subject's first 30 years of life. Under typical circumstances, adult human beta-cells and their cellular precursors are established by young adulthood.

  3. Properties of single potassium channels modulated by glucose in rat pancreatic beta-cells.

    PubMed Central

    Ashcroft, F M; Ashcroft, S J; Harrison, D E

    1988-01-01

    1. The patch clamp method has been used to examine the effect of glucose on single K+ channel currents recorded from cell-attached patches on dissociated rat pancreatic beta-cells. Patch pipettes contained a 140 mM-K+ solution. 2. In glucose-free solution three types of K+ channels were observed. Two of these, having conductances of around 50 pS (G-channel) and 20 pS when the external K+ concentration, [K+]0, was 140 mM, were active at the resting potential of the cell. The G-channel was observed in more patches and showed higher activity; it therefore appears to contribute the major fraction of the resting K+ permeability of the beta-cell. At membrane potentials positive to about +20 mV a third type of K+ channel, having a mean conductance of 120 pS, was activated. The open probability of this channel was strongly voltage dependent and increased with depolarization. 3. The reversal potential of the G-channel current was shifted 59 mV by a 10-fold change in external K+ (Na+ substitution) indicating the channel is highly K+ selective. The single-channel conductance varied with [K+]o as predicted from the Goldman-Hodgkin-Katz equation; at physiological [K+]o (5 mM-K+) an inward conductance of around 10 pS is predicted. The amplitude of the single-channel current showed a tendency to saturate with increasing [K+]o. 4. Single G-channel currents show burst kinetics indicating at least two closed states. The open and closed (gap) times within the bursts were distributed exponentially with time constants of 2.5 ms (tau o) and 0.5 ms (tau c1) respectively at the resting potential of the cell. There was little change in tau c1 over the voltage range -40 to 60 mV (pipette potential) but tau o increased slightly with membrane depolarization. 5. The addition of glucose to the bath solution produced a reversible, dose-dependent decrease in G-channel activity. This decrease results principally from a reduction in the frequency and duration of the bursts of openings with

  4. MicroRNA-29a is up-regulated in beta-cells by glucose and decreases glucose-stimulated insulin secretion

    SciTech Connect

    Bagge, Annika; Clausen, Trine R.; Larsen, Sylvester; Ladefoged, Mette; Rosenstierne, Maiken W.; Larsen, Louise; Vang, Ole; Nielsen, Jens H.; Dalgaard, Louise T.

    2012-09-21

    Highlights: Black-Right-Pointing-Pointer MicroRNA-29a (miR-29a) levels are increased by glucose in human and rat islets and INS-1E cells. Black-Right-Pointing-Pointer miR-29a increases proliferation of INS-1E beta-cells. Black-Right-Pointing-Pointer Forced expression of miR-29a decreases glucose-stimulated insulin secretion (GSIS). Black-Right-Pointing-Pointer Depletion of beta-cell miR-29a improves GSIS. Black-Right-Pointing-Pointer miR-29a may be a mediator of glucose toxicity in beta-cells. -- Abstract: Chronically elevated levels of glucose impair pancreatic beta-cell function while inducing beta-cell proliferation. MicroRNA-29a (miR-29a) levels are increased in several tissues in diabetic animals and mediate decreased insulin-stimulated glucose-transport of adipocytes. The aim was to investigate the impact of glucose on miR-29a levels in INS-1E beta-cells and in human islets of Langerhans and furthermore to evaluate the impact of miR-29a on beta-cell function and proliferation. Increased glucose levels up-regulated miR-29a in beta-cells and human and rat islets of Langerhans. Glucose-stimulated insulin-secretion (GSIS) of INS-1E beta-cells was decreased by forced expression of miR-29a, while depletion of endogenous miR-29a improved GSIS. Over-expression of miR-29a increased INS-1E proliferation. Thus, miR-29a up-regulation is involved in glucose-induced proliferation of beta-cells. Furthermore, as depletion of miR-29a improves beta-cell function, miR-29a is a mediator of glucose-induced beta-cell dysfunction. Glucose-induced up-regulation of miR-29a in beta-cells could be implicated in progression from impaired glucose tolerance to type 2 diabetes.

  5. Dietary Intervention in Infancy and Later Signs of Beta-Cell Autoimmunity

    PubMed Central

    Knip, Mikael; Virtanen, Suvi M.; Seppä, Karri; Ilonen, Jorma; Savilahti, Erkki; Vaarala, Outi; Reunanen, Antti; Teramo, Kari; Hämäläinen, Anu-Maaria; Paronen, Johanna; Dosch, Hans-Michael; Hakulinen, Timo; Åkerblom, Hans K.

    2014-01-01

    BACKGROUND Early exposure to complex dietary proteins may increase the risk of beta-cell autoimmunity and type 1 diabetes in children with genetic susceptibility. We tested the hypothesis that supplementing breast milk with highly hydrolyzed milk formula would decrease the cumulative incidence of diabetes-associated autoantibodies in such children. METHODS In this double-blind, randomized trial, we assigned 230 infants with HLA-conferred susceptibility to type 1 diabetes and at least one family member with type 1 diabetes to receive either a casein hydrolysate formula or a conventional, cow’s-milk–based formula (control) whenever breast milk was not available during the first 6 to 8 months of life. Autoantibodies to insulin, glutamic acid decarboxylase (GAD), the insulinoma-associated 2 molecule (IA-2), and zinc transporter 8 were analyzed with the use of radiobinding assays, and islet-cell antibodies were analyzed with the use of immunofluorescence, during a median observation period of 10 years (mean, 7.5). The children were monitored for incident type 1 diabetes until they were 10 years of age. RESULTS The unadjusted hazard ratio for positivity for one or more autoantibodies in the casein hydrolysate group, as compared with the control group, was 0.54 (95% confidence interval [CI], 0.29 to 0.95), and the hazard ratio adjusted for an observed difference in the duration of exposure to the study formula was 0.51 (95% CI, 0.28 to 0.91). The unadjusted hazard ratio for positivity for two or more autoantibodies was 0.52 (95% CI, 0.21 to 1.17), and the adjusted hazard ratio was 0.47 (95% CI, 0.19 to 1.07). The rate of reported adverse events was similar in the two groups. CONCLUSIONS Dietary intervention during infancy appears to have a long-lasting effect on markers of beta-cell autoimmunity — markers that may reflect an autoimmune process leading to type 1 diabetes. (Funded by the European Commission and others; ClinicalTrials.gov number, NCT00570102.) PMID

  6. Role for c-Jun N-terminal kinase in beta-cell recovery from nitric oxide-mediated damage.

    PubMed

    Scarim, Anna L; Nishimoto, Sheri Y; Weber, Sarah M; Corbett, John A

    2003-08-01

    Treatment of rat islets with the cytokine IL-1 results in the inhibition of mitochondrial function and insulin secretion, events that are mediated by beta-cell expression of iNOS [inducible nitric oxide (NO) synthase] and production of NO. beta-Cells recover from the inhibitory actions of NO, produced following 24 h incubation with IL-1, on islet oxidative metabolism and insulin secretion if iNOS enzymatic activity is inhibited and the islets are cultured (in the presence of IL-1 and iNOS inhibitors) for a brief period of 8 h. Islet recovery from cytokine- and NO-mediated damage is an active process that requires new gene expression, and NO itself is one activator of this recovery process. In this study, the mechanism by which NO stimulates islet recovery has been examined. Incubation of rat islets or RINm5F cells with the NO donor compound, sodium (Z)-1(N,N-diethylamino) diazen-1-ium-1,2-diolate (DEA-NO) for 1 h results in a 60% inhibition of mitochondrial aconitase activity. beta-Cells completely recover aconitase activity if the cells are washed to remove the NO donor compound and incubated for an additional 5 h in the absence of DEA-NO. The recovery of mitochondrial aconitase activity correlates with a 4-fold increase in cyclic GMP accumulation and is prevented by the inhibition of guanylate cyclase. The recovery of aconitase activity also correlates with the activation of members of the MAPKs, p38, c-Jun N-terminal kinase (JNK) and ERK, and the activation p38 and JNK is attenuated by inhibition of guanylate cyclase. ERK and p38 do not appear to participate in the recovery process as selective inhibition of these kinases fails to prevent recovery of aconitase activity; however, transduction of beta-cells with a dominant negative mutant JNK prevents beta-cell recovery from NO-mediated damage. These findings support a role for guanylate cyclase and JNK in the recovery of beta-cells from NO-mediated damage. PMID:12865320

  7. Microbial phenolic metabolites improve glucose-stimulated insulin secretion and protect pancreatic beta cells against tert-butyl hydroperoxide-induced toxicity via ERKs and PKC pathways.

    PubMed

    Fernández-Millán, Elisa; Ramos, Sonia; Alvarez, Carmen; Bravo, Laura; Goya, Luis; Martín, María Ángeles

    2014-04-01

    Oxidative stress is accepted as one of the causes of beta cell failure in type 2 diabetes. Therefore, identification of natural antioxidant agents that preserve beta cell mass and function is considered an interesting strategy to prevent or treat diabetes. Recent evidences indicated that colonic metabolites derived from flavonoids could possess beneficial effects on various tissues. The aim of this work was to establish the potential anti-diabetic properties of the microbial-derived flavonoid metabolites 3,4-dihydroxyphenylacetic acid (DHPAA), 2,3-dihydroxybenzoic acid (DHBA) and 3-hydroxyphenylpropionic acid (HPPA). To this end, we tested their ability to influence beta cell function and to protect against tert-butyl hydroperoxide-induced beta cell toxicity. DHPAA and HPPA were able to potentiate glucose-stimulated insulin secretion (GSIS) in a beta cell line INS-1E and in rat pancreatic islets. Moreover, pre-treatment of cells with both compounds protected against beta cell dysfunction and death induced by the pro-oxidant. Finally, experiments with pharmacological inhibitors indicate that these effects were mediated by the activation of protein kinase C and the extracellular regulated kinases pathways. Altogether, these findings strongly suggest that the microbial-derived flavonoid metabolites DHPAA and HPPA may have anti-diabetic potential by promoting survival and function of pancreatic beta cells. PMID:24491264

  8. CTLs are targeted to kill beta cells in patients with type 1 diabetes through recognition of a glucose-regulated preproinsulin epitope.

    PubMed

    Skowera, Ania; Ellis, Richard J; Varela-Calviño, Ruben; Arif, Sefina; Huang, Guo Cai; Van-Krinks, Cassie; Zaremba, Anna; Rackham, Chloe; Allen, Jennifer S; Tree, Timothy I M; Zhao, Min; Dayan, Colin M; Sewell, Andrew K; Unger, Wendy W; Unger, Wendy; Drijfhout, Jan W; Ossendorp, Ferry; Roep, Bart O; Peakman, Mark

    2008-10-01

    The final pathway of beta cell destruction leading to insulin deficiency, hyperglycemia, and clinical type 1 diabetes is unknown. Here we show that circulating CTLs can kill beta cells via recognition of a glucose-regulated epitope. First, we identified 2 naturally processed epitopes from the human preproinsulin signal peptide by elution from HLA-A2 (specifically, the protein encoded by the A*0201 allele) molecules. Processing of these was unconventional, requiring neither the proteasome nor transporter associated with processing (TAP). However, both epitopes were major targets for circulating effector CD8+ T cells from HLA-A2+ patients with type 1 diabetes. Moreover, cloned preproinsulin signal peptide-specific CD8+ T cells killed human beta cells in vitro. Critically, at high glucose concentration, beta cell presentation of preproinsulin signal epitope increased, as did CTL killing. This study provides direct evidence that autoreactive CTLs are present in the circulation of patients with type 1 diabetes and that they can kill human beta cells. These results also identify a mechanism of self-antigen presentation that is under pathophysiological regulation and could expose insulin-producing beta cells to increasing cytotoxicity at the later stages of the development of clinical diabetes. Our findings suggest that autoreactive CTLs are important targets for immune-based interventions in type 1 diabetes and argue for early, aggressive insulin therapy to preserve remaining beta cells.

  9. Molecular mechanisms involved in secretory vesicle recruitment to the plasma membrane in beta-cells.

    PubMed

    Varadi, Aniko; Ainscow, E K; Allan, V J; Rutter, G A

    2002-04-01

    Glucose stimulates the release of insulin in part by activating the recruitment of secretory vesicles to the cell surface. While this movement is known to be microtubule-dependent, the molecular motors involved are undefined. Active kinesin was found to be essential for vesicle translocation in live beta-cells, since microinjection of cDNA encoding dominant-negative KHC(mut) (motor domain of kinesin heavy chain containing a Thr(93)-->Asn point mutation) blocked vesicular movements. Moreover, expression of KHC(mut) strongly inhibited the sustained, but not acute, stimulation of secretion by glucose. Thus, vesicles released during the first phase of insulin secretion exist largely within a translocation-independent pool. Kinesin-driven anterograde movement of vesicles is then necessary for the sustained (second phase) of insulin release. Kinesin may, therefore, represent a novel target for increases in intracellular ATP concentrations in response to elevated extracellular glucose and may be involved in the ATP-sensitive K+channel-independent stimulation of secretion by the sugar.

  10. Antibodies to new beta cell antigen ICA12 in Latvian diabetes patients.

    PubMed

    Shtauvere-Brameus, A; Hagopian, W; Rumba, I; Sanjeevi, C B

    2002-04-01

    In Latvia diabetes mellitus is diagnosed using the WHO's clinical criteria, and assays for the detection of autoantibodies are not available. In consequence, slowly progressive autoimmune diabetes or LADA is likely to be missed. Antibodies to GAD65 and IA-2 are the major immunological markers in autoimmune diabetes. Recently, a new beta cell antigen, called ICA12, has been identified, which has a homology to the SOX family of transcription factors. The aim of the study was to analyze the prevalence of ICA12 antibodies in diabetes mellitus patients and controls from Latvia and to see whether this antigen is important in revealing autoimmunity when antibodies against major antigens are not present. We studied 88 IDDM patients and 100 NIDDM patients as well as controls for the prevalence of GAD65, IA-2, and ICA12 antibodies by radioligand binding assay (RIA) using (35)S-labeled islet antigens. We found ICA12Abs in 26 of 88 IDDM patients (30%) vs. 4% in healthy controls (4/100) and in 9 of 100 NIDDM patients (9%) vs. 2% controls (2/100). ICA12Abs alone are present in only 3% (3/88) of the patients with IDDM and 1% (1/100) of the NIDDM patients. We conclude that ICA12 represents the minor antigens in autoimmune diabetes and that, as a minor antigen, ICA12 alone does not contribute significantly in revealing new cases of autoimmunity.

  11. GTP- and GDP-Dependent Rab27a Effectors in Pancreatic Beta-Cells.

    PubMed

    Yamaoka, Mami; Ishizaki, Toshimasa; Kimura, Toshihide

    2015-01-01

    Small guanosine triphosphatases (GTPases) participate in a wide variety of cellular functions including proliferation, differentiation, adhesion, and intracellular transport. Conventionally, only the guanosine 5'-triphosphate (GTP)-bound small GTPase interacts with effector proteins, and the resulting downstream signals control specific cellular functions. Therefore, the GTP-bound form is regarded as active, and the focus has been on searching for proteins that bind the GTP form to look for their effectors. The Rab family small GTPase Rab27a is highly expressed in some secretory cells and is involved in the control of membrane traffic. The present study reviews recent progress in our understanding of the roles of Rab27a and its effectors in pancreatic beta-cells. In the basal state, GTP-bound Rab27a controls insulin secretion at pre-exocytic stages via its GTP-dependent effectors. We previously identified novel guanosine 5'-diphosphate (GDP)-bound Rab27-interacting proteins. Interestingly, GDP-bound Rab27a controls endocytosis of the secretory membrane via its interaction with these proteins. We also demonstrated that the insulin secretagogue glucose converts Rab27a from its GTP- to GDP-bound forms. Thus, GTP- and GDP-bound Rab27a regulate pre-exocytic and endocytic stages in membrane traffic, respectively. Since the physiological importance of GDP-bound GTPases has been largely overlooked, we consider that the investigation of GDP-dependent effectors for other GTPases is necessary for further understanding of cellular function.

  12. Evidence that an isoform of calpain-10 is a regulator of exocytosis in pancreatic beta-cells.

    PubMed

    Marshall, Catriona; Hitman, Graham A; Partridge, Christopher J; Clark, Anne; Ma, Hong; Shearer, Thomas R; Turner, Mark D

    2005-01-01

    Calpain-10 (CAPN10) is the first type 2 diabetes susceptibility gene to be identified through a genome scan, with polymorphisms being associated with altered CAPN10 expression. Functional data have been hitherto elusive, but we report here a corresponding increase between CAPN10 expression level and regulated insulin secretion. Pancreatic beta-cell secretory granule exocytosis is mediated by the soluble N-ethylmaleimide-sensitive fusion protein attachment receptor protein complex of synaptosomal-associated protein of 25 kDa (SNAP-25), syntaxin 1, and vesicle-associated membrane protein 2. We report, for the first time, direct binding of a calpain-10 isoform with members of this complex. Furthermore, SNAP-25 undergoes a Ca2+-dependent partial proteolysis during exocytosis, with calpain protease inhibitor similarly suppressing both insulin secretion and SNAP-25 proteolysis. Based upon these findings, we postulate that an isoform of calpain-10 is a Ca2+-sensor that functions to trigger exocytosis in pancreatic beta-cells.

  13. Measuring phospholipase D activity in insulin-secreting pancreatic beta-cells and insulin-responsive muscle cells and adipocytes.

    PubMed

    Cazzolli, Rosanna; Huang, Ping; Teng, Shuzhi; Hughes, William E

    2009-01-01

    Phospholipase D (PLD) is an enzyme producing phosphatidic acid and choline through hydrolysis of phosphatidylcholine. The enzyme has been identified as a member of a variety of signal transduction cascades and as a key regulator of numerous intracellular vesicle trafficking processes. A role for PLD in regulating glucose homeostasis is emerging as the enzyme has recently been identified in events regulating exocytosis of insulin from pancreatic beta-cells and also in insulin-stimulated glucose uptake through controlling GLUT4 vesicle exocytosis in muscle and adipose tissue. We present methodologies for assessing cellular PLD activity in secretagogue-stimulated insulin-secreting pancreatic beta-cells and also insulin-stimulated adipocyte and muscle cells, two of the principal insulin-responsive cell types controlling blood glucose levels. PMID:19160674

  14. Measuring phospholipase D activity in insulin-secreting pancreatic beta-cells and insulin-responsive muscle cells and adipocytes.

    PubMed

    Cazzolli, Rosanna; Huang, Ping; Teng, Shuzhi; Hughes, William E

    2009-01-01

    Phospholipase D (PLD) is an enzyme producing phosphatidic acid and choline through hydrolysis of phosphatidylcholine. The enzyme has been identified as a member of a variety of signal transduction cascades and as a key regulator of numerous intracellular vesicle trafficking processes. A role for PLD in regulating glucose homeostasis is emerging as the enzyme has recently been identified in events regulating exocytosis of insulin from pancreatic beta-cells and also in insulin-stimulated glucose uptake through controlling GLUT4 vesicle exocytosis in muscle and adipose tissue. We present methodologies for assessing cellular PLD activity in secretagogue-stimulated insulin-secreting pancreatic beta-cells and also insulin-stimulated adipocyte and muscle cells, two of the principal insulin-responsive cell types controlling blood glucose levels.

  15. Toxins from Physalia physalis (Cnidaria) raise the intracellular Ca(2+) of beta-cells and promote insulin secretion.

    PubMed

    Diaz-Garcia, C M; Fuentes-Silva, D; Sanchez-Soto, C; Domínguez-Pérez, D; García-Delgado, N; Varela, C; Mendoza-Hernández, G; Rodriguez-Romero, A; Castaneda, O; Hiriart, M

    2012-01-01

    Physalia physalis is a marine cnidarian from which high molecular weight toxins with hemolytic and neurotoxic effects have been isolated. In the present work, two novel toxins, PpV9.4 and PpV19.3 were purified from P. physalis by bioactive guideline isolation. It involved two steps of column chromatography, gel filtration and RP-HPLC. The molecular weights were 550.7 and 4720.9 Da for PpV9.4 and PpV19.3, respectively. In the light of the Edman sequencing results, the structure of these toxins included the presence of modified amino acids. Both toxins increased the percentage of insulin secreting beta-cells and induced cytosolic Ca2+ elevation. To date, this is the first report of low molecular weight toxins increasing insulin secretion purified from cnidarians, by constituting a new approach to the study of beta-cells physiology. PMID:22830340

  16. XIAP discriminates between type I and type II FAS-induced apoptosis.

    PubMed

    Jost, Philipp J; Grabow, Stephanie; Gray, Daniel; McKenzie, Mark D; Nachbur, Ueli; Huang, David C S; Bouillet, Philippe; Thomas, Helen E; Borner, Christoph; Silke, John; Strasser, Andreas; Kaufmann, Thomas

    2009-08-20

    FAS (also called APO-1 and CD95) and its physiological ligand, FASL, regulate apoptosis of unwanted or dangerous cells, functioning as a guardian against autoimmunity and cancer development. Distinct cell types differ in the mechanisms by which the 'death receptor' FAS triggers their apoptosis. In type I cells, such as lymphocytes, activation of 'effector caspases' by FAS-induced activation of caspase-8 suffices for cell killing, whereas in type II cells, including hepatocytes and pancreatic beta-cells, caspase cascade amplification through caspase-8-mediated activation of the pro-apoptotic BCL-2 family member BID (BH3 interacting domain death agonist) is essential. Here we show that loss of XIAP (X-chromosome linked inhibitor of apoptosis protein) function by gene targeting or treatment with a second mitochondria-derived activator of caspases (SMAC, also called DIABLO; direct IAP-binding protein with low pI) mimetic drug in mice rendered hepatocytes and beta-cells independent of BID for FAS-induced apoptosis. These results show that XIAP is the critical discriminator between type I and type II apoptosis signalling and suggest that IAP inhibitors should be used with caution in cancer patients with underlying liver conditions.

  17. Inter-domain tagging implicates caveolin-1 in insulin receptor trafficking and Erk signaling bias in pancreatic beta-cells

    PubMed Central

    Boothe, Tobias; Lim, Gareth E.; Cen, Haoning; Skovsø, Søs; Piske, Micah; Li, Shu Nan; Nabi, Ivan R.; Gilon, Patrick; Johnson, James D.

    2016-01-01

    Objective The role and mechanisms of insulin receptor internalization remain incompletely understood. Previous trafficking studies of insulin receptors involved fluorescent protein tagging at their termini, manipulations that may be expected to result in dysfunctional receptors. Our objective was to determine the trafficking route and molecular mechanisms of functional tagged insulin receptors and endogenous insulin receptors in pancreatic beta-cells. Methods We generated functional insulin receptors tagged with pH-resistant fluorescent proteins between domains. Confocal, TIRF and STED imaging revealed a trafficking pattern of inter-domain tagged insulin receptors and endogenous insulin receptors detected with antibodies. Results Surprisingly, interdomain-tagged and endogenous insulin receptors in beta-cells bypassed classical Rab5a- or Rab7-mediated endocytic routes. Instead, we found that removal of insulin receptors from the plasma membrane involved tyrosine-phosphorylated caveolin-1, prior to trafficking within flotillin-1-positive structures to lysosomes. Multiple methods of inhibiting caveolin-1 significantly reduced Erk activation in vitro or in vivo, while leaving Akt signaling mostly intact. Conclusions We conclude that phosphorylated caveolin-1 plays a role in insulin receptor internalization towards lysosomes through flotillin-1-positive structures and that caveolin-1 helps bias physiological beta-cell insulin signaling towards Erk activation. PMID:27110488

  18. GeneSpeed Beta Cell: An Online Genomics Data Repository and Analysis Resource Tailored for the Islet Cell Biologist

    PubMed Central

    Quayum, Nayeem; Kutchma, Alecksandr; Sarkar, Suparna A.; Juhl, Kirstine; Gradwohl, Gerard; Mellitzer, Georg; Hutton, John C.; Jensen, Jan

    2008-01-01

    Objective. We here describe the development of a freely available online database resource, GeneSpeed Beta Cell, which has been created for the pancreatic islet and pancreatic developmental biology investigator community. Research Design and Methods. We have developed GeneSpeed Beta Cell as a separate component of the GeneSpeed database, providing a genomics-type data repository of pancreas and islet-relevant datasets interlinked with the domain-oriented GeneSpeed database. Results. GeneSpeed Beta Cell allows the query of multiple published and unpublished select genomics datasets in a simultaneous fashion (multiexperiment viewing) and is capable of defining intersection results from precomputed analysis of such datasets (multidimensional querying). Combined with the protein-domain categorization/assembly toolbox provided by the GeneSpeed database, the user is able to define spatial expression constraints of select gene lists in a relatively rigid fashion within the pancreatic expression space. We provide several demonstration case studies of relevance to islet cell biology and development of the pancreas that provide novel insight into islet biology. Conclusions. The combination of an exhaustive domain-based compilation of the transcriptome with gene array data of interest to the islet biologist affords novel methods for multidimensional querying between individual datasets in a rapid fashion, presently not available elsewhere. PMID:18795106

  19. Usurping the mitochondrial supremacy: extramitochondrial sources of reactive oxygen intermediates and their role in beta cell metabolism and insulin secretion.

    PubMed

    Gray, Joshua P; Heart, Emma

    2010-05-01

    Insulin secretion from pancreatic beta cells is a process dependent on metabolism. While oxidative stress is a well-known inducer of beta cell toxicity and impairs insulin secretion, recent studies suggest that low levels of metabolically-derived reactive oxygen intermediates (ROI) also play a positive role in insulin secretion. Glucose metabolism is directly correlated with ROI production, particularly in beta cells in which glucose uptake is proportional to the extracellular concentration of glucose. Low levels of exogenously added ROI or quinones, which stimulate ROI production, positively affect insulin secretion, while antioxidants block insulin secretion, suggesting that ROI activate unidentified redox-sensitive signal transduction components within these cells. The mitochondria are one source of ROI: increased metabolic flux increases mitochondrial membrane potential resulting in electron leakage and adventitious ROI production. A second source of ROI are cytosolic and plasma membrane oxidoreductases which oxidize NAD(P)H and directly produce ROI through the reduction of molecular oxygen. The mechanism of ROI-mediated potentiation of insulin secretion remains an important topic for future study.

  20. Impaired beta-cell functions induced by chronic exposure of cultured human pancreatic islets to high glucose.

    PubMed

    Marshak, S; Leibowitz, G; Bertuzzi, F; Socci, C; Kaiser, N; Gross, D J; Cerasi, E; Melloul, D

    1999-06-01

    In type 2 diabetes, chronic hyperglycemia has been suggested to be detrimental to beta-cell function, causing reduced glucose-stimulated insulin secretion and disproportionately elevated proinsulin. In the present study, we investigated the effect on several beta-cell functions of prolonged in vitro exposure of human pancreatic islet cultures to high glucose concentrations. Islets exposed to high glucose levels (33 mmol/l) for 4 and 9 days showed dramatic decreases in glucose-induced insulin release and in islet insulin content, with increased proportion of proinsulin-like peptides relative to insulin. The depletion in insulin stores correlated with the reduction in insulin mRNA levels and human insulin promoter transcriptional activity. We also demonstrated that high glucose dramatically lowered the binding activity of pancreatic duodenal homeobox 1 (the glucose-sensitive transcription factor), whereas the transcription factor rat insulin promoter element 3b1 activator was less influenced and insulin enhancer factor 1 remained unaffected. Most of these beta-cell impairments were partially reversible when islets first incubated for 6 days in high glucose were transferred to normal glucose (5.5 mmol/l) concentrations for 3 days. We conclude that cultured human islets are sensitive to the deleterious effect of high glucose concentrations at multiple functional levels, and that such mechanisms may play an important role in the decreased insulin production and secretion of type 2 diabetic patients. PMID:10342809

  1. Induced ICER I{gamma} down-regulates cyclin A expression and cell proliferation in insulin-producing {beta} cells

    SciTech Connect

    Inada, Akari; Weir, Gordon C.; Bonner-Weir, Susan . E-mail: susan.bonner-weir@joslin.harvard.edu

    2005-04-15

    We have previously found that cyclin A expression is markedly reduced in pancreatic {beta}-cells by cell-specific overexpression of repressor inducible cyclic AMP early repressor (ICER I{gamma}) in transgenic mice. Here we further examined regulatory effects of ICER I{gamma} on cyclin A gene expression using Min6 cells, an insulin-producing cell line. The cyclin A promoter luciferase assay showed that ICER I{gamma} directly repressed cyclin A gene transcription. In addition, upon ICER I{gamma} overexpression, cyclin A mRNA levels markedly decreased, thereby confirming an inhibitory effect of ICER I{gamma} on cyclin A expression. Suppression of cyclin A results in inhibition of BrdU incorporation. Under normal culture conditions endogenous cyclin A is abundant in these cells, whereas ICER is hardly detectable. However, serum starvation of Min6 cells induces ICER I{gamma} expression with a concomitant very low expression level of cyclin A. Cyclin A protein is not expressed unless the cells are in active DNA replication. These results indicate a potentially important anti-proliferative effect of ICER I{gamma} in pancreatic {beta} cells. Since ICER I{gamma} is greatly increased in diabetes as well as in FFA- or high glucose-treated islets, this effect may in part exacerbate diabetes by limiting {beta}-cell proliferation.

  2. TRPM4 impacts on Ca2+ signals during agonist-induced insulin secretion in pancreatic beta-cells.

    PubMed

    Marigo, V; Courville, K; Hsu, W H; Feng, J M; Cheng, H

    2009-02-27

    TRPM4 is a Ca(2+)-activated non-selective cation (CAN) channel that functions in cell depolarization, which is important for Ca(2+) influx and insulin secretion in pancreatic beta-cells. We investigated TRPM4 expression and function in the beta-cell lines HIT-T15 (hamster), RINm5F (rat), beta-TC3 (mouse), MIN-6 (mouse) and the alpha-cell line INR1G9 (hamster). By RT-PCR, we identified TRPM4 transcripts in alpha- and beta-cells. Patch-clamp recordings with increasing Ca(2+) concentrations resulted in a dose-dependent activation of TRPM4 with the greatest depolarizing currents recorded from hamster-derived cells. Further, Ca(2+) imaging experiments revealed that inhibition of TRPM4 by a dominant-negative effect significantly decreased the magnitude of the Ca(2+) signals generated by agonist stimulation compared to control cells. The decrease in the [Ca(2+)](i) resulted in reduced insulin secretion. Our data suggest that depolarizing currents generated by TRPM4 are an important component in the control of intracellular Ca(2+) signals necessary for insulin secretion and perhaps glucagon from alpha-cells.

  3. Canine Fibroblast Growth Factor 21 Ameliorates Hyperglycemia Associated with Inhibiting Hepatic Gluconeogenesis and Improving Pancreatic Beta-Cell Survival in Diabetic Mice and Dogs.

    PubMed

    Xu, Pengfei; Zhang, Yingjie; Jiang, Xinghao; Li, Junyan; Song, Liying; Khoso, Mir Hasson; Liu, Yunye; Wu, Qiang; Ren, Guiping; Li, Deshan

    2016-01-01

    Diabetes mellitus is a common endocrinopathy in dog. Fibroblast growth factor 21 (FGF-21) is a secreted protein, which is involved in glucose homeostasis. We speculate that the recombinant canine FGF-21 (cFGF-21) has the potential to become a powerful therapeutics to treat canine diabetes. The cFGF-21 gene was cloned and expressed in E. coli Rosetta (DE3). After purification, a cFGF-21 protein with the purity exceeding 95% was obtained. Mouse 3T3-L1 adipocytes and type 1 diabetic mice/dogs induced by STZ were used to examine the biological activity of cFGF-21 in vitro and in vivo, respectively. Results showed that cFGF-21 stimulated glucose uptake in adipocytes significantly in a dose-dependent manner, and reduced plasma glucose significantly in diabetic mice/dogs. After treatment with cFGF-21, the serum insulin level, glycosylated hemoglobin (HbA1c) level and the expressions of the hepatic gluconeogenesis genes (glucose-6-phosphatase, G6Pase and phosphoenolpyruvate carboxykinase, PCK) of the diabetic mice/dogs were attenuated significantly. In the mouse experiment, we also found that the phosphorylation of signal transducer and activator of transcription 3 (STAT3) and the expression of suppressor of cytokine signaling 3 (SOCS3) were up-regulated significantly in the livers after treatment. Histopathological and immunohistochemical results showed that treatment with cFGF-21 promoted recovery of pancreatic islets from STZ-induced apoptosis. Besides, we also found that treatment with cFGF-21 protected liver against STZ or hyperglycemia induced damage and the mechanism of this action associated with inhibiting oxidative stress. In conclusion, cFGF-21 represents a promising candidate for canine diabetes therapeutics. The mechanism of cFGF-21 ameliorates hyperglycemia associated with inhibiting hepatic gluconeogenesis by regulation of STAT3 signal pathway and improving pancreatic beta-cell survival. PMID:27203422

  4. Canine Fibroblast Growth Factor 21 Ameliorates Hyperglycemia Associated with Inhibiting Hepatic Gluconeogenesis and Improving Pancreatic Beta-Cell Survival in Diabetic Mice and Dogs

    PubMed Central

    Xu, Pengfei; Zhang, Yingjie; Jiang, Xinghao; Li, Junyan; Song, Liying; Khoso, Mir Hasson; Liu, Yunye; Wu, Qiang; Ren, Guiping; Li, Deshan

    2016-01-01

    Diabetes mellitus is a common endocrinopathy in dog. Fibroblast growth factor 21 (FGF-21) is a secreted protein, which is involved in glucose homeostasis. We speculate that the recombinant canine FGF-21 (cFGF-21) has the potential to become a powerful therapeutics to treat canine diabetes. The cFGF-21 gene was cloned and expressed in E. coli Rosetta (DE3). After purification, a cFGF-21 protein with the purity exceeding 95% was obtained. Mouse 3T3-L1 adipocytes and type 1 diabetic mice/dogs induced by STZ were used to examine the biological activity of cFGF-21 in vitro and in vivo, respectively. Results showed that cFGF-21 stimulated glucose uptake in adipocytes significantly in a dose-dependent manner, and reduced plasma glucose significantly in diabetic mice/dogs. After treatment with cFGF-21, the serum insulin level, glycosylated hemoglobin (HbA1c) level and the expressions of the hepatic gluconeogenesis genes (glucose-6-phosphatase, G6Pase and phosphoenolpyruvate carboxykinase, PCK) of the diabetic mice/dogs were attenuated significantly. In the mouse experiment, we also found that the phosphorylation of signal transducer and activator of transcription 3 (STAT3) and the expression of suppressor of cytokine signaling 3 (SOCS3) were up-regulated significantly in the livers after treatment. Histopathological and immunohistochemical results showed that treatment with cFGF-21 promoted recovery of pancreatic islets from STZ-induced apoptosis. Besides, we also found that treatment with cFGF-21 protected liver against STZ or hyperglycemia induced damage and the mechanism of this action associated with inhibiting oxidative stress. In conclusion, cFGF-21 represents a promising candidate for canine diabetes therapeutics. The mechanism of cFGF-21 ameliorates hyperglycemia associated with inhibiting hepatic gluconeogenesis by regulation of STAT3 signal pathway and improving pancreatic beta-cell survival. PMID:27203422

  5. The Glucotoxicity Protecting Effect of Ezetimibe in Pancreatic Beta Cells via Inhibition of CD36

    PubMed Central

    2016-01-01

    Inhibition of CD36, a fatty acid transporter, has been reported to prevent glucotoxicity and ameliorate high glucose induced beta cell dysfunction. Ezetimibe is a selective cholesterol absorption inhibitor that blocks Niemann Pick C1-like 1 protein, but may exert its effect through suppression of CD36. We attempted to clarify the beneficial effect of ezetimibe on insulin secreting cells and to determine whether this effect is related to change of CD36 expression. mRNA expression of insulin and CD36, intracellular peroxide level and glucose stimulated insulin secretion (GSIS) under normal (5.6 mM) or high glucose (30 mM) condition in INS-1 cells and primary rat islet cells were compared. Changes of the aforementioned factors with treatment with ezetimibe (20 μM) under normal or high glucose condition were also assessed. mRNA expression of insulin was decreased with high glucose, which was reversed by ezetimibe in both INS-1 cells and primary rat islets. CD36 mRNA expression was increased with high glucose, but decreased by ezetimibe in INS-1 cells and primary rat islets. Three-day treatment with high glucose resulted in an increase in intracellular peroxide level; however, it was decreased by treatment with ezetimibe. Decrease in GSIS by three-day treatment with high glucose was reversed by ezetimibe. Palmitate uptake following exposure to high glucose conditions for three days was significantly elevated, which was reversed by ezetimibe in INS-1 cells. Ezetimibe may prevent glucotoxicity in pancreatic β-cells through a decrease in fatty acid influx via inhibition of CD36. PMID:27051238

  6. Distinct glucose lowering and beta cell protective effects of vanadium and food restriction in streptozotocin-diabetes.

    PubMed

    Cam, M C; Rodrigues, B; McNeill, J H

    1999-11-01

    Vanadium is an oral insulin-mimetic agent that diminishes hyperglycemia, improves beta-cell insulin store and secretory function, and can reverse the diabetic state chronically after withdrawal from treatment. As food restriction has been reported to enhance insulin sensitivity and reduce insulin demand, we assessed the contribution of a reduced food intake to the glucose lowering and beta-cell protective effects of vanadium. Streptozotocin (STZ)-diabetic rats were untreated (D) or administered vanadyl sulfate in the drinking water (DT) at one week prior to and for 5 weeks following the administration of STZ. An additional group was pair-fed (DP) with an equal amount of food as that consumed by the DT group. Shortly after the induction of diabetes, hyperglycemic D rats demonstrated a significant rise in plasma insulin to levels that initially exceeded that of the controls. This was followed by a steady reduction over several weeks, suggesting a gradual depletion of functional beta-cells. Both vanadium treatment and pair-feeding abolished the insulin hypersecretory response following STZ administration. Glucose lowering was enhanced in DT animals when administered higher concentrations of vanadium, despite no further reduction in food intake, and all DT animals (10/10) were normoglycemic by 5 weeks. Mean pancreatic insulin content in DT rats was improved fourfold and was associated with a greater number of granulated beta-cells. Conversely, food restriction only modestly improved glycemia and the pancreatic insulin store and, unlike DT, DP rats remained highly glucose-intolerant. At 5 weeks of diabetes, fed circulating glucose and insulin levels were strongly correlated (P=0.0002) in the D and DP groups, supporting the notion that glucose lowering with food restriction is dependent on improved plasma insulin levels. A separate correlation was observed in DT animals within a lower range of plasma insulin, suggesting that vanadium, unlike food restriction, reduced

  7. Cell type-specific activation of metabolism reveals that beta-cell secretion suppresses glucagon release from alpha-cells in rat pancreatic islets.

    PubMed

    Takahashi, Rui; Ishihara, Hisamitsu; Tamura, Akira; Yamaguchi, Suguru; Yamada, Takahiro; Takei, Daisuke; Katagiri, Hideki; Endou, Hitoshi; Oka, Yoshitomo

    2006-02-01

    Abnormal glucagon secretion is often associated with diabetes mellitus. However, the mechanisms by which nutrients modulate glucagon secretion remain poorly understood. Paracrine modulation by beta- or delta-cells is among the postulated mechanisms. Herein we present further evidence of the paracrine mechanism. First, to activate cellular metabolism and thus hormone secretion in response to specific secretagogues, we engineered insulinoma INS-1E cells using an adenovirus-mediated expression system. Expression of the Na+-dependent dicarboxylate transporter (NaDC)-1 resulted in 2.5- to 4.6-fold (P < 0.01) increases in insulin secretion in response to various tricarboxylic acid cycle intermediates. Similarly, expression of glycerol kinase (GlyK) increased insulin secretion 3.8- or 4.2-fold (P < 0.01) in response to glycerol or dihydroxyacetone, respectively. This cell engineering method was then modified, using the Cre-loxP switching system, to activate beta-cells and non-beta-cells separately in rat islets. NaDC-1 expression only in non-beta-cells, among which alpha-cells are predominant, caused an increase (by 1.8-fold, P < 0.05) in glucagon secretion in response to malate or succinate. However, the increase in glucagon release was prevented when NaDC-1 was expressed in whole islets, i.e., both beta-cells and non-beta-cells. Similarly, an increase in glucagon release with glycerol was observed when GlyK was expressed only in non-beta-cells but not when it was expressed in whole islets. Furthermore, dicarboxylates suppressed basal glucagon secretion by 30% (P < 0.05) when NaDC-1 was expressed only in beta-cells. These data demonstrate that glucagon secretion from rat alpha-cells depends on beta-cell activation and provide insights into the coordinated mechanisms underlying hormone secretion from pancreatic islets.

  8. Increased secretion of insulin and proliferation of islet {beta}-cells in rats with mesenteric lymph duct ligation

    SciTech Connect

    Nagino, Ko; Yokozawa, Junji; Sasaki, Yu; Matsuda, Akiko; Takeda, Hiroaki; Kawata, Sumio

    2012-08-24

    Highlights: Black-Right-Pointing-Pointer Insulin secretion was increased during the OGTT or IVGTT in mesenteric lymph duct-ligated rats. Black-Right-Pointing-Pointer Proliferation of islet {beta}-cells was upregulated in lymph duct-ligated rats. Black-Right-Pointing-Pointer Mesenteric lymph duct flow has a role in glucose metabolism. -- Abstract: Background and aims: It has been suggested that intestinal lymph flow plays an important role in insulin secretion and glucose metabolism after meals. In this study, we investigated the influence of ligation of the mesenteric lymph duct on glucose metabolism and islet {beta}-cells in rats. Methods: Male Sprague-Dawley rats (10 weeks old) were divided into two groups: one underwent ligation of the mesenteric lymph duct above the cistern (ligation group), and the other underwent a sham operation (sham group). After 1 and 2 weeks, fasting plasma concentrations of glucose, insulin, triglyceride, glucose-dependent insulinotropic polypeptide (GIP), and the active form of glucagon-like peptide-1 (GLP-1) were measured. At 2 weeks after the operation, the oral glucose tolerance test (OGTT) and intravenous glucose tolerance test (IVGTT) were performed. After the rats had been sacrificed, the insulin content of the pancreas was measured and the proliferation of {beta}-cells was assessed immunohistochemically using antibodies against insulin and Ki-67. Results: During the OGTT, the ligation group showed a significant decrease in the plasma glucose concentration at 120 min (p < 0.05) and a significant increase in the plasma insulin concentration by more than 2-fold at 15 min (p < 0.01). On the other hand, the plasma GIP concentration was significantly decreased at 60 min (p < 0.01) in the ligated group, while the active form of GLP-1 showed a significantly higher level at 90 min (1.7-fold; p < 0.05) and 120 min (2.5-fold; p < 0.01). During the IVGTT, the plasma insulin concentration in the ligation group was significantly higher at 2

  9. Evidence that down-regulation of. beta. -cell glucose transporters in non-insulin-dependent diabetes may be the cause of diabetic hyperglycemia

    SciTech Connect

    Orci, L.; Ravazzola, M.; Baetens, D.; Amherdt, M. ); Inman, L.; Johnson, J.H.; Unger, R.H. Dept. of Veterans Affairs Medical Center, Dallas, TX ); Peterson, R.G. ); Newgard, C.B. )

    1990-12-01

    Non-insulin-dependent diabetes mellitus (NIDDM) is attributed to a failure of pancreatic {beta} cells to maintain insulin secretion at a level sufficient to compensate for underlying insulin resistance. In the ZDF rat, a model of NIDDM that closely resembles the human syndrome, the authors have previously reported profound underexpression of GLUT-2, the high-K{sub m} facilitative glucose transporter expressed by {beta} cells of normal animals. Here they report that islets of diabetic rats exhibit a marked decrease in the volume density of GLUT-2-positive {beta} cells and a reduction at the electron-microscopic level in the number of GLUT-2-immunoreactive sites per unit of {beta}-cell plasma membrane. The deficiency of GLUT-2 cannot be induced in normal {beta} cells by in vivo or in vitro exposure to high levels of glucose nor can it be prevented in {beta} cells of prediabetic ZDF rats by elimination of hyperglycemia. They conclude that this dearth of immunodetectable GLUT-2 in NIDDM is not secondary to hyperglycemia and therefore that it may well play a causal role in the development of hyperglycemia.

  10. Neogenesis and proliferation of {beta}-cells induced by human betacellulin gene transduction via retrograde pancreatic duct injection of an adenovirus vector

    SciTech Connect

    Tokui, Yae . E-mail: ytokui@imed2.med.osaka-u.ac.jp; Kozawa, Junji; Yamagata, Kazuya; Zhang, Jun; Ohmoto, Hiroshi; Tochino, Yoshihiro; Okita, Kohei; Iwahashi, Hiromi; Namba, Mitsuyoshi; Shimomura, Iichiro; Miyagawa, Jun-ichiro |

    2006-12-01

    Betacellulin (BTC) has been shown to have a role in the differentiation and proliferation of {beta}-cells both in vitro and in vivo. We administered a human betacellulin (hBTC) adenovirus vector to male ICR mice via retrograde pancreatic duct injection. As a control, we administered a {beta}-galactosidase adenovirus vector. In the mice, hBTC protein was mainly overexpressed by pancreatic duct cells. On immunohistochemical analysis, we observed features of {beta}-cell neogenesis as newly formed insulin-positive cells in the duct cell lining or islet-like cell clusters (ICCs) closely associated with the ducts. The BrdU labeling index of {beta}-cells was also increased by the betacellulin vector compared with that of control mice. These results indicate that hBTC gene transduction into adult pancreatic duct cells promoted {beta}-cell differentiation (mainly from duct cells) and proliferation of pre-existing {beta}-cells, resulting in an increase of the {beta}-cell mass that improved glucose tolerance in diabetic mice.

  11. MicroRNA Expression in Alpha and Beta Cells of Human Pancreatic Islets

    PubMed Central

    Vargas, Nancy; Rosero, Samuel; Piroso, Julieta; Ichii, Hirohito; Umland, Oliver; Zhijie, Jiang; Tsinoremas, Nicholas; Ricordi, Camillo; Inverardi, Luca; Domínguez-Bendala, Juan; Pastori, Ricardo L.

    2013-01-01

    microRNAs (miRNAs) play an important role in pancreatic development and adult β-cell physiology. Our hypothesis is based on the assumption that each islet cell type has a specific pattern of miRNA expression. We sought to determine the profile of miRNA expression in α-and β-cells, the main components of pancreatic islets, because this analysis may lead to a better understanding of islet gene regulatory pathways. Highly enriched (>98%) subsets of human α-and β-cells were obtained by flow cytometric sorting after intracellular staining with c-peptide and glucagon antibody. The method of sorting based on intracellular staining is possible because miRNAs are stable after fixation. MiRNA expression levels were determined by quantitative high throughput PCR-based miRNA array platform screening. Most of the miRNAs were preferentially expressed in β-cells. From the total of 667 miRNAs screened, the Significant Analysis of Microarray identified 141 miRNAs, of which only 7 were expressed more in α-cells (α-miRNAs) and 134 were expressed more in β-cells (β-miRNAs). Bioinformatic analysis identified potential targets of β-miRNAs analyzing the Beta Cell Gene Atlas, described in the T1Dbase, the web platform, supporting the type 1 diabetes (T1D) community. cMaf, a transcription factor regulating glucagon expression expressed selectively in α-cells (TFα) is targeted by β-miRNAs; miR-200c, miR-125b and miR-182. Min6 cells treated with inhibitors of these miRNAs show an increased expression of cMaf RNA. Conversely, over expression of miR-200c, miR-125b or miR-182 in the mouse alpha cell line αTC6 decreases the level of cMAF mRNA and protein. MiR-200c also inhibits the expression of Zfpm2, a TFα that inhibits the PI3K signaling pathway, at both RNA and protein levels. In conclusion, we identified miRNAs differentially expressed in pancreatic α- and β-cells and their potential transcription factor targets that could add new insights into different aspects of islet

  12. Surviving native beta-cells determine outcome of syngeneic intraportal islet transplantation.

    PubMed

    Hughes, S J; Powis, S H; Press, M

    2001-01-01

    In moderately diabetic rats (plasma glucose 20-30 mmol/L), where there is some residual pancreatic islet function, normoglycemia can be restored by transplantation of pancreatic islets into the liver via the portal vein. To examine whether normoglycemia can also be achieved in more severely diabetic animals (which more closely resemble human type I diabetes), we have compared the effect of transplanting 1000 islets intraportally in Lewis rats made moderately diabetic (55 mg/kg streptozotocin injected IP while nonfasting) or severely diabetic (65 mg/kg streptozotocin injected IP while fasting). In the moderately diabetic rats in which residual pancreatic insulin was 128 +/- 40 mU insulin (2.0% of control), plasma glucose stabilized (32 +/- 2.8 mmol/L at 1 week, 34 +/- 2 mmol/L at 3 weeks) as did body weight (falling from 290 +/- 5 to 265 +/- 5 g at 1 week and 253 +/- 6 g at 3 weeks). In contrast, in severely diabetic rats in which residual pancreatic insulin was only 13.5 +/- 4.2 mU insulin (0.21% of control), there was a progressive rise in plasma glucose (30 +/- 1.3 mmol/L at 1 week, 49 +/- 4 mmol/L at 2 weeks, and 67 +/- 7 mmol/L at 3 weeks) and a progressive fall in body weight (from 304 +/- 10 to 260 +/- 5 g by week 1 and to 209 +/- 6 g by week 3). Following islet transplantation, nonfasting plasma glucose normalized in moderately diabetic rats (10.5 +/- 0.6 vs. 9.1 +/- 0.6 mmol/L in nondiabetic controls, NS) after 23 +/- 5 days. In contrast, in the severely diabetic rats plasma glucose stabilized at 32 +/- 5 mmol/L (p < 0.05 compared to moderately diabetic group) but did not normalize. This difference was not attributable to different plasma glucose levels at the time of transplantation (35.1 +/- 1.8 in moderately diabetic vs. 32.5 +/- 2.5 mmol/L in severely diabetic rats). These observations demonstrate that residual native beta-cells (equivalent to only 60-80 islets) contribute to the survival or function of intraportally transplanted islets. PMID:11332628

  13. Dynamics of glucose-induced membrane recruitment of protein kinase C beta II in living pancreatic islet beta-cells.

    PubMed

    Pinton, Paolo; Tsuboi, Takashi; Ainscow, Edward K; Pozzan, Tullio; Rizzuto, Rosario; Rutter, Guy A

    2002-10-01

    The mechanisms by which glucose may affect protein kinase C (PKC) activity in the pancreatic islet beta-cell are presently unclear. By developing adenovirally expressed chimeras encoding fusion proteins between green fluorescent protein and conventional (betaII), novel (delta), or atypical (zeta) PKCs, we show that glucose selectively alters the subcellular localization of these enzymes dynamically in primary islet and MIN6 beta-cells. Examined by laser scanning confocal or total internal reflection fluorescence microscopy, elevated glucose concentrations induced oscillatory translocations of PKCbetaII to spatially confined regions of the plasma membrane. Suggesting that increases in free cytosolic Ca(2+) concentrations ([Ca(2+)](c)) were primarily responsible, prevention of [Ca(2+)](c) increases with EGTA or diazoxide completely eliminated membrane recruitment, whereas elevation of cytosolic [Ca(2+)](c) with KCl or tolbutamide was highly effective in redistributing PKCbetaII both to the plasma membrane and to the surface of dense core secretory vesicles. By contrast, the distribution of PKCdelta.EGFP, which binds diacylglycerol but not Ca(2+), was unaffected by glucose. Measurement of [Ca(2+)](c) immediately beneath the plasma membrane with a ratiometric "pericam," fused to synaptic vesicle-associated protein-25, revealed that depolarization induced significantly larger increases in [Ca(2+)](c) in this domain. These data demonstrate that nutrient stimulation of beta-cells causes spatially and temporally complex changes in the subcellular localization of PKCbetaII, possibly resulting from the generation of Ca(2+) microdomains. Localized changes in PKCbetaII activity may thus have a role in the spatial control of insulin exocytosis.

  14. Membrane Potential and Calcium Dynamics in Beta Cells from Mouse Pancreas Tissue Slices: Theory, Experimentation, and Analysis.

    PubMed

    Dolenšek, Jurij; Špelič, Denis; Klemen, Maša Skelin; Žalik, Borut; Gosak, Marko; Rupnik, Marjan Slak; Stožer, Andraž

    2015-10-28

    Beta cells in the pancreatic islets of Langerhans are precise biological sensors for glucose and play a central role in balancing the organism between catabolic and anabolic needs. A hallmark of the beta cell response to glucose are oscillatory changes of membrane potential that are tightly coupled with oscillatory changes in intracellular calcium concentration which, in turn, elicit oscillations of insulin secretion. Both membrane potential and calcium changes spread from one beta cell to the other in a wave-like manner. In order to assess the properties of the abovementioned responses to physiological and pathological stimuli, the main challenge remains how to effectively measure membrane potential and calcium changes at the same time with high spatial and temporal resolution, and also in as many cells as possible. To date, the most wide-spread approach has employed the electrophysiological patch-clamp method to monitor membrane potential changes. Inherently, this technique has many advantages, such as a direct contact with the cell and a high temporal resolution. However, it allows one to assess information from a single cell only. In some instances, this technique has been used in conjunction with CCD camera-based imaging, offering the opportunity to simultaneously monitor membrane potential and calcium changes, but not in the same cells and not with a reliable cellular or subcellular spatial resolution. Recently, a novel family of highly-sensitive membrane potential reporter dyes in combination with high temporal and spatial confocal calcium imaging allows for simultaneously detecting membrane potential and calcium changes in many cells at a time. Since the signals yielded from both types of reporter dyes are inherently noisy, we have developed complex methods of data denoising that permit for visualization and pixel-wise analysis of signals. Combining the experimental approach of high-resolution imaging with the advanced analysis of noisy data enables novel

  15. Membrane Potential and Calcium Dynamics in Beta Cells from Mouse Pancreas Tissue Slices: Theory, Experimentation, and Analysis

    PubMed Central

    Dolenšek, Jurij; Špelič, Denis; Skelin Klemen, Maša; Žalik, Borut; Gosak, Marko; Slak Rupnik, Marjan; Stožer, Andraž

    2015-01-01

    Beta cells in the pancreatic islets of Langerhans are precise biological sensors for glucose and play a central role in balancing the organism between catabolic and anabolic needs. A hallmark of the beta cell response to glucose are oscillatory changes of membrane potential that are tightly coupled with oscillatory changes in intracellular calcium concentration which, in turn, elicit oscillations of insulin secretion. Both membrane potential and calcium changes spread from one beta cell to the other in a wave-like manner. In order to assess the properties of the abovementioned responses to physiological and pathological stimuli, the main challenge remains how to effectively measure membrane potential and calcium changes at the same time with high spatial and temporal resolution, and also in as many cells as possible. To date, the most wide-spread approach has employed the electrophysiological patch-clamp method to monitor membrane potential changes. Inherently, this technique has many advantages, such as a direct contact with the cell and a high temporal resolution. However, it allows one to assess information from a single cell only. In some instances, this technique has been used in conjunction with CCD camera-based imaging, offering the opportunity to simultaneously monitor membrane potential and calcium changes, but not in the same cells and not with a reliable cellular or subcellular spatial resolution. Recently, a novel family of highly-sensitive membrane potential reporter dyes in combination with high temporal and spatial confocal calcium imaging allows for simultaneously detecting membrane potential and calcium changes in many cells at a time. Since the signals yielded from both types of reporter dyes are inherently noisy, we have developed complex methods of data denoising that permit for visualization and pixel-wise analysis of signals. Combining the experimental approach of high-resolution imaging with the advanced analysis of noisy data enables novel

  16. Islet-selectivity of G-protein coupled receptor ligands evaluated for PET imaging of pancreatic {beta}-cell mass

    SciTech Connect

    Cline, Gary W.; Zhao, Xiaojian; Jakowski, Amy B.; Soeller, Walter C.; Treadway, Judith L.

    2011-09-02

    Highlights: {yields} We screened G-protein coupled receptors for imaging pancreatic. {yields} Database mining and immunohistochemistry identified GPCRs enriched in {beta}-cells. {yields} In vitro and in vivo assays were used to determine exocrine vs endocrine specificity. {yields} GPCR candidates for imaging of {beta}-cell mass are Prokineticin-1R, mGluR5, and GLP-1R. -- Abstract: A critical unmet need exists for methods to quantitatively measure endogenous pancreatic {beta}-cell mass (BCM) for the clinical evaluation of therapies to prevent or reverse loss of BCM and diabetes progression. Our objective was to identify G-protein coupled receptors (GPCRs) that are expressed with a high degree of specificity to islet {beta}-cells for receptor-targeted imaging of BCM. GPCRs enriched in pancreatic islets relative to pancreas acinar and hepatic tissue were identified using a database screen. Islet-specific expression was confirmed by human pancreas immunohistochemistry (IHC). In vitro selectivity assessment was determined from the binding and uptake of radiolabeled ligands to the rat insulinoma INS-1 832/13 cell line and isolated rat islets relative to the exocrine pancreas cell-type, PANC-1. Tail-vein injections of radioligands into rats were used to determine favorable image criteria of in vivo biodistribution to the pancreas relative to other internal organs (i.e., liver, spleen, stomach, and lungs). Database and IHC screening identified four candidate receptors for further in vitro and in vivo evaluation for PET imaging of BCM: prokineticin-1 receptor (PK-1R), metabotropic glutamate receptor type-5 (mGluR5), neuropeptide Y-2 receptor (NPY-2R), and glucagon-like peptide 1 receptor (GLP-1R). In vitro specificity ratios gave the following receptor rank order: PK-1R > GLP-1R > NPY-2R > mGluR5. The biodistribution rank order of selectivity to the pancreas was found to be PK-1R > VMAT2 {approx} GLP-1R > mGluR5. Favorable islet selectivity and biodistribution

  17. Characterization of Stimulus-Secretion Coupling in the Human Pancreatic EndoC-βH1 Beta Cell Line

    PubMed Central

    Andersson, Lotta E.; Valtat, Bérengère; Bagge, Annika; Sharoyko, Vladimir V.; Nicholls, David G.; Ravassard, Philippe; Scharfmann, Raphael; Spégel, Peter; Mulder, Hindrik

    2015-01-01

    Aims/Hypothesis Studies on beta cell metabolism are often conducted in rodent beta cell lines due to the lack of stable human beta cell lines. Recently, a human cell line, EndoC-βH1, was generated. Here we investigate stimulus-secretion coupling in this cell line, and compare it with that in the rat beta cell line, INS-1 832/13, and human islets. Methods Cells were exposed to glucose and pyruvate. Insulin secretion and content (radioimmunoassay), gene expression (Gene Chip array), metabolite levels (GC/MS), respiration (Seahorse XF24 Extracellular Flux Analyzer), glucose utilization (radiometric), lactate release (enzymatic colorimetric), ATP levels (enzymatic bioluminescence) and plasma membrane potential and cytoplasmic Ca2+ responses (microfluorometry) were measured. Metabolite levels, respiration and insulin secretion were examined in human islets. Results Glucose increased insulin release, glucose utilization, raised ATP production and respiratory rates in both lines, and pyruvate increased insulin secretion and respiration. EndoC-βH1 cells exhibited higher insulin secretion, while plasma membrane depolarization was attenuated, and neither glucose nor pyruvate induced oscillations in intracellular calcium concentration or plasma membrane potential. Metabolite profiling revealed that glycolytic and TCA-cycle intermediate levels increased in response to glucose in both cell lines, but responses were weaker in EndoC-βH1 cells, similar to those observed in human islets. Respiration in EndoC-βH1 cells was more similar to that in human islets than in INS-1 832/13 cells. Conclusions/Interpretation Functions associated with early stimulus-secretion coupling, with the exception of plasma membrane potential and Ca2+ oscillations, were similar in the two cell lines; insulin secretion, respiration and metabolite responses were similar in EndoC-βH1 cells and human islets. While both cell lines are suitable in vitro models, with the caveat of replicating key findings

  18. The transcription factors Nkx6.1 and Nkx6.2 possess equivalent activities in promoting beta-cell fate specification in Pdx1+ pancreatic progenitor cells.

    PubMed

    Nelson, Shelley B; Schaffer, Ashleigh E; Sander, Maike

    2007-07-01

    Despite much progress in identifying transcriptional regulators that control the specification of the different pancreatic endocrine cell types, the spatiotemporal aspects of endocrine subtype specification have remained largely elusive. Here, we address the mechanism by which the transcription factors Nkx6.1 (Nkx6-1) and Nkx6.2 (Nkx6-2) orchestrate development of the endocrine alpha- and beta-cell lineages. Specifically, we assayed for the rescue of insulin-producing beta-cells in Nkx6.1 mutant mice upon restoring Nkx6 activity in select progenitor cell populations with different Nkx6-expressing transgenes. Beta-cell formation and maturation was restored when Nkx6.1 was expressed in multipotential Pdx1(+) pancreatic progenitors, whereas no rescue was observed upon expression in committed Ngn3(+) (Neurog3(+)) endocrine progenitors. Although not excluding additional roles downstream of Ngn3, this finding suggests a first requirement for Nkx6.1 in specifying beta-cell progenitors prior to Ngn3 activation. Surprisingly, although Nkx6.2 only compensates for Nkx6.1 in alpha-but not in beta-cell development in Nkx6.1(-/-) mice, a Pdx1-promoter-driven Nkx6.2 transgene had the same ability to rescue beta-cells as the Pdx1-Nkx6.1 transgene. This demonstrates that the distinct requirements for Nkx6.1 and Nkx6.2 in endocrine differentiation are a consequence of their divergent spatiotemporal expression domains rather than their biochemical activities and implies that both Nkx6.1 and Nkx6.2 possess alpha- and beta-cell-specifying activities. PMID:17537793

  19. Arsenic Exposure and Calpain-10 Polymorphisms Impair the Function of Pancreatic Beta-Cells in Humans: A Pilot Study of Risk Factors for T2DM

    PubMed Central

    Díaz-Villaseñor, Andrea; Cruz, Laura; Cebrián, Arturo; Hernández-Ramírez, Raúl U.; Hiriart, Marcia; García-Vargas, Gonzálo; Bassol, Susana; Sordo, Monserrat; Gandolfi, A. Jay; Klimecki, Walter T.; López-Carillo, Lizbeth; Cebrián, Mariano E.; Ostrosky-Wegman, Patricia

    2013-01-01

    The incidence of type 2 diabetes mellitus (T2DM) is increasing worldwide and diverse environmental and genetic risk factors are well recognized. Single nucleotide polymorphisms (SNPs) in the calpain-10 gene (CAPN-10), which encodes a protein involved in the secretion and action of insulin, and chronic exposure to inorganic arsenic (iAs) through drinking water have been independently associated with an increase in the risk for T2DM. In the present work we evaluated if CAPN-10 SNPs and iAs exposure jointly contribute to the outcome of T2DM. Insulin secretion (beta-cell function) and insulin sensitivity were evaluated indirectly through validated indexes (HOMA2) in subjects with and without T2DM who have been exposed to a gradient of iAs in their drinking water in northern Mexico. The results were analyzed taking into account the presence of the risk factor SNPs SNP-43 and -44 in CAPN-10. Subjects with T2DM had significantly lower beta-cell function and insulin sensitivity. An inverse association was found between beta-cell function and iAs exposure, the association being more pronounced in subjects with T2DM. Subjects without T2DM who were carriers of the at-risk genotype SNP-43 or -44, also had significantly lower beta-cell function. The association of SNP-43 with beta-cell function was dependent on iAs exposure, age, gender and BMI, whereas the association with SNP-44 was independent of all of these factors. Chronic exposure to iAs seems to be a risk factor for T2DM in humans through the reduction of beta-cell function, with an enhanced effect seen in the presence of the at-risk genotype of SNP-43 in CAPN-10. Carriers of CAPN-10 SNP-44 have also shown reduced beta-cell function. PMID:23349674

  20. Perforin facilitates beta cell killing and regulates autoreactive CD8+ T-cell responses to antigen in mouse models of type 1 diabetes.

    PubMed

    Trivedi, Prerak; Graham, Kate L; Krishnamurthy, Balasubramaninan; Fynch, Stacey; Slattery, Robyn M; Kay, Thomas W H; Thomas, Helen E

    2016-04-01

    In type 1 diabetes, cytotoxic CD8(+) T lymphocytes (CTLs) directly interact with pancreatic beta cells through major histocompatibility complex class I. An immune synapse facilitates delivery of cytotoxic granules, comprised mainly of granzymes and perforin. Perforin deficiency protects the majority of non-obese diabetic (NOD) mice from autoimmune diabetes. Intriguingly perforin deficiency does not prevent diabetes in CD8(+) T-cell receptor transgenic NOD8.3 mice. We therefore investigated the importance of perforin-dependent killing via CTL-beta cell contact in autoimmune diabetes. Perforin-deficient CTL from NOD mice or from NOD8.3 mice were significantly less efficient at adoptive transfer of autoimmune diabetes into NODRag1(-/-) mice, confirming that perforin is essential to facilitate beta cell destruction. However, increasing the number of transferred in vitro-activated perforin-deficient 8.3 T cells reversed the phenotype and resulted in diabetes. Perforin-deficient NOD8.3 T cells were present in increased proportion in islets, and proliferated more in response to antigen in vivo indicating that perforin may regulate the activation of CTLs, possibly by controlling cytokine production. This was confirmed when we examined the requirement for direct interaction between beta cells and CD8(+) T cells in NOD8.3 mice, in which beta cells specifically lack major histocompatibility complex (MHC) class I through conditional deletion of β2-microglobulin. Although diabetes was significantly reduced, 40% of these mice developed diabetes, indicating that NOD8.3 T cells can kill beta cells in the absence of direct interaction. Our data indicate that although perforin delivery is the main mechanism that CTL use to destroy beta cells, they can employ alternative mechanisms to induce diabetes in a perforin-independent manner.

  1. Perforin facilitates beta cell killing and regulates autoreactive CD8+ T-cell responses to antigen in mouse models of type 1 diabetes.

    PubMed

    Trivedi, Prerak; Graham, Kate L; Krishnamurthy, Balasubramaninan; Fynch, Stacey; Slattery, Robyn M; Kay, Thomas W H; Thomas, Helen E

    2016-04-01

    In type 1 diabetes, cytotoxic CD8(+) T lymphocytes (CTLs) directly interact with pancreatic beta cells through major histocompatibility complex class I. An immune synapse facilitates delivery of cytotoxic granules, comprised mainly of granzymes and perforin. Perforin deficiency protects the majority of non-obese diabetic (NOD) mice from autoimmune diabetes. Intriguingly perforin deficiency does not prevent diabetes in CD8(+) T-cell receptor transgenic NOD8.3 mice. We therefore investigated the importance of perforin-dependent killing via CTL-beta cell contact in autoimmune diabetes. Perforin-deficient CTL from NOD mice or from NOD8.3 mice were significantly less efficient at adoptive transfer of autoimmune diabetes into NODRag1(-/-) mice, confirming that perforin is essential to facilitate beta cell destruction. However, increasing the number of transferred in vitro-activated perforin-deficient 8.3 T cells reversed the phenotype and resulted in diabetes. Perforin-deficient NOD8.3 T cells were present in increased proportion in islets, and proliferated more in response to antigen in vivo indicating that perforin may regulate the activation of CTLs, possibly by controlling cytokine production. This was confirmed when we examined the requirement for direct interaction between beta cells and CD8(+) T cells in NOD8.3 mice, in which beta cells specifically lack major histocompatibility complex (MHC) class I through conditional deletion of β2-microglobulin. Although diabetes was significantly reduced, 40% of these mice developed diabetes, indicating that NOD8.3 T cells can kill beta cells in the absence of direct interaction. Our data indicate that although perforin delivery is the main mechanism that CTL use to destroy beta cells, they can employ alternative mechanisms to induce diabetes in a perforin-independent manner. PMID:26446877

  2. Enhanced glucose-induced intracellular signaling promotes insulin hypersecretion: pancreatic beta-cell functional adaptations in a model of genetic obesity and prediabetes.

    PubMed

    Irles, Esperanza; Ñeco, Patricia; Lluesma, Mónica; Villar-Pazos, Sabrina; Santos-Silva, Junia Carolina; Vettorazzi, Jean F; Alonso-Magdalena, Paloma; Carneiro, Everardo M; Boschero, Antonio C; Nadal, Ángel; Quesada, Ivan

    2015-03-15

    Obesity is associated with insulin resistance and is known to be a risk factor for type-2 diabetes. In obese individuals, pancreatic beta-cells try to compensate for the increased insulin demand in order to maintain euglycemia. Most studies have reported that this adaptation is due to morphological changes. However, the involvement of beta-cell functional adaptations in this process needs to be clarified. For this purpose, we evaluated different key steps in the glucose-stimulated insulin secretion (GSIS) in intact islets from female ob/ob obese mice and lean controls. Obese mice showed increased body weight, insulin resistance, hyperinsulinemia, glucose intolerance and fed hyperglycemia. Islets from ob/ob mice exhibited increased glucose-induced mitochondrial activity, reflected by enhanced NAD(P)H production and mitochondrial membrane potential hyperpolarization. Perforated patch-clamp examination of beta-cells within intact islets revealed several alterations in the electrical activity such as increased firing frequency and higher sensitivity to low glucose concentrations. A higher intracellular Ca(2+) mobilization in response to glucose was also found in ob/ob islets. Additionally, they displayed a change in the oscillatory pattern and Ca(2+) signals at low glucose levels. Capacitance experiments in intact islets revealed increased exocytosis in individual ob/ob beta-cells. All these up-regulated processes led to increased GSIS. In contrast, we found a lack of beta-cell Ca(2+) signal coupling, which could be a manifestation of early defects that lead to beta-cell malfunction in the progression to diabetes. These findings indicate that beta-cell functional adaptations are an important process in the compensatory response to obesity.

  3. Reduced insulin secretion in protein malnourished mice is associated with multiple changes in the beta-cell stimulus-secretion coupling.

    PubMed

    Soriano, Sergi; Gonzalez, Alejandro; Marroquí, Laura; Tudurí, Eva; Vieira, Elaine; Amaral, Andressa G; Batista, Thiago M; Rafacho, Alex; Boschero, Antonio C; Nadal, Angel; Carneiro, Everardo M; Quesada, Ivan

    2010-08-01

    The mechanism by which protein malnutrition impairs glucose-stimulated insulin secretion in the pancreatic beta-cell is not completely known but may be related to alterations in the signaling events involved in insulin release. Here, we aimed to study the stimulus-secretion coupling of beta-cells from mice fed with low-protein (LP) diet or normal-protein (NP) diet for 8 wk after weaning. Patch-clamp measurements in isolated cells showed that beta-cells from LP mice had a resting membrane potential that was more hyperpolarized than controls. Additionally, depolarization and generation of action potentials in response to stimulatory glucose concentrations were also impaired in beta-cells of LP mice. All these alterations in the LP group were most likely attributed to higher ATP-dependent K(+) (K(ATP)) channel activity in resting conditions and lower efficiency of glucose to induce the closure of these channels. Moreover, a Western blot analysis revealed higher protein levels of the sulphonylurea receptor of the K(ATP) channel in islets of LP mice. Because beta-cell Ca(2+) signals depend on electrical activity, intracellular Ca(2+) oscillations were measured by fluorescence microscopy in intact islets, indicating a lower response to glucose in the LP group. Finally, cell-to-cell synchrony of Ca(2+) signals was analyzed by confocal microscopy. Islets from LP mice exhibited a decreased level of coupling among beta-cells, which was probably due to the low expression levels of connexin 36. Therefore, low-protein diet leads to several alterations in the stimulus-secretion coupling of pancreatic beta-cells that might explain the diminished insulin secretion in response to glucose in this malnutrition state.

  4. Home Use of an Artificial Beta Cell in Type 1 Diabetes

    PubMed Central

    Allen, J.M.; Leelarathna, L.; Hartnell, S.; Wilinska, M.E.; Acerini, C.L.; Dellweg, S.; Benesch, C.; Heinemann, L.; Mader, J.K.; Holzer, M.; Kojzar, H.; Exall, J.; Yong, J.; Pichierri, J.; Barnard, K.D.; Kollman, C.; Cheng, P.; Hindmarsh, P.C.; Campbell, F.M.; Arnolds, S.; Pieber, T.R.; Evans, M.L.; Dunger, D.B.; Hovorka, R.

    2015-01-01

    BACKGROUND The feasibility, safety, and efficacy of prolonged use of an artificial beta cell (closed-loop insulin-delivery system) in the home setting have not been established. METHODS In two multicenter, crossover, randomized, controlled studies conducted under free-living home conditions, we compared closed-loop insulin delivery with sensor-augmented pump therapy in 58 patients with type 1 diabetes. The closed-loop system was used day and night by 33 adults and overnight by 25 children and adolescents. Participants used the closed-loop system for a 12-week period and sensor-augmented pump therapy (control) for a similar period. The primary end point was the proportion of time that the glucose level was between 70 mg and 180 mg per deciliter for adults and between 70 mg and 145 mg per deciliter for children and adolescents. RESULTS Among adults, the proportion of time that the glucose level was in the target range was 11.0 percentage points (95% confidence interval [CI], 8.1 to 13.8) greater with the use of the closed-loop system day and night than with control therapy (P<0.001). The mean glucose level was lower during the closed-loop phase than during the control phase (difference, −11 mg per deciliter; 95% CI, −17 to −6; P<0.001), as were the area under the curve for the period when the glucose level was less than 63 mg per deciliter (39% lower; 95% CI, 24 to 51; P<0.001) and the mean glycated hemoglobin level (difference, −0.3%; 95% CI, −0.5 to −0.1; P=0.002). Among children and adolescents, the proportion of time with the nighttime glucose level in the target range was higher during the closed-loop phase than during the control phase (by 24.7 percentage points; 95% CI, 20.6 to 28.7; P<0.001), and the mean nighttime glucose level was lower (difference, −29 mg per deciliter; 95% CI, −39 to −20; P<0.001). The area under the curve for the period in which the day-and-night glucose levels were less than 63 mg per deciliter was lower by 42% (95% CI

  5. Glucose-stimulated oscillations in free cytosolic ATP concentration imaged in single islet beta-cells: evidence for a Ca2+-dependent mechanism.

    PubMed

    Ainscow, Edward K; Rutter, Guy A

    2002-02-01

    Normal glucose-stimulated insulin secretion is pulsatile, but the molecular mechanisms underlying this pulsatility are poorly understood. Oscillations in the intracellular free [ATP]/[ADP] ratio represent one possible mechanism because they would be expected to cause fluctuations in ATP-sensitive K(+) channel activity and hence oscillatory Ca(2+) influx. After imaging recombinant firefly luciferase, expressed via an adenoviral vector in single human or mouse islet beta-cells, we report here that cytosolic free ATP concentrations oscillate and that these oscillations are affected by glucose. In human beta-cells, oscillations were observed at both 3 and 15 mmol/l glucose, but the oscillations were of a longer wavelength at the higher glucose concentration (167 vs. 66 s). Mouse beta-cells displayed oscillations in both cytosolic free [Ca(2+)] and [ATP] only at elevated glucose concentrations, both with a period of 120 s. To explore the causal relationship between [Ca(2+)] and [ATP] oscillations, the regulation of each was further investigated in populations of MIN6 beta-cells. Incubation in Ca(2+)-free medium lowered cytosolic [Ca(2+)] but increased [ATP] in MIN6 cells at both 3 and 30 mmol/l glucose. Removal of external Ca(2+) increased [ATP], possibly by decreasing ATP consumption by endoplasmic reticulum Ca(2+)-ATPases. These results allow a model to be constructed of the beta-cell metabolic oscillator that drives nutrient-induced insulin secretion.

  6. Loss of connexin36 channels alters beta-cell coupling, islet synchronization of glucose-induced Ca2+ and insulin oscillations, and basal insulin release.

    PubMed

    Ravier, Magalie A; Güldenagel, Martin; Charollais, Anne; Gjinovci, Asllan; Caille, Dorothée; Söhl, Goran; Wollheim, Claes B; Willecke, Klaus; Henquin, Jean-Claude; Meda, Paolo

    2005-06-01

    Normal insulin secretion requires the coordinated functioning of beta-cells within pancreatic islets. This coordination depends on a communications network that involves the interaction of beta-cells with extracellular signals and neighboring cells. In particular, adjacent beta-cells are coupled via channels made of connexin36 (Cx36). To assess the function of this protein, we investigated islets of transgenic mice in which the Cx36 gene was disrupted by homologous recombination. We observed that compared with wild-type and heterozygous littermates that expressed Cx36 and behaved as nontransgenic controls, mice homozygous for the Cx36 deletion (Cx36(-/-)) featured beta-cells devoid of gap junctions and failing to exchange microinjected Lucifer yellow. During glucose stimulation, islets of Cx36(-/-) mice did not display the regular oscillations of intracellular calcium concentrations ([Ca(2+)](i)) seen in controls due to the loss of cell-to-cell synchronization of [Ca(2+)](i) changes. The same islets did not release insulin in a pulsatile fashion, even though the overall output of the hormone in response to glucose stimulation was normal. However, under nonstimulatory conditions, islets lacking Cx36 showed increased basal release of insulin. These data show that Cx36-dependent signaling is essential for the proper functioning of beta-cells, particularly for the pulsatility of [Ca(2+)](i) and insulin secretion during glucose stimulation.

  7. Involvement of conventional kinesin in glucose-stimulated secretory granule movements and exocytosis in clonal pancreatic beta-cells.

    PubMed

    Varadi, Aniko; Ainscow, Edward K; Allan, Victoria J; Rutter, Guy A

    2002-11-01

    Recruitment of secretory vesicles to the cell surface is essential for the sustained secretion of insulin in response to glucose. At present, the molecular motors involved in this movement, and the mechanisms whereby they may be regulated, are undefined. To investigate the role of kinesin family members, we labelled densecore vesicles in clonal beta-cells using an adenovirally expressed, vesicle-targeted green fluorescent protein (phogrin.EGFP), and employed immunoadsorption to obtain highly purified insulin-containing vesicles. Whereas several kinesin family members were expressed in this cell type, only conventional kinesin heavy chain (KHC) was detected in vesicle preparations. Expression of a dominant-negative KHC motor domain (KHC(mut)) blocked all vesicular movements with velocity >0.4 micro m second(-1), which demonstrates that kinesin activity was essential for vesicle motility in live beta-cells. Moreover, expression of KHC(mut) strongly inhibited the sustained, but not acute, stimulation of secretion by glucose. Finally, vesicle movement was stimulated by ATP dose-dependently in permeabilized cells, which suggests that glucose-induced increases in cytosolic [ATP] mediate the effects of the sugar in vivo, by enhancing kinesin activity. These data therefore provide evidence for a novel mechanism whereby glucose may enhance insulin release.

  8. Glucose metabolism and glutamate analog acutely alkalinize pH of insulin secretory vesicles of pancreatic beta-cells.

    PubMed

    Eto, Kazuhiro; Yamashita, Tokuyuki; Hirose, Kenzo; Tsubamoto, Yoshiharu; Ainscow, Edward K; Rutter, Guy A; Kimura, Satoshi; Noda, Mitsuhiko; Iino, Masamitsu; Kadowaki, Takashi

    2003-08-01

    We studied acute changes of secretory vesicle pH in pancreatic beta-cells with a fluorescent pH indicator, lysosensor green DND-189. Fluorescence was decreased by 0.66 +/- 0.10% at 149 +/- 16 s with 22.2 mM glucose stimulation, indicating that vesicular pH was alkalinized by approximately 0.016 unit. Glucose-responsive pH increase was observed when cytosolic Ca2+ influx was blocked but disappeared when an inhibitor of glycolysis or mitochondrial ATP synthase was present. Glutamate dimethyl ester (GME), a plasma membrane-permeable analog of glutamate, potentiated glucose-stimulated insulin secretion at 5 mM without changing cellular ATP content or cytosolic Ca2+ concentration ([Ca2+]). Application of GME at basal glucose concentration decreased DND-189 fluorescence by 0.83 +/- 0.19% at 38 +/- 2 s. These results indicated that the acutely alkalinizing effect of glucose on beta-cell secretory vesicle pH was dependent on glucose metabolism but independent of modulations of cytosolic [Ca2+]. Moreover, glutamate derived from glucose may be one of the mediators of this alkalinizing effect of glucose, which may have potential relevance to the alteration of secretory function by glutamate.

  9. Activation of the Wnt/β-catenin pathway in pancreatic beta cells during the compensatory islet hyperplasia in prediabetic mice.

    PubMed

    Maschio, D A; Oliveira, R B; Santos, M R; Carvalho, C P F; Barbosa-Sampaio, H C L; Collares-Buzato, C B

    2016-09-30

    The Wnt/β-catenin signaling pathway, also known as the canonical Wnt pathway, plays a role in cell proliferation and differentiation in several tissues/organs. It has been recently described in humans a relationship between type 2 diabetes (T2DM) and mutation in the gene encoding the transcription factor TCF7L2 associated to the Wnt/β-catenin pathway. In the present study, we demonstrated that hyperplastic pancreatic islets from prediabetic mice fed a high-fat diet (HFD) for 60 d displayed nuclear translocation of active β-catenin associated with significant increases in protein content and gene expression of β-catenin as well as of cyclins D1, D2 and c-Myc (target genes of the Wnt pathway) but not of Tcf7l2 (the transcription factor). Meanwhile, these alterations were not observed in pancreatic islets from 30 d HFD-fed mice, that do not display significant beta cell hyperplasia. These data suggest that the Wnt/β-catenin pathway is activated in pancreatic islets during prediabetes and may play a role in the induction of the compensatory beta cell hyperplasia observed at early phase of T2DM. PMID:27576200

  10. Sulforaphane protects against cytokine- and streptozotocin-induced {beta}-cell damage by suppressing the NF-{kappa}B pathway

    SciTech Connect

    Song, Mi-Young; Kim, Eun-Kyung; Moon, Woo-Sung; Park, Jin-Woo; Kim, Hyung-Jin; So, Hong-Seob; Park, Raekil; Kwon, Kang-Beom Park, Byung-Hyun

    2009-02-15

    Sulforaphane (SFN) is an indirect antioxidant that protects animal tissues from chemical or biological insults by stimulating the expression of several NF-E2-related factor-2 (Nrf2)-regulated phase 2 enzymes. Treatment of RINm5F insulinoma cells with SFN increases Nrf2 nuclear translocation and expression of phase 2 enzymes. In this study, we investigated whether the activation of Nrf2 by SFN treatment or ectopic overexpression of Nrf2 inhibited cytokine-induced {beta}-cell damage. Treatment of RIN cells with IL-1{beta} and IFN-{gamma} induced {beta}-cell damage through a NF-{kappa}B-dependent signaling pathway. Activation of Nrf2 by treatment with SFN and induction of Nrf2 overexpression by transfection with Nrf2 prevented cytokine toxicity. The mechanism by which Nrf2 activation inhibited NF-{kappa}B-dependent cell death signals appeared to involve the reduction of oxidative stress, as demonstrated by the inhibition of cytokine-induced H{sub 2}O{sub 2} production. The protective effect of SFN was further demonstrated by the restoration of normal insulin secreting responses to glucose in cytokine-treated rat pancreatic islets. Furthermore, pretreatment with SFN blocked the development of type 1 diabetes in streptozotocin-treated mice.

  11. Untangling the interplay of genetic and metabolic influences on beta-cell function: Examples of potential therapeutic implications involving TCF7L2 and FFAR1☆

    PubMed Central

    Wagner, Robert; Staiger, Harald; Ullrich, Susanne; Stefan, Norbert; Fritsche, Andreas; Häring, Hans-Ulrich

    2014-01-01

    Deteriorating beta-cell function is a common feature of type 2 diabetes. In this review, we briefly address the regulation of beta-cell function, and discuss some of the main determinants of beta-cell failure. We will focus on the role of interactions between the genetic background and metabolic environment (insulin resistance, fuel supply and flux as well as metabolic signaling). We present data on the function of the strongest common diabetes risk variant, the single nucleotide polymorphism (SNP) rs7903146 in TCF7L2. As also mirrored by its interaction with glycemia on insulin secretion, this SNP in large part confers resistance against the incretin effect. Genetic influence on insulin secretion also interacts with free fatty acids, as evidenced by data on rs1573611 in FFAR1. Several medications marketed by now or currently under development for diabetes treatment engage these pathways, and therapeutic implications from these findings are soon to be expected. PMID:24749055

  12. Metformin prevents endoplasmic reticulum stress-induced apoptosis through AMPK-PI3K-c-Jun NH2 pathway

    USGS Publications Warehouse

    Jung, T.W.; Lee, M.W.; Lee, Y.-J.; Kim, S.M.

    2012-01-01

    Type 2 diabetes mellitus is thought to be partially associated with endoplasmic reticulum (ER) stress toxicity on pancreatic beta cells and the result of decreased insulin synthesis and secretion. In this study, we showed that a well-known insulin sensitizer, metformin, directly protects against dysfunction and death of ER stress-induced NIT-1 cells (a mouse pancreatic beta cell line) via AMP-activated protein kinase (AMPK) and phosphatidylinositol-3 (PI3) kinase activation. We also showed that exposure of NIT-1 cells to metformin (5mM) increases cellular resistance against ER stress-induced NIT-1 cell dysfunction and death. AMPK and PI3 kinase inhibitors abolished the effect of metformin on cell function and death. Metformin-mediated protective effects on ER stress-induced apoptosis were not a result of an unfolded protein response or the induced inhibitors of apoptotic proteins. In addition, we showed that exposure of ER stressed-induced NIT-1 cells to metformin decreases the phosphorylation of c-Jun NH(2) terminal kinase (JNK). These data suggest that metformin is an important determinant of ER stress-induced apoptosis in NIT-1 cells and may have implications for ER stress-mediated pancreatic beta cell destruction via regulation of the AMPK-PI3 kinase-JNK pathway.

  13. Selective destruction of mouse islet beta cells by human T lymphocytes in a newly-established humanized type 1 diabetic model

    SciTech Connect

    Zhao, Yong; Guo, Chengshan; Hwang, David; Lin, Brian; Dingeldein, Michael; Mihailescu, Dan; Sam, Susan; Sidhwani, Seema; Zhang, Yongkang; Jain, Sumit; Skidgel, Randal A.; Prabhakar, Bellur S.; Mazzone, Theodore; Holterman, Mark J.

    2010-09-03

    Research highlights: {yields} Establish a human immune-mediated type 1 diabetic model in NOD-scid IL2r{gamma}{sup null} mice. {yields} Using the irradiated diabetic NOD mouse spleen mononuclear cells as trigger. {yields} The islet {beta} cells were selectively destroyed by infiltrated human T cells. {yields} The model can facilitate translational research to find a cure for type 1 diabetes. -- Abstract: Type 1 diabetes (T1D) is caused by a T cell-mediated autoimmune response that leads to the loss of insulin-producing {beta} cells. The optimal preclinical testing of promising therapies would be aided by a humanized immune-mediated T1D model. We develop this model in NOD-scid IL2r{gamma}{sup null} mice. The selective destruction of pancreatic islet {beta} cells was mediated by human T lymphocytes after an initial trigger was supplied by the injection of irradiated spleen mononuclear cells (SMC) from diabetic nonobese diabetic (NOD) mice. This resulted in severe insulitis, a marked loss of total {beta}-cell mass, and other related phenotypes of T1D. The migration of human T cells to pancreatic islets was controlled by the {beta} cell-produced highly conserved chemokine stromal cell-derived factor 1 (SDF-1) and its receptor C-X-C chemokine receptor (CXCR) 4, as demonstrated by in vivo blocking experiments using antibody to CXCR4. The specificity of humanized T cell-mediated immune responses against islet {beta} cells was generated by the local inflammatory microenvironment in pancreatic islets including human CD4{sup +} T cell infiltration and clonal expansion, and the mouse islet {beta}-cell-derived CD1d-mediated human iNKT activation. The selective destruction of mouse islet {beta} cells by a human T cell-mediated immune response in this humanized T1D model can mimic those observed in T1D patients. This model can provide a valuable tool for translational research into T1D.

  14. Partial preservation of pancreatic beta-cells by vanadium: evidence for long-term amelioration of diabetes.

    PubMed

    Cam, M C; Li, W M; McNeill, J H

    1997-07-01

    Streptozotocin (STZ)-diabetic rats treated with vanadium can remain euglycemic for up to 20 weeks following withdrawal from vanadium treatment. In this study, we examined the effects of short-term vanadium treatment in preventing or reversing the STZ-induced diabetic state. Male Wistar rats were untreated (D) or treated (DT) with vanadyl sulfate for 1 week before administering STZ. Treatment was subsequently maintained for 3 days (DT3) or 14 days (DT14) post-STZ, after which vanadium was withdrawn. At 4 to 5 weeks post-STZ and following long-term withdrawal from vanadium, DT14 rats demonstrated levels of food and fluid intake and glucose tolerance that were not significantly different from those of age-matched untreated nondiabetic rats, and had significantly reduced glycemic levels in the fed state compared with D and DT3 groups. The proportion of animals that were euglycemic (fed plasma glucose < 9.0 mmol/L) was significant in DT14 (five of 10) relative to D (one of 10) and DT3 (one of 10) (P = .01). All euglycemic animals had an improved pancreatic insulin content that, albeit low (12% of control), was strongly linked to euglycemia in the fed state (r = -.91, P < .0001). Moreover, the highly significant correlation persisted with the analysis of untreated STZ-rats alone (r = -.95, P < .0001). Similarly, improvements in glucose tolerance and insulin secretory function in euglycemic rats were strongly correlated with small changes in residual insulin content. Hence, as vanadium pretreatment did not prevent STZ-induced beta-cytotoxicity, the vanadium-induced amelioration of the diabetic state appears to be secondary to the preservation of a functional portion of pancreatic beta cells that initially survived STZ toxicity. The partial preservation of pancreatic beta cells, albeit small in proportion to the total insulin store, was both critical and sufficient for a long-term reversal of the diabetic state. These results suggest that apparently modest effects in

  15. A survival Kit for pancreatic beta cells: stem cell factor and c-Kit receptor tyrosine kinase.

    PubMed

    Feng, Zhi-Chao; Riopel, Matthew; Popell, Alex; Wang, Rennian

    2015-04-01

    The interactions between c-Kit and its ligand, stem cell factor (SCF), play an important role in haematopoiesis, pigmentation and gametogenesis. c-Kit is also found in the pancreas, and recent studies have revealed that c-Kit marks a subpopulation of highly proliferative pancreatic endocrine cells that may harbour islet precursors. c-Kit governs and maintains pancreatic endocrine cell maturation and function via multiple signalling pathways. In this review we address the importance of c-Kit signalling within the pancreas, including its profound role in islet morphogenesis, islet vascularisation, and beta cell survival and function. We also discuss the impact of c-Kit signalling in pancreatic disease and the use of c-Kit as a potential target for the development of cell-based and novel drug therapies in the treatment of diabetes.

  16. Pattern of rise in subplasma membrane Ca2+ concentration determines type of fusing insulin granules in pancreatic beta cells.

    PubMed

    Ohara-Imaizumi, Mica; Aoyagi, Kyota; Nakamichi, Yoko; Nishiwaki, Chiyono; Sakurai, Takashi; Nagamatsu, Shinya

    2009-07-31

    We simultaneously analyzed insulin granule fusion with insulin fused to green fluorescent protein and the subplasma membrane Ca2+ concentration ([Ca2+](PM)) with the Ca2+ indicator Fura Red in rat beta cells by dual-color total internal reflection fluorescence microscopy. We found that rapid and marked elevation in [Ca2+](PM) caused insulin granule fusion mostly from previously docked granules during the high KCl-evoked release and high glucose-evoked first phase release. In contrast, the slow and sustained elevation in [Ca2+](PM) induced fusion from newcomers translocated from the internal pool during the low KCl-evoked release and glucose-evoked second phase release. These data suggest that the pattern of the [Ca2+](PM) rise directly determines the types of fusing granules.

  17. Glucose generates sub-plasma membrane ATP microdomains in single islet beta-cells. Potential role for strategically located mitochondria.

    PubMed

    Kennedy, H J; Pouli, A E; Ainscow, E K; Jouaville, L S; Rizzuto, R; Rutter, G A

    1999-05-01

    Increases in the concentration of free ATP within the islet beta-cell may couple elevations in blood glucose to insulin release by closing ATP-sensitive K+ (KATP) channels and activating Ca2+ influx. Here, we use recombinant targeted luciferases and photon counting imaging to monitor changes in free [ATP] in subdomains of single living MIN6 and primary beta-cells. Resting [ATP] in the cytosol ([ATP]c), in the mitochondrial matrix ([ATP]m), and beneath the plasma membrane ([ATP]pm) were similar ( approximately 1 mM). Elevations in extracellular glucose concentration (3-30 mM) increased free [ATP] in each domain with distinct kinetics. Thus, sustained increases in [ATP]m and [ATP]pm were observed, but only a transient increase in [ATP]c. However, detectable increases in [ATP]c and [ATP]pm, but not [ATP]m, required extracellular Ca2+. Enhancement of glucose-induced Ca2+ influx with high [K+] had little effect on the apparent [ATP]c and [ATP]m increases but augmented the [ATP]pm increase. Underlying these changes, glucose increased the mitochondrial proton motive force, an effect mimicked by high [K+]. These data support a model in which glucose increases [ATP]m both through enhanced substrate supply and by progressive Ca2+-dependent activation of mitochondrial enzymes. This may then lead to a privileged elevation of [ATP]pm, which may be essential for the sustained closure of KATP channels. Luciferase imaging would appear to be a useful new tool for dynamic in vivo imaging of free ATP concentration.

  18. Pancreatic Beta Cell G-Protein Coupled Receptors and Second Messenger Interactions: A Systems Biology Computational Analysis.

    PubMed

    Fridlyand, Leonid E; Philipson, Louis H

    2016-01-01

    Insulin secretory in pancreatic beta-cells responses to nutrient stimuli and hormonal modulators include multiple messengers and signaling pathways with complex interdependencies. Here we present a computational model that incorporates recent data on glucose metabolism, plasma membrane potential, G-protein-coupled-receptors (GPCR), cytoplasmic and endoplasmic reticulum calcium dynamics, cAMP and phospholipase C pathways that regulate interactions between second messengers in pancreatic beta-cells. The values of key model parameters were inferred from published experimental data. The model gives a reasonable fit to important aspects of experimentally measured metabolic and second messenger concentrations and provides a framework for analyzing the role of metabolic, hormones and neurotransmitters changes on insulin secretion. Our analysis of the dynamic data provides support for the hypothesis that activation of Ca2+-dependent adenylyl cyclases play a critical role in modulating the effects of glucagon-like peptide 1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP) and catecholamines. The regulatory properties of adenylyl cyclase isoforms determine fluctuations in cytoplasmic cAMP concentration and reveal a synergistic action of glucose, GLP-1 and GIP on insulin secretion. On the other hand, the regulatory properties of phospholipase C isoforms determine the interaction of glucose, acetylcholine and free fatty acids (FFA) (that act through the FFA receptors) on insulin secretion. We found that a combination of GPCR agonists activating different messenger pathways can stimulate insulin secretion more effectively than a combination of GPCR agonists for a single pathway. This analysis also suggests that the activators of GLP-1, GIP and FFA receptors may have a relatively low risk of hypoglycemia in fasting conditions whereas an activator of muscarinic receptors can increase this risk. This computational analysis demonstrates that study of second messenger

  19. Pancreatic Beta Cell G-Protein Coupled Receptors and Second Messenger Interactions: A Systems Biology Computational Analysis

    PubMed Central

    Fridlyand, Leonid E.; Philipson, Louis H.

    2016-01-01

    Insulin secretory in pancreatic beta-cells responses to nutrient stimuli and hormonal modulators include multiple messengers and signaling pathways with complex interdependencies. Here we present a computational model that incorporates recent data on glucose metabolism, plasma membrane potential, G-protein-coupled-receptors (GPCR), cytoplasmic and endoplasmic reticulum calcium dynamics, cAMP and phospholipase C pathways that regulate interactions between second messengers in pancreatic beta-cells. The values of key model parameters were inferred from published experimental data. The model gives a reasonable fit to important aspects of experimentally measured metabolic and second messenger concentrations and provides a framework for analyzing the role of metabolic, hormones and neurotransmitters changes on insulin secretion. Our analysis of the dynamic data provides support for the hypothesis that activation of Ca2+-dependent adenylyl cyclases play a critical role in modulating the effects of glucagon-like peptide 1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP) and catecholamines. The regulatory properties of adenylyl cyclase isoforms determine fluctuations in cytoplasmic cAMP concentration and reveal a synergistic action of glucose, GLP-1 and GIP on insulin secretion. On the other hand, the regulatory properties of phospholipase C isoforms determine the interaction of glucose, acetylcholine and free fatty acids (FFA) (that act through the FFA receptors) on insulin secretion. We found that a combination of GPCR agonists activating different messenger pathways can stimulate insulin secretion more effectively than a combination of GPCR agonists for a single pathway. This analysis also suggests that the activators of GLP-1, GIP and FFA receptors may have a relatively low risk of hypoglycemia in fasting conditions whereas an activator of muscarinic receptors can increase this risk. This computational analysis demonstrates that study of second messenger

  20. Restoring Mitochondrial Function: A Small Molecule-mediated Approach to Enhance Glucose Stimulated Insulin Secretion in Cholesterol Accumulated Pancreatic beta cells

    PubMed Central

    Asalla, Suman; Girada, Shravan Babu; Kuna, Ramya S.; Chowdhury, Debabrata; Kandagatla, Bhaskar; Oruganti, Srinivas; Bhadra, Utpal; Bhadra, Manika Pal; Kalivendi, Shasi Vardhan; Rao, Swetha Pavani; Row, Anupama; Ibrahim, A; Ghosh, Partha Pratim; Mitra, Prasenjit

    2016-01-01

    Dyslipidemia, particularly the elevated serum cholesterol levels, aggravate the pathophysiology of type 2 diabetes. In the present study we explored the relationship between fasting blood sugar and serum lipid parameters in human volunteers which revealed a significant linear effect of serum cholesterol on fasting blood glucose. Short term feeding of cholesterol enriched diet to rodent model resulted in elevated serum cholesterol levels, cholesterol accumulation in pancreatic islets and hyperinsulinemia with modest increase in plasma glucose level. To explore the mechanism, we treated cultured BRIN-BD11 pancreatic beta cells with soluble cholesterol. Our data shows that cholesterol treatment of cultured pancreatic beta cells enhances total cellular cholesterol. While one hour cholesterol exposure enhances insulin exocytosis, overnight cholesterol accumulation in cultured pancreatic beta cells affects cellular respiration, and inhibits Glucose stimulated insulin secretion. We further report that (E)-4-Chloro-2-(1-(2-(2,4,6-trichlorophenyl) hydrazono) ethyl) phenol (small molecule M1) prevents the cholesterol mediated blunting of cellular respiration and potentiates Glucose stimulated insulin secretion which was abolished in pancreatic beta cells on cholesterol accumulation. PMID:27282931

  1. Npas4 Transcription Factor Expression Is Regulated by Calcium Signaling Pathways and Prevents Tacrolimus-induced Cytotoxicity in Pancreatic Beta Cells.

    PubMed

    Speckmann, Thilo; Sabatini, Paul V; Nian, Cuilan; Smith, Riley G; Lynn, Francis C

    2016-02-01

    Cytosolic calcium influx activates signaling pathways known to support pancreatic beta cell function and survival by modulating gene expression. Impaired calcium signaling leads to decreased beta cell mass and diabetes. To appreciate the causes of these cytotoxic perturbations, a more detailed understanding of the relevant signaling pathways and their respective gene targets is required. In this study, we examined the calcium-induced expression of the cytoprotective beta cell transcription factor Npas4. Pharmacological inhibition implicated the calcineurin, Akt/protein kinase B, and Ca(2+)/calmodulin-dependent protein kinase signaling pathways in the regulation of Npas4 transcription and translation. Both Npas4 mRNA and protein had high turnover rates, and, at the protein level, degradation was mediated via the ubiquitin-proteasome pathway. Finally, beta cell cytotoxicity of the calcineurin inhibitor and immunosuppressant tacrolimus (FK-506) was prevented by Npas4 overexpression. These results delineate the pathways regulating Npas4 expression and stability and demonstrate its importance in clinical settings such as islet transplantation.

  2. The Relationship between Membrane Potential and Calcium Dynamics in Glucose-Stimulated Beta Cell Syncytium in Acute Mouse Pancreas Tissue Slices

    PubMed Central

    Miller, Evan W.; Slak Rupnik, Marjan

    2013-01-01

    Oscillatory electrical activity is regarded as a hallmark of the pancreatic beta cell glucose-dependent excitability pattern. Electrophysiologically recorded membrane potential oscillations in beta cells are associated with in-phase oscillatory cytosolic calcium activity ([Ca2+]i) measured with fluorescent probes. Recent high spatial and temporal resolution confocal imaging revealed that glucose stimulation of beta cells in intact islets within acute tissue slices produces a [Ca2+]i change with initial transient phase followed by a plateau phase with highly synchronized [Ca2+]i oscillations. Here, we aimed to correlate the plateau [Ca2+]i oscillations with the oscillations of membrane potential using patch-clamp and for the first time high resolution voltage-sensitive dye based confocal imaging. Our results demonstrated that the glucose-evoked membrane potential oscillations spread over the islet in a wave-like manner, their durations and wave velocities being comparable to the ones for [Ca2+]i oscillations and waves. High temporal resolution simultaneous records of membrane potential and [Ca2+]i confirmed tight but nevertheless limited coupling of the two processes, with membrane depolarization preceding the [Ca2+]i increase. The potassium channel blocker tetraethylammonium increased the velocity at which oscillations advanced over the islet by several-fold while, at the same time, emphasized differences in kinetics of the membrane potential and the [Ca2+]i. The combination of both imaging techniques provides a powerful tool that will help us attain deeper knowledge of the beta cell network. PMID:24324777

  3. Partial regeneration of beta-cells in the islets of Langerhans by Nymphayol a sterol isolated from Nymphaea stellata (Willd.) flowers.

    PubMed

    Subash-Babu, P; Ignacimuthu, S; Agastian, P; Varghese, Babu

    2009-04-01

    Reduction of the beta-cell mass is critical in the pathogenesis of diabetes mellitus. The discovery of agents which induce regeneration of pancreatic beta-cells would be useful to develop new therapeutic approaches to treat diabetes. The present study was aimed at identifying a new agent for the control of diabetes through regeneration of pancreatic beta cells and insulin secretory potential. Nymphaea stellata flower chloroform extract (NSFCExt) showed significant plasma glucose lowering effect. Further NSFCExt was utilized to isolate and identify the lead compound based on bioassay guided fractionation; we found Nymphayol (25,26-dinorcholest-5-en-3beta-ol) a new crystal [space group P2(1) (No. 4), a=9.618(5), b=7.518(5), c=37.491(5)]. It was purified by repeat column. The structure was determined on the basis of X-ray crystallography and spectral data. Oral administration of Nymphayol for 45 days significantly (p<0.05) lowered the blood glucose level and more importantly it effectively increased the insulin content in diabetic rats. In addition, Nymphayol increased the number of beta cell mass enormously. Islet-like cell clusters in the islets of Langerhans were clearly observed based on histochemical and immunohistochemical study. PMID:19272781

  4. Differences in beta-cell function and insulin secretion in Black vs. White obese adolescents: Do incretin hormones play a role?

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Black youth are at higher risk for type 2 diabetes (T2D) than their White peers. Previously we demonstrated that for the same degree of insulin sensitivity, Black youth have an upregulated beta-cell function and insulin hypersecretion, in response to intravenous (IV) glucose, compared with Whites. T...

  5. Effect of coriander seed (Coriandrum sativum L.) ethanol extract on insulin release from pancreatic beta cells in streptozotocin-induced diabetic rats.

    PubMed

    Eidi, Maryam; Eidi, Akram; Saeidi, Ali; Molanaei, Saadat; Sadeghipour, Alireza; Bahar, Massih; Bahar, Kamal

    2009-03-01

    Coriander (Coriandrum sativum L.) is grown as a spice crop all over the world. The seeds have been used to treat indigestion, diabetes, rheumatism and pain in the joints. In the present study, an ethanol extract of the seeds was investigated for effects on insulin release from the pancreatic beta cells in streptozotocin-induced diabetic rats. Blood samples were drawn from the retro-orbital sinus before and 1.5, 3 and 5 h after administration of the seed extract. Serum glucose levels were determined by the glucose oxidase method. To determine the insulin releasing activity, after extract treatment the animals were anaesthetized by diethyl ether, the pancreas was excised, fixed in 10% formaldehyde and embedded in paraffin for sectioning. Pancreatic sections of 5 microm were processed for examination of insulin-releasing activity using an immunocytochemistry kit. The results showed that administration of the ethanol extract (200 and 250 mg/kg, i.p.) exhibited a significant reduction in serum glucose. Administration of streptozotocin decreased the number of beta cells with insulin secretory activity in comparison with intact rats, but treatment with the coriander seed extract (200 mg/kg) increased significantly the activity of the beta cells in comparison with the diabetic control rats. The extract decreased serum glucose in streptozotocin-induced diabetic rats and increased insulin release from the beta cells of the pancreas.

  6. Over-expression of ZnT7 increases insulin synthesis and secretion in pancreatic beta-cells by promoting insulin gene transcription

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The mechanism by which zinc regulates insulin synthesis and secretion in pancreatic beta-cells is still unclear. Cellular zinc homeostasis is largely maintained by zinc transporters and intracellular zinc binding proteins. In this study, we demonstrated that zinc transporter 7 (ZnT7, Slc30a7) was co...

  7. VAMP-2 and cellubrevin are expressed in pancreatic beta-cells and are essential for Ca(2+)-but not for GTP gamma S-induced insulin secretion.

    PubMed Central

    Regazzi, R; Wollheim, C B; Lang, J; Theler, J M; Rossetto, O; Montecucco, C; Sadoul, K; Weller, U; Palmer, M; Thorens, B

    1995-01-01

    VAMP proteins are important components of the machinery controlling docking and/or fusion of secretory vesicles with their target membrane. We investigated the expression of VAMP proteins in pancreatic beta-cells and their implication in the exocytosis of insulin. cDNA cloning revealed that VAMP-2 and cellubrevin, but not VAMP-1, are expressed in rat pancreatic islets and that their sequence is identical to that isolated from rat brain. Pancreatic beta-cells contain secretory granules that store and secrete insulin as well as synaptic-like microvesicles carrying gamma-aminobutyric acid. After subcellular fractionation on continuous sucrose gradients, VAMP-2 and cellubrevin were found to be associated with both types of secretory vesicle. The association of VAMP-2 with insulin-containing granules was confirmed by confocal microscopy of primary cultures of rat pancreatic beta-cells. Pretreatment of streptolysin-O permeabilized insulin-secreting cells with tetanus and botulinum B neurotoxins selectively cleaved VAMP-2 and cellubrevin and abolished Ca(2+)-induced insulin release (IC50 approximately 15 nM). By contrast, the pretreatment with tetanus and botulinum B neurotoxins did not prevent GTP gamma S-stimulated insulin secretion. Taken together, our results show that pancreatic beta-cells express VAMP-2 and cellubrevin and that one or both of these proteins selectively control Ca(2+)-mediated insulin secretion. Images PMID:7796801

  8. Preptin derived from proinsulin-like growth factor II (proIGF-II) is secreted from pancreatic islet beta-cells and enhances insulin secretion.

    PubMed Central

    Buchanan, C M; Phillips, A R; Cooper, G J

    2001-01-01

    Pancreatic islet beta-cells secrete the hormones insulin, amylin and pancreastatin. To search for further beta-cell hormones, we purified peptides from secretory granules isolated from cultured murine beta TC6-F7 beta-cells. We identified a 34-amino-acid peptide (3948 Da), corresponding to Asp(69)-Leu(102) of the proinsulin-like growth factor II E-peptide, which we have termed 'preptin'. Preptin, is present in islet beta-cells and undergoes glucose-mediated co-secretion with insulin. Synthetic preptin increases insulin secretion from glucose-stimulated beta TC6-F7 cells in a concentration-dependent and saturable manner. Preptin infusion into the isolated, perfused rat pancreas increases the second phase of glucose-mediated insulin secretion by 30%, while anti-preptin immunoglobulin infusion decreases the first and second phases of insulin secretion by 29 and 26% respectively. These findings suggest that preptin is a physiological amplifier of glucose-mediated insulin secretion. PMID:11716772

  9. Primary prevention of beta-cell autoimmunity and type 1 diabetes – The Global Platform for the Prevention of Autoimmune Diabetes (GPPAD) perspectives

    PubMed Central

    Ziegler, A.G.; Danne, T.; Dunger, D.B.; Berner, R.; Puff, R.; Kiess, W.; Agiostratidou, G.; Todd, J.A.; Bonifacio, E.

    2016-01-01

    Objective Type 1 diabetes can be identified by the presence of beta-cell autoantibodies that often arise in the first few years of life. The purpose of this perspective is to present the case for primary prevention of beta-cell autoimmunity and to provide a study design for its implementation in Europe. Methods We examined and summarized recruitment strategies, enrollment rates, and outcomes in published TRIGR, FINDIA and BABYDIET primary prevention trials, and the TEDDY intensive observational study. A proposal for a recruitment and implementation strategy to perform a phase II/III primary prevention randomized controlled trial in infants with genetic risk for developing beta-cell autoimmunity is outlined. Results Infants with a family history of type 1 diabetes (TRIGR, BABYDIET, TEDDY) and infants younger than age 3 months from the general population (FINDIA, TEDDY) were enrolled into these studies. All studies used HLA genotyping as part of their eligibility criteria. Predicted beta-cell autoimmunity risk in the eligible infants ranged from 3% (FINDIA, TEDDY general population) up to 12% (TRIGR, BABYDIET). Amongst eligible infants, participation was between 38% (TEDDY general population) and 97% (FINDIA). Outcomes, defined as multiple beta-cell autoantibodies, were consistent with predicted risks. We subsequently modeled recruitment into a randomized controlled trial (RCT) that could assess the efficacy of oral insulin treatment as adapted from the Pre-POINT pilot trial. The RCT would recruit infants with and without a first-degree family history of type 1 diabetes and be based on general population genetic risk testing. HLA genotyping and, for the general population, genotyping at additional type 1 diabetes susceptibility SNPs would be used to identify children with around 10% risk of beta-cell autoimmunity. The proposed RCT would have 80% power to detect a 50% reduction in multiple beta-cell autoantibodies by age 4 years at a two-tailed alpha of 0.05, and

  10. [Protective effects of glucagon-like peptide-1 on beta-cells: preclinical and clinical data].

    PubMed

    Consoli, Agostino; Di Biagio, Rosamaria

    2011-12-01

    Dipartimento di Medicina Interna e Scienze dell'Invecchiamento, Università degli Studi "G. d'Annunzio", Chieti Continuing b-cell mass and function loss represents the key mechanism for the pathogenesis and the progression of type 2 diabetes mellitus. Drugs capable of arresting b-cell loss and eventually able to bring b-cell function close to be back to normal would then be a formidable help in type 2 diabetes mellitus treatment. The glucagon-like peptide-1 (GLP-1) receptor agonists exenatide and liraglutide can stimulate in vitro neogenesis and prevent apoptosis in b-cell-like cell lines. Consistently, treatment with GLP-1 receptor agonists ameliorates glucose metabolism, preserves b-cell mass and improves b-cell function in several animal models of diabetes. For instance, in the db/db mice, liraglutide protects the b-cell from oxidative stress and endoplasmic reticulum stress-related damage. Data in humans, in vivo, are less definitive and often based on scarcely reliable indexes of b-cell function. However, short-term treatment (14 weeks) with liraglutide increased b-cell maximal response capacity in a dose-response fashion. A longer (1 year) exenatide treatment also was able to increase b-cell maximal response capacity, but the effect was no longer there after a 4-week washout period. However, a marginal, although significant as compared to glargine treatment, improvement in another b-cell function index (disposition index) was observed after a 4-week washout period following 3-year exenatide treatment. Finally, although no clinical trials with a long enough follow-up period are presently available, durable glucose control has been obtained during 2 years of liraglutide treatment in monotherapy. Since the durability of good control is strictly dependent upon a lack of further b-cell function deterioration, these clinical data may foster hope that GLP-1 receptor antagonist treatment might help preserving b-cell function also in individuals affected by type 2

  11. The cytokine interleukin-1beta reduces the docking and fusion of insulin granules in pancreatic beta-cells, preferentially decreasing the first phase of exocytosis.

    PubMed

    Ohara-Imaizumi, Mica; Cardozo, Alessandra K; Kikuta, Toshiteru; Eizirik, Decio L; Nagamatsu, Shinya

    2004-10-01

    The prediabetic period in type I diabetes mellitus is characterized by the loss of first phase insulin release. This might be due to islet infiltration mediated by mononuclear cells and local release of cytokines, but the mechanisms involved are unknown. To determine the role of cytokines in insulin exocytosis, we have presently utilized total internal reflection fluorescence microscopy (TIRFM) to image and analyze the dynamic motion of single insulin secretory granules near the plasma membrane in live beta-cells exposed for 24 h to interleukin (IL)-1beta or interferon (IFN)-gamma. Immunohistochemistry observed via TIRFM showed that the number of docked insulin granules was decreased by 60% in beta-cells treated with IL-1beta, while it was not affected by exposure to IFN-gamma. This effect of IL-1beta was paralleled by a 50% reduction in the mRNA and the number of clusters of SNAP-25 in the plasma membrane. TIRF images of single insulin granule motion during a 15-min stimulation by 22 mm glucose in IL-1beta-treated beta-cells showed a marked reduction in the fusion events from previously docked granules during the first phase insulin release. Fusion from newcomers, however, was well preserved during the second phase of insulin release of IL-1beta-treated beta-cells. The present observations indicate that IL-1beta, but not IFN-gamma, has a preferential inhibitory effect on the first phase of glucose-induced insulin release, mostly via an action on previously docked granules. This suggests that beta-cell exposure to immune mediators during the course of insulitis might be responsible for the loss of first phase insulin release.

  12. Trypanosomatid apoptosis: 'Apoptosis' without the canonical regulators.

    PubMed

    Smirlis, Despina; Soteriadou, Ketty

    2011-01-01

    Apoptosis is a regulated process of cell death originally described in multicelullar organisms contributing to their development and functionality. There is now increasing experimental evidence that a similar form of cell death is operative in unicellular eukaryotes, including trypanosomatids of the genera Trypanosoma and Leishmania. The determination of ancestral executors and regulators of 'apoptosis' in these protozoa belonging to the most primitive eukaryotes that appeared on earth 1.5 billion years ago, provide an exciting challenge in the understanding of the evolution of apoptosis-regulating processes. A review of the present knowledge of trypanosomatid apoptosis points to the fact that these dying protozoa acquire common apoptotic morphological features as metazoan cells, although they lack many of the molecules accepted today as canonical apoptosis mediators (Bcl-2 family members, caspases, TNF related family of receptors). Herein, we discuss how the knowledge of regulators and executors of trypanosomatid apoptosis may provide answers to the gaps concerning the origin of apoptosis. The aim of this addendum is to emphasize the need for classifying the ancestral death program and to discuss how this relates to the complex death programs in multicellular lineages, with the hope to stimulate further enquiry and research into this area.

  13. Effects of intracellular pH on ATP-sensitive K+ channels in mouse pancreatic beta-cells.

    PubMed Central

    Proks, P; Takano, M; Ashcroft, F M

    1994-01-01

    1. The effects of intracellular pH (pHi) on the ATP-sensitive K+ channel (K+ATP channel) from mouse pancreatic beta-cells were examined in inside-out patches exposed to symmetrical 140 mM K+ solutions. 2. The relationship between channel activity and pHi was described by the Hill equation with half-maximal inhibition (Ki) at pHi 6.25 and a Hill coefficient of 3.7. 3. Following exposure to pHi < 6.8, channel activity did not recover to its original level. Subsequent application of trypsin to the intracellular membrane surface restored channel activity to its initial level or above. 4. At -60 mV the relationship between pHi and the single-channel current amplitude was described by a modified Hill equation with a Hill coefficient of 2.1, half-maximal inhibition at pHi 6.48 and a maximum inhibition of 18.5%. 5. A decrease in pHi reduced the extent of channel inhibition by ATP: Ki was 18 microM at pH 7.2 and 33 microM at pH 6.4. The Hill coefficient was also reduced, being 1.65 at pH 7.2 and 1.17 at pH 6.4. 6. When channel activity was plotted as a function of ATP4- (rather than total ATP) there was no effect of pHi on the relationship. This suggests that ATP4- is the inhibitory ion species and that the effects of reducing pHi are due to the lowered concentration of ATP4-. 7. Changes in external pH had little effect on either single-channel or whole-cell K+ATP currents. 8. The effects of pHi do not support a role for H+ in linking glucose metabolism to K+ATP channel inhibition in pancreatic beta-cells. PMID:8189391

  14. Profound defects in pancreatic beta-cell function in mice with combined heterozygous mutations in Pdx-1, Hnf-1alpha, and Hnf-3beta.

    PubMed

    Shih, David Q; Heimesaat, Markus; Kuwajima, Satoru; Stein, Roland; Wright, Christopher V E; Stoffel, Markus

    2002-03-19

    Defects in pancreatic beta-cell function contribute to the development of type 2 diabetes, a polygenic disease that is characterized by insulin resistance and compromised insulin secretion. Hepatocyte nuclear factors (HNFs) -1alpha, -3beta, -4alpha, and Pdx-1 contribute in the complex transcriptional circuits within the pancreas that are involved in beta-cell development and function. In mice, a heterozygous mutation in Pdx-1 alone, but not Hnf-1alpha(+/-), Hnf-3beta(+/-), or Hnf-4alpha(+/-), causes impaired glucose-stimulated insulin secretion in mice. To investigate the possible functional relationships between these transcription factors on beta-cell activity in vivo, we generated mice with the following combined heterozygous mutations: Pdx-1(+/-)/Hnf-1alpha(+/-), Pdx-1(+/-)/Hnf-3beta(+/-), Pdx-1(+/-)/Hnf-4alpha(+/-), Hnf-1alpha(+/-)/Hnf-4alpha(+/-), and Hnf-3beta(+/-)/Hnf-4alpha(+/-). The greatest loss in function was in combined heterozygous null alleles of Pdx-1 and Hnf-1alpha (Pdx-1(+/-)/Hnf-1alpha(+/-)), or Pdx-1 and Hnf-3beta (Pdx-1(+/-)/Hnf-3beta(+/-)). Both double mutants develop progressively impaired glucose tolerance and acquire a compromised first- and second-phase insulin secretion profile in response to glucose compared with Pdx-1(+/-) mice alone. The loss in beta-cell function in Pdx-1(+/-)/Hnf-3beta(+/-) mice was associated with decreased expression of Nkx-6.1, glucokinase (Gck), aldolase B (aldo-B), and insulin, whereas Nkx2.2, Nkx-6.1, Glut-2, Gck, aldo-B, the liver isoform of pyruvate kinase, and insulin expression was reduced in Pdx-1(+/-)/Hnf-1alpha(+/-) mice. The islet cell architecture was also abnormal in Pdx-1(+/-)/Hnf-3beta(+/-) and Pdx-1(+/-)/Hnf-1alpha(+/-) mice, with glucagon-expressing cells scattered throughout the islet, a defect that may be connected to decreased E-cadherin expression. Our data suggest that functional interactions between key islet regulatory factors play an important role in maintaining islet architecture and

  15. Customized cell-based treatment options to combat autoimmunity and restore beta-cell function in type 1 diabetes mellitus: current protocols and future perspectives.

    PubMed

    Fändrich, Fred; Ungefroren, Hendrik

    2010-01-01

    Type 1 diabetes mellitus (T1D) is considered a classical autoimmune disease which commonly starts during childhood but may appear later in adulthood in a proportion of 30-40% of affected individuals. Its development is based on a combination of a genetic predisposition and autoimmune processes that result in gradual destruction of the beta-cells of the pancreas and cause absolute insulin deficiency. Evidence for an autoimmune origin of T1D results from measurable islet beta-cell autoantibody directed against various autoantigens such as proinsulin or insulin itself, glutamic acid decarboxylase 65, the islet tyrosine phosphatase IA-2, and the islet-specific glucose-6-phosphatase catalytic subunit-related protein. In addition, T-cell lines with specificity for insulin or glutamic acid decarboxylase have been identified within peripheral blood lymphocytes. Importantly, in most instances the pathogenesis of T1D comprises a slowly progressive destruction of beta-cell tissue in the pancreas preceded by several years of a prediabetic phase where autoimmunity has already developed but with no clinically apparent insulin dependency. Unless immunological tolerance to pancreatic autoantigens is re-established, diabetes treated by islet cell transplantation or stimulation/regeneration of endogenous beta-cells would remain a chronic disease secondary to immune suppression related morbidity. Hence, if islet cell tolerance could be re-induced, a major clinical hurdle to curing diabetes by islet cell neogenesis may be overcome. Targeted immunotherapies are currently explored in a variety of clinical studies and hold great promise for causative treatment to readjust the underlying immunologic imbalance with the goal to cure the disease. This chapter will outline possible treatment options to stop or reverse the beta-cell-specific autoimmune and inflammatory process within pancreatic islets. Special emphasis is given to stem cells of embryonic, mesenchymal, and haematopoietic origin

  16. Structural basis for the killing of human beta cells by CD8(+) T cells in type 1 diabetes.

    PubMed

    Bulek, Anna M; Cole, David K; Skowera, Ania; Dolton, Garry; Gras, Stephanie; Madura, Florian; Fuller, Anna; Miles, John J; Gostick, Emma; Price, David A; Drijfhout, Jan W; Knight, Robin R; Huang, Guo C; Lissin, Nikolai; Molloy, Peter E; Wooldridge, Linda; Jakobsen, Bent K; Rossjohn, Jamie; Peakman, Mark; Rizkallah, Pierre J; Sewell, Andrew K

    2012-03-01

    The structural characteristics of the engagement of major histocompatibility complex (MHC) class II-restricted self antigens by autoreactive T cell antigen receptors (TCRs) is established, but how autoimmune TCRs interact with complexes of self peptide and MHC class I has been unclear. Here we examined how CD8(+) T cells kill human islet beta cells in type 1 diabetes via recognition of a human leukocyte antigen HLA-A*0201-restricted glucose-sensitive preproinsulin peptide by the autoreactive TCR 1E6. Rigid 'lock-and-key' binding underpinned the 1E6-HLA-A*0201-peptide interaction, whereby 1E6 docked similarly to most MHC class I-restricted TCRs. However, this interaction was extraordinarily weak because of limited contacts with MHC class I. TCR binding was highly peptide centric, dominated by two residues of the complementarity-determining region 3 (CDR3) loops that acted as an 'aromatic-cap' over the complex of peptide and MHC class I (pMHCI). Thus, highly focused peptide-centric interactions associated with suboptimal TCR-pMHCI binding affinities might lead to thymic escape and potential CD8(+) T cell-mediated autoreactivity. PMID:22245737

  17. Targeted genetic inactivation of N-acetylglucosaminyltransferase-IVa impairs insulin secretion from pancreatic beta cells and evokes type 2 diabetes.

    PubMed

    Ohtsubo, Kazuaki

    2010-01-01

    The biological significance of protein N-glycosylation has been elucidated using a mouse model bearing a genetic mutation of N-acetylglucosaminyltransferases (GnTs), which initiate the formation of specific branch structures on the mannose core of N-glycans. These glycosylation defects evoked a variety of abnormalities and disorders in specific cell types, tissues, and the whole body, reflecting functional requirements. N-Acetylglucosaminyltransferase-IVa (GnT-IVa) initiates the GlcNAcbeta1-4 branch synthesis on the Manalpha1-3 arm of the N-glycan core thereby increasing N-glycan branch complexity. To investigate the physiological function of GnT-IVa, we engineered and characterized GnT-IVa-deficient mice. GnT-IVa-deficient mice showed a metabolic disorder subsequently diagnosed as type 2 diabetes. In this chapter, methods for characterizing GnT-IVa-deficient mice by physiological analyses to detect metabolic alterations and biochemical analyses using primary isolated pancreatic beta cells are summarized and discussed.

  18. Extracellular ATP stimulates exocytosis via localized Ca(2+) release from acidic stores in rat pancreatic beta cells.

    PubMed

    Xie, Li; Zhang, Ming; Zhou, Wei; Wu, Zhengxing; Ding, Jiuping; Chen, Liangyi; Xu, Tao

    2006-04-01

    Three different methods, membrane capacitance (C(m)) measurement, amperometry and FM dye labeling were used to investigate the role of extracellular ATP in insulin secretion from rat pancreatic beta cells. We found that extracellular application of ATP mobilized intracellular Ca(2+) stores and synchronously triggered vigorous exocytosis. No influence of ATP on the readily releasable pool of vesicles was observed, which argues against a direct modulation of the secretory machinery at a level downstream of Ca(2+) elevation. The stimulatory effects of ATP were greatly reduced by intracellular perfusion of BAPTA but not EGTA, suggesting a close spatial association of fusion sites with intracellular Ca(2+) releasing sites. ATP-induced Ca(2+) transients and exocytosis were not blocked by thapsigargin (TG), by a ryanodine receptor antagonist or by dissipation of pH in acidic stores by monensin alone, but they were greatly attenuated by IP(3) receptor inhibition as well as ionomycin plus monensin, suggesting involvement of IP(3)-sensitive acidic Ca(2+) stores. Taken together, our data suggest that extracellular ATP triggers exocytosis by mobilizing spatially limited acidic Ca(2+) stores through IP(3) receptors. This mechanism may explain how insulin secretion from the pancreas is coordinated through diffusible ATP that is co-released with insulin. PMID:16536741

  19. Determinants of glucose toxicity and its reversibility in the pancreatic islet beta-cell line, HIT-T15.

    PubMed

    Gleason, C E; Gonzalez, M; Harmon, J S; Robertson, R P

    2000-11-01

    HIT-T15 cells, a clonal beta-cell line, were cultured and passaged weekly for 6 mo in RPMI 1640 media containing various concentrations of glucose. Insulin content decreased in the intermediate- and late-passage cells as a continuous rather than a threshold glucose concentration effect. In a second series of experiments, cells were grown in media containing either 0.8 or 16.0 mM glucose from passages 76 through 105. Subcultures of passages 86, 92, and 99 that had been grown in media containing 16.0 mM glucose were switched to media containing 0.8 mM glucose and also carried forward to passage 105. Dramatic increases in insulin content and secretion and insulin gene expression were observed when the switches were made at passages 86 and 92 but not when the switch was made at passage 99. These findings suggest that glucose toxicity of insulin-secreting cells is a continuous rather than a threshold function of glucose concentration and that the shorter the period of antecedent glucose toxicity, the more likely that full recovery of cell function will occur. PMID:11052953

  20. Determinants of glucose toxicity and its reversibility in the pancreatic islet beta-cell line, HIT-T15.

    PubMed

    Gleason, C E; Gonzalez, M; Harmon, J S; Robertson, R P

    2000-11-01

    HIT-T15 cells, a clonal beta-cell line, were cultured and passaged weekly for 6 mo in RPMI 1640 media containing various concentrations of glucose. Insulin content decreased in the intermediate- and late-passage cells as a continuous rather than a threshold glucose concentration effect. In a second series of experiments, cells were grown in media containing either 0.8 or 16.0 mM glucose from passages 76 through 105. Subcultures of passages 86, 92, and 99 that had been grown in media containing 16.0 mM glucose were switched to media containing 0.8 mM glucose and also carried forward to passage 105. Dramatic increases in insulin content and secretion and insulin gene expression were observed when the switches were made at passages 86 and 92 but not when the switch was made at passage 99. These findings suggest that glucose toxicity of insulin-secreting cells is a continuous rather than a threshold function of glucose concentration and that the shorter the period of antecedent glucose toxicity, the more likely that full recovery of cell function will occur.

  1. DPP-4 inhibitor des-F-sitagliptin treatment increased insulin exocytosis from db/db mice {beta} cells

    SciTech Connect

    Nagamatsu, Shinya; Ohara-Imaizumi, Mica; Nakamichi, Yoko; Aoyagi, Kyota; Nishiwaki, Chiyono

    2011-09-09

    Highlights: {yields} Anti-diabetic new drug, DPP-4 inhibitor, can affect the insulin exocytosis. {yields} DPP-4 inhibitor treatment altered syntaxin 1 expression. {yields} Treatment of db/db mice with DPP-4 inhibitor increased insulin release. -- Abstract: Incretin promotes insulin secretion acutely. Recently, orally-administered DPP-4 inhibitors represent a new class of anti-hyperglycemic agents. Indeed, inhibitors of dipeptidyl peptidase-IV (DPP-4), sitagliptin, has just begun to be widely used as therapeutics for type 2 diabetes. However, the effects of sitagliptin-treatment on insulin exocytosis from single {beta}-cells are yet unknown. We therefore investigated how sitagliptin-treatment in db/db mice affects insulin exocytosis by treating db/db mice with des-F-sitagliptin for 2 weeks. Perfusion studies showed that 2 weeks-sitagliptin treatment potentiated insulin secretion. We then analyzed insulin granule motion and SNARE protein, syntaxin 1, by TIRF imaging system. TIRF imaging of insulin exocytosis showed the increased number of docked insulin granules and increased fusion events from them during first-phase release. In accord with insulin exocytosis data, des-F-sitagliptin-treatment increased the number of syntaxin 1 clusters on the plasma membrane. Thus, our data demonstrated that 2-weeks des-F-sitagliptin-treatment increased the fusion events of insulin granules, probably via increased number of docked insulin granules and that of syntaxin 1 clusters.

  2. Deletion of glutamate dehydrogenase in beta-cells abolishes part of the insulin secretory response not required for glucose homeostasis.

    PubMed

    Carobbio, Stefania; Frigerio, Francesca; Rubi, Blanca; Vetterli, Laurène; Bloksgaard, Maria; Gjinovci, Asllan; Pournourmohammadi, Shirin; Herrera, Pedro L; Reith, Walter; Mandrup, Susanne; Maechler, Pierre

    2009-01-01

    Insulin exocytosis is regulated in pancreatic ss-cells by a cascade of intracellular signals translating glucose levels into corresponding secretory responses. The mitochondrial enzyme glutamate dehydrogenase (GDH) is regarded as a major player in this process, although its abrogation has not been tested yet in animal models. Here, we generated transgenic mice, named betaGlud1(-/-), with ss-cell-specific GDH deletion. Our results show that GDH plays an essential role in the full development of the insulin secretory response. In situ pancreatic perfusion revealed that glucose-stimulated insulin secretion was reduced by 37% in betaGlud1(-/-). Furthermore, isolated islets with either constitutive or acute adenovirus-mediated knock-out of GDH showed a 49 and 38% reduction in glucose-induced insulin release, respectively. Adenovirus-mediated re-expression of GDH in betaGlud1(-/-) islets fully restored glucose-induced insulin release. Thus, GDH appears to account for about 40% of glucose-stimulated insulin secretion and to lack redundant mechanisms. In betaGlud1(-/-) mice, the reduced secretory capacity resulted in lower plasma insulin levels in response to both feeding and glucose load, while body weight gain was preserved. The results demonstrate that GDH is essential for the full development of the secretory response in beta-cells. However, maximal secretory capacity is not required for maintenance of glucose homeostasis in normo-caloric conditions.

  3. The Influence of Exposure to Maternal Diabetes In Utero on the Rate Of Decline in Beta-Cell Function Among Youth with Diabetes

    PubMed Central

    Crume, T.L.; Andrews, J.S.; D’Agostino, R.; Pettitt, D.J.; Mayer-Davis, E.J.; Law, J.R.; Dolan, L.; Lawrence, J.M.; Saydah, S.; Greenbaum, C.; Rodriguez, B.L.; Dabelea, D.

    2014-01-01

    A relationship between exposure to maternal diabetes in utero and a younger age at diagnosis of type 2 diabetes was detected in SEARCH for Diabetes in Youth Study, while no significant association was detected with paternal diabetes status, suggesting an independent effect of the intrauterine exposure to hyperglycemia. We assessed the influence of exposure to maternal diabetes in utero on beta cell decline measured using fasting C-peptide (FCP) among 1079 youth with diabetes, including 941 with type 1 and 138 with type 2, who were followed post-diagnosis for an average of 58 months. No significant relationship was detected between exposure to maternal diabetes in utero and change in FCP levels in youth with type 1 or type 2 diabetes. These findings suggest that exposure to maternal diabetes in utero may not be an important determinant of short-term beta-cell function decline in youth with type 1 or type 2 diabetes. PMID:23645121

  4. Activation of ATP-sensitive potassium channels in rat pancreatic beta-cells by linoleic acid through both intracellular metabolites and membrane receptor signalling pathway.

    PubMed

    Zhao, Yu-Feng; Pei, Jianming; Chen, Chen

    2008-09-01

    ATP-sensitive potassium channels (K(ATP) channels) determine the excitability of pancreatic beta-cells and importantly regulate glucose-stimulated insulin secretion (GSIS). Long-chain free fatty acids (FFAs) decrease GSIS after long-term exposure to beta-cells, but the effects of exogenous FFAs on K(ATP) channels are not yet well clarified. In this study, the effects of linoleic acid (LA) on membrane potential (MP) and K(ATP) channels were observed in primary cultured rat pancreatic beta-cells. LA (20 microM) induced hyperpolarization of MP and opening of K(ATP) channels, which was totally reversed and inhibited by tolbutamide, a K(ATP) channel blocker. Inhibition of LA metabolism by acyl-CoA synthetase inhibitor, triacsin C (10 microM), partially inhibited LA-induced opening of K(ATP) channels by 64%. The non-FFA G protein-coupled receptor (GPR) 40 agonist, GW9508 (40 microM), induced an opening of K(ATP) channels, which was similar to that induced by LA under triacsin C treatment. Blockade of protein kinases A and C did not influence the opening of K(ATP) channels induced by LA and GW9508, indicating that these two protein kinase pathways are not involved in the action of LA on K(ATP) channels. The present study demonstrates that LA induces hyperpolarization of MP by activating K(ATP) channels via both intracellular metabolites and activation of GPR40. It indicates that not only intracellular metabolites of FFAs but also GPR40-mediated pathways take part in the inhibition of GSIS and beta-cell dysfunction induced by FFAs.

  5. Distinct roles for insulin and insulin-like growth factor-1 receptors in pancreatic beta-cell glucose sensing revealed by RNA silencing.

    PubMed Central

    Da Silva Xavier, Gabriela; Qian, Qingwen; Cullen, Peter J; Rutter, Guy A

    2004-01-01

    The importance of the insulin receptor (IR) and the insulin-like growth factor-1 receptor (IGF-1R) for glucose-regulated insulin secretion and gene expression in pancreatic islet beta-cells is at present unresolved. Here, we have used small interfering RNAs (siRNAs) to silence the expression of each receptor selectively in clonal MIN6 beta-cells. Reduction of IR levels by >90% completely inhibited glucose (30 mM compared with 3 mM)-induced insulin secretion, but had no effect on depolarization-stimulated secretion. IR depletion also blocked the accumulation of preproinsulin (PPI), pancreatic duodenum homoeobox-1 (PDX-1) and glucokinase (GK) mRNAs at elevated glucose concentrations, as assessed by quantitative real-time PCR analysis (TaqMan). Similarly, depletion of IGF-1R inhibited glucose-induced insulin secretion but, in contrast with the effects of IR silencing, had little impact on the regulation of gene expression by glucose. Moreover, loss of IGF-1R, but not IR, markedly inhibited glucose-stimulated increases in cytosolic and mitochondrial ATP, suggesting a role for IGF-1R in the maintenance of oxidative metabolism and in the generation of mitochondrial coupling factors. RNA silencing thus represents a useful tool for the efficient and selective inactivation of receptor tyrosine kinases in isolated beta-cells. By inhibiting glucose-stimulated insulin secretion through the inactivation of IGF-1R, this approach also demonstrates the existence of insulin-independent mechanisms whereby elevated glucose concentrations regulate PPI, PDX-1 and GK gene expression in beta-cells. PMID:14563207

  6. Activation of NF-kappaB by extracellular matrix is involved in spreading and glucose-stimulated insulin secretion of pancreatic beta cells.

    PubMed

    Hammar, Eva B; Irminger, Jean-Claude; Rickenbach, Katharina; Parnaud, Géraldine; Ribaux, Pascale; Bosco, Domenico; Rouiller, Dominique G; Halban, Philippe A

    2005-08-26

    Laminin-5-rich extracellular matrix derived from 804G cells (804G-ECM) engages beta1 integrins to induce spreading, improve glucose-stimulated insulin secretion (GSIS), and increase survival of pancreatic beta cells. The present study examines whether 804G-ECM activates the transcriptional activity of NF-kappaB and the involvement of NF-kappaB in those effects of 804G-ECM on pancreatic beta cells. 804G-ECM induces nuclear translocation and the DNA binding activity of the p65 subunit of NF-kappaB. 804G-ECM-induced nuclear translocation of NF-kappaB was weak as compared with that induced by interleukin-1beta. Transient 804G-ECM-induced DNA binding activity of NF-kappaB (peak at 2 h) and overexpression of NF-kappaB target genes IkappaB alpha and NF-kappaB1(p105) (peak at 4 h) were observed. When NF-kappaB was inhibited by an inhibitor of IkappaB alpha phosphorylation (Bay 11-7082) or by a recombinant adenovirus expressing the nonphosphorylatable form of IkappaB alpha, 804G-ECM-induced cell spreading and actin cytoskeleton organization were reduced. GSIS from cells on 804G-ECM was inhibited 5-fold, whereas cell survival was not affected. In summary, the results indicate that 804G-ECM induces a transient and moderate NF-kappaB activity. This study shows for the first time that ECM-induced NF-kappaB activity is necessary in maintaining GSIS, although it does not affect survival of pancreatic beta cells. The effects of ECM-induced NF-kappaB activity contrast with the deleterious effects of cytokine-induced NF-kappaB activity. It is proposed that transient and moderate NF-kappaB activity is essential for proper function of the pancreatic beta cell.

  7. Beneficial effect of 17{beta}-estradiol on hyperglycemia and islet {beta}-cell functions in a streptozotocin-induced diabetic rat model

    SciTech Connect

    Yamabe, Noriko; Kang, Ki Sung; Zhu Baoting

    2010-11-15

    The modulating effect of estrogen on glucose homeostasis remains a controversial issue at present. In this study, we sought to determine the beneficial effect of 17{beta}-estradiol (E{sub 2}) on hyperglycemia and islet {beta}-cell functions in streptozotocin (STZ)-induced diabetic rats. Male Sprague-Dawley rats were injected i.p. with STZ to induce a relatively mild diabetic condition. The rats were then treated with E{sub 2} orally at 500 {mu}g/kg body weight/day for 15 days to evaluate the modulating effect on hyperglycemia, insulin secretion, and islet {beta}-cell proliferation. E{sub 2} administration for 10 days significantly lowered plasma glucose levels, increased plasma insulin levels, and improved glucose tolerance by attenuating insulin response to oral glucose loading. These beneficial effects of E{sub 2} were accompanied by increases in islet number and volume, rate of islet cell proliferation, and the amount of insulin secreted. The growth-stimulatory effect of E{sub 2} on islet cells was linked to the functions of the estrogen receptor {alpha}. Notably, these protective effects of E{sub 2} on diabetic conditions were basically not observed when the STZ-treated rats had a more severe degree of islet damage and hyperglycemia. Taken together, we conclude that E{sub 2} can promote the regeneration of damaged pancreatic islets by stimulating {beta}-cell proliferation in diabetic rats, and this effect is accompanied by improvements in glucose tolerance and a decrease in plasma glucose levels. These findings suggest that oral administration of E{sub 2} may be beneficial in diabetic patients with an accelerated loss of islet {beta}-cells.

  8. The MODY1 gene for hepatocyte nuclear factor 4alpha and a feedback loop control COUP-TFII expression in pancreatic beta cells.

    PubMed

    Perilhou, Anaïs; Tourrel-Cuzin, Cécile; Zhang, Pili; Kharroubi, Ilham; Wang, Haiyan; Fauveau, Véronique; Scott, Donald K; Wollheim, Claes B; Vasseur-Cognet, Mireille

    2008-07-01

    Pancreatic islet beta cell differentiation and function are dependent upon a group of transcription factors that maintain the expression of key genes and suppress others. Knockout mice with the heterozygous deletion of the gene for chicken ovalbumin upstream promoter-transcription factor II (COUP-TFII) or the complete disruption of the gene for hepatocyte nuclear factor 4alpha (HNF4alpha) in pancreatic beta cells have similar insulin secretion defects, leading us to hypothesize that there is transcriptional cross talk between these two nuclear receptors. Here, we demonstrate specific HNF4alpha activation of a reporter plasmid containing the COUP-TFII gene promoter region in transfected pancreatic beta cells. The stable association of the endogenous HNF4alpha with a region of the COUP-TFII gene promoter that contains a direct repeat 1 (DR-1) binding site was revealed by chromatin immunoprecipitation. Mutation experiments showed that this DR-1 site is essential for HNF4alpha transactivation of COUP-TFII. The dominant negative suppression of HNF4alpha function decreased endogenous COUP-TFII expression, and the specific inactivation of COUP-TFII by small interfering RNA caused HNF4alpha mRNA levels in 832/13 INS-1 cells to decrease. This positive regulation of HNF4alpha by COUP-TFII was confirmed by the adenovirus-mediated overexpression of human COUP-TFII (hCOUP-TFII), which increased HNF4alpha mRNA levels in 832/13 INS-1 cells and in mouse pancreatic islets. Finally, hCOUP-TFII overexpression showed that there is direct COUP-TFII autorepression, as COUP-TFII occupies the proximal DR-1 binding site of its own gene in vivo. Therefore, COUP-TFII may contribute to the control of insulin secretion through the complex HNF4alpha/maturity-onset diabetes of the young 1 (MODY1) transcription factor network operating in beta cells. PMID:18474611

  9. The effects of a HTR2B stop codon and testosterone on energy metabolism and beta cell function among antisocial Finnish males.

    PubMed

    Tikkanen, Roope; Saukkonen, Tero; Fex, Malin; Bennet, Hedvig; Rautiainen, Marja-Riitta; Paunio, Tiina; Koskinen, Mika; Panarsky, Rony; Bevilacqua, Laura; Sjöberg, Rickard L; Tiihonen, Jari; Virkkunen, Matti

    2016-10-01

    Herein, we examined insulin resistance (IR), insulin sensitivity (IS), beta cell activity, and glucose metabolism in subjects with antisocial personality disorder (ASPD), and whether the serotonin 2B (5-HT2B) receptor and testosterone have a role in energy metabolism. A cohort of subjects belonging to a founder population that included 98 ASPD males, aged 25-30, was divided into groups based on the presence of a heterozygous 5-HT2B receptor loss-of-function gene mutation (HTR2B Q20*; n = 9) or not (n = 89). Serum glucose and insulin levels were measured in a 5 h oral glucose tolerance test (75 g) and indices describing IR, IS, and beta cell activity were calculated. Body mass index (BMI) was also determined. Concentrations of the serotonin metabolite 5-hydroxyindoleacetic acid were measured in cerebrospinal fluid, and testosterone levels from serum. An IR-like state comprising high IR, low IS, and high beta cell activity indices was observed among ASPD subjects without the HTR2B Q20* allele. By contrast, being an ASPD HTR2B Q20* carrier appeared to be preventive of these pathophysiologies. The HTR2B Q20* allele and testosterone predicted lower BMI independently, but an interaction between HTR2B Q20* and testosterone lead to increased insulin sensitivity among HTR2B Q20* carriers with low testosterone levels. The HTR2B Q20* allele also predicted reduced beta cell activity and enhanced glucose metabolism. Reduced 5-HT2B receptor function at low or normal testosterone levels may be protective of obesity. Results were observed among Finnish males having an antisocial personality disorder, which limits the generality.

  10. Male-specific beta-cell dysfunction and diabetes resulting from increased expression of a syngeneic MHC class I protein in the pancreata of transgenic mice.

    PubMed

    Martin, L; Keller, G A; Liggitt, D; Oakley, H; Pitts-Meek, S L; Siegel, M W; Terrell, T; Stewart, T A

    1990-12-01

    It is well established that insulin-dependent diabetes (IDDM) is an autoimmune disease with a strong genetic link to the HLA locus. It is less well understood, however, how the destruction of the insulin-producing beta cells is effected and why neighboring non-beta islet cells are spared. Also incompletely explained are the observations that, unlike other autoimmune diseases such as multiple sclerosis, IDDM does not preferentially affect females, the incidence of the disease is highest among young adults, and there are temporal correlations between the onset of the disease and emotional trauma. We have addressed some of these questions by using transgenic mice that constitutively express the MHC class I antigen Dd in the beta cells of the pancreas. Although both male and female Ins.Dd mice expressed equivalent amounts of the Dd protein only the males developed diabetes. The diabetes in the males could be reversed by castration, and the normoglycemic females became diabetic following either ovariectomy and the implantation of a slow-release pellet containing testosterone or the inclusion of dexamethasone in the drinking water. In contrast, transgenic mice that expressed the herpes simplex virus type 1 glycoprotein D in the pancreatic beta cells were normoglycemic and showed no obvious histopathological consequences. The observation that the beta-cell dysfunction by the increased expression of the MHC class I protein Dd cannot be induced by the herpes viral protein suggests that the cellular damage is related to a specific structure or function of the MHC proteins.(ABSTRACT TRUNCATED AT 250 WORDS)

  11. The effects of a HTR2B stop codon and testosterone on energy metabolism and beta cell function among antisocial Finnish males.

    PubMed

    Tikkanen, Roope; Saukkonen, Tero; Fex, Malin; Bennet, Hedvig; Rautiainen, Marja-Riitta; Paunio, Tiina; Koskinen, Mika; Panarsky, Rony; Bevilacqua, Laura; Sjöberg, Rickard L; Tiihonen, Jari; Virkkunen, Matti

    2016-10-01

    Herein, we examined insulin resistance (IR), insulin sensitivity (IS), beta cell activity, and glucose metabolism in subjects with antisocial personality disorder (ASPD), and whether the serotonin 2B (5-HT2B) receptor and testosterone have a role in energy metabolism. A cohort of subjects belonging to a founder population that included 98 ASPD males, aged 25-30, was divided into groups based on the presence of a heterozygous 5-HT2B receptor loss-of-function gene mutation (HTR2B Q20*; n = 9) or not (n = 89). Serum glucose and insulin levels were measured in a 5 h oral glucose tolerance test (75 g) and indices describing IR, IS, and beta cell activity were calculated. Body mass index (BMI) was also determined. Concentrations of the serotonin metabolite 5-hydroxyindoleacetic acid were measured in cerebrospinal fluid, and testosterone levels from serum. An IR-like state comprising high IR, low IS, and high beta cell activity indices was observed among ASPD subjects without the HTR2B Q20* allele. By contrast, being an ASPD HTR2B Q20* carrier appeared to be preventive of these pathophysiologies. The HTR2B Q20* allele and testosterone predicted lower BMI independently, but an interaction between HTR2B Q20* and testosterone lead to increased insulin sensitivity among HTR2B Q20* carriers with low testosterone levels. The HTR2B Q20* allele also predicted reduced beta cell activity and enhanced glucose metabolism. Reduced 5-HT2B receptor function at low or normal testosterone levels may be protective of obesity. Results were observed among Finnish males having an antisocial personality disorder, which limits the generality. PMID:27420381

  12. O-Linked β-N-acetylglucosamine (O-GlcNAc) Acts as a Glucose Sensor to Epigenetically Regulate the Insulin Gene in Pancreatic Beta Cells.

    PubMed

    Durning, Sean P; Flanagan-Steet, Heather; Prasad, Nripesh; Wells, Lance

    2016-01-29

    The post-translational protein modification O-linked β-N-acetylglucosamine (O-GlcNAc) is a proposed nutrient sensor that has been shown to regulate multiple biological pathways. This dynamic and inducible enzymatic modification to intracellular proteins utilizes the end product of the nutrient sensing hexosamine biosynthetic pathway, UDP-GlcNAc, as its substrate donor. Type II diabetic patients have elevated O-GlcNAc-modified proteins within pancreatic beta cells due to chronic hyperglycemia-induced glucose overload, but a molecular role for O-GlcNAc within beta cells remains unclear. Using directed pharmacological approaches in the mouse insulinoma-6 (Min6) cell line, we demonstrate that elevating nuclear O-GlcNAc increases intracellular insulin levels and preserves glucose-stimulated insulin secretion during chronic hyperglycemia. The molecular mechanism for these observed changes appears to be, at least in part, due to elevated O-GlcNAc-dependent increases in Ins1 and Ins2 mRNA levels via elevations in histone H3 transcriptional activation marks. Furthermore, RNA deep sequencing reveals that this mechanism of altered gene transcription is restricted and that the majority of genes regulated by elevated O-GlcNAc levels are similarly regulated by a shift from euglycemic to hyperglycemic conditions. These findings implicate the O-GlcNAc modification as a potential mechanism for hyperglycemic-regulated gene expression in the beta cell.

  13. Beneficial effects of candesartan, an angiotensin II type 1 receptor blocker, on beta-cell function and morphology in db/db mice.

    PubMed

    Shao, Jiaqing; Iwashita, Noseki; Ikeda, Fuki; Ogihara, Takeshi; Uchida, Toyoyoshi; Shimizu, Tomoaki; Uchino, Hiroshi; Hirose, Takahisa; Kawamori, Ryuzo; Watada, Hirotaka

    2006-06-16

    Several epidemiological studies suggested that treatment with angiotensin II type 1 receptor blocker (ARB) provided a risk reduction of developing type 2 diabetes. In this study, we investigated whether and how ARB treatment can improve abnormalities of pancreatic islets in diabetes state. We randomized db/db mice, a model of type 2 diabetes with obesity, at the age of 8 weeks to receive candesartan, an ARB, for 6 weeks. We also studied age-matched db/misty mice as control. Glucose tolerance test revealed that candesartan treatment improved glucose tolerance with the modest increase in serum insulin level in db/db mice. Concurrently, candesartan increased beta-cell mass, increased staining intensity of insulin, and decreased staining intensity of components of NAD(P)H oxidase, p22phox and gp91phox, and those of oxidative stress markers in beta-cells. These changes were accompanied by reduction of mitochondrial volume. Treatment with candesartan also reduced fibrosis in and around the islets and prevented the loss of endothelial cells in islets. Our results showed that candesartan partially prevented deterioration of glucose tolerance by providing protection against progressive beta-cell damage in diabetes. PMID:16650382

  14. Beneficial effects of Murraya koenigii leaves on antioxidant defense system and ultra structural changes of pancreatic beta-cells in experimental diabetes in rats.

    PubMed

    Arulselvan, Palanisamy; Subramanian, Sorimuthu Pillai

    2007-01-30

    Oxidative stress and oxidative damage to tissues are common end points of chronic diseases such as atherosclerosis, diabetes, and rheumatoid arthritis. Oxidative stress in diabetes coexists with a reduction in the antioxidant status, which can further increase the deleterious effects of free radicals. The aim of the present study was to evaluate the possible protective effects of Murraya koenigii leaves extract against beta-cell damage and antioxidant defense systems of plasma and pancreas in streptozotocin induced diabetes in rats. The levels of glucose and glycosylated hemoglobin in blood and insulin, Vitamin C, Vitamin E, ceruloplasmin, reduced glutathione and TBARS were estimated in plasma of control and experimental groups of rats. To assess the changes in the cellular antioxidant defense system such as the level of reduced glutathione and activities of superoxide dismutase, catalase and glutathione peroxidase were assayed in pancreatic tissue homogenate. The levels of glucose, glycosylated hemoglobin, insulin, TBARS, enzymatic and non-enzymatic antioxidants were altered in diabetic rats. These alterations were reverted back to near control levels after the treatment of M. koenigii leaves extract. Transmission electron microscopic studies also revealed the protective nature of M. koenigii leaves on pancreatic beta-cells. These findings suggest that M. koenigii treatment exerts a therapeutic protective nature in diabetes by decreasing oxidative stress and pancreatic beta-cell damage. The antioxidant effect of the M. koenigii extract was compared with glibenclamide, a well-known hypoglycemic drug. PMID:17188670

  15. Glucose and insulin synergistically activate phosphatidylinositol 3-kinase to trigger oscillations of phosphatidylinositol 3,4,5-trisphosphate in beta-cells.

    PubMed

    Hagren, Olof Idevall; Tengholm, Anders

    2006-12-22

    In insulin-secreting beta-cells, activation of phosphatidylinositol 3'-OH-kinase with resulting formation of phosphatidylinositol 3,4,5-trisphosphate (PIP(3)) has been implicated in the regulation of ion channels, insulin secretion, and gene transcription as well as in cell growth and survival, but the kinetics of PIP(3) signals following physiological stimulation of insulin secretion is unknown. Using evanescent wave microscopy and a green fluorescent protein-tagged PIP(3)-binding protein domain for real-time monitoring of plasma membrane PIP(3) concentration in single MIN6 beta-cells, we now demonstrate that glucose stimulation of insulin secretion results in pronounced PIP(3) oscillations via autocrine stimulation of insulin receptors. Glucose lacked effect when insulin secretion was prevented with the hyperpolarizing agent diazoxide, but the sugar dose dependently enhanced the PIP(3) response to maximal insulin stimulation without affecting the rate of PIP(3) degradation. We conclude that glucose is an important co-activator of phosphatidylinositol-3'-OH-kinase and that the plasma membrane PIP(3) concentration in beta-cells undergoes oscillations due to pulsatile release of insulin.

  16. In vivo synchronous membrane potential oscillations in mouse pancreatic beta-cells: lack of co-ordination between islets.

    PubMed Central

    Valdeolmillos, M; Gomis, A; Sánchez-Andrés, J V

    1996-01-01

    1. The properties of the oscillations in electrical activity of different beta-cells within the same islet of Langerhans, and of different islets within the same pancreas, recorded in vivo, are described. 2. Simultaneous recordings of two cells within the same islet showed that the oscillations were synchronous. A rapid increase in blood glucose led to the simultaneous appearance of a transitory phase of continuous electrical activity in both cells. These results indicate that under physiological conditions, the islets operate as a functional syncytium. 3. Simultaneous recordings of cells from two different islets within the same pancreas showed that the oscillations in the electrical activity were not synchronous, which suggests that each islet is a functionally independent unit. Rapid changes in blood glucose led to the appearance of a transitory phase of increased electrical activity in both islets, although of different duration. These results suggest that the endocrine pancreas lacks a pacemaker driving the electrical activity of all the islets. 4. The comparison of the degree of activation of different islets, simultaneously recorded at different glucose concentrations, indicated that all the islets had a similar sensitivity to glucose. Furthermore, when the glucose concentration was increased, the electrical activity in both islets increased in parallel, suggesting that the amount of insulin released due to the increase in glycaemia was produced by the simultaneous response of all the islets and not by the recruitment of islets with different sensitivities to glucose. 5. Our results predict that the synchronous electrical activity of all the cells within an islet will result in widespread intracellular calcium oscillations and pulsatile insulin secretion. The periodicity of the pulses of insulin secretion in different islets is suggested to be of slightly different length and asynchronous. PMID:8735691

  17. Pancreatic duodenal homeobox 1 protein is a novel beta-cell-specific autoantigen for type I diabetes.

    PubMed

    Li, Shi-Wu; Koya, Vijay; Li, Yi; Donelan, William; Lin, Peng; Reeves, Westley H; Yang, Li-Jun

    2010-01-01

    Pancreatic duodenal homeobox 1 (Pdx1) protein is a key transcription factor involved in the regulation of insulin gene expression that is expressed at high levels in the beta-cells of the pancreatic islets. We asked whether Pdx1 is a target of anti-islet autoimmunity in type I diabetes (T1D). Pdx1 autoantibodies (PAAs) were detected in non-obese diabetic (NOD) mice using ELISA, western blotting, and radioimmunoprecipitation of [(35)S]-labeled insulinoma cell line-derived Pdx1 protein. PAAs were detected as early as at 5 weeks of age, and generally peaked before the onset of clinically overt diabetes in diabetes-prone female NOD mice. Levels declined substantially after the onset of diabetes. PAAs were not detected in the sera of NOD-scid, C57BL/6, or BALB/c mice. The titers of PAAs in NOD mouse sera were as high as 1/93 750 by ELISA. The fine specificity of PAAs was determined by western blotting using a series of truncated recombinant Pdx1 proteins. The immunodominant epitopes were located to the C-terminus of the Pdx1 (p200-283) in NOD mice. PAAs also were detected in sera from human T1D patients, but the major epitopes were localized to amino acids 159-200 as well as the same region (p200-283) recognized by PAAs from NOD mice. Using [(3)H]thymidine incorporation, the p83 fragment of Pdx1 specifically stimulated proliferation of splenic T cells from recent-onset diabetic NOD mice. The presence of PAAs in prediabetic NOD mice and human T1D patients, and Pdx1-specific T-cell proliferation in NOD mice provide a strong rationale for further investigation of the pathogenic role of immune responses against Pdx1 in T1D.

  18. Lipodystrophy in human immunodeficiency virus patients impairs insulin action and induces defects in beta-cell function.

    PubMed

    Andersen, Ove; Haugaard, Steen B; Andersen, Ulrik B; Friis-Møller, Nina; Storgaard, Heidi; Vølund, Aage; Nielsen, Jens Ole; Iversen, Johan; Madsbad, Sten

    2003-10-01

    The pathophysiology of insulin resistance in human immunodeficiency virus (HIV)-associated lipodystrophy syndrome (HALS) is not fully clarified. We investigated 18 men with HALS and 18 HIV-positive males without lipodystrophy (control subjects). Duration and modality of antiretroviral therapy were similar between study groups. A hyperinsulinemic euglycemic clamp showed an impaired glucose disposal rate (GDR) in HALS patients (5.6 v 8.3 mg glucose/min. kg(FFM), P =.0006). As demonstrated by indirect calorimetry, HALS patients showed an impaired nonoxidative glucose metabolism (NOGM, 2.2 v 4.2, P =.006), whereas levels of basal and insulin-stimulated oxidative glucose metabolism (OGM) (2.4 v 2.3, P =.55, and 3.3 v 4.0, P =.064, respectively) were not significantly different between groups. Despite comparable total fat masses, dual energy x-ray absorptiometry (DEXA) scans showed that the percentage of limb fat (ie, peripheral-fat-mass/[peripheral-fat-mass + trunk-fat-mass]. 100%) was reduced in HALS patients (36% v 46%, P =.0002). Multiple linear regression analysis indicated that percentage of limb fat explained 53% of the variability of GDR and 45% of the variability of NOGM in HALS patients. In HALS patients, leg fat mass correlated positively with NOGM (r =.51, P <.05), whereas abdominal fat mass and NOGM did not correlate (P =.91). Analyzing the relationship between first phase insulin secretion and insulin sensitivity, 6 HALS patients compared with none of the control subjects exhibited impaired insulin secretion (P <.05). Our data suggest that fat redistribution independently of antiretroviral therapy is highly related to insulin resistance in HALS patients. Furthermore, in HALS patients, impaired glucose metabolism most likely relates to decreased NOGM and to defects in beta-cell function.

  19. Delayed-rectifier (KV2.1) regulation of pancreatic beta-cell calcium responses to glucose: inhibitor specificity and modeling.

    PubMed

    Tamarina, Natalia A; Kuznetsov, Andrey; Fridlyand, Leonid E; Philipson, Louis H

    2005-10-01

    The delayed-rectifier (voltage-activated) K(+) conductance (K(V)) in pancreatic islet beta-cells has been proposed to regulate plasma membrane repolarization during responses to glucose, thereby determining bursting and Ca(2+) oscillations. Here, we verified the expression of K(V)2.1 channel protein in mouse and human islets of Langerhans. We then probed the function of K(V)2.1 channels in islet glucose responses by comparing the effect of hanatoxin (HaTx), a specific blocker of K(V)2.1 channels, with a nonspecific K(+) channel blocker, tetraethylammonium (TEA). Application of HaTx (1 microM) blocked delayed-rectifier currents in mouse beta-cells, resulting in a 40-mV rightward shift in threshold of activation of the voltage-dependent outward current. In the presence of HaTx, there was negligible voltage-activated outward current below 0 mV, suggesting that K(V)2.1 channels form the predominant part of this current in the physiologically relevant range. We then employed HaTx to study the role of K(V)2.1 in the beta-cell Ca(2+) responses to elevated glucose in comparison with TEA. Only HaTx was able to induce slow intracellular Ca(2+) concentration ([Ca(2+)](i)) oscillations in cells stimulated with 20 mM glucose, whereas TEA induced an immediate rise in [Ca(2+)](i) followed by rapid oscillations. In human islets, HaTx acted in a similar fashion. The data were analyzed using a detailed mathematical model of ionic flux and Ca(2+) regulation in beta-cells. The results can be explained by a specific HaTx effect on the K(V) current, whereas TEA affects multiple K(+) conductances. The results underscore the importance of K(V)2.1 channel in repolarization of the pancreatic beta-cell plasma membrane and its role in regulating insulin secretion. PMID:16014354

  20. Calpains, mitochondria, and apoptosis

    PubMed Central

    Smith, Matthew A.; Schnellmann, Rick G.

    2012-01-01

    Mitochondrial activity is critical for efficient function of the cardiovascular system. In response to cardiovascular injury, mitochondrial dysfunction occurs and can lead to apoptosis and necrosis. Calpains are a 15-member family of Ca2+-activated cysteine proteases localized to the cytosol and mitochondria, and several have been shown to regulate apoptosis and necrosis. For example, in endothelial cells, Ca2+ overload causes mitochondrial calpain 1 cleavage of the Na+/Ca2+ exchanger leading to mitochondrial Ca2+ accumulation. Also, activated calpain 1 cleaves Bid, inducing cytochrome c release and apoptosis. In renal cells, calpains 1 and 2 promote apoptosis and necrosis by cleaving cytoskeletal proteins, which increases plasma membrane permeability and cleavage of caspases. Calpain 10 cleaves electron transport chain proteins, causing decreased mitochondrial respiration and excessive activation, or inhibition of calpain 10 activity induces mitochondrial dysfunction and apoptosis. In cardiomyocytes, calpain 1 activates caspase 3 and poly-ADP ribose polymerase during tumour necrosis factor-α-induced apoptosis, and calpain 1 cleaves apoptosis-inducing factor after Ca2+ overload. Many of these observations have been elucidated with calpain inhibitors, but most calpain inhibitors are not specific for calpains or a specific calpain family member, creating more questions. The following review will discuss how calpains affect mitochondrial function and apoptosis within the cardiovascular system. PMID:22581845

  1. Mitochondrial regulation of apoptosis.

    PubMed

    Mayer, Bernd; Oberbauer, Rainer

    2003-06-01

    Mitochondria play a central part in cellular survival and apoptotic death. These processes are highly regulated by pro- and antiapoptotic Bcl-2 superfamily members. A key feature within apoptosis cascades is disruption of mitochondrial transmembrane potential and apoptogenic protein release, caused by opening of the permeability transition pore (PT). New data, however, indicate that mitochondrial apoptosis may occur without PT involvement.

  2. The apoptosis database.

    PubMed

    Doctor, K S; Reed, J C; Godzik, A; Bourne, P E

    2003-06-01

    The apoptosis database is a public resource for researchers and students interested in the molecular biology of apoptosis. The resource provides functional annotation, literature references, diagrams/images, and alternative nomenclatures on a set of proteins having 'apoptotic domains'. These are the distinctive domains that are often, if not exclusively, found in proteins involved in apoptosis. The initial choice of proteins to be included is defined by apoptosis experts and bioinformatics tools. Users can browse through the web accessible lists of domains, proteins containing these domains and their associated homologs. The database can also be searched by sequence homology using basic local alignment search tool, text word matches of the annotation, and identifiers for specific records. The resource is available at http://www.apoptosis-db.org and is updated on a regular basis.

  3. IL-1beta-induced nitric oxide release from insulin-secreting beta-cells: further evidence for the involvement of GTP-binding proteins.

    PubMed

    Tannous, M; Veluthakal, R; Amin, R; Kowluru, A

    2002-12-01

    Recently, we have demonstrated regulatory roles for G-proteins (e.g., H-Ras) in IL-1beta induced NO release from HIT-T15 cells. Herein, we report a similar regulatory mechanism for IL-1beta induced NO release from RIN5F and INS-1 cells. Our data indicate that functional inactivation of Ras, either by Clostridial toxins or by specific inhibitors of Ras function, results in a significant inhibition in IL-1beta induced NO release, suggesting that activation of specific G-proteins is essential for IL-1beta induced NO release. In the present study, we report possible loci where IL-1beta treatment might result in functional activation of these G-proteins. For example, IL-1beta treatment resulted in significant reduction in (high-and low-affinity) GTPase activities in lysates derived from normal rat islets; such a scenario might lead to retention of candidate G-proteins in GTP-bound, active conformation. Further, IL-1beta treatment increased the G-protein carboxyl methyl transferase activity as well as carboxyl methylation of endogenous beta-cell proteins; such a modification has been shown to increase the membrane association and interaction of these G-proteins with their respective effector proteins. Also, we report immunologic localization of H-Ras regulatory proteins including its nucleotide exchange factor (GRF-1) and its effector protein (eg., Raf-1) in isolated beta-cells. Together, our data indicate localization, and regulation by IL-1beta, of specific enzymes that are critical to activation of G-proteins. Based on these preliminary findings, we propose a model for the involvement of G-proteins in IL-1beta induced NO release and subsequent demise of the pancreatic beta-cell. PMID:12688637

  4. Trophic effects of vanadium on beta-cells of STZ-induced insulin dependent diabetic rats & evidence for long-term relief of diabetes mellitus.

    PubMed

    Dehghani, G A; Sotoodeh, M; Omrani, G R

    1999-08-01

    The effects of one year combined vanadium and insulin treatment (VIT) on blood glucose levels of insulin dependent diabetic (IDD) rats were studied. Rats made diabetic by an i.v. injection of 55-60 mg/kg streptozotocin (STZ), divided into two groups and treated with a low dose of NPH insulin (2-4 U/rat) for two months to survive from hyperglycaemic shock. In group A, hyperglycaemia ameliorated during one year by increasing the daily dose of insulin to 8.2 +/- 0.4 U/100 g (IT) and in group B by switching over to hydrated vanadium solution (1 mg/ml vanadyl oxide sulphate pentahydrate in drinking water; VIT). The results of the study indicated that one year VIT regenerated new beta-cells, and relieved diabetes both during treatment and after withdrawal. However, one year IT showed no trophic effects on the destroyed beta-cells, hence no improvement in the glycaemic status of the animal was seen after withdrawal. The action of VIT was such that in group B normoglycaemeia persisted in 90 per cent of diabetic rats two weeks after insulin withdrawal. But in the same group, 45 days after combined vanadium and insulin withdrawal blood glucose was normal in 60 per cent of the rats, it was between 250-300 mg/dl in 18 per cent and between 350-400 mg/dl in 24 per cent of the rats. In conclusion it appears that long term VIT regenerates pancreatic beta-cells of IDD rats and possibly by improving their secretory functions it relieves diabetes mellitus.

  5. Prolonged L-alanine exposure induces changes in metabolism, Ca(2+) handling and desensitization of insulin secretion in clonal pancreatic beta-cells.

    PubMed

    McClenaghan, Neville H; Scullion, Siobhan M; Mion, Brian; Hewage, Chandralal; Malthouse, J Paul G; Flatt, Peter R; Newsholme, Philip; Brennan, Lorraine

    2009-02-01

    Acute insulin-releasing actions of amino acids have been studied in detail, but comparatively little is known about the beta-cell effects of long-term exposure to amino acids. The present study examined the effects of prolonged exposure of beta-cells to the metabolizable amino acid L-alanine. Basal insulin release or cellular insulin content were not significantly altered by alanine culture, but acute alanine-induced insulin secretion was suppressed by 74% (P<0.001). Acute stimulation of insulin secretion with glucose, KCl or KIC (2-oxoisocaproic acid) following alanine culture was not affected. Acute alanine exposure evoked strong cellular depolarization after control culture, whereas AUC (area under the curve) analysis revealed significant (P<0.01) suppression of this action after culture with alanine. Compared with control cells, prior exposure to alanine also markedly decreased (P<0.01) the acute elevation of [Ca(2+)](i) (intracellular [Ca(2+)]) induced by acute alanine exposure. These diminished stimulatory responses were partially restored after 18 h of culture in the absence of alanine, indicating reversible amino-acid-induced desensitization. (13)C NMR spectra revealed that alanine culture increased glutamate labelling at position C4 (by 60%; P<0.01), as a result of an increase in the singlet peak, indicating increased flux through pyruvate dehydrogenase. Consistent with this, protein expression of the pyruvate dehydrogenase kinases PDK2 and PDK4 was significantly reduced. This was accompanied by a decrease in cellular ATP (P<0.05), consistent with diminished insulin-releasing actions of this amino acid. Collectively, these results illustrate the phenomenon of beta-cell desensitization by amino acids, indicating that prolonged exposure to alanine can induce reversible alterations to metabolic flux, Ca(2+) handling and insulin secretion. PMID:18702613

  6. Differentiation and transplantation of functional pancreatic beta cells generated from induced pluripotent stem cells derived from a type 1 diabetes mouse model.

    PubMed

    Jeon, Kilsoo; Lim, Hyejin; Kim, Jung-Hyun; Thuan, Nguyen Van; Park, Seung Hwa; Lim, Yu-Mi; Choi, Hye-Yeon; Lee, Eung-Ryoung; Kim, Jin-Hoi; Lee, Myung-Shik; Cho, Ssang-Goo

    2012-09-20

    The nonobese diabetic (NOD) mouse is a classical animal model for autoimmune type 1 diabetes (T1D), closely mimicking features of human T1D. Thus, the NOD mouse presents an opportunity to test the effectiveness of induced pluripotent stem cells (iPSCs) as a therapeutic modality for T1D. Here, we demonstrate a proof of concept for cellular therapy using NOD mouse-derived iPSCs (NOD-iPSCs). We generated iPSCs from NOD mouse embryonic fibroblasts or NOD mouse pancreas-derived epithelial cells (NPEs), and applied directed differentiation protocols to differentiate the NOD-iPSCs toward functional pancreatic beta cells. Finally, we investigated whether the NPE-iPSC-derived insulin-producing cells could normalize hyperglycemia in transplanted diabetic mice. The NOD-iPSCs showed typical embryonic stem cell-like characteristics such as expression of markers for pluripotency, in vitro differentiation, teratoma formation, and generation of chimeric mice. We developed a method for stepwise differentiation of NOD-iPSCs into insulin-producing cells, and found that NPE-iPSCs differentiate more readily into insulin-producing cells. The differentiated NPE-iPSCs expressed diverse pancreatic beta cell markers and released insulin in response to glucose and KCl stimulation. Transplantation of the differentiated NPE-iPSCs into diabetic mice resulted in kidney engraftment. The engrafted cells responded to glucose by secreting insulin, thereby normalizing blood glucose levels. We propose that NOD-iPSCs will provide a useful tool for investigating genetic susceptibility to autoimmune diseases and generating a cellular interaction model of T1D, paving the way for the potential application of patient-derived iPSCs in autologous beta cell transplantation for treating diabetes. PMID:22512788

  7. Apoptosis, autophagy, and more.

    PubMed

    Lockshin, Richard A; Zakeri, Zahra

    2004-12-01

    Cell death has been subdivided into the categories apoptosis (Type I), autophagic cell death (Type II), and necrosis (Type III). The boundary between Type I and II has never been completely clear and perhaps does not exist due to intrinsic factors among different cell types and the crosstalk among organelles within each type. Apoptosis can begin with autophagy, autophagy can end with apoptosis, and blockage of caspase activity can cause a cell to default to Type II cell death from Type I. Furthermore, autophagy is a normal physiological process active in both homeostasis (organelle turnover) and atrophy. "Autophagic cell death" may be interpreted as the process of autophagy that, unlike other situations, does not terminate before the cell collapses. Since switching among the alternative pathways to death is relatively common, interpretations based on knockouts or inhibitors, and therapies directed at controlling apoptosis must include these considerations.

  8. Development of a novel beta-cell specific promoter system for the identification of insulin-producing cells in in vitro cell cultures.

    PubMed

    Fukazawa, Takuya; Matsuoka, Junji; Naomoto, Yoshio; Nakai, Toru; Durbin, Mary L; Kojima, Itaru; Lakey, Jonathan R T; Tanaka, Noriaki

    2006-10-15

    Recently, it has been reported that islet transplantation into patients with Type 1 diabetes may achieve insulin independence for a year or longer [Shapiro et al., Islet transplantation in seven patients with type 1 diabetes mellitus using a glucocorticoid-free immunosuppressive regimen, N Engl J Med. 343 (2000) 230-238]. However, the amount of donor islet tissue is limited, therefore, multiple approaches are being explored to generate insulin-producing cells in vitro. Some promising results have been obtained using mouse and human stem cells and progenitor cells [Soria et al., From stem cells to beta cells: new strategies in cell therapy of diabetes mellitus, Diabetologia. 4 (2001) 407-415; Lechner et al., Stem/progenitor cells derived from adult tissues: potential for the treatment of diabetes mellitus, Am J Physiol Endocrinol Metab. 284 (2003) 259-266; Bonner-Weir et al., In vitro cultivation of human islets from expanded ductal tissue, Proc Natl Acad Sci U S A, 97 (2000) 7999-8004; Assady et al., Insulin production by human embryonic stem cells, 50 (2001) Diabetes 1691-1697]. However, the efficiency of obtaining populations with high numbers of differentiated cells has been poor. In order to improve the efficiency of producing and selecting insulin-producing cells from undifferentiated cells, we have designed a novel beta-cell specific and glucose responsive promoter system designated pGL3.hINS-363 3x. This artificial promoter system exhibits significant luciferase activity not only in insulin-producing MIN6 m9 cells but also in isolated human islets. The pGL3.hINS-363 3x construct shows no activity in non-insulin-producing cells in low glucose conditions (2 mM glucose) but demonstrates significant activity and beta-cell specificity in high glucose conditions (16 mM glucose). Furthermore, pGL3.hINS-363 3x shows significant promoter activity in differentiated AR42J cells that can produce insulin after activin A and betacellulin treatment. Here, we describe a

  9. Pancreatic beta cell function following liraglutide-augmented weight loss in individuals with prediabetes: analysis of a randomised, placebo-controlled study

    PubMed Central

    Liu, Alice; Ariel, Danit; Abbasi, Fahim; Lamendola, Cindy; Grove, Kaylene; Tomasso, Vanessa; Reaven, Gerald

    2016-01-01

    Aims/hypothesis Liraglutide can modulate insulin secretion by directly stimulating beta cells or indirectly through weight loss and enhanced insulin sensitivity. Recently, we showed that liraglutide treatment in overweight individuals with prediabetes (impaired fasting glucose and/or impaired glucose tolerance) led to greater weight loss (−7.7% vs −3.9%) and improvement in insulin resistance compared with placebo. The current study evaluates the effects on beta cell function of weight loss augmented by liraglutide compared with weight loss alone. Methods This was a parallel, randomised study conducted in a single academic centre. Both participants and study administrators were blinded to treatment assignment. Individuals who were 40–70 years old, overweight (BMI 27–40 kg/m2) and with prediabetes were randomised (via a computerised system) to receive liraglutide (n = 35) or matching placebo (n = 33), and 49 participants were analysed. All were instructed to follow an energy-restricted diet. Primary outcome was insulin secretory function, which was evaluated in response to graded infusions of glucose and day-long mixed meals. Results Liraglutide treatment (n = 24) significantly (p ≤0.03) increased the insulin secretion rate (% mean change [95% CI]; 21% [12, 31] vs −4% [−11, 3]) and pancreatic beta cell sensitivity to intravenous glucose (229% [161, 276] vs −0.5% (−15, 14]), and decreased insulin clearance rate (−3.5% [−11, 4] vs 8.2 [0.2, 16]) as compared with placebo (n = 25). The liraglutide-treated group also had significantly (p ≤0.03) lower day-long glucose (−8.2% [−11, −6] vs −0.1 [−3, 2]) and NEFA concentrations (−14 [−20, −8] vs −2.1 [−10, 6]) following mixed meals, whereas day-long insulin concentrations did not significantly differ as compared with placebo. In a multivariate regression analysis, weight loss was associated with a decrease in insulin secretion rate and day-long glucose and insulin concentrations in

  10. Regulation of gene expression by glucose in pancreatic beta -cells (MIN6) via insulin secretion and activation of phosphatidylinositol 3'-kinase.

    PubMed

    da Silva Xavier, G; Varadi, A; Ainscow, E K; Rutter, G A

    2000-11-17

    Increases in glucose concentration control the transcription of the preproinsulin (PPI) gene and several other genes in the pancreatic islet beta-cell. Although recent data have demonstrated that secreted insulin may regulate the PPI gene (Leibiger, I. B., Leibiger, B., Moede, T., and Berggren, P. O. (1998) Mol. Cell 1, 933-938), the role of insulin in the control of other beta-cell genes is unexplored. To study the importance of insulin secretion in the regulation of the PPI and liver-type pyruvate kinase (L-PK) genes by glucose, we have used intranuclear microinjection of promoter-luciferase constructs into MIN6 beta-cells and photon-counting imaging. The activity of each promoter was increased either by 30 (versus 3) mm glucose or by 1-20 nm insulin. These effects of insulin were not due to enhanced glucose metabolism since culture with the hormone had no impact on the stimulation of increases in intracellular ATP concentration caused by 30 mm glucose. Furthermore, the islet-specific glucokinase promoter and cellular glucokinase immunoreactivity were unaffected by 30 mm glucose or 20 nm insulin. Inhibition of insulin secretion with the Ca(2+) channel blocker verapamil, the ATP-sensitive K(+) channel opener diazoxide, or the alpha(2)-adrenergic agonist clonidine blocked the effects of glucose on L-PK gene transcription. Similarly, 30 mm glucose failed to induce the promoter after inhibition of phosphatidylinositol 3'-kinase activity with LY294002 and the expression of dominant negative-acting phosphatidylinositol 3'-kinase (Deltap85) or the phosphoinositide 3'-phosphatase PTEN (phosphatase and tensin homologue). LY294002 also diminished the activation of the L-PK gene caused by inhibition of 5'-AMP-activated protein kinase with anti-5'-AMP-activated protein kinase alpha2 antibodies. Conversely, stimulation of insulin secretion with 13 mm KCl or 10 microm tolbutamide strongly activated the PPI and L-PK promoters. These data indicate that, in MIN6 beta-cells

  11. Induction of Apoptosis.

    PubMed

    2016-01-01

    The apoptotic activity of plants is checked to confirm its anti-tumour and anti-cancer activity. Apoptosis is a specific process that leads to intrinsic programmed cell death which is essential in the homeostasis of normal tissues of the body and occurs in various physiological and pathological situations. Method to check apoptosis in EAC cells and DNA analysis are featured in this chapter as a preliminary test manner. PMID:26939284

  12. Age related changes in pancreatic beta cells: A putative extra-cerebral site of Alzheimer’s pathology

    PubMed Central

    Maj, Magdalena; Ilhan, Aysegul; Neziri, Dashurie; Gartner, Wolfgang; Berggard, Tord; Attems, Johannes; Base, Wolfgang; Wagner, Ludwig

    2011-01-01

    Frequent concomitant manifestation of type 2 diabetes mellitus (T2DM) and Alzheimer’s disease (AD) has been recently demonstrated by epidemiological studies. This might be due to functional similarities between β-cells and neurons, such as secretion on demand of highly specific molecules in a tightly controlled fashion. An additional similarity represents the age-related alteration of hyperphosphorylated tau in AD patients. Similarly, alterations have been identified in β-cells of T2DM patients. The islet amyloid polypeptide has been associated with β-cell apoptosis. As a consequence of increasing age, the accumulation of highly modified proteins together with decreased regenerative potential might lead to increasing rates of apoptosis. Moreover, reduction of β-cell replication capabilities results in reduction of β-cell mass in mammals, simultaneously with impaired glucose tolerance. The new challenge is to learn much more about age-related protein modifications. This can lead to new treatment strategies for reducing the incidence of T2DM and AD. PMID:21537460

  13. Hepatocyte nuclear factor 4 alpha isoforms originated from the P1 promoter are expressed in human pancreatic beta-cells and exhibit stronger transcriptional potentials than P2 promoter-driven isoforms.

    PubMed

    Eeckhoute, J; Moerman, E; Bouckenooghe, T; Lukoviak, B; Pattou, F; Formstecher, P; Kerr-Conte, J; Vandewalle, B; Laine, B

    2003-05-01

    The nuclear receptor hepatocyte nuclear factor (HNF) 4 alpha is involved in a transcriptional network and plays an important role in pancreatic beta-cells. Mutations in the HNF4 alpha gene are correlated with maturity-onset diabetes of the young 1. HNF4 alpha isoforms result from both alternative splicing and alternate usage of promoters P1 and P2. It has recently been reported that HNF4 alpha transcription is driven almost exclusively by the P2 promoter in pancreatic islets. We observed that transcripts from both P1 and P2 promoters were expressed in human pancreatic beta-cells and in the pancreatic beta-cell lines RIN m5F and HIT-T15. Expression of HNF4 alpha proteins originating from the P1 promoter was confirmed by immunodetection. Due to the presence of the activation function module AF-1, HNF4 alpha isoforms originating from the P1 promoter exhibit stronger transcriptional activities and recruit coactivators more efficiently than isoforms driven by the P2 promoter. Conversely, activities of isoforms produced by both promoters were similarly repressed by the corepressor small heterodimer partner. These behaviors were observed on the promoter of HNF1 alpha that is required for beta-cell function. Our results highlight that expression of P1 promoter-driven isoforms is important in the control of pancreatic beta-cell function.

  14. Metformin ameliorates lipotoxicity-induced mesangial cell apoptosis partly via upregulation of glucagon like peptide-1 receptor (GLP-1R).

    PubMed

    Kim, Dong-il; Park, Min-jung; Heo, Young-ran; Park, Soo-hyun

    2015-10-15

    Glucagon like peptide-1 receptor (GLP-1R), known to be expressed in pancreatic beta cells, is also expressed in glomerular mesangial cells and its agonist has protective effects in diabetic nephropathy. However, its regulatory mechanisms by lipotoxicity in glomerular mesangial cells are not understood. We found that palmitate-mediated lipotoxicity increased apoptosis and decreased GLP-1R expression in a rat mesangial cell line. Silencing GLP-1R expression also increased mesangial cell apoptosis. Interestingly, metformin, one of the biguanide drugs that has anti-diabetic effects, attenuated lipotoxicity-induced mesangial cell apoptosis and restored GLP-1R expression. Moreover, this treatment alleviated GLP-1R knockdown-induced mesangial cell apoptosis. To further evaluate in vivo, diabetic obese db/db mice were administered metformin. Glomerular GLP-1R expression was diminished in db/db mice, as compared with db/m control mice. However, this decrease significantly recovered on metformin administration. Together, these data provide novel evidence that lipotoxicity decreases the mesangial GLP-1R expression in intact cells and in vivo. The decrease induced mesangial cell apoptosis. Furthermore, we provided the evidence that metformin treatment has a renal protective effect partly via increased mesangial GLP-1R expression. Our data suggested that regulation of GLP-1R expression could be a promising approach to treat diabetic nephropathy and the novel mechanism of metformin mediated GLP-1R regulation.

  15. PTBP1 is required for glucose-stimulated cap-independent translation of insulin granule proteins and Coxsackieviruses in beta cells

    PubMed Central

    Knoch, Klaus-Peter; Nath-Sain, Suchita; Petzold, Antje; Schneider, Hendryk; Beck, Mike; Wegbrod, Carolin; Sönmez, Anke; Münster, Carla; Friedrich, Anne; Roivainen, Merja; Solimena, Michele

    2014-01-01

    Glucose and GLP-1 stimulate not only insulin secretion, but also the post-transcriptional induction of insulin granule biogenesis. This process involves the nucleocytoplasmic translocation of the RNA binding protein PTBP1. Binding of PTBP1 to the 3′-UTRs of mRNAs for insulin and other cargoes of beta cell granules increases their stability. Here we show that glucose enhances also the binding of PTBP1 to the 5′-UTRs of these transcripts, which display IRES activity, and their translation exclusively in a cap-independent fashion. Accordingly, glucose-induced biosynthesis of granule cargoes was unaffected by pharmacological, genetic or Coxsackievirus-mediated inhibition of cap-dependent translation. Infection with Coxsackieviruses, which also depend on PTBP1 for their own cap-independent translation, reduced instead granule stores and insulin release. These findings provide insight into the mechanism for glucose-induction of insulin granule production and on how Coxsackieviruses, which have been implicated in the pathogenesis of type 1 diabetes, can foster beta cell failure. PMID:25061557

  16. The relationship between node degree and dissipation rate in networks of diffusively coupled oscillators and its significance for pancreatic beta cells.

    PubMed

    Gosak, Marko; Stožer, Andraž; Markovič, Rene; Dolenšek, Jurij; Marhl, Marko; Rupnik, Marjan Slak; Perc, Matjaž

    2015-07-01

    Self-sustained oscillatory dynamics is a motion along a stable limit cycle in the phase space, and it arises in a wide variety of mechanical, electrical, and biological systems. Typically, oscillations are due to a balance between energy dissipation and generation. Their stability depends on the properties of the attractor, in particular, its dissipative characteristics, which in turn determine the flexibility of a given dynamical system. In a network of oscillators, the coupling additionally contributes to the dissipation, and hence affects the robustness of the oscillatory solution. Here, we therefore investigate how a heterogeneous network structure affects the dissipation rate of individual oscillators. First, we show that in a network of diffusively coupled oscillators, the dissipation is a linearly decreasing function of the node degree, and we demonstrate this numerically by calculating the average divergence of coupled Hopf oscillators. Subsequently, we use recordings of intracellular calcium dynamics in pancreatic beta cells in mouse acute tissue slices and the corresponding functional connectivity networks for an experimental verification of the presented theory. We use methods of nonlinear time series analysis to reconstruct the phase space and calculate the sum of Lyapunov exponents. Our analysis reveals a clear tendency of cells with a higher degree, that is, more interconnected cells, having more negative values of divergence, thus confirming our theoretical predictions. We discuss these findings in the context of energetic aspects of signaling in beta cells and potential risks for pathological changes in the tissue. PMID:26232966

  17. Glucose enhances insulin promoter activity in MIN6 beta-cells independently of changes in intracellular Ca2+ concentration and insulin secretion.

    PubMed Central

    Kennedy, H J; Rafiq, I; Pouli, A E; Rutter, G A

    1999-01-01

    Recent studies have suggested that glucose may activate insulin gene transcription through increases in intracellular Ca(2+) concentration, possibly acting via the release of stored insulin. We have investigated this question by dynamic photon-counting imaging of insulin- and c-fos-promoter-firefly luciferase reporter construct activity. Normalized to constitutive viral promoter activity, insulin promoter activity in MIN6 beta-cells was increased 1.6-fold after incubation at 30 mM compared with 3 mM glucose, but was unaltered at either glucose concentration by the presence of insulin (100 nM) or the Ca(2+) channel inhibitor, verapamil (100 microM). Increases in intracellular [Ca(2+)] achieved by plasma membrane depolarization with KCl failed to enhance either insulin or c-fos promoter activity in MIN6 cells, but increased c-fos promoter activity 5-fold in AtT20 cells. Together, these results demonstrate that glucose can exert a direct effect on insulin promoter activity in islet beta-cells, via a signalling pathway which does not require increases in intracellular [Ca(2+)] nor insulin release and insulin receptor activation. PMID:10455011

  18. Relationship between homeostasis model assessment of insulin resistance and beta cell function and serum 25-hydroxyvitamin D in non-diabetic Korean adults

    PubMed Central

    Yoon, Hyun; Jeon, Dae Jung; Park, Chang Eun; You, Hye Sook; Moon, Ae Eun

    2016-01-01

    The purpose of this study is to look at these relationships in non-diabetic Korean adults. This study was based on data from the KNHANES V-1, which is representative of the population of Korea. A total of 5,492 participants (≥20 years in age) without type 1 or type 2 diabetes, assessed for serum 25-hydroxyvitamin D [25(OH)D], fasting blood glucose and insulin, as well as anthropometric variables, were included in the analyses. The key study results were as follows: First, vitamin D status [vitamin D deficient, 25(OH)D <25 nM; vitamin D insufficient, 25(OH)D ≥25, <50 nM; vitamin D sufficient, 25(OH)D ≥50 nM] was inversely associated with homeostasis model assessment of insulin resistance (HOMA-IR) and beta cell function (HOMA-B) in model 2 (adjusted for age and gender) and 3 (further adjusted for smoking, alcohol drinking, regular exercise, systolic and diastolic blood pressure, waist circumference, and body mass index). Second, in model 4, when further adjusted for total cholesterol, triglycerides, and HDL-C, vitamin D status was inversely associated with HOMA-B. However, association of vitamin D status and HOMA-IR was no longer significant. In conclusion, vitamin D was inversely associated with beta cell function in non-diabetic Korean adults but was not associated with insulin resistance.

  19. The diabetogenic effects of excessive ethanol: reducing beta-cell mass, decreasing phosphatidylinositol 3-kinase activity and GLUT-4 expression in rats.

    PubMed

    Zhao, Li-Na; Hao, Li-Ping; Yang, Xue-Feng; Ying, Chen-Jiang; Yu, Dong; Sun, Xiu-Fa

    2009-05-01

    The diabetogenic impact of ethanol remains as a focal point of basic and clinical investigations. In this study, Wistar rats were subjected to daily intragastric ethanol administration (10 ml/kg body weight injection with 0 (control), 10, 20 and 33 % (v/v) ethanol in the injections, respectively) for 19 weeks. At the end of the administration, we found that the fasting plasma glucose level of the 33 % (v/v) ethanol-loaded group was 18 % higher than the control. Insulin sensitivity was decreased in a dose-dependent manner in all the ethanol-loaded groups (r - 0.842, P < 0.001) during intraperitoneal insulin tolerance test. Necrotic/haemorrhagic injury was detected in the pancreas and islet beta-cell mass was significantly reduced in the 33 % (v/v) ethanol-loaded rats by immunohistochemical and morphometric analysis. At the molecular level, we detected a dose-dependent attenuation of phosphatidylinositol 3-kinase activity (r - 0.956, P < 0.001) and GLUT-4 expression (GLUT-4 mRNA, r - 0.899, P < 0.001; GLUT-4 protein, r - 0.964, P < 0.001) in skeletal muscle. These results demonstrated that drinking is a conditional aetiological factor for diabetes and excessive ethanol intake is negatively associated with both insulin sensitivity and beta-cell mass. The whole-body insulin resistance might result from the ethanol-induced insulin signalling defects in muscle.

  20. Effects of glass ionomers and dental resin composites on viability of beta-cells and insulin release in isolated islets of Langerhans.

    PubMed

    Persson-Sjögren, Solveig; Sjögren, Göran

    2003-09-01

    Information on the biocompatibility of glass ionomers and resin composites is sparse. To extend the scale of biological testing we evaluated the influence of those materials on insulin secretion at whole organ level in vitro. The effects on insulin secretion of three glass ionomers and two resin composites, aged for 1 week, were studied in isolated mouse islets of Langerhans at basal (5.5mM) and at stimulatory (11.1mM) D-glucose concentrations. In addition, viability of single mouse beta-cells was evaluated. The effect of glass ionomer specimens aged for 1 and 4 months on insulin secretion at 11.1mM D-glucose was also studied. None of the materials affected the viability of the beta-cells. At 5.5mM D-glucose none of the materials affected the insulin secretion. At 11.1mM D-glucose, the glass ionomers only decreased the secretion and glass ionomers aged for 1 month still decreased insulin release whereas after 4 months ageing only one of the glass ionomers affected the release. The result shows a dynamic effect on insulin release of the elements and/or compounds released from the specimens.

  1. Anti-diabetic actions of carbon monoxide-releasing molecule (CORM)-A1: Immunomodulation and regeneration of islet beta cells.

    PubMed

    Nikolic, Ivana; Saksida, Tamara; Vujicic, Milica; Stojanovic, Ivana; Stosic-Grujicic, Stanislava

    2015-05-01

    We have recently shown that carbon monoxide releasing molecule (CORM)-A1 prevents type 1 diabetes induced in C57BL/6 mice with multiple low doses of streptozotocin (MLDS) by shifting the Th1/Th17/M1 balance towards a Th2/M2 response. In the present work we tested the hypothesis that CORM-A1 might influence regulatory arm of the immune response, as well as beta cell regeneration. CORM-A1 (2 mg/kg/day) was administered for 10 days to mice induced with MLDS and/or depleted of low dose cyclophosphamide (CY)-sensitive FoxP3+ T regulatory (Treg) cells. Besides monitoring hyperglycaemia, ex vivo analysis of spleen, pancreatic lymph nodes (PLN) and pancreas was performed at the end of treatment. In CORM-A1-treated MLDS-induced mice the improvement of hyperglycaemia was observed only without depletion of CY-sensitive FoxP3+ Treg cells. This was accompanied by decreased levels of interleukin (IL)-12, IL-2 and early activation marker CD25 in the spleen and PLN and increased transforming growth factor (TGF)-β, resulting in reduced lymphocyte proliferation in both organs. In parallel, decreased transcript levels of IL-2, but increased mRNA expression of TGF-β, accompanied with up-regulation of Ki-67 protein expression was observed within pancreas. Together, the data suggested that besides the immunomodulatory potential, CORM-A1 probably induces beta cell regeneration.

  2. The relationship between node degree and dissipation rate in networks of diffusively coupled oscillators and its significance for pancreatic beta cells.

    PubMed

    Gosak, Marko; Stožer, Andraž; Markovič, Rene; Dolenšek, Jurij; Marhl, Marko; Rupnik, Marjan Slak; Perc, Matjaž

    2015-07-01

    Self-sustained oscillatory dynamics is a motion along a stable limit cycle in the phase space, and it arises in a wide variety of mechanical, electrical, and biological systems. Typically, oscillations are due to a balance between energy dissipation and generation. Their stability depends on the properties of the attractor, in particular, its dissipative characteristics, which in turn determine the flexibility of a given dynamical system. In a network of oscillators, the coupling additionally contributes to the dissipation, and hence affects the robustness of the oscillatory solution. Here, we therefore investigate how a heterogeneous network structure affects the dissipation rate of individual oscillators. First, we show that in a network of diffusively coupled oscillators, the dissipation is a linearly decreasing function of the node degree, and we demonstrate this numerically by calculating the average divergence of coupled Hopf oscillators. Subsequently, we use recordings of intracellular calcium dynamics in pancreatic beta cells in mouse acute tissue slices and the corresponding functional connectivity networks for an experimental verification of the presented theory. We use methods of nonlinear time series analysis to reconstruct the phase space and calculate the sum of Lyapunov exponents. Our analysis reveals a clear tendency of cells with a higher degree, that is, more interconnected cells, having more negative values of divergence, thus confirming our theoretical predictions. We discuss these findings in the context of energetic aspects of signaling in beta cells and potential risks for pathological changes in the tissue.

  3. The Incretins and Pancreatic beta-Cells: Use of Glucagon-Like Peptide-1 and Glucose-Dependent Insulinotropic Polypeptide to Cure Type 2 Diabetes Mellitus.

    PubMed

    Kim, Mi-Hyun; Lee, Moon-Kyu

    2010-02-01

    Type 2 diabetes mellitus (T2DM) is increasing in prevalence worldwide. The complications associated with T2DM result in increased mortality and financial cost for those affected. T2DM has long been known to be associated with insulin resistance in peripheral tissues and a relative degree of insulin deficiency. However, the concept that insulin secretion and insulin sensitivity are not linked through a hyperbolic relationship in T2DM has continuously been demonstrated in many clinical trials. Thus, in order to prevent and treat T2DM, it is necessary to identify the substance(s) that will improve the function and survival of the pancreatic beta-cells in both normal and pathologic conditions, so that production and secretion of insulin can be enhanced. Incretin hormones, such as glucagon-like peptide (GLP)-1 and glucose-dependent insulinotropic polypeptide (GIP), have been shown to lower the postprandial and fasting glucose and the glycated hemoglobin levels, suppress the elevated glucagon level, and stimulate glucose-dependent insulin synthesis and secretion. In this report, we will review the biological actions and mechanisms associated with the actions of incretin hormones, GLP-1 and GIP, on beta-cell health and compare the differences between GLP-1 and GIP.

  4. Spaceflight Associated Apoptosis

    NASA Technical Reports Server (NTRS)

    Ichiki, Albert T.; Gibson, Linda A.; Allebban, Zuhair

    1996-01-01

    Lymphoid tissues have been shown to atrophy in rats flown on Russian spaceflights. Histological examination indicated evidence for cell degradation. Lymphoid tissues from rats flown on Spacelab Life Sciences-2 mission were analyzed for apoptosis by evidence of fragmented lymphocytes, which could be engulfed by macrophages, or DNA strand breaks using the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay. Apoptosis was not detected in the thymus and spleen collected inflight or from the synchronous ground rats but was detected in the thymus, spleen and inguinal lymph node of the flight animals on recovery. These results indicate that the apoptosis observed in the lymphatic tissues of the rats on recovery could have been induced by the gravitational stress of reentry, corroborating the findings from the early space-flight observations.

  5. Myocardial apoptosis and SIDS.

    PubMed

    Grasmeyer, Sarah; Madea, Burkhard

    2015-01-01

    Apoptosis mediates cardiac damage in severe forms of myocarditis. In fatal myocarditis, large amounts of cardiomyocytes show apoptotic DNA fragmentation, while in human controls, few apoptotic cardiomyocytes are found. In the present study the frequency of apoptosis in 88 SIDS cases (category 1b according to the San Diego Classification) and 15 control cases was investigated. In every case myocardial samples from 8 standard locations were collected. Detection of apoptotic cardiomyocytes was performed by TUNEL method. Furthermore the myocardial tissue was stained with HE and immunohistochemical methods (LCA, CD68, CD45-R0). More than 90% of the slides did not contain apoptotic cardiomyocytes at all. The detection rate of apoptotic cardiomyocytes was almost equal in control group (26.7%) and SIDS group (23.86%). A quantification of apoptotic cardiomyocytes per mm(2) revealed no significant difference between both groups either. Altogether there is no evidence for a higher rate of apoptosis in SIDS.

  6. Apoptosis-an introduction.

    PubMed

    Lawen, Alfons

    2003-09-01

    Apoptosis has become a major research area in the biomedical sciences. As there are more than 13,000 papers published annually on the topic, it is impossible to keep track on all developments in the area. The individual aspects of molecular control of apoptosis are well reviewed, but more general, introductory recent reviews into the field are lacking. This review aims to give a brief overview of the field, providing an introduction into the literature for students and newcomers; as it is written for the un-initiated, wherever possible, review articles will be cited rather than original papers.

  7. Oxidative stress-mediated, post-translational loss of MafA protein as a contributing mechanism to loss of insulin gene expression in glucotoxic beta cells.

    PubMed

    Harmon, Jamie S; Stein, Roland; Robertson, R Paul

    2005-03-25

    Glucose toxicity in pancreatic islet beta cells causes loss of insulin gene expression, content, and secretion due to loss of binding of transcription factors, most notably PDX-1 and RIPE-3b1 activator, to the promoter region of the insulin gene. Recently, RIPE-3b1 activator was cloned and identified as the mammalian homologue of avian MafA/Maf-L (MafA). This enabled us to carry out more extensive studies of the role of MafA in glucotoxicity than were hitherto possible. Northern analysis of glucotoxic HIT-T15 cells revealed normal amounts of MafA mRNA, but Western analysis demonstrated a 97 +/- 1% reduction in MafA protein (p < 0.0001). The proteasome is a likely site for MafA degradation as lactacystin, an irreversible proteasome inhibitor, caused an accumulation of MafA protein. Antioxidants have previously been shown to prevent the adverse effects of glucose toxicity on beta cell function both in vivo and in vitro. In the current study, chronic culturing of HIT-T15 cells with the antioxidant N-acetylcysteine (NAC) prevented loss of MafA protein (late passage = 18.9 +/- 10.4% of early passage, p < 0.001; late passage with NAC = 68.7 +/- 19.7% of early passage, p = not significant) and loss of DNA binding (late passage = 63.7 +/- 9% of early passage, p < 0.02; late passage with NAC = 116 +/- 10% of early passage, p = not significant). Additionally, transient transfection of PDX-1 or MafA cDNA into glucotoxic cells increased PDX-1 and MafA protein levels and individually increased insulin promoter activity (untreated = 34%, PDX-1 = 70%, MafA = 78%; percentage of activity of early passage cells), whereas the combined transfection of MafA and PDX-1 completely restored insulin promoter activity. This recovery of promoter activity following transient transfection had no effect on endogenous insulin mRNA. However, adenoviral infection of MafA and PDX-1 significantly increased endogenous insulin mRNA levels by 93% (121 +/- 9 versus 233 +/- 18 density light units; n = 5

  8. Variation in the FFAR1 Gene Modifies BMI, Body Composition and Beta-Cell Function in Overweight Subjects: An Exploratory Analysis

    PubMed Central

    Walker, Celia G.; Goff, Louise; Bluck, Les J.; Griffin, Bruce A.; Jebb, Susan A.; Lovegrove, Julie A.; Sanders, Thomas A. B.; Frost, Gary S.

    2011-01-01

    Background FFAR1 receptor is a long chain fatty acid G-protein coupled receptor which is expressed widely, but found in high density in the pancreas and central nervous system. It has been suggested that FFAR1 may play a role in insulin sensitivity, lipotoxicity and is associated with type 2 diabetes. Here we investigate the effect of three common SNPs of FFAR1 (rs2301151; rs16970264; rs1573611) on pancreatic function, BMI, body composition and plasma lipids. Methodology/Principal Findings For this enquiry we used the baseline RISCK data, which provides a cohort of overweight subjects at increased cardiometabolic risk with detailed phenotyping. The key findings were SNPs of the FFAR1 gene region were associated with differences in body composition and lipids, and the effects of the 3 SNPs combined were cumulative on BMI, body composition and total cholesterol. The effects on BMI and body fat were predominantly mediated by rs1573611 (1.06 kg/m2 higher (P = 0.009) BMI and 1.53% higher (P = 0.002) body fat per C allele). Differences in plasma lipids were also associated with the BMI-increasing allele of rs2301151 including higher total cholesterol (0.2 mmol/L per G allele, P = 0.01) and with the variant A allele of rs16970264 associated with lower total (0.3 mmol/L, P = 0.02) and LDL (0.2 mmol/L, P<0.05) cholesterol, but also with lower HDL-cholesterol (0.09 mmol/L, P<0.05) although the difference was not apparent when controlling for multiple testing. There were no statistically significant effects of the three SNPs on insulin sensitivity or beta cell function. However accumulated risk allele showed a lower beta cell function on increasing plasma fatty acids with a carbon chain greater than six. Conclusions/Significance Differences in body composition and lipids associated with common SNPs in the FFAR1 gene were apparently not mediated by changes in insulin sensitivity or beta-cell function. PMID:21552566

  9. Oxidative stress-mediated, post-translational loss of MafA protein as a contributing mechanism to loss of insulin gene expression in glucotoxic beta cells.

    PubMed

    Harmon, Jamie S; Stein, Roland; Robertson, R Paul

    2005-03-25

    Glucose toxicity in pancreatic islet beta cells causes loss of insulin gene expression, content, and secretion due to loss of binding of transcription factors, most notably PDX-1 and RIPE-3b1 activator, to the promoter region of the insulin gene. Recently, RIPE-3b1 activator was cloned and identified as the mammalian homologue of avian MafA/Maf-L (MafA). This enabled us to carry out more extensive studies of the role of MafA in glucotoxicity than were hitherto possible. Northern analysis of glucotoxic HIT-T15 cells revealed normal amounts of MafA mRNA, but Western analysis demonstrated a 97 +/- 1% reduction in MafA protein (p < 0.0001). The proteasome is a likely site for MafA degradation as lactacystin, an irreversible proteasome inhibitor, caused an accumulation of MafA protein. Antioxidants have previously been shown to prevent the adverse effects of glucose toxicity on beta cell function both in vivo and in vitro. In the current study, chronic culturing of HIT-T15 cells with the antioxidant N-acetylcysteine (NAC) prevented loss of MafA protein (late passage = 18.9 +/- 10.4% of early passage, p < 0.001; late passage with NAC = 68.7 +/- 19.7% of early passage, p = not significant) and loss of DNA binding (late passage = 63.7 +/- 9% of early passage, p < 0.02; late passage with NAC = 116 +/- 10% of early passage, p = not significant). Additionally, transient transfection of PDX-1 or MafA cDNA into glucotoxic cells increased PDX-1 and MafA protein levels and individually increased insulin promoter activity (untreated = 34%, PDX-1 = 70%, MafA = 78%; percentage of activity of early passage cells), whereas the combined transfection of MafA and PDX-1 completely restored insulin promoter activity. This recovery of promoter activity following transient transfection had no effect on endogenous insulin mRNA. However, adenoviral infection of MafA and PDX-1 significantly increased endogenous insulin mRNA levels by 93% (121 +/- 9 versus 233 +/- 18 density light units; n = 5

  10. Calbindin-D(28k) controls [Ca(2+)](i) and insulin release. Evidence obtained from calbindin-d(28k) knockout mice and beta cell lines

    NASA Technical Reports Server (NTRS)

    Sooy, K.; Schermerhorn, T.; Noda, M.; Surana, M.; Rhoten, W. B.; Meyer, M.; Fleischer, N.; Sharp, G. W.; Christakos, S.

    1999-01-01

    The role of the calcium-binding protein, calbindin-D(28k) in potassium/depolarization-stimulated increases in the cytosolic free Ca(2+) concentration ([Ca(2+)](i)) and insulin release was investigated in pancreatic islets from calbindin-D(28k) nullmutant mice (knockouts; KO) or wild type mice and beta cell lines stably transfected and overexpressing calbindin. Using single islets from KO mice and stimulation with 45 mM KCl, the peak of [Ca(2+)](i) was 3.5-fold greater in islets from KO mice compared with wild type islets (p < 0.01) and [Ca(2+)](i) remained higher during the plateau phase. In addition to the increase in [Ca(2+)](i) in response to KCl there was also a significant increase in insulin release in islets isolated from KO mice. Evidence for modulation by calbindin of [Ca(2+)](i) and insulin release was also noted using beta cell lines. Rat calbindin was stably expressed in betaTC-3 and betaHC-13 cells. In response to depolarizing concentrations of K(+), insulin release was decreased by 45-47% in calbindin expressing betaTC cells and was decreased by 70-80% in calbindin expressing betaHC cells compared with insulin release from vector transfected betaTC or betaHC cells (p < 0.01). In addition, the K(+)-stimulated intracellular calcium peak was markedly inhibited in calbindin expressing betaHC cells compared with vector transfected cells (225 nM versus 1,100 nM, respectively). Buffering of the depolarization-induced rise in [Ca(2+)](i) was also observed in calbindin expressing betaTC cells. In summary, our findings, using both isolated islets from calbindin-D(28k) KO mice and beta cell lines, establish a role for calbindin in the modulation of depolarization-stimulated insulin release and suggest that calbindin can control the rate of insulin release via regulation of [Ca(2+)](i).

  11. Apoptosis during intramembranous ossification

    PubMed Central

    Palumbo, Carla; Ferretti, Marzia; De Pol, Anto

    2003-01-01

    This paper concerns the role of apoptosis during the onset of bone histogenesis. Previous investigations by us performed on intramembranous ossification revealed the existence of two types of osteogenesis: static (SBF) and dynamic bone formation (DBF). During SBF, the first to occur, stationary osteoblasts transform into osteocytes in the same location where they differentiated, forming the primary spongiosa. DBF takes place later, when movable osteoblastic laminae differentiate along the surface of the primary trabeculae. The main distinctive feature between SBF and DBF is that the latter involves the invasion of pre-existing adjacent tissue, whereas the former does not. To ascertain whether programmed cell death during the invasive DBF process determines the fate of surrounding pre-existing mesenchyme differently from that occurring during the non-invasive SBF process, we studied apoptosis in ossification centres of tibial diaphysis in chick embryos and newborn rabbits with TUNEL and TEM. It emerged that, in both SBF and DBF, apoptosis affects mesenchymal cells located between the forming trabeculae and capillaries. However, apoptotic cells were observed more frequently during DBF than during SBF. This suggests that, during bone histogenesis, apoptosis, which is mostly associated with the invasive process of DBF, is probably dedicated to making space for advancing bone growth. PMID:14686694

  12. Pattern of rise in subplasma membrane Ca{sup 2+} concentration determines type of fusing insulin granules in pancreatic {beta} cells

    SciTech Connect

    Ohara-Imaizumi, Mica; Aoyagi, Kyota; Nakamichi, Yoko; Nishiwaki, Chiyono; Sakurai, Takashi; Nagamatsu, Shinya

    2009-07-31

    We simultaneously analyzed insulin granule fusion with insulin fused to green fluorescent protein and the subplasma membrane Ca{sup 2+} concentration ([Ca{sup 2+}]{sub PM}) with the Ca{sup 2+} indicator Fura Red in rat {beta} cells by dual-color total internal reflection fluorescence microscopy. We found that rapid and marked elevation in [Ca{sup 2+}]{sub PM} caused insulin granule fusion mostly from previously docked granules during the high KCl-evoked release and high glucose-evoked first phase release. In contrast, the slow and sustained elevation in [Ca{sup 2+}]{sub PM} induced fusion from newcomers translocated from the internal pool during the low KCl-evoked release and glucose-evoked second phase release. These data suggest that the pattern of the [Ca{sup 2+}]{sub PM} rise directly determines the types of fusing granules.

  13. Long-term pancreatic beta cell exposure to high levels of glucose but not palmitate induces DNA methylation within the insulin gene promoter and represses transcriptional activity.

    PubMed

    Ishikawa, Kota; Tsunekawa, Shin; Ikeniwa, Makoto; Izumoto, Takako; Iida, Atsushi; Ogata, Hidetada; Uenishi, Eita; Seino, Yusuke; Ozaki, Nobuaki; Sugimura, Yoshihisa; Hamada, Yoji; Kuroda, Akio; Shinjo, Keiko; Kondo, Yutaka; Oiso, Yutaka

    2015-01-01

    Recent studies have implicated epigenetics in the pathophysiology of diabetes. Furthermore, DNA methylation, which irreversibly deactivates gene transcription, of the insulin promoter, particularly the cAMP response element, is increased in diabetes patients. However, the underlying mechanism remains unclear. We aimed to investigate insulin promoter DNA methylation in an over-nutrition state. INS-1 cells, the rat pancreatic beta cell line, were cultured under normal-culture-glucose (11.2 mmol/l) or experimental-high-glucose (22.4 mmol/l) conditions for 14 days, with or without 0.4 mmol/l palmitate. DNA methylation of the rat insulin 1 gene (Ins1) promoter was investigated using bisulfite sequencing and pyrosequencing analysis. Experimental-high-glucose conditions significantly suppressed insulin mRNA and increased DNA methylation at all five CpG sites within the Ins1 promoter, including the cAMP response element, in a time-dependent and glucose concentration-dependent manner. DNA methylation under experimental-high-glucose conditions was unique to the Ins1 promoter; however, palmitate did not affect DNA methylation. Artificial methylation of Ins1 promoter significantly suppressed promoter-driven luciferase activity, and a DNA methylation inhibitor significantly improved insulin mRNA suppression by experimental-high-glucose conditions. Experimental-high-glucose conditions significantly increased DNA methyltransferase activity and decreased ten-eleven-translocation methylcytosine dioxygenase activity. Oxidative stress and endoplasmic reticulum stress did not affect DNA methylation of the Ins1 promoter. High glucose but not palmitate increased ectopic triacylglycerol accumulation parallel to DNA methylation. Metformin upregulated insulin gene expression and suppressed DNA methylation and ectopic triacylglycerol accumulation. Finally, DNA methylation of the Ins1 promoter increased in isolated islets from Zucker diabetic fatty rats. This study helps to clarify the

  14. Ryanodine receptor type I and nicotinic acid adenine dinucleotide phosphate receptors mediate Ca2+ release from insulin-containing vesicles in living pancreatic beta-cells (MIN6).

    PubMed

    Mitchell, Kathryn J; Lai, F Anthony; Rutter, Guy A

    2003-03-28

    We have demonstrated recently (Mitchell, K. J., Pinton, P., Varadi, A., Tacchetti, C., Ainscow, E. K., Pozzan, T., Rizzuto, R., and Rutter, G. A. (2001) J. Cell Biol. 155, 41-51) that ryanodine receptors (RyR) are present on insulin-containing secretory vesicles. Here we show that pancreatic islets and derived beta-cell lines express type I and II, but not type III, RyRs. Purified by subcellular fractionation and membrane immuno-isolation, dense core secretory vesicles were found to possess a similar level of type I RyR immunoreactivity as Golgi/endoplasmic reticulum (ER) membranes but substantially less RyR II than the latter. Monitored in cells expressing appropriately targeted aequorins, dantrolene, an inhibitor of RyR I channels, elevated free Ca(2+) concentrations in the secretory vesicle compartment from 40.1 +/- 6.7 to 90.4 +/- 14.8 microm (n = 4, p < 0.01), while having no effect on ER Ca(2+) concentrations. Furthermore, nicotinic acid adenine dinucleotide phosphate (NAADP), a novel Ca(2+)-mobilizing agent, decreased dense core secretory vesicle but not ER free Ca(2+) concentrations in permeabilized MIN6 beta-cells, and flash photolysis of caged NAADP released Ca(2+) from a thapsigargin-insensitive Ca(2+) store in single MIN6 cells. Because dantrolene strongly inhibited glucose-stimulated insulin secretion (from 3.07 +/- 0.51-fold stimulation to no significant glucose effect; n = 3, p < 0.01), we conclude that RyR I-mediated Ca(2+)-induced Ca(2+) release from secretory vesicles, possibly potentiated by NAADP, is essential for the activation of insulin secretion.

  15. Relationship of beta-cell function and autoantibodies to progression and nonprogression of subclinical type 1 diabetes: follow-up of the Seattle Family Study.

    PubMed

    Greenbaum, C J; Sears, K L; Kahn, S E; Palmer, J P

    1999-01-01

    A total of 85 islet cell antibody (ICA)+ or insulin autoantibody (IAA)+ relatives of patients with type 1 diabetes have been followed as part of the Seattle Family Study for a mean of 2.8 years. Of the subjects followed, 10 developed diabetes during this time period. The presence of GAD antibodies was strongly associated with the development of diabetes. In contrast, the presence of IAAs did not influence the risk of diabetes among ICA+ GAD+ subjects. When either the initial absolute acute insulin response to glucose (AIRg) or the AIR percentile, which accounts for the individual's insulin sensitivity, was below the 10th percentile of normal subjects, the risk of diabetes approached 50% at 5 years. However, impaired beta-cell function did not influence the risk of diabetes among those who were GAD+. There were 13 subjects with low AIRg and 13 subjects with two or more antibodies who had not progressed to diabetes during the course of the study. Other measurements of beta-cell function or demographic characteristics were not different in this group of nonprogressors compared with those with low AIRg who did not progress to diabetes. We conclude that ICA+ relatives with GAD antibodies or low AIRg have a high risk for development of diabetes, but among ICA+ GAD+ relatives, the addition of IAA or a single determination of AIRg does not enhance the prediction of diabetes. We suggest that prediction of diabetes risk depends on both the type and the number of antibodies present. In addition, there are a group of ICA+ relatives with low AIRg and/or multiple antibodies who have not progressed to diabetes over the course of the study.

  16. Arsenite reduces insulin secretion in rat pancreatic {beta}-cells by decreasing the calcium-dependent calpain-10 proteolysis of SNAP-25

    SciTech Connect

    Diaz-Villasenor, Andrea; Burns, Anna L.; Salazar, Ana Maria; Sordo, Monserrat; Hiriart, Marcia; Cebrian, Mariano E.; Ostrosky-Wegman, Patricia

    2008-09-15

    An increase in the prevalence of type 2 diabetes has been consistently observed among residents of high arsenic exposure areas. We have previously shown that in rat pancreatic {beta}-cells, low arsenite doses impair the secretion of insulin without altering its synthesis. To further study the mechanism by which arsenite reduces insulin secretion, we evaluated the effects of arsenite on the calcium-calpain pathway that triggers insulin exocytosis in RINm5F cells. Cell cycle and proliferation analysis were also performed to complement the characterization. Free [Ca{sup 2+}]i oscillations needed for glucose-stimulated insulin secretion were abated in the presence of subchronic low arsenite doses (0.5-2 {mu}M). The global activity of calpains increased with 2 {mu}M arsenite. However, during the secretion of insulin stimulated with glucose (15.6 mM), 1 {mu}M arsenite decreased the activity of calpain-10, measured as SNAP-25 proteolysis. Both proteins are needed to fuse insulin granules with the membrane to produce insulin exocytosis. Arsenite also induced a slowdown in the {beta} cell line proliferation in a dose-dependent manner, reflected by a reduction of dividing cells and in their arrest in G2/M. Data obtained showed that one of the mechanisms by which arsenite impairs insulin secretion is by decreasing the oscillations of free [Ca{sup 2+}]i, thus reducing calcium-dependent calpain-10 partial proteolysis of SNAP-25. The effects in cell division and proliferation observed with arsenite exposure can be an indirect consequence of the decrease in insulin secretion.

  17. Unprotected daily sun exposure is differently associated with central adiposity and beta-cell dysfunction by gender: The Korean national health and nutrition examination survey (KNHANES) V

    SciTech Connect

    Ohn, Jung Hun; Kwon, In Ho; Park, Juri; Ryu, Ohk Hyun; Lee, Seong Jin; Kim, Doo-Man; Ihm, Sung-Hee; Choi, Moon-Gi; Yoo, Hyung Joon; Hong, Eun-Gyoung

    2014-08-15

    Background: Ultraviolet irradiation by sun exposure has been associated with both harms and benefits to metabolic health. Objective: The objective of this study was to determine whether unprotected daily sun exposure is associated with the prevalence of diabetes and explore the underlying mechanism. Methods: We analyzed the Korean National Health and Nutrition Survey V from 2010 to 2011. Participants 19–60 years of age were asked about the average amount of time they had been exposed to direct sunlight per day since the age of 19. We categorized participants into three groups with different levels of lifetime daily sun exposure and explored the association of sun exposure with the prevalence of diabetes. Results: The risk of diabetes was higher in subjects with more than 5 h of unprotected sun exposure per day, with an odds ratio of 2.39 (95% CI 1.75–3.25), compared to those with less than 2 h of sun exposure, and the association remained significant after adjusting for diabetes risk factors. Long-term sun exposure was associated with increased central obesity and the possibility of an increase in visceral adiposity, especially among women, and with decrease in beta cell function and peripheral adiposity or percent body fat in men. Conclusions: Our study provides a cutoff for upper limit of sun exposure and suggests unprotected daily sun exposure for more than 5 h should be avoided to prevent diabetes. Increased central adiposity and decreased beta cell function were observed in women and men, respectively, who had long-term unprotected daily sun exposure. - Highlights: • Sun exposure for more than 5 h per day is associated with diabetes risk. • Insulin resistance associated with visceral adiposity may play a role in women. • Insulin secretory defect may explain diabetes risk in men.

  18. Ca2+-dependent dephosphorylation of kinesin heavy chain on beta-granules in pancreatic beta-cells. Implications for regulated beta-granule transport and insulin exocytosis

    NASA Technical Reports Server (NTRS)

    Donelan, Matthew J.; Morfini, Gerardo; Julyan, Richard; Sommers, Scott; Hays, Lori; Kajio, Hiroshi; Briaud, Isabelle; Easom, Richard A.; Molkentin, Jeffery D.; Brady, Scott T.; Rhodes, Christopher J.

    2002-01-01

    The specific biochemical steps required for glucose-regulated insulin exocytosis from beta-cells are not well defined. Elevation of glucose leads to increases in cytosolic [Ca2+]i and biphasic release of insulin from both a readily releasable and a storage pool of beta-granules. The effect of elevated [Ca2+]i on phosphorylation of isolated beta-granule membrane proteins was evaluated, and the phosphorylation of four proteins was found to be altered by [Ca2+]i. One (a 18/20-kDa doublet) was a Ca2+-dependent increase in phosphorylation, and, surprisingly, three others (138, 42, and 36 kDa) were Ca2+-dependent dephosphorylations. The 138-kDa beta-granule phosphoprotein was found to be kinesin heavy chain (KHC). At low levels of [Ca2+]i KHC was phosphorylated by casein kinase 2, but KHC was rapidly dephosphorylated by protein phosphatase 2B beta (PP2Bbeta) as [Ca2+]i increased. Inhibitors of PP2B specifically reduced the second, microtubule-dependent, phase of insulin secretion, suggesting that dephosphorylation of KHC was required for transport of beta-granules from the storage pool to replenish the readily releasable pool of beta-granules. This is distinct from synaptic vesicle exocytosis, because neurotransmitter release from synaptosomes did not require a Ca2+-dependent KHC dephosphorylation. These results suggest a novel mechanism for regulating KHC function and beta-granule transport in beta-cells that is mediated by casein kinase 2 and PP2B. They also implicate a novel regulatory role for PP2B/calcineurin in the control of insulin secretion downstream of a rise in [Ca2+]i.

  19. Cooperation between HMGA1, PDX-1, and MafA is Essential for Glucose-Induced Insulin Transcription in Pancreatic Beta Cells.

    PubMed

    Arcidiacono, Biagio; Iiritano, Stefania; Chiefari, Eusebio; Brunetti, Francesco S; Gu, Guoqiang; Foti, Daniela Patrizia; Brunetti, Antonio

    2014-01-01

    The high-mobility group AT-hook 1 (HMGA1) protein is a nuclear architectural factor that can organize chromatin structures. It regulates gene expression by controlling the formation of stereospecific multiprotein complexes called "enhanceosomes" on the AT-rich regions of target gene promoters. Previously, we reported that defects in HMGA1 caused decreased insulin receptor expression and increased susceptibility to type 2 diabetes mellitus in humans and mice. Interestingly, mice with disrupted HMGA1 gene had significantly smaller islets and decreased insulin content in their pancreata, suggesting that HMGA1 may have a direct role in insulin transcription and secretion. Herein, we investigate the regulatory roles of HMGA1 in insulin transcription. We provide evidence that HMGA1 physically interacts with PDX-1 and MafA, two critical transcription factors for insulin gene expression and beta-cell function, both in vitro and in vivo. We then show that the overexpression of HMGA1 significantly improves the transactivating activity of PDX-1 and MafA on human and mouse insulin promoters, while HMGA1 knockdown considerably decreased this transactivating activity. Lastly, we demonstrate that high glucose stimulus significantly increases the binding of HMGA1 to the insulin (INS) gene promoter, suggesting that HMGA1 may act as a glucose-sensitive element controlling the transcription of the INS gene. Together, our findings provide evidence that HMGA1, by regulating PDX-1- and MafA-induced transactivation of the INS gene promoter, plays a critical role in pancreatic beta-cell function and insulin production.

  20. Correlations between beta-cells' calcium dynamics reveal differences in functional connectivity patterns in islets of Langerhans from pancreas tissue slices under low and high levels of glucose

    NASA Astrophysics Data System (ADS)

    Stožer, Andraž; Gosak, Marko; Korošak, Dean; Yakubo, Kousuke; Dolenšek, Jure; Marhl, Marko; Rupnik, Marjan Slak

    2012-08-01

    In the last decade, approach developed in the frame of complex network theory has presented very successful and popular tools for studying the structure and functioning of complex systems. A particularly attractive avenue in this context is the analysis of biological systems, since structural principles of complex networks have been identified at all scales of functioning of living organisms. In the present paper, we propose the construction of a complex network representation of a pancreatic islet. In this compact microorgan, under physiological conditions the release of the single most important anabolic hormone insulin is robustly controlled by an efficient cell-to-cell communication mediated by gap junctions. Here, we extract networks of insulin releasing beta-cells from experimentally measured time series data on calcium dynamics and from positional information obtained by confocal laser-scanning functional imaging of islets in acute pancreatic tissue slices. In particular, connectivity patterns are determined on the basis of correlations between calcium dynamics in the islet. The extracted networks are then scrutinized with conventional tools for network analysis, whereby particular importance is devoted to comparison of the network structure under low and high glucose levels, i.e. physiologically resting and stimulating conditions, respectively. We show that the cellular dynamics is more correlated under stimulation and that the networks obtained in both regimes display a different organization. The range of interactions among beta cells is significantly shorter in the case of a higher stimulation. Our results thus provide novel insights into the relationship between network topology and functional organization of pancreatic islets.

  1. Geniposide promotes beta-cell regeneration and survival through regulating β-catenin/TCF7L2 pathway

    PubMed Central

    Yao, D D; Yang, L; Wang, Y; Liu, C; Wei, Y J; Jia, X B; Yin, W; Shu, L

    2015-01-01

    T-cell factor 7-like 2 (TCF7L2) is an important transcription factor of Wnt/β-catenin signaling, which has critical roles in β-cell survival and regeneration. In preliminary screening assay, we found geniposide, a naturally occurring compound, was able to increase TCF7L2 mRNA level in Min6 cells. Here we aimed to investigate the role of geniposide in β-cell and underlying mechanism involved. Geniposide was found to promote β-cell survival by increasing β-cell proliferation and decreasing β-cell apoptosis in cultured mouse islets after challenge with diabetic stimuli. Geniposide protected β-cell through activating Wnt signaling, enhanced expressions of TCF7L2 and GLP-1R, activated AKT, inhibited GSK3β activity, and promoted β-catenin nuclear translocation. The protective effect of geniposide was remarkably suppressed by siRNAs against β-catenin, or by ICG001 (β-catenin/TCF-mediated transcription inhibitor). Moreover, geniposide promoted β-cell regeneration in vivo to normalize blood glucose in high-fat diet and db/db mice. Increased β-cell proliferation was observed in pancreatic sections of geniposide-treated diabetic mice. Most importantly, geniposide triggered small islet-like cell clusters formation as a result of β-cell neogenesis from ductal epithelium, which was well correlated with the increase in TCF7L2 expression. In exocrine cells isolated from mouse pancreas, geniposide could induce duct cell differentiation through upregulating TCF7L2 expression and activating JAK2/STAT3 pathway. Taken together, we identified a novel role of geniposide in promoting β-cell survival and regeneration by mechanisms involving the activation of β-catenin/TCF7L2 signaling. Our finding highlights the potential value of geniposide as a possible treatment for type 2 diabetes. PMID:25950476

  2. Cell death and impairment of glucose-stimulated insulin secretion induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the {beta}-cell line INS-1E

    SciTech Connect

    Piaggi, Simona; Novelli, Michela; Martino, Luisa; Masini, Matilde; Raggi, Chiara; Orciuolo, Enrico; Masiello, Pellegrino; Casini, Alessandro; De Tata, Vincenzo . E-mail: v.detata@ipg.med.unipi.it

    2007-05-01

    The aim of this research was to characterize 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxicity on the insulin-secreting {beta}-cell line INS-1E. A sharp decline of cell survival (below 20%) was observed after 1 h exposure to TCDD concentrations between 12.5 and 25 nM. Ultrastructurally, {beta}-cell death was characterized by extensive degranulation, appearance of autophagic vacuoles, and peripheral nuclear condensation. Cytotoxic concentrations of TCDD rapidly induced a dose-dependent increase in intracellular calcium concentration. Blocking calcium entry by EGTA significantly decreased TCDD cytotoxicity. TCDD was also able to rapidly induce mitochondrial depolarization. Interestingly, 1 h exposition of INS-1E cells to very low TCDD concentrations (0.05-1 nM) dramatically impaired glucose-stimulated but not KCl-stimulated insulin secretion. In conclusion, our results clearly show that TCDD exerts a direct {beta}-cell cytotoxic effect at concentrations of 15-25 nM, but also markedly impairs glucose-stimulated insulin secretion at concentrations 20 times lower than these. On the basis of this latter observation we suggest that pancreatic {beta}-cells could be considered a specific and sensitive target for dioxin toxicity.

  3. TRAIL upregulates decoy receptor 1 and mediates resistance to apoptosis in insulin-secreting INS-1 cells.

    PubMed

    Kang, Soojeong; Park, So-Young; Lee, Hye-Jeong; Yoo, Young Hyun

    2010-06-01

    TRAIL/Apo2L (tumor necrosis factor-related apoptosis-inducing ligand) is a multifunctional protein regulating the homeostasis of the immune system, infection, autoimmune diseases, and apoptosis. In particular, the potential role of TRAIL in type 1 diabetes (T1D) has been studied by several research groups. A previous study found that TRAIL did not have significant cytotoxic effects on the insulin-secreting pancreatic beta cell line, INS-1. However, the mechanism was not clear. Here we demonstrate that INS-1 cells are resistant to TRAIL-induced apoptosis and show alteration in the expression of death and decoy receptors upon TRAIL treatment. To compare TRAIL-resistant INS-1 cells with TRAIL-sensitive cells, we utilized U87MG cells, which are known to be TRAIL-sensitive. TRAIL treatment showed NF-kappaB translocation to the nucleus in TRAIL-resistant INS-1 cells, and TRAIL-induced NF-kappaB activation was preceded by IkappaBalpha degradation. A pharmacological inhibitor of NF-kappaB, Bay 11-7082, blocked TRAIL-induced NF-kappaB translocation to the nucleus and IkappaBalpha degradation. Four related receptors bind TRAIL: two death receptors (DR4 and DR5) that promote apoptosis, and two decoy receptors (DcR1 and DcR2) that act as dominant-negative inhibitors of TRAIL-mediated apoptosis. In the present study, TRAIL treatment in INS-1 cells upregulated DcR1 and downregulated DR5 without altering the expression of DcR2 and DR4. The resistance to apoptosis in INS-1 cells might therefore, be a consequence of DcR1 upregulation and DR5 downregulation, and the transcription factor, NF-kappaB, could regulate the sensitivity of cells to TRAIL by controlling the ratio of decoy to death receptors. Thus, TRAIL may play an important role in the survival of pancreatic beta cells by regulating receptor expression in an NF-kappaB-dependent manner.

  4. Mortalin, Apoptosis, and Neurodegeneration

    PubMed Central

    Londono, Carolina; Osorio, Cristina; Gama, Vivian; Alzate, Oscar

    2012-01-01

    Mortalin is a highly conserved heat-shock chaperone usually found in multiple subcellular locations. It has several binding partners and has been implicated in various functions ranging from stress response, control of cell proliferation, and inhibition/prevention of apoptosis. The activity of this protein involves different structural and functional mechanisms, and minor alterations in its expression level may lead to serious biological consequences, including neurodegeneration. In this article we review the most current data associated with mortalin’s binding partners and how these protein-protein interactions may be implicated in apoptosis and neurodegeneration. A complete understanding of the molecular pathways in which mortalin is involved is important for the development of therapeutic strategies for cancer and neurodegenerative diseases. PMID:24970131

  5. Pathophysiological Significance of Hepatic Apoptosis

    PubMed Central

    Wang, Kewei; Lin, Bingliang

    2013-01-01

    Apoptosis is a classical pathological feature in liver diseases caused by various etiological factors such as drugs, viruses, alcohol, and cholestasis. Hepatic apoptosis and its deleterious effects exacerbate liver function as well as involvement in fibrosis/cirrhosis and carcinogenesis. An imbalance between apoptotic and antiapoptotic capabilities is a prominent characteristic of liver injury. The regulation of apoptosis and antiapoptosis can be a pivotal step in the treatment of liver diseases. PMID:27335822

  6. Apoptosis pathways and neuroblastoma therapy.

    PubMed

    Fulda, S

    2009-01-01

    Evasion of apoptosis, the cell's intrinsic death program, is a hallmark of human cancers including neuroblastoma. Also, failure to undergo apoptosis may cause treatment resistance, since the cytotoxic activity of anticancer therapies commonly used in the clinic, e.g. chemotherapy, gamma-irradiation or immunotherapy, is predominantly mediated by triggering apoptosis in tumor cells. Therefore, a better understanding of the signaling pathways and molecules that govern apoptosis in neuroblastoma cells is expected to open new avenues for the design of molecular targeted therapies for neuroblastoma.

  7. Role of Apoptosis in disease

    PubMed Central

    Favaloro, B.; Allocati, N.; Graziano, V.; Di Ilio, C.; De Laurenzi, V.

    2012-01-01

    Since the initial description of apoptosis, a number of different forms of cell death have been described. In this review we will focus on classic caspase-dependent apoptosis and its variations that contribute to diseases. Over fifty years of research have clarified molecular mechanisms involved in apoptotic signaling as well and shown that alterations of these pathways lead to human diseases. Indeed both reduced and increased apoptosis can result in pathology. More recently these findings have led to the development of therapeutic approaches based on regulation of apoptosis, some of which are in clinical trials or have entered medical practice. PMID:22683550

  8. Microcephaly with Simplified Gyration, Epilepsy, and Infantile Diabetes Linked to Inappropriate Apoptosis of Neural Progenitors

    PubMed Central

    Poulton, Cathryn J.; Schot, Rachel; Kia, Sima Kheradmand; Jones, Marta; Verheijen, Frans W.; Venselaar, Hanka; de Wit, Marie-Claire Y.; de Graaff, Esther; Bertoli-Avella, Aida M.; Mancini, Grazia M.S.

    2011-01-01

    We describe a syndrome of primary microcephaly with simplified gyral pattern in combination with severe infantile epileptic encephalopathy and early-onset permanent diabetes in two unrelated consanguineous families with at least three affected children. Linkage analysis revealed a region on chromosome 18 with a significant LOD score of 4.3. In this area, two homozygous nonconserved missense mutations in immediate early response 3 interacting protein 1 (IER3IP1) were found in patients from both families. IER3IP1 is highly expressed in the fetal brain cortex and fetal pancreas and is thought to be involved in endoplasmic reticulum stress response. We reported one of these families previously in a paper on Wolcott-Rallison syndrome (WRS). WRS is characterized by increased apoptotic cell death as part of an uncontrolled unfolded protein response. Increased apoptosis has been shown to be a cause of microcephaly in animal models. An autopsy specimen from one patient showed increased apoptosis in the cerebral cortex and pancreas beta cells, implicating premature cell death as the pathogenetic mechanism. Both patient fibroblasts and control fibroblasts treated with siRNA specific for IER3IP1 showed an increased susceptibility to apoptotic cell death under stress conditions in comparison to controls. This directly implicates IER3IP1 in the regulation of cell survival. Identification of IER3IP1 mutations sheds light on the mechanisms of brain development and on the pathogenesis of infantile epilepsy and early-onset permanent diabetes. PMID:21835305

  9. Aqueous leaf extract of Passiflora alata Curtis promotes antioxidant and anti-inflammatory effects and consequently preservation of NOD mice beta cells (non-obese diabetic).

    PubMed

    Figueiredo, D; Colomeu, Talita Cristina; Schumacher, Nayara Simon Gonzalez; Stivanin-Silva, L G; Cazarin, Cinthia Baú Betim; Meletti, Laura Maria Molina; Fernandes, Luís Gustavo Romani; Prado, Marcelo Alexandre; Zollner, R L

    2016-06-01

    Passiflora alata Curtis (P. alata) leaves have anti-inflammatory properties; the present study aimed to investigate the anti-diabetogenic properties of P. alata aqueous leaf extract. HPLC analysis identified the phenolic compounds catechin, epicatechin and rutin. The aqueous extract was administered for 30weeks to non-obese diabetic (NOD) mice presenting a decrease of 28.6% in diabetes incidence and the number of inflammatory cells in pancreatic islets, when compared with the control group (water). The P. alata group presented an antioxidant effect and decreased lipid peroxidation in the serum of NOD mice. Increased numbers of insulin-positive cells were also observed in the pancreatic islets of the treated group. The diabetic group exhibited higher levels in the glucose tolerance test and glycemic index, in comparison to the P. alata-treated group and non-diabetic control BALB/c mice. In addition, the P. alata extract reduced the percentage and the proliferation index of NOD mice lymphocytes submitted to in vitro dose/response mitogenic stimulation assays. These results suggest that the aqueous extract of P. alata has anti-inflammatory properties, contributing to the protection of beta cells in pancreatic islets in NOD mice, and presents potential for use a supporting approach to treat type 1 diabetes.

  10. A new approach for pancreatic tissue engineering: human endometrial stem cells encapsulated in fibrin gel can differentiate to pancreatic islet beta-cell.

    PubMed

    Niknamasl, Azadeh; Ostad, Seyed Nasser; Soleimani, Mansoureh; Azami, Mahmoud; Salmani, Maryam Kabir; Lotfibakhshaiesh, Nasrin; Ebrahimi-Barough, Somayeh; Karimi, Roya; Roozafzoon, Reza; Ai, Jafar

    2014-10-01

    Metabolic diabetes mellitus as the most serious and prevalent metabolic disease in the world has various complications. The most effective treatment of type I diabetes seems to be islet cell transplantation. Shortage of donors and difficult procedures and high rate of rejection have always restricted this approach. Tissue engineering is a novel effective solution to many medical problems such as diabetes. Endometrial mesenchymal stem cells as a lineage which have the potential to differentiate to mesodermal and endodermal tissues seem to be suitable for this purpose. Fibrin hydrogel with a high degree of biocompatibility and specific properties making it similar to normal pancreas seems to be an ideal scaffold. After successfully isolating stem cells (hEnSCs) from human endometrium, a three-step protocol was used to differentiate them into pancreatic beta cells. Fibrin was used as 3D scaffold. After 2 weeks, cells formed clusters like islets cells, and secretion of insulin was measured by chemiluminescence. PDX1, proinsulin, and c-peptide as special markers of β cells were detected by immunofluorescence. Expression of glucagon, PDX1, and insulin genes in mRNA level was detected by Real time PCR and gel electrophoresis. The former showed higher levels of gene expression in 3D cultures. SEM analysis showed good integrity between cells and scaffold. No toxicity was detected with fibrin scaffold by MTT assay.

  11. Aqueous leaf extract of Passiflora alata Curtis promotes antioxidant and anti-inflammatory effects and consequently preservation of NOD mice beta cells (non-obese diabetic).

    PubMed

    Figueiredo, D; Colomeu, Talita Cristina; Schumacher, Nayara Simon Gonzalez; Stivanin-Silva, L G; Cazarin, Cinthia Baú Betim; Meletti, Laura Maria Molina; Fernandes, Luís Gustavo Romani; Prado, Marcelo Alexandre; Zollner, R L

    2016-06-01

    Passiflora alata Curtis (P. alata) leaves have anti-inflammatory properties; the present study aimed to investigate the anti-diabetogenic properties of P. alata aqueous leaf extract. HPLC analysis identified the phenolic compounds catechin, epicatechin and rutin. The aqueous extract was administered for 30weeks to non-obese diabetic (NOD) mice presenting a decrease of 28.6% in diabetes incidence and the number of inflammatory cells in pancreatic islets, when compared with the control group (water). The P. alata group presented an antioxidant effect and decreased lipid peroxidation in the serum of NOD mice. Increased numbers of insulin-positive cells were also observed in the pancreatic islets of the treated group. The diabetic group exhibited higher levels in the glucose tolerance test and glycemic index, in comparison to the P. alata-treated group and non-diabetic control BALB/c mice. In addition, the P. alata extract reduced the percentage and the proliferation index of NOD mice lymphocytes submitted to in vitro dose/response mitogenic stimulation assays. These results suggest that the aqueous extract of P. alata has anti-inflammatory properties, contributing to the protection of beta cells in pancreatic islets in NOD mice, and presents potential for use a supporting approach to treat type 1 diabetes. PMID:27039211

  12. Long-term glycemic control using polymer-encapsulated human stem cell-derived beta cells in immune-competent mice.

    PubMed

    Vegas, Arturo J; Veiseh, Omid; Gürtler, Mads; Millman, Jeffrey R; Pagliuca, Felicia W; Bader, Andrew R; Doloff, Joshua C; Li, Jie; Chen, Michael; Olejnik, Karsten; Tam, Hok Hei; Jhunjhunwala, Siddharth; Langan, Erin; Aresta-Dasilva, Stephanie; Gandham, Srujan; McGarrigle, James J; Bochenek, Matthew A; Hollister-Lock, Jennifer; Oberholzer, Jose; Greiner, Dale L; Weir, Gordon C; Melton, Douglas A; Langer, Robert; Anderson, Daniel G

    2016-03-01

    The transplantation of glucose-responsive, insulin-producing cells offers the potential for restoring glycemic control in individuals with diabetes. Pancreas transplantation and the infusion of cadaveric islets are currently implemented clinically, but these approaches are limited by the adverse effects of immunosuppressive therapy over the lifetime of the recipient and the limited supply of donor tissue. The latter concern may be addressed by recently described glucose-responsive mature beta cells that are derived from human embryonic stem cells (referred to as SC-β cells), which may represent an unlimited source of human cells for pancreas replacement therapy. Strategies to address the immunosuppression concerns include immunoisolation of insulin-producing cells with porous biomaterials that function as an immune barrier. However, clinical implementation has been challenging because of host immune responses to the implant materials. Here we report the first long-term glycemic correction of a diabetic, immunocompetent animal model using human SC-β cells. SC-β cells were encapsulated with alginate derivatives capable of mitigating foreign-body responses in vivo and implanted into the intraperitoneal space of C57BL/6J mice treated with streptozotocin, which is an animal model for chemically induced type 1 diabetes. These implants induced glycemic correction without any immunosuppression until their removal at 174 d after implantation. Human C-peptide concentrations and in vivo glucose responsiveness demonstrated therapeutically relevant glycemic control. Implants retrieved after 174 d contained viable insulin-producing cells.

  13. Low dose monoethyl phthalate (MEP) exposure triggers proliferation by activating PDX-1 at 1.1B4 human pancreatic beta cells.

    PubMed

    Güven, Celal; Dal, Fulya; Aydoğan Ahbab, Müfide; Taskin, Eylem; Ahbab, Süleyman; Adin Çinar, Suzan; Sırma Ekmekçi, Sema; Güleç, Çağrı; Abacı, Neslihan; Akçakaya, Handan

    2016-07-01

    Phthalate plasticizers used in a wide range of common plastic products are released into the environment and may pose a risk of increased incidence of type 2 diabetes. In this work, we studied the effects of monoethyl phthalate (MEP), the metabolite of diethyl phthalate, exposure on 1.1B4 human pancreatic beta cells at low doses (1-1000 nM). We showed that MEP treatment induced proliferation in 1.1B4 cells. Also PCNA protein expression levels were increased related to proliferation induction. It has been noted that phthalates can exert estrogen mediated response by interacting with ER. In our study 24 h MEP treatment decreased ERα protein expression level conversely it increased the same protein expression level after 72 h treatment. Also MEP treatment decreased ERβ expression after 72 h at 1.1B4 cells. Our results further show that insulin content of 1.1B4 cells were increased with low dose MEP treatment. Along with our insulin content results, PDX- 1 expression levels were also increased at 1.1B4 cells with MEP treatment. These findings suggest that MEP acts as an estrogenic compound and PPARγ agonist at lower concentrations. Also it should be noted that PDX-1 may be a critical regulator of 1.1B4 cells treated with MEP.

  14. Regulatory roles for Tiam1, a guanine nucleotide exchange factor for Rac1, in glucose-stimulated insulin secretion in pancreatic beta-cells.

    PubMed

    Veluthakal, Rajakrishnan; Madathilparambil, Suresh Vasu; McDonald, Phillip; Olson, Lawrence Karl; Kowluru, Anjaneyulu

    2009-01-01

    Using various biochemical, pharmacological and molecular biological approaches, we have recently reported regulatory roles for Rac1, a small G-protein, in glucose-stimulated insulin secretion (GSIS). However, little is understood with respect to localization of, and regulation by, specific regulatory factors of Rac1 in GSIS. Herein, we investigated regulatory roles for Tiam1, a specific nucleotide exchange factor (GEF) for Rac1, in GSIS in pancreatic beta-cells. Western blot analysis indicated that Tiam1 is predominantly cytosolic in distribution. NSC23766, a specific inhibitor of Tiam1-mediated activation of Rac1, markedly attenuated glucose-induced, but not KCl-induced insulin secretion in INS 832/13 cells and normal rat islets. Further, NSC23766 significantly reduced glucose-induced activation (i.e. GTP-bound form) and membrane association of Rac1 in INS 832/13 cells and rat islets. Moreover, siRNA-mediated knock-down of Tiam1 markedly inhibited glucose-induced membrane trafficking and activation of Rac1 in INS 832/13 cells. Interestingly, however, in contrast to the inhibitory effects of NSC23766, Tiam1 gene depletion potentiated GSIS in these cells; such a potentiation of GSIS was sensitive to extracellular calcium. Together, our studies present the first evidence for a regulatory role for Tiam1/Rac1-sensitive signaling step in GSIS. They also provide evidence for the existence of a potential Rac1/Tiam1-independent, but calcium-sensitive component for GSIS in these cells.

  15. Adipose Tissue-Derived Mesenchymal Stem Cells Exert In Vitro Immunomodulatory and Beta Cell Protective Functions in Streptozotocin-Induced Diabetic Mice Model

    PubMed Central

    Rahavi, Hossein; Hashemi, Seyed Mahmoud; Soleimani, Masoud; Mohammadi, Jamal; Tajik, Nader

    2015-01-01

    Regenerative and immunomodulatory properties of mesenchymal stem cells (MSCs) might be applied for type 1 diabetes mellitus (T1DM) treatment. Thus, we proposed in vitro assessment of adipose tissue-derived MSCs (AT-MSCs) immunomodulation on autoimmune response along with beta cell protection in streptozotocin- (STZ-) induced diabetic C57BL/6 mice model. MSCs were extracted from abdominal adipose tissue of normal mice and cultured to proliferate. Diabetic mice were prepared by administration of multiple low-doses of streptozotocin. Pancreatic islets were isolated from normal mice and splenocytes prepared from normal and diabetic mice. Proliferation, cytokine production, and insulin secretion assays were performed in coculture experiments. AT-MSCs inhibited splenocytes proliferative response to specific (islet lysate) and nonspecific (PHA) triggers in a dose-dependent manner (P < 0.05). Decreased production of proinflammatory cytokines, such as IFN-γ, IL-2, and IL-17, and increased secretion of regulatory cytokines such as TGF-β, IL-4, IL-10, and IL-13 by stimulated splenocytes were also shown in response to islet lysate or PHA stimulants (P < 0.05). Finally, we demonstrated that AT-MSCs could effectively sustain viability as well as insulin secretion potential of pancreatic islets in the presence of reactive splenocytes (P < 0.05). In conclusion, it seems that MSCs may provide a new horizon for T1DM cell therapy and islet transplantation in the future. PMID:25893202

  16. Apoptosis Evaluation by Electrochemical Techniques.

    PubMed

    Yin, Jian; Miao, Peng

    2016-03-01

    Apoptosis has close relevance to pathology, pharmacology, and toxicology. Accurate and convenient detection of apoptosis would be beneficial for biological study, clinical diagnosis, and drug development. Based on distinct features of apoptotic cells, a diversity of analytical techniques have been exploited for sensitive analysis of apoptosis, such as surface plasmon resonance, electrochemical methods, flow cytometry, and some imaging assays. Among them, the features of simplicity, easy operation, low cost, and high sensitivity make electrochemical techniques powerful tools to investigate electron-transfer processes of in vitro biological systems. In this contribution, a general overview of current knowledge on various technical approaches for apoptosis evaluation is provided. Furthermore, recently developed electrochemical biosensors for detecting apoptotic cells and their advantages over traditional methods are summarized. One of the main considerations focuses on designing the recognition elements based on various biochemical events during apoptosis.

  17. Apoptosis and the Airway Epithelium

    PubMed Central

    White, Steven R.

    2011-01-01

    The airway epithelium functions as a barrier and front line of host defense in the lung. Apoptosis or programmed cell death can be elicited in the epithelium as a response to viral infection, exposure to allergen or to environmental toxins, or to drugs. While apoptosis can be induced via activation of death receptors on the cell surface or by disruption of mitochondrial polarity, epithelial cells compared to inflammatory cells are more resistant to apoptotic stimuli. This paper focuses on the response of airway epithelium to apoptosis in the normal state, apoptosis as a potential regulator of the number and types of epithelial cells in the airway, and the contribution of epithelial cell apoptosis in important airways diseases. PMID:22203854

  18. Anesthesia and cerebral apoptosis.

    PubMed

    Brée, B; Gourdin, M; De Kock, M

    2008-01-01

    General anesthetics interact with targets at the cellular and molecular levels. They have the potential to induce changes in the body and the brain. Usually, these interactions are thought to be short lasting. In contrast, recent evidences suggest that alcohol, a toxic sharing many mechanisms with general anesthetics, induces long term effect at these levels. This is particularly evident in the period of synaptogenesis during which alcohol can induce excessive cerebral apoptosis (histopathologic changes) in juvenile animal models. Even if the vast majority of our patients seems to completely restore homeostasis after general anesthesia, we don't know if the changes induced at the brain level in animal models exist in human. This article intends to supply biological, pharmacological and experimental basis for a possible long term effect of general anesthetics on the human developing brain. PMID:19051443

  19. Antibody Response to Serpin B13 Induces Adaptive Changes in Mouse Pancreatic Islets and Slows Down the Decline in the Residual Beta Cell Function in Children with Recent Onset of Type 1 Diabetes Mellitus.

    PubMed

    Kryvalap, Yury; Lo, Chi-Wen; Manuylova, Ekaterina; Baldzizhar, Raman; Jospe, Nicholas; Czyzyk, Jan

    2016-01-01

    Type 1 diabetes mellitus (T1D) is characterized by a heightened antibody (Ab) response to pancreatic islet self-antigens, which is a biomarker of progressive islet pathology. We recently identified a novel antibody to clade B serpin that reduces islet-associated T cell accumulation and is linked to the delayed onset of T1D. As natural immunity to clade B arises early in life, we hypothesized that it may influence islet development during that time. To test this possibility healthy young Balb/c male mice were injected with serpin B13 mAb or IgG control and examined for the number and cellularity of pancreatic islets by immunofluorescence and FACS. Beta cell proliferation was assessed by measuring nucleotide analog 5-ethynyl-2'-deoxyuridine (5-EdU) incorporation into the DNA and islet Reg gene expression was measured by real time PCR. Human studies involved measuring anti-serpin B13 autoantibodies by Luminex. We found that injecting anti-serpin B13 monoclonal Ab enhanced beta cell proliferation and Reg gene expression, induced the generation of ∼80 pancreatic islets per animal, and ultimately led to increase in the beta cell mass. These findings are relevant to human T1D because our analysis of subjects just diagnosed with T1D revealed an association between baseline anti-serpin activity and slower residual beta cell function decline in the first year after the onset of diabetes. Our findings reveal a new role for the anti-serpin immunological response in promoting adaptive changes in the endocrine pancreas and suggests that enhancement of this response could potentially help impede the progression of T1D in humans.

  20. Positron emission tomography study on pancreatic somatostatin receptors in normal and diabetic rats with {sup 68}Ga-DOTA-octreotide: A potential PET tracer for beta cell mass measurement

    SciTech Connect

    Sako, Takeo; Hasegawa, Koki; Nishimura, Mie; Kanayama, Yousuke; Wada, Yasuhiro; Hayashinaka, Emi; Cui, Yilong; Kataoka, Yosky; Senda, Michio; Watanabe, Yasuyoshi

    2013-12-06

    Highlights: •PET images showed high uptake of {sup 68}Ga-DOTA-octreotide in the normal pancreas. •{sup 68}Ga-DOTA-octreotide specifically binds to somatostatin receptors in the pancreas. •The pancreatic uptake of {sup 68}Ga-DOTA-octreotide was decreased in the diabetic rats. •{sup 68}Ga-DOTA-octreotide could be a candidate PET probe to measure the beta cell mass. -- Abstract: Diabetes mellitus (DM) is a metabolic disorder characterized by hyperglycemia, and the loss or dysfunction of pancreatic beta cells has been reported before the appearance of clinical symptoms and hyperglycemia. To evaluate beta cell mass (BCM) for improving the detection and treatment of DM at earlier stages, we focused on somatostatin receptors that are highly expressed in the pancreatic beta cells, and developed a positron emission tomography (PET) probe derived from octreotide, a metabolically stable somatostatin analog. Octreotide was conjugated with 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), a chelating agent, and labeled with {sup 68}Gallium ({sup 68}Ga). After intravenous injection of {sup 68}Ga-DOTA-octreotide, a 90-min emission scan of the abdomen was performed in normal and DM model rats. The PET studies showed that {sup 68}Ga-DOTA-octreotide radioactivity was highly accumulated in the pancreas of normal rats and that the pancreatic accumulation was significantly reduced in the rats administered with an excess amount of unlabeled octreotide or after treatment with streptozotocin, which was used for the chemical induction of DM in rats. These results were in good agreement with the ex vivo biodistribution data. These results indicated that the pancreatic accumulation of {sup 68}Ga-DOTA-octreotide represented specific binding to the somatostatin receptors and reflected BCM. Therefore, PET imaging with {sup 68}Ga-DOTA-octreotide could be a potential tool for evaluating BCM.

  1. Antibody Response to Serpin B13 Induces Adaptive Changes in Mouse Pancreatic Islets and Slows Down the Decline in the Residual Beta Cell Function in Children with Recent Onset of Type 1 Diabetes Mellitus.

    PubMed

    Kryvalap, Yury; Lo, Chi-Wen; Manuylova, Ekaterina; Baldzizhar, Raman; Jospe, Nicholas; Czyzyk, Jan

    2016-01-01

    Type 1 diabetes mellitus (T1D) is characterized by a heightened antibody (Ab) response to pancreatic islet self-antigens, which is a biomarker of progressive islet pathology. We recently identified a novel antibody to clade B serpin that reduces islet-associated T cell accumulation and is linked to the delayed onset of T1D. As natural immunity to clade B arises early in life, we hypothesized that it may influence islet development during that time. To test this possibility healthy young Balb/c male mice were injected with serpin B13 mAb or IgG control and examined for the number and cellularity of pancreatic islets by immunofluorescence and FACS. Beta cell proliferation was assessed by measuring nucleotide analog 5-ethynyl-2'-deoxyuridine (5-EdU) incorporation into the DNA and islet Reg gene expression was measured by real time PCR. Human studies involved measuring anti-serpin B13 autoantibodies by Luminex. We found that injecting anti-serpin B13 monoclonal Ab enhanced beta cell proliferation and Reg gene expression, induced the generation of ∼80 pancreatic islets per animal, and ultimately led to increase in the beta cell mass. These findings are relevant to human T1D because our analysis of subjects just diagnosed with T1D revealed an association between baseline anti-serpin activity and slower residual beta cell function decline in the first year after the onset of diabetes. Our findings reveal a new role for the anti-serpin immunological response in promoting adaptive changes in the endocrine pancreas and suggests that enhancement of this response could potentially help impede the progression of T1D in humans. PMID:26578518

  2. 3,5,3'-Triiodo-L-thyronine enhances the differentiation of a human pancreatic duct cell line (hPANC-1) towards a beta-cell-Like phenotype.

    PubMed

    Misiti, Silvia; Anastasi, Emanuela; Sciacchitano, Salvatore; Verga Falzacappa, Cecilia; Panacchia, Laura; Bucci, Barbara; Khouri, Daniele; D'Acquarica, Ilaria; Brunetti, Ercole; Di Mario, Umberto; Toscano, Vincenzo; Perfetti, Riccardo

    2005-07-01

    The thyroid hormone, 3,5,3'-Triiodo-L-thyronine (T3), is essential for growth, differentiation, and regulation of metabolic functions in multicellular organisms, although the specific mechanisms of this control are still unknown. In this study, treatment of a human pancreatic duct cell line (hPANC-1) with T3 blocks cell growth by an increase of cells in G(0)/G(1) cell cycle phase and enhances morphological and functional changes as indicated by the marked increase in the synthesis of insulin and the parallel decrease of the ductal differentiation marker cytokeratin19. Expression analysis of some of the genes regulating pancreatic beta-cell differentiation revealed a time-dependent increase in insulin and glut2 mRNA levels in response to T3. As last step of the acquisition of a beta-cell-like phenotype, we present evidence that thyroid hormones are able to increase the release of insulin into the culture medium. In conclusion, our results suggest, for the first time, that thyroid hormones induce cell cycle perturbations and play an important role in the process of transdifferentiation of a human pancreatic duct line (hPANC-1) into pancreatic-beta-cell-like cells. These findings have important implications in cell-therapy based treatment of diabetes and may provide important insights in the designing of novel therapeutic agents to restore normal glycemia in subjects with diabetes.

  3. Restitution of defective glucose-stimulated insulin secretion in diabetic GK rat by acetylcholine uncovers paradoxical stimulatory effect of beta-cell muscarinic receptor activation on cAMP production.

    PubMed

    Dolz, Manuel; Bailbé, Danielle; Giroix, Marie-Hélène; Calderari, Sophie; Gangnerau, Marie-Noelle; Serradas, Patricia; Rickenbach, Katharina; Irminger, Jean-Claude; Portha, Bernard

    2005-11-01

    Because acetylcholine (ACh) is a recognized potentiator of glucose-stimulated insulin release in the normal beta-cell, we have studied ACh's effect on islets of the Goto-Kakizaki (GK) rat, a spontaneous model of type 2 diabetes. We first verified that ACh was able to restore the insulin secretory glucose competence of the GK beta-cell. Then, we demonstrated that in GK islets 1) ACh elicited a first-phase insulin release at low glucose, whereas it had no effect in Wistar; 2) total phospholipase C activity, ACh-induced inositol phosphate production, and intracellular free calcium concentration ([Ca2+]i) elevation were normal; 3) ACh triggered insulin release, even in the presence of thapsigargin, which induced a reduction of the ACh-induced [Ca2+]i response (suggesting that ACh produces amplification signals that augment the efficacy of elevated [Ca2+]i on GK exocytosis); 4) inhibition of protein kinase C did not affect [Ca2+]i nor the insulin release responses to ACh; and 5) inhibition of cAMP-dependent protein kinases (PKAs), adenylyl cyclases, or cAMP generation, while not affecting the [Ca2+]i response, significantly lowered the insulinotropic response to ACh (at low and high glucose). In conclusion, ACh acts mainly through activation of the cAMP/PKA pathway to potently enhance Ca2+-stimulated insulin release in the GK beta-cell and, in doing so, normalizes its defective glucose responsiveness.

  4. Mitochondrial control of nuclear apoptosis

    PubMed Central

    1996-01-01

    Anucleate cells can be induced to undergo programmed cell death (PCD), indicating the existence of a cytoplasmic PCD pathway that functions independently from the nucleus. Cytoplasmic structures including mitochondria have been shown to participate in the control of apoptotic nuclear disintegration. Before cells exhibit common signs of nuclear apoptosis (chromatin condensation and endonuclease-mediated DNA fragmentation), they undergo a reduction of the mitochondrial transmembrane potential (delta psi m) that may be due to the opening of mitochondrial permeability transition (PT) pores. Here, we present direct evidence indicating that mitochondrial PT constitutes a critical early event of the apoptotic process. In a cell-free system combining purified mitochondria and nuclei, mitochondria undergoing PT suffice to induce chromatin condensation and DNA fragmentation. Induction of PT by pharmacological agents augments the apoptosis-inducing potential of mitochondria. In contrast, prevention of PT by pharmacological agents impedes nuclear apoptosis, both in vitro and in vivo. Mitochondria from hepatocytes or lymphoid cells undergoing apoptosis, but not those from normal cells, induce disintegration of isolated Hela nuclei. A specific ligand of the mitochondrial adenine nucleotide translocator (ANT), bongkreik acid, inhibits PT and reduces apoptosis induction by mitochondria in a cell-free system. Moreover, it inhibits the induction of apoptosis in intact cells. Several pieces of evidence suggest that the proto-oncogene product Bcl-2 inhibits apoptosis by preventing mitochondrial PT. First, to inhibit nuclear apoptosis, Bcl-2 must be localized in mitochondrial but not nuclear membranes. Second, transfection-enforced hyperexpression of Bcl-2 directly abolishes the induction of mitochondrial PT in response to a protonophore, a pro- oxidant, as well as to the ANT ligand atractyloside, correlating with its apoptosis-inhibitory effect. In conclusion, mitochondrial PT appears

  5. Regulation of Apoptosis by Inhibitors of Apoptosis (IAPs).

    PubMed

    Berthelet, Jean; Dubrez, Laurence

    2013-03-14

    Inhibitors of Apoptosis (IAPs) are a family of proteins with various biological functions including regulation of innate immunity and inflammation, cell proliferation, cell migration and apoptosis. They are characterized by the presence of at least one N-terminal baculoviral IAP repeat (BIR) domain involved in protein-protein interaction. Most of them also contain a C-terminal RING domain conferring an E3-ubiquitin ligase activity. In drosophila, IAPs are essential to ensure cell survival, preventing the uncontrolled activation of the apoptotic protease caspases. In mammals, IAPs can also regulate apoptosis through controlling caspase activity and caspase-activating platform formation. Mammalian IAPs, mainly X-linked IAP (XIAP) and cellular IAPs (cIAPs) appeared to be important determinants of the response of cells to endogenous or exogenous cellular injuries, able to convert the survival signal into a cell death-inducing signal. This review highlights the role of IAP in regulating apoptosis in Drosophila and Mammals.

  6. [Apoptosis and its biomedical significance].

    PubMed

    Ortega-Camarillo, C; Díaz-Flores, M; Avalos-Rodríguez, A; Vergara-Onofre, M; Rosales-Torres, A M

    2001-01-01

    Cell death can occur through apoptotic or necrotic death pathways. Membrane disruption leads to inflammation, a typical feature of necrosis. Apoptosis constitutes a genetically controlled physiologic process of cell removal. It is characterized by cell shrinkage, chromatin condensation, and DNA cleavage. Apoptotic cells are rapidly recognized and engulfed by phagocytes thus inhibiting an inflammatory response following necrosis. Apoptosis has been proposed as a basic event to protect tissue homeostasis. This paper analyzes the genetic, biochemical, and morphologic characteristics related to apoptosis, as well as its relationship to certain illnesses. PMID:11766462

  7. Glucagon-like peptide-1 mobilizes intracellular Ca2+ and stimulates mitochondrial ATP synthesis in pancreatic MIN6 beta-cells.

    PubMed Central

    Tsuboi, Takashi; da Silva Xavier, Gabriela; Holz, George G; Jouaville, Laurence S; Thomas, Andrew P; Rutter, Guy A

    2003-01-01

    Glucagon-like peptide-1 (GLP-1) is a potent regulator of glucose-stimulated insulin secretion whose mechanisms of action are only partly understood. In the present paper, we show that at low (3 mM) glucose concentrations, GLP-1 increases the free intramitochondrial concentrations of both Ca(2+) ([Ca(2+)](m)), and ATP ([ATP](m)) in clonal MIN6 beta-cells. Suggesting that cAMP-mediated release of Ca(2+) from intracellular stores is responsible for these effects, increases in [ATP](m) that were induced by GLP-1 were completely blocked by the Rp isomer of adenosine-3',5'-cyclic monophosphothioate (Rp-cAMPS), or by chelation of intracellular Ca(2+). Furthermore, inhibition of Ins(1,4,5) P (3) (IP(3)) receptors with xestospongin C, or application of ryanodine, partially inhibited GLP-1-induced [ATP](m) increases, and the simultaneous blockade of both IP(3) and ryanodine receptors (RyR) completely eliminated the rise in [ATP](m). GLP-1 appeared to prompt Ca(2+)-induced Ca(2+) release through IP(3) receptors via a protein kinase A (PKA)-mediated phosphorylation event, since ryanodine-insensitive [ATP](m) increases were abrogated with the PKA inhibitor, H89. In contrast, the effects of GLP-1 on RyR-mediated [ATP](m) increases were apparently mediated by the cAMP-regulated guanine nucleotide exchange factor cAMP-GEFII, since xestospongin C-insensitive [ATP](m) increases were blocked by a dominant-negative form of cAMP-GEFII (G114E,G422D). Taken together, these results demonstrate that GLP-1 potentiates glucose-stimulated insulin release in part via the mobilization of intracellular Ca(2+), and the stimulation of mitochondrial ATP synthesis. PMID:12410638

  8. Apoptosis pathways in neuroblastoma therapy.

    PubMed

    Fulda, Simone; Debatin, Klaus Michael

    2003-07-18

    Apoptosis, the cell's intrinsic death program, plays a crucial role in the regulation of tissue homeostasis, and an imbalance between cell death and proliferation may result in tumor formation. Also, killing of tumor cells by diverse cytotoxic approaches such as anticancer drugs, gamma-irradiation, suicide genes or immunotherapy, is predominantly mediated through induction of apoptosis. Failure to activate apoptotic pathways in response to drug treatment may lead to resistance of neuroblastoma cells to anticancer therapies. Understanding the molecular events that regulate apoptosis induced by cytotoxic therapies and how neuroblastoma cells evade apoptotic events may provide a new paradigm for neuroblastoma therapy. Thus, novel strategies targeting resistance of neuroblastoma cells will be based on insights into the molecular mechanisms of apoptosis as well as other forms of cell death.

  9. Protooncogenes as mediators of apoptosis.

    PubMed

    Teng, C S

    2000-01-01

    Apoptosis has been well established as a vital biological phenomenon that is important in the maintenance of cellular homeostasis. Three major protooncogene families and their encoded proteins function as mediators of apoptosis in various cell types and are the subject of this chapter. Protooncogenic proteins such as c-Myc/Max, c-Fos/c-Jun, and Bcl-2/Bax utilize a synergetic effect to enhance their roles in the pro- or antiapoptotic action. These family members activate and repress the expression of their target genes, control cell cycle progression, and execute programmed cell death. Repression or overproduction of these protooncogenic proteins induces apoptosis, which may vary as a result of either cell type specificity or the nature of the apoptotic stimuli. The proapoptotic and antiapoptotic proteins exert their effects in the membrane of cellular organelles. Here they generate cell-type-specific signals that activate the caspase family of proteases and their regulators for the execution of apoptosis.

  10. Apoptosis and acute kidney injury

    PubMed Central

    Havasi, Andrea; Borkan, Steven C.

    2015-01-01

    Improved mechanistic understanding of renal cell death in acute kidney injury (AKI) has generated new therapeutic targets. Clearly, the classic lesion of acute tubular necrosis is not adequate to describe the consequences of renal ischemia, nephrotoxin exposure, or sepsis on glomerular filtration rate. Experimental evidence supports a pathogenic role for apoptosis in AKI. Interestingly, proximal tubule epithelial cells are highly susceptible to apoptosis, and injury at this site contributes to organ failure. During apoptosis, well-orchestrated events converge at the mitochondrion, the organelle that integrates life and death signals generated by the BCL2 (B-cell lymphoma 2) protein family. Death requires the ‘perfect storm’ for outer mitochondrial membrane injury to release its cellular ‘executioners’. The complexity of this process affords new targets for effective interventions, both before and after renal insults. Inhibiting apoptosis appears to be critical, because circulating factors released by the injured kidney induce apoptosis and inflammation in distant organs including the heart, lung, liver, and brain, potentially contributing to the high morbidity and mortality associated with AKI. Manipulation of known stress kinases upstream of mitochondrial injury, induction of endogenous, anti-apoptotic proteins, and improved understanding of the timing and consequences of renal cell apoptosis will inevitably improve the outcome of human AKI. PMID:21562469

  11. Interferon-alpha reduces insulin resistance and beta-cell secretion in responders among patients with chronic hepatitis B and C.

    PubMed

    Tai, T-Y; Lu, J-Y; Chen, C-L; Lai, M-Y; Chen, P-J; Kao, J-H; Lee, C-Z; Lee, H-S; Chuang, L-M; Jeng, Y-M

    2003-09-01

    This study aimed at elucidating the effects of interferon (IFN)-alpha on glucose metabolism in patients with chronic hepatitis B and C infections. Twenty-eight biopsy-proven patients with chronic hepatitis B (ten cases) and hepatitis C (18 cases) were given IFN-alpha for a total of 24 weeks. The patients received a 75 g oral glucose tolerance test (OGTT), glucagon stimulation test, tests for type 1 diabetes-related autoantibodies and an insulin suppression test before and after IFN-alpha therapy. Ten of the 28 patients responded to IFN-alpha therapy. Steady-state plasma glucose of the insulin suppression test decreased significantly in responders (13.32+/-1.48 (S.E.M.) vs 11.33+/-1.19 mmol/l, P=0.0501) but not in non-responders (12.29+/-1.24 vs 11.11+/-0.99 mmol/l, P=0.2110) immediately after completion of IFN-alpha treatment. In the oral glucose tolerance test, no significant difference was observed in plasma glucose in either responders (10.17+/-0.23 vs 10.03+/-0.22 mmol/l) or non-responders (10.11+/-0.22 vs 9.97+/-0.21 mmol/l) 3 Months after completion of IFN-alpha treatment. However, significant differences were noted in C-peptide in both responders (2.90+/-0.13 vs 2.20+/-0.09 nmol/l, P=0.0040) and non-responders (2.45+/-0.11 vs 2.22+/-0.08 nmol/l, P=0.0287) before vs after treatment. The changes of C-peptide in an OGTT between responders and non-responders were also significantly different (P=0.0028), with responders reporting a greater reduction in C-peptide. No case developed autoantibodies during the treatment. In patients who were successfully treated with IFN-alpha, insulin sensitivity improved and their plasma glucose stayed at the same level without secreting as much insulin from islet beta-cells.

  12. Viral Control of Mitochondrial Apoptosis

    PubMed Central

    Morselli, Eugenia; Touat, Zahia; Kroemer, Guido

    2008-01-01

    Throughout the process of pathogen–host co-evolution, viruses have developed a battery of distinct strategies to overcome biochemical and immunological defenses of the host. Thus, viruses have acquired the capacity to subvert host cell apoptosis, control inflammatory responses, and evade immune reactions. Since the elimination of infected cells via programmed cell death is one of the most ancestral defense mechanisms against infection, disabling host cell apoptosis might represent an almost obligate step in the viral life cycle. Conversely, viruses may take advantage of stimulating apoptosis, either to kill uninfected cells from the immune system, or to induce the breakdown of infected cells, thereby favoring viral dissemination. Several viral polypeptides are homologs of host-derived apoptosis-regulatory proteins, such as members of the Bcl-2 family. Moreover, viral factors with no homology to host proteins specifically target key components of the apoptotic machinery. Here, we summarize the current knowledge on the viral modulation of mitochondrial apoptosis, by focusing in particular on the mechanisms by which viral proteins control the host cell death apparatus. PMID:18516228

  13. Molecular mechanisms of hepatic apoptosis

    PubMed Central

    Wang, K

    2014-01-01

    Apoptosis is a prominent feature of liver diseases. Causative factors such as alcohol, viruses, toxic bile acids, fatty acids, drugs, and immune response, can induce apoptotic cell death via membrane receptors and intracellular stress. Apoptotic signaling network, including membrane death receptor-mediated cascade, reactive oxygen species (ROS) generation, endoplasmic reticulum (ER) stress, lysosomal permeabilization, and mitochondrial dysfunction, is intermixed each other, but one mechanism may dominate at a particular stage. Mechanisms of hepatic apoptosis are complicated by multiple signaling pathways. The progression of liver disease is affected by the balance between apoptotic and antiapoptotic capabilities. Therapeutic options of liver injury are impacted by the clear understanding toward mechanisms of hepatic apoptosis. PMID:24434519

  14. Inhibition of palmitate-induced GADD34 expression augments apoptosis in mouse insulinoma cells (MIN6).

    PubMed

    Fransson, Liselotte; Sjöholm, Ake; Ortsäter, Henrik

    2014-07-01

    Saturated fatty acids like palmitate induce endoplasmic reticulum (ER) stress in pancreatic beta-cells, an event linked to apoptotic loss of β-cells in type 2 diabetes. Sustained activation of the ER stress response leads to expression of growth arrest and DNA damage-inducible protein 34 (GADD34), a regulatory subunit of protein phosphatase 1. In the present study, we have used small interfering RNA in order to knockdown GADD34 expression in insulin-producing MIN6 cells prior to induction of ER stress by palmitate and evaluated its consequences on RNA-activated protein kinase-like ER-localized eIF2alpha kinase (PERK) signalling and apoptosis. Salubrinal, a specific inhibitor of eukaryotic initiation factor 2α (eIF2α) dephosphorylation, was used as a comparison. Salubrinal treatment augmented palmitate-induced ER stress and increased GADD34 levels. Both GADD34 knockdown and salubrinal treatment potentiated the cytotoxic effects of palmitate as evidenced by increased DNA fragmentation and activation of caspase 3, with the fundamental difference that the former did not involve enhanced levels of GADD34. The data from this study suggest that sustained activation of PERK signalling and eIF2α phosphorylation sensitizes insulin-producing MIN6 cells to lipoapoptosis independently of GADD34 expression levels. PMID:24633916

  15. Pancreatic carcinogenesis: apoptosis and angiogenesis.

    PubMed

    Onizuka, Shinya; Kawakami, Shunsuke; Taniguchi, Ken; Fujioka, Hikaru; Miyashita, Kosei

    2004-04-01

    Apoptosis and angiogenesis are critical biologic processes that are altered during carcinogenesis. Both apoptosis and angiogenesis may play an important role in pancreatic carcinogenesis. Despite numerous advances in the diagnosis and treatment of pancreatic cancer, its prognosis remains dismal and a new therapeutic approach is much needed. Recent research has revealed that apoptosis and angiogenesis are closely interrelated. Several reports show that a tumor suppresser gene that is expressed in pancreatic carcinoma and related to malignant potential can induce apoptosis and also inhibit angiogenesis. At present, it is generally accepted that tumor growth in cancers, including pancreatic cancer, depends on angiogenesis. We have identified 2 new angiogenesis inhibitors from a conditioned medium of human pancreatic carcinoma cell line (BxPC-3): antiangiogenic antithrombin III (aaAT-III) and vitamin D binding protein-macrophage activating factor (DBP-maf). These molecules were able to regress tumors in severe combined immunodeficiency disease (SCID) mice, demonstrating potent inhibition of endothelial cell proliferation. Moreover, the angiogenesis inhibitors induced tumor dormancy in the animal model. These results suggest that antiangiogenic therapy using angiogenesis inhibitors may become a new strategy for treatment of pancreatic cancer in the near future. PMID:15084979

  16. APOPTOSIS IN WHOLE MOUSE OVARIES

    EPA Science Inventory

    Apoptosis in Whole Mouse Ovaries
    Robert M. Zucker Susan C. Jeffay and Sally D. Perreault
    Reproductive Toxicology Division, National Health and Environmental Effects Research Laboratory, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina, 27711.

  17. Fluorescence Lifetime Imaging of Apoptosis

    PubMed Central

    Xiao, Annie; Gibbons, Anne E.; Luker, Kathryn E.; Luker, Gary D.

    2015-01-01

    Genetically-encoded fluorescence resonance energy transfer (FRET) reporters are powerful tools to analyze cell signaling and function at single cell resolution in standard two-dimensional cell cultures, but these reporters rarely have been applied to three-dimensional environments. FRET interactions between donor and acceptor molecules typically are determined by changes in relative fluorescence intensities, but wavelength-dependent differences in absorption of light complicate this analysis method in three-dimensional settings. Here we report fluorescence lifetime imaging microscopy (FLIM) with phasor analysis, a method that displays fluorescence lifetimes on a pixel-wise basis in real time, to quantify apoptosis in breast cancer cells stably expressing a genetically encoded FRET reporter. This microscopic imaging technology allowed us to identify treatment-induced apoptosis in single breast cancer cells in environments ranging from two-dimensional cell culture, spheroids with cancer and bone marrow stromal cells, and living mice with orthotopic human breast cancer xenografts. Using this imaging strategy, we showed that combined metabolic therapy targeting glycolysis and glutamine pathways significantly reduced overall breast cancer metabolism and induced apoptosis. We also determined that distinct subpopulations of bone marrow stromal cells control resistance of breast cancer cells to chemotherapy, suggesting heterogeneity of treatment responses of malignant cells in different bone marrow niches. Overall, this study establishes FLIM with phasor analysis as an imaging tool for apoptosis in cell-based assays and living mice, enabling real-time, cellular-level assessment of treatment efficacy and heterogeneity. PMID:26771007

  18. Analysis of fumonisin B1-induced apoptosis.

    PubMed Central

    Jones, C; Ciacci-Zanella, J R; Zhang, Y; Henderson, G; Dickman, M

    2001-01-01

    Fumonisins are mycotoxins produced by Fusarium moniliforme, a prevalent fungus that infects corn and other cereal grains. Fumonisin B1(FB1 is the most common mycotoxin produced by F. moniliforme, suggesting it has toxicologic significance. The structure of FB1 resembles sphingoid bases, and it inhibits ceramide synthase. Because sphingoid bases regulate cell growth, differentiation, transformation, and apoptosis, it is not surprising to find that FB1 can alter growth of certain mammalian cells. Previous studies concluded FB1-induced apoptosis, or cell cycle arrest, in African green monkey kidney fibroblasts (CV-1). In this study we have identified genes that inhibit FB1 induced apoptosis in CV-1 cells and two mouse embryo fibroblasts (MEF). A baculovirus gene, inhibitor of apoptosis (CpIAP), protected these cells from apoptosis. CpIAP blocks apoptosis induced by the tumor necrosis factor (TNF) pathway as well as other mechanisms. Further support for the involvement of the TNF signal transduction pathway in FB1 induced apoptosis was the cleavage of caspase 8. Inhibition of caspases by the baculovirus gene (italic)p35 also inhibited FB1-induced apoptosis. The tumor suppressor gene p53 was not required for FB1 induced apoptosis because p53-/- MEF undergo apoptosis following FB1 treatment. Furthermore, Bcl-2 was not an effective inhibitor of FB1-induced apoptosis in CV-1 cells or p53+/+ MEF. In summary, these results provide new information to help understand the mechanism by which FB1 induces apoptosis. PMID:11359701

  19. Combination high-dose omega-3 fatty acids and high-dose cholecalciferol in new onset type 1 diabetes: a potential role in preservation of beta-cell mass.

    PubMed

    Baidal, D A; Ricordi, C; Garcia-Contreras, M; Sonnino, A; Fabbri, A

    2016-07-01

    Several studies have evaluated the role of inflammation in type 1 diabetes (T1D). The safety profile and anti-inflammatory properties of high dose omega-3 fatty acids combined with Vitamin D supplementation make this therapy a possible candidate for T1D intervention trials. Herein, we describe the case of a 14-year-old boy with new onset T1D treated with high dose Omega-3 and vitamin D3. By 12 months, peak C-peptide increased to 0.55 nmol/L (1.66 ng/mL) corresponding to a 20% increment from baseline and AUC C-peptide was slightly higher compared to 9 months (0.33 vs. 0.30 nmol/L/min) although remaining slightly lower than baseline. Combination high-dose Omega-3 fatty acids and high-dose vitamin D3 therapy was well tolerated and may have beneficial effects on beta-cell function. Randomized controlled trials could be of assistance to determine whether this therapy may result in the preservation of beta-cell function in patients with new onset T1D.

  20. Mechanisms of p53-induced apoptosis.

    PubMed

    Bennett, M R

    1999-10-01

    The p53 tumour suppressor gene functions in both cell cycle arrest and apoptosis. Despite considerable advances in understanding as to how p53 regulates growth arrest, the mechanisms by which p53 regulates apoptosis are only just emerging. In particular, there appears to be a structural and functional separation between the ability of p53 to induce growth arrest and apoptosis. This review examines the interactions between p53-induced growth arrest and apoptosis, and the mechanisms of p53-induced apoptosis, both via induction of p53 transcriptional targets and via nontranscriptional mechanisms.

  1. [Apoptosis modulation by human papillomavirus].

    PubMed

    Jave-Suárez, Luis Felipe; Ratkovich-González, Sarah; Olimón-Andalón, Vicente; Aguilar-Lemarroy, Adriana

    2015-01-01

    One of the most important processes to keep the homeostasis in organisms is the apoptosis, also called programmed cell death. This mechanism works through two pathways: The intrinsic or mitochondrial, which responds to DNA damage and extern agents like UV radiation; and the extrinsic or receptor-mediated, which binds to their ligands to initiate the apoptotic trail. The evasion of apoptosis is one of the main causes of cellular transformation to malignity. Many viruses had shown capacity to modify the apoptotic process; among them is the human papillomavirus, which, by means of its oncoproteins, interferes in pathways, reacting with the receptors and molecules and participating in the death mechanism. This creates ideal conditions for cancer development.

  2. Apoptosis in Cryopreserved Eukaryotic Cells.

    PubMed

    Savitskaya, M A; Onishchenko, G E

    2016-05-01

    This review considers apoptosis mechanisms that have been revealed in cryopreserved cells and which can be controlled using different chemical agents, thereby improving the viability of cells after their return to normal conditions. The role of oxidative stress as of the most significant damaging factor is discussed, as well as the reasonability of including antioxidants into cryopreservation/thawing protocols as independent agents or in combination with other compounds.

  3. Apoptosis in irradiated murine tumors.

    PubMed

    Stephens, L C; Ang, K K; Schultheiss, T E; Milas, L; Meyn, R E

    1991-09-01

    Early radiation responses of transplantable murine ovarian (OCaI) and hepatocellular (HCaI) carcinomas were examined at 6, 24, 48, 96, and 144 h after single photon doses of 25, 35, or 45 Gy. Previous studies using tumor growth delay and tumor radiocurability assays had shown OCaI tumors to be relatively radiosensitive and HCaI tumors to be radioresistant. At 6 h, approximately 20% of nuclei in OCaI tumors showed aberrations characteristic of cell death by apoptosis. This contrasted to an incidence of 3% in HCaI tumors. Mitotic activity was eliminated in OCaI tumors but was only transiently suppressed in HCaI tumors. At 24-96 h, OCaI tumors continued to display apoptosis and progressive necrosis, whereas HCaI tumors responded by exhibiting marked pleomorphism. Factors other than mitotic activity may influence tumor radiosensitivity, and one of these may be susceptibility to induction of apoptosis (programmed cell death), because this was a prominent early radiation response by the radiosensitive OCaI tumors.

  4. Apoptosis in irradiated murine tumors.

    PubMed

    Stephens, L C; Ang, K K; Schultheiss, T E; Milas, L; Meyn, R E

    1991-09-01

    Early radiation responses of transplantable murine ovarian (OCaI) and hepatocellular (HCaI) carcinomas were examined at 6, 24, 48, 96, and 144 h after single photon doses of 25, 35, or 45 Gy. Previous studies using tumor growth delay and tumor radiocurability assays had shown OCaI tumors to be relatively radiosensitive and HCaI tumors to be radioresistant. At 6 h, approximately 20% of nuclei in OCaI tumors showed aberrations characteristic of cell death by apoptosis. This contrasted to an incidence of 3% in HCaI tumors. Mitotic activity was eliminated in OCaI tumors but was only transiently suppressed in HCaI tumors. At 24-96 h, OCaI tumors continued to display apoptosis and progressive necrosis, whereas HCaI tumors responded by exhibiting marked pleomorphism. Factors other than mitotic activity may influence tumor radiosensitivity, and one of these may be susceptibility to induction of apoptosis (programmed cell death), because this was a prominent early radiation response by the radiosensitive OCaI tumors. PMID:1886987

  5. Glucose- and GTP-dependent stimulation of the carboxyl methylation of CDC42 in rodent and human pancreatic islets and pure beta cells. Evidence for an essential role of GTP-binding proteins in nutrient-induced insulin secretion.

    PubMed Central

    Kowluru, A; Seavey, S E; Li, G; Sorenson, R L; Weinhaus, A J; Nesher, R; Rabaglia, M E; Vadakekalam, J; Metz, S A

    1996-01-01

    Several GTP-binding proteins (G-proteins) undergo post-translational modifications (isoprenylation and carboxyl methylation) in pancreatic beta cells. Herein, two of these were identified as CDC42 and rap 1, using Western blotting and immunoprecipitation. Confocal microscopic data indicated that CDC42 is localized only in islet endocrine cells but not in acinar cells of the pancreas. CDC42 undergoes a guanine nucleotide-specific membrane association and carboxyl methylation in normal rat islets, human islets, and pure beta (HIT or INS-1) cells. GTPgammaS-dependent carboxyl methylation of a 23-kD protein was also demonstrable in secretory granule fractions from normal islets or beta cells. AFC (a specific inhibitor of prenyl-cysteine carboxyl methyl transferases) blocked the carboxyl methylation of CDC42 in five types of insulin-secreting cells, without blocking GTPgammaS-induced translocation, implying that methylation is a consequence (not a cause) of transfer to membrane sites. High glucose (but not a depolarizing concentration of K+) induced the carboxyl methylation of CDC42 in intact cells, as assessed after specific immunoprecipitation. This effect was abrogated by GTP depletion using mycophenolic acid and was restored upon GTP repletion by coprovision of guanosine. In contrast, although rap 1 was also carboxyl methylated, it was not translocated to the particulate fraction by GTPgammaS; furthermore, its methylation was also stimulated by 40 mM K+ (suggesting a role which is not specific to nutrient stimulation). AFC also impeded nutrient-induced (but not K+-induced) insulin secretion from islets and beta cells under static or perifusion conditions, whereas an inactive structural analogue of AFC failed to inhibit insulin release. These effects were reproduced not only by S-adenosylhomocysteine (another methylation inhibitor), but also by GTP depletion. Thus, the glucose- and GTP-dependent carboxyl methylation of G-proteins such as CDC42 is an obligate step in

  6. Human papillomavirus oncoproteins and apoptosis (Review)

    PubMed Central

    JIANG, PEIYUE; YUE, YING

    2014-01-01

    The aim of this study was to review the literature and identify the association between human papillomavirus (HPV) oncoproteins and apoptosis. HPV-associated apoptosis may be primarily blocked by a number of oncoproteins, including E5, E6 and E7. E5 protein protects cells from tumor necrosis factor-associated apoptosis; the oncoprotein E6 predominantly inhibits apoptosis through the p53 pathway; and oncoprotein E7 is involved in apoptosis activation and inhibition. In addition, HPV oncoproteins are involved in activating or repressing the transcription of E6/E7. In conclusion, HPV oncoproteins, including E5, E6 and E7 protein, may interfere with apoptosis via certain regulatory principles. PMID:24348754

  7. Islet-Specific CTL Cloned from a Type 1 Diabetes Patient Cause Beta-Cell Destruction after Engraftment into HLA-A2 Transgenic NOD/SCID/IL2RG Null Mice

    PubMed Central

    Unger, Wendy W. J.; Pearson, Todd; Abreu, Joana R. F.; Laban, Sandra; van der Slik, Arno R.; der Kracht, Sacha Mulder-van; Kester, Michel G. D.; Serreze, Dave V.; Shultz, Leonard D.; Griffioen, Marieke; Drijfhout, Jan Wouter

    2012-01-01

    Despite increasing evidence that autoreactive CD8 T-cells are involved in both the initiation of type 1 diabetes (T1D) and the destruction of beta-cells, direct evidence for their destructive role in-vivo is lacking. To address a destructive role for autoreactive CD8 T-cells in human disease, we assessed the pathogenicity of a CD8 T-cell clone derived from a T1D donor and specific for an HLA-A2-restricted epitope of islet-specific glucose-6-phosphatase catalytic-subunit related protein (IGRP). HLA-A2/IGRP tetramer staining revealed a higher frequency of IGRP-specific CD8 T-cells in the peripheral blood of recent onset human individuals than of healthy donors. IGRP265–273-specific CD8 T-cells that were cloned from the peripheral blood of a recent onset T1D individual were shown to secrete IFNγ and Granzyme B after antigen-specific activation and lyse HLA-A2-expressing murine islets in-vitro. Lytic capacity was also demonstrated in-vivo by specific killing of peptide-pulsed target cells. Using the HLA-A2 NOD-scid IL2rγnull mouse model, HLA-A2-restricted IGRP-specific CD8 T-cells induced a destructive insulitis. Together, this is the first evidence that human HLA-restricted autoreactive CD8 T-cells target HLA-expressing beta-cells in-vivo, demonstrating the translational value of humanized mice to study mechanisms of disease and therapeutic intervention strategies. PMID:23155466

  8. Apoptosis in cancer: from pathogenesis to treatment

    PubMed Central

    2011-01-01

    Apoptosis is an ordered and orchestrated cellular process that occurs in physiological and pathological conditions. It is also one of the most studied topics among cell biologists. An understanding of the underlying mechanism of apoptosis is important as it plays a pivotal role in the pathogenesis of many diseases. In some, the problem is due to too much apoptosis, such as in the case of degenerative diseases while in others, too little apoptosis is the culprit. Cancer is one of the scenarios where too little apoptosis occurs, resulting in malignant cells that will not die. The mechanism of apoptosis is complex and involves many pathways. Defects can occur at any point along these pathways, leading to malignant transformation of the affected cells, tumour metastasis and resistance to anticancer drugs. Despite being the cause of problem, apoptosis plays an important role in the treatment of cancer as it is a popular target of many treatment strategies. The abundance of literature suggests that targeting apoptosis in cancer is feasible. However, many troubling questions arise with the use of new drugs or treatment strategies that are designed to enhance apoptosis and critical tests must be passed before they can be used safely in human subjects. PMID:21943236

  9. Curcumin enhances recovery of pancreatic islets from cellular stress induced inflammation and apoptosis in diabetic rats

    SciTech Connect

    Rashid, Kahkashan; Sil, Parames C.

    2015-02-01

    The phytochemical, curcumin, has been reported to play many beneficial roles. However, under diabetic conditions, the detail mechanism of its beneficial action in the glucose homeostasis regulatory organ, pancreas, is poorly understood. The present study has been designed and carried out to explore the role of curcumin in the pancreatic tissue of STZ induced and cellular stress mediated diabetes in eight weeks old male Wistar rats. Diabetes was induced with a single intraperitoneal dose of STZ (65 mg/kg body weight). Post to diabetes induction, animals were treated with curcumin at a dose of 100 mg/kg body weight for eight weeks. Underlying molecular and cellular mechanism was determined using various biochemical assays, DNA fragmentation, FACS, histology, immunoblotting and ELISA. Treatment with curcumin reduced blood glucose level, increased plasma insulin and mitigated oxidative stress related markers. In vivo and in vitro experimental results revealed increased levels of proinflammatory cytokines (TNF-α, IL1-β and IFN-γ), reduced level of cellular defense proteins (Nrf-2 and HO-1) and glucose transporter (GLUT-2) along with enhanced levels of signaling molecules of ER stress dependent and independent apoptosis (cleaved Caspase-12/9/8/3) in STZ administered group. Treatment with curcumin ameliorated all the adverse changes and helps the organ back to its normal physiology. Results suggest that curcumin protects pancreatic beta-cells by attenuating inflammatory responses, and inhibiting ER/mitochondrial dependent and independent pathways of apoptosis and crosstalk between them. This uniqueness and absence of any detectable adverse effect proposes the possibility of using this molecule as an effective protector in the cellular stress mediated diabetes mellitus. - Highlights: • STZ induced cellular stress plays a vital role in pancreatic dysfunction. • Cellular stress causes inflammation, pancreatic islet cell death and diabetes. • Deregulation of Nrf-2

  10. Apoptosis in myocardial ischaemia and infarction.

    PubMed

    Krijnen, P A J; Nijmeijer, R; Meijer, C J L M; Visser, C A; Hack, C E; Niessen, H W M

    2002-11-01

    Recent studies indicate that, in addition to necrosis, apoptosis also plays a role in the process of tissue damage after myocardial infarction, which has pathological and therapeutic implications. This review article will discuss studies in which the role and mechanisms of apoptosis in myocardial infarction were analysed in vivo and in vitro in humans and in animals.

  11. Proteases in Fas-mediated apoptosis.

    PubMed

    Zhivotovsky, B; Burgess, D H; Schlegel, J; Pörn, M I; Vanags, D; Orrenius, S

    1997-01-01

    Involvement of a unique family of cysteine proteases in the multistep apoptotic process has been documented. Cloning of several mammalian genes identifies some components of this cellular response. However, it is currently unclear which protease plays a role as a signal and/or effector of apoptosis. We summarize contributions to the data concerning proteases in Fas-mediated apoptosis.

  12. CHCHD2 connects mitochondrial metabolism to apoptosis.

    PubMed

    Liu, Yong; Zhang, Yanping

    2015-01-01

    As the powerhouse of cells and gatekeeper for apoptosis, mitochondria control life and death. CHCHD2, a mitochondrial protein previously known to regulate metabolism, has recently been identified as an apoptosis inhibitor. New data suggest a model in which CHCHD2 performs a prosurvival function by acting as both a reactive oxygen species scavenger and BCL-XL activator. PMID:27308501

  13. THE ROLE OF APOPTOSIS IN NEUROTOXICOLOGY.

    EPA Science Inventory

    The role of apoptosis in neurodegeneration in developing animals and in adults has become increasingly apparent in the past ten years. Normal apoptosis occurs in the CNS from the embryonic stage through senescence, with different cells in each region of the nervous system having ...

  14. Induction of apoptosis by Shiga toxins

    PubMed Central

    Tesh, Vernon L

    2010-01-01

    Shiga toxins comprise a family of structurally and functionally related protein toxins expressed by Shigella dysenteriae serotype 1 and multiple serotypes of Escherichia coli. While the capacity of Shiga toxins to inhibit protein synthesis by catalytic inactivation of eukaryotic ribosomes has been well described, it is also apparent that Shiga toxins trigger apoptosis in many cell types. This review presents evidence that Shiga toxins induce apoptosis of epithelial, endothelial, leukocytic, lymphoid and neuronal cells. Apoptotic signaling pathways activated by the toxins are reviewed with an emphasis on signaling mechanisms that are shared among different cell types. Data suggesting that Shiga toxins induce apoptosis through the endoplasmic reticulum stress response and clinical evidence demonstrating apoptosis in humans infected with Shiga toxin-producing bacteria are briefly discussed. The potential for use of Shiga toxins to induce apoptosis in cancer cells is briefly reviewed. PMID:20210553

  15. Switch to anaerobic glucose metabolism with NADH accumulation in the beta-cell model of mitochondrial diabetes. Characteristics of betaHC9 cells deficient in mitochondrial DNA transcription.

    PubMed

    Noda, Mitsuhiko; Yamashita, Shigeo; Takahashi, Noriko; Eto, Kazuhiro; Shen, Lin-Ming; Izumi, Kazuo; Daniel, Samira; Tsubamoto, Yoshiharu; Nemoto, Tomomi; Iino, Masamitsu; Kasai, Haruo; Sharp, Geoffrey W G; Kadowaki, Takashi

    2002-11-01

    To elucidate the mechanism underlying diabetes caused by mitochondrial gene mutations, we created a model by applying 0.4 microg/ml ethidium bromide (EtBr) to the murine pancreatic beta cell line betaH