| 1. |
- Elksnis, Andris, et al.
(författare)
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Imatinib protects against human beta-cell death via inhibition of mitochondrial respiration and activation of AMPK
- 2021
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Ingår i: Clinical Science. - : Portland Press. - 0143-5221 .- 1470-8736. ; 135:19, s. 2243-2263
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Tidskriftsartikel (refereegranskat)abstract
- The protein tyrosine kinase inhibitor imatinib is used in the treatment of various malignancies but may also promote beneficial effects in the treatment of diabetes. The aim of the present investigation was to characterize the mechanisms by which imatinib protects insulin producing cells. Treatment of non-obese diabetic (NOD) mice with imatinib resulted in increased beta-cell AMP-activated kinase (AMPK) phosphorylation. Imatinib activated AMPK also in vitro, resulting in decreased ribosomal protein S6 phosphorylation and protection against islet amyloid polypeptide (IAPP)-aggregation, thioredoxin interacting protein (TXNIP) up-regulation and beta-cell death. 5-Aminoimidazole-4-carboxamide ribonucleotide (AICAR) mimicked and compound C counteracted the effect of imatinib on beta-cell survival. Imatinib-induced AMPK activation was preceded by reduced glucose/pyruvate-dependent respiration, increased glycolysis rates, and a lowered ATP/AMP ratio. Imatinib augmented the fractional oxidation of fatty acids/malate, possibly via a direct interaction with the beta-oxidation enzyme enoyl coenzyme A hydratase, short chain, 1, mitochondrial (ECHS1). In non-beta cells, imatinib reduced respiratory chain complex I and II-mediated respiration and acyl-CoA carboxylase (ACC) phosphorylation, suggesting that mitochondrial effects of imatinib are not beta-cell specific. In conclusion, tyrosine kinase inhibitors modestly inhibit mitochondrial respiration, leading to AMPK activation and TXNIP down-regulation, which in turn protects against beta-cell death.
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| 2. |
- Elksnis, Andris, 1993-
(författare)
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Pharmaceutical Protection of Beta-Cells in Diabetes : Using Tyrosine Kinase Inhibition and NOX4 Inhibitors
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Diabetes mellitus is a complex and heterogenous disease, with loss of beta-cell function and mass being a characteristic of not only type 1 diabetes (T1D), but also type 2 diabetes (T2D). In T1D, inappropriate inflammatory signaling is thought to participate in the autoimmune suppression and destruction of beta-cells. In T2D progressive insulin resistance with resulting glucolipotoxicity, increased inflammation and oxidative stress, drives islet amyloid formation and subsequent beta-cell exhaustion and failure. Even under best managed care, disease progression and eventual complications are unavoidable. New interventions that aim to improve beta-cell survival are highly needed. This thesis investigates two such possible interventions: the tyrosine kinase inhibitor Imatinib, and selective NADPH-oxidase inhibition.Imatinib mesylate, used in treatment of chronic myeloid leukemia and other malignancies, was soon after its introduction reported to possess anti-diabetic properties in both T1D and T2D patients undergoing treatment. Imatinib has been shown to prevent and reverse diabetes in NOD mice and improve glucose tolerance in high fat diet treated rats. In paper I, we aimed to characterize the mechanisms by which imatinib protects beta-cells. We found that imatinib inhibits complex I and II of the respiratory chain, leading to improved beta-cell survival through AMPK activation, reduced amyloid formation and protection against TXNIP upregulation.Oxidative stress may play a pivotal role in the development of beta-cell dysfunction and failure in T2D. The NADPH-oxidases are a family of 7 enzymes (NOX1-5 and DUOX 1-2), that produce reactive oxygen species that are important in various physiological processes but may, if excessively activated, also be a source for oxidative stress in T2D. In paper II, we evaluate novel selective NOX inhibitors as protective agents against in vitro induced human beta-cell stress. Selective NOX4 inhibition protected beta-cells against both cytokines and high-glucose + palmitate. In paper III we found that NOX4 inhibition increased mitochondrial membrane potential, mitochondrial reactive oxygen species and ATP/ADP ratio in a human beta-cell line, and this was paralleled with protection against human islet cell death when challenged with high-glucose and palmitate. Finally, in paper IV, we attempt to apply these findings in vivo, by transplanting athymic diabetic mice with human islets and treating them with a NOX4 inhibitor over a period of 4 weeks. Treated mice achieved lower blood glucose levels and water consumption throughout the treatment period, and apoptotic rates of insulin-positive human cells, measured as co-localization of insulin and cleaved caspase-3, were greatly reduced.
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| 3. |
- Elksnis, Andris, et al.
(författare)
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Pharmacological Inhibition of NOX4 Improves Mitochondrial Function and Survival in Human Beta-Cells
- 2021
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Ingår i: Biomedicines. - : MDPI. - 2227-9059. ; 9:12
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Tidskriftsartikel (refereegranskat)abstract
- Previous studies have reported beneficial effects of NADPH oxidase 4 (NOX4) inhibition on beta-cell survival in vitro and in vivo. The mechanisms by which NOX4 inhibition protects insulin producing cells are, however, not known. The aim of the present study was to investigate the effects of a pharmacological NOX4 inhibitor (GLX7013114) on human islet and EndoC-beta H1 cell mitochondrial function, and to correlate such effects with survival in islets of different size, activity, and glucose-stimulated insulin release responsiveness. We found that maximal oxygen consumption rates, but not the rates of acidification and proton leak, were increased in islets after acute NOX4 inhibition. In EndoC-beta H1 cells, NOX4 inhibition increased the mitochondrial membrane potential, as estimated by JC-1 fluorescence; mitochondrial reactive oxygen species (ROS) production, as estimated by MitoSOX fluorescence; and the ATP/ADP ratio, as assessed by a bioluminescent assay. Moreover, the insulin release from EndoC-beta H1 cells at a high glucose concentration increased with NOX4 inhibition. These findings were paralleled by NOX4 inhibition-induced protection against human islet cell death when challenged with high glucose and sodium palmitate. The NOX4 inhibitor protected equally well islets of different size, activity, and glucose responsiveness. We conclude that pharmacological alleviation of NOX4-induced inhibition of beta-cell mitochondria leads to increased, and not decreased, mitochondrial ROS, and this was associated with protection against cell death occurring in different types of heterogeneous islets. Thus, NOX4 inhibition or modulation may be a therapeutic strategy in type 2 diabetes that targets all types of islets.
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| 4. |
- Elksnis, Andris, et al.
(författare)
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The selective NOX4 inhibitor GLX7013159 decreases blood glucose concentrations and human beta-cell apoptotic rates in diabetic NMRI nu/nu mice transplanted with human islets
- 2023
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Ingår i: Free radical research. - : Taylor & Francis. - 1071-5762 .- 1029-2470. ; 57:6-12, s. 460-469
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Tidskriftsartikel (refereegranskat)abstract
- NADPH oxidase 4 (NOX4) inhibition has been reported to mitigate diabetes-induced beta-cell dysfunction and improve survival in vitro, as well as counteract high-fat diet-induced glucose intolerance in mice. We investigated the antidiabetic effects of the selective NOX4 inhibitor GLX7013159 in vivo in athymic diabetic mice transplanted with human islets over a period of 4 weeks. The GLX7013159-treated mice achieved lower blood glucose and water consumption throughout the treatment period. Furthermore, GLX7013159 treatment resulted in improved insulin and c-peptide levels, better insulin secretion capacity, as well as in greatly reduced apoptotic rates of the insulin-positive human cells, measured as colocalization of insulin and cleaved caspase-3. We conclude that the antidiabetic effects of NOX4 inhibition by GLX7013159 are observed also during a prolonged study period in vivo and are likely to be due to an improved survival and function of the human beta-cells.
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| 5. |
- Ngamjariyawat, Anongnad, 1976-, et al.
(författare)
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GDF15 Protects Insulin-Producing Beta Cells against Pro-Inflammatory Cytokines and Metabolic Stress via Increased Deamination of Intracellular Adenosine
- 2024
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Ingår i: International Journal of Molecular Sciences. - : MDPI. - 1661-6596 .- 1422-0067. ; 25:2
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Tidskriftsartikel (refereegranskat)abstract
- It has been proposed that antidiabetic drugs, such as metformin and imatinib, at least in part, promote improved glucose tolerance in type 2 diabetic patients via increased production of the inflammatory cytokine GDF15. This is supported by studies, performed in rodent cell lines and mouse models, in which the addition or production of GDF15 improved beta-cell function and survival. The aim of the present study was to determine whether human beta cells produce GDF15 in response to antidiabetic drugs and, if so, to further elucidate the mechanisms by which GDF15 modulates the function and survival of such cells. The effects and expression of GDF15 were analyzed in human insulin-producing EndoC-betaH1 cells and human islets. We observed that alpha and beta cells exhibit considerable heterogeneity in GDF15 immuno-positivity. The predominant form of GDF15 present in islet and EndoC-betaH1 cells was pro-GDF15. Imatinib, but not metformin, increased pro-GDF15 levels in EndoC-betaH1 cells. Under basal conditions, exogenous GDF15 increased human islet oxygen consumption rates. In EndoC-betaH1 cells and human islets, exogenous GDF15 partially ameliorated cytokine- or palmitate + high-glucose-induced loss of function and viability. GDF15-induced cell survival was paralleled by increased inosine levels, suggesting a more efficient disposal of intracellular adenosine. Knockdown of adenosine deaminase, the enzyme that converts adenosine to inosine, resulted in lowered inosine levels and loss of protection against cytokine- or palmitate + high-glucose-induced cell death. It is concluded that imatinib-induced GDF15 production may protect human beta cells partially against inflammatory and metabolic stress. Furthermore, it is possible that the GDF15-mediated activation of adenosine deaminase and the increased disposal of intracellular adenosine participate in protection against beta-cell death.
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| 6. |
- Ngamjariyawat, Anongnad, 1976-, et al.
(författare)
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Metabolic stress-induced human beta-cell death is mediated by increased intracellular levels of adenosine
- 2023
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Ingår i: Frontiers in Endocrinology. - : Frontiers Media S.A.. - 1664-2392. ; 14
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Tidskriftsartikel (refereegranskat)abstract
- Introduction: High intracellular concentrations of adenosine and 2'-deoxyadenosine have been suggested to be an important mediator of cell death. The aim of the present study was to characterize adenosine-induced death in insulin-producing beta-cells, at control and high glucose + palmitate-induced stress conditions.Methods: Human insulin-producing EndoC-betaH1 cells were treated with adenosine, 2'-deoxyadenosine, inosine and high glucose + sodium palmitate, and death rates using flow cytometry were studied.Results: We observed that adenosine and the non-receptor-activating analogue 2-deoxyadenosine, but not the adenosine deamination product inosine, promoted beta-cell apoptosis at concentrations exceeding maximal adenosine-receptor stimulating concentrations. Both adenosine and inosine were efficiently taken up by EndoC-betaH1 cells, and inosine counteracted the cell death promoting effect of adenosine by competing with adenosine for uptake. Both adenosine and 2'-deoxyadenosine promptly reduced insulin-stimulated production of plasma membrane PI(3,4,5)P-3, an effect that was reversed upon wash out of adenosine. In line with this, adenosine, but not inosine, rapidly diminished Akt phosphorylation. Both pharmacological Bax inhibition and Akt activation blocked adenosine-induced beta-cell apoptosis, indicating that adenosine/2'-deoxyadenosine inhibits the PI3K/Akt/BAD anti-apoptotic pathway. High glucose + palmitate-induced cell death was paralleled by increased intracellular adenosine and inosine levels. Overexpression of adenosine deaminase-1 (ADA1) in EndoC-betaH1 cells, which increased Akt phosphorylation, prevented both adenosine-induced apoptosis and high glucose + palmitate-induced necrosis. ADA2 overexpression not only failed to protect against adenosine and high glucose + palmitate-activated cell death, but instead potentiated the apoptosis-stimulating effect of adenosine. In line with this, ADA1 overexpression increased inosine production from adenosine-exposed cells, whereas ADA2 did not. Knockdown of ADA1 resulted in increased cell death rates in response to both adenosine and high glucose + palmitate. Inhibition of miR-30e-3p binding to the ADA1 mRNA 3'-UTR promoted the opposite effects on cell death rates and reduced intracellular adenosine contents.Discussion: It is concluded that intracellular adenosine/2'-deoxyadenosine regulates negatively the PI3K pathway and is therefore an important mediator of beta-cell apoptosis. Adenosine levels are controlled, at least in part, by ADA1, and strategies to upregulate ADA1 activity, during conditions of metabolic stress, could be useful in attempts to preserve beta-cell mass in diabetes.
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| 7. |
- Shi, Ruifeng, et al.
(författare)
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CLEC11A improves insulin secretion and promotes cell proliferation in human beta-cells
- 2023
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Ingår i: Journal of Molecular Endocrinology. - : Bioscientifica. - 0952-5041 .- 1479-6813. ; 71:1
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Tidskriftsartikel (refereegranskat)abstract
- Beta-cell dysfunction is a hallmark of disease progression in patients with diabetes. Research has been focused on maintaining and restoring beta-cell function during diabetes development. The aims of this study were to explore th e expression of C-type lectin domain containing 11A (CLEC11A), a secreted sulphated glycoprotein, in human islets and to evaluate the effects of CLEC11A on beta-cell funct ion and proliferation in vitro. To test these hypotheses, human islets and human EndoC-beta H1 cell line were used in this study. We identified that CLEC11A was expressed in beta-cells and alpha-cells in human islets but not in EndoC-beta H1 cells, whereas the receptor of CLEC11A called integrin subunit alpha 11 was found in both human islets and En doC-beta H1 cells. Long-term treatment with exogenous recombinant human CLEC11A (rhCLEC11A) accentuated glucose-stimulated insulin secretion, insulin content, and proliferation from human islets and EndoC-beta H1 cells, which was partially due to the accentuated expression levels of transcription factors MAFA and PDX1. However, the impaired beta-cell function and reduced mRNA expression of INS and MAFA in EndoC-beta H1 cells that were caused by chronic palmitate exposure could only be partially improved by the introduction of rhCLEC11A. Based on these results, we conclude that rhCLEC11A promotes insulin secretion, insulin content, and proliferation in human beta-cells, which are associated with the accentuated expression levels of transcription factors MAFA and PDX1. CLEC11A, therefore, may provide a novel therapeutic target for maintaining beta-cell function in patients with diabetes.
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| 8. |
- Wang, Xuan, 1984-, et al.
(författare)
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ZBED6 counteracts high-fat diet-induced glucose intolerance by maintaining beta cell area and reducing excess mitochondrial activation
- 2021
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Ingår i: Diabetologia. - : Springer Nature. - 0012-186X .- 1432-0428. ; 64:10, s. 2292-2305
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Tidskriftsartikel (refereegranskat)abstract
- Aims/hypothesisZBED6 (zinc finger, BED-type containing 6) is known to regulate muscle mass by suppression of Igf2 gene transcription. In insulin-producing cell lines, ZBED6 maintains proliferative capacity at the expense of differentiation and beta cell function. The aim was to study the impact of Zbed6 knockout on beta cell function and glucose tolerance in C57BL/6 mice.MethodsBeta cell area and proliferation were determined in Zbed6 knockout mice using immunohistochemical analysis. Muscle and fat distribution were assessed using micro-computed tomography. Islet gene expression was assessed by RNA sequencing. Effects of a high-fat diet were analysed by glucose tolerance and insulin tolerance tests. ZBED6 was overexpressed in EndoC-βH1 cells and human islet cells using an adenoviral vector. Beta cell cell-cycle analysis, insulin release and mitochondrial function were studied in vitro using propidium iodide staining and flow cytometry, ELISA, the Seahorse technique, and the fluorescent probes JC-1 and MitoSox.ResultsIslets from Zbed6 knockout mice showed lowered expression of the cell cycle gene Pttg1, decreased beta cell proliferation and decreased beta cell area, which occurred independently from ZBED6 effects on Igf2 gene expression. Zbed6 knockout mice, but not wild-type mice, developed glucose intolerance when given a high-fat diet. The high-fat diet Zbed6 knockout islets displayed upregulated expression of oxidative phosphorylation genes and genes associated with beta cell differentiation. In vitro, ZBED6 overexpression resulted in increased EndoC-βH1 cell proliferation and a reduced glucose-stimulated insulin release in human islets. ZBED6 also reduced mitochondrial JC-1 J-aggregate formation, mitochondrial oxygen consumption rates (OCR) and mitochondrial reactive oxygen species (ROS) production, both at basal and palmitate + high glucose-stimulated conditions. ZBED6-induced inhibition of OCR was not rescued by IGF2 addition. ZBED6 reduced levels of the mitochondrial regulator PPAR-γ related coactivator 1 protein (PRC) and bound its promoter/enhancer region. Knockdown of PRC resulted in a lowered OCR.Conclusions/interpretationIt is concluded that ZBED6 is required for normal beta cell replication and also limits excessive beta cell mitochondrial activation in response to an increased functional demand. ZBED6 may act, at least in part, by restricting PRC-mediated mitochondrial activation/ROS production, which may lead to protection against beta cell dysfunction and glucose intolerance in vivo.
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| 9. |
- Welsh, Nils
(författare)
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Are off-target effects of imatinib the key to improving beta-cell function in diabetes?
- 2022
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Ingår i: Upsala Journal of Medical Sciences. - : Upsala Medical Society. - 0300-9734 .- 2000-1967. ; 127
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Forskningsöversikt (refereegranskat)abstract
- The small tyrosine kinase (TK) inhibitor imatinib mesylate (Gleevec, STI571) protects against both type 1 and type 2 diabetes, but as it inhibits many TKs and other proteins, it is not clear by which mechanisms it acts. This present review will focus on the possibility that imatinib acts, at least in part, by improving beta-cell function and survival via off-target effects on beta-cell signaling/metabolic flow events. Particular attention will be given to the possibility that imatinib and other TK inhibitors function as inhibitors of mitochondrial respiration. A better understanding of how imatinib counteracts diabetes will possibly help to clarify the pathogenic role of beta-cell signaling events and mitochondrial function, and hopefully leading to improved treatment of the disease.
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| 10. |
- Younis, Shady, et al.
(författare)
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The importance of the ZBED6-IGF2 axis for metabolic regulation in mouse myoblast cells
- 2020
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Ingår i: The FASEB Journal. - 0892-6638 .- 1530-6860. ; 34:8, s. 10250-10266
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Tidskriftsartikel (refereegranskat)abstract
- The transcription factor ZBED6 acts as a repressor of Igf2 and affects directly or indirectly the transcriptional regulation of thousands of genes. Here, we use gene editing in mouse C2C12 myoblasts and show that ZBED6 regulates Igf2 exclusively through its binding site 5'-GGCTCG-3' in intron 1 of Igf2. Deletion of this motif (Igf2ΔGGCT ) or complete ablation of Zbed6 leads to ~20-fold upregulation of the IGF2 protein. Quantitative proteomics revealed an activation of Ras signaling pathway in both Zbed6-/- and Igf2ΔGGCT myoblasts, and a significant enrichment of mitochondrial membrane proteins among proteins showing altered expression in Zbed6-/- myoblasts. Both Zbed6-/- and Igf2ΔGGCT myoblasts showed a faster growth rate and developed myotube hypertrophy. These cells exhibited an increased O2 consumption rate, due to IGF2 upregulation. Transcriptome analysis revealed ~30% overlap between differentially expressed genes in Zbed6-/- and Igf2ΔGGCT myotubes, with an enrichment of upregulated genes involved in muscle development. In contrast, ZBED6-overexpression in myoblasts led to cell apoptosis, cell cycle arrest, reduced mitochondrial activities, and ceased myoblast differentiation. The similarities in growth and differentiation phenotypes observed in Zbed6-/- and Igf2ΔGGCT myoblasts demonstrates that ZBED6 affects mitochondrial activity and myogenesis largely through its regulation of IGF2 expression. This study adds new insights how the ZBED6-Igf2 axis affects muscle metabolism.
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