| 1. |
- Addinsall, Alex B., et al.
(author)
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Ruxolitinib : A new hope for ventilator-induced diaphragm dysfunction
- 2024
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In: Acta Physiologica. - : John Wiley & Sons. - 1748-1708 .- 1748-1716. ; 240:5
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Journal article (peer-reviewed)abstract
- Aim: Mechanical ventilation (MV) results in diminished diaphragm size and strength, termed ventilator-induced diaphragm dysfunction (VIDD). VID increases dependence, prolongs weaning, and increases discharge mortality rates. The Janus kinase (JAK)/Signal Transducer and Activator of Transcription (STAT) pathway is implicated in VIDD, upregulated following MV. JAK/STAT inhibition alleviates chronic muscle wasting conditions. This study aimed to explore the therapeutic potential of Ruxolitinib, an FDA approved JAK1/2 inhibitor (JI) for the treatment of VIDD. Methods: Rats were subjected to 5 days controlled MV (CMV) with and without daily Ruxolitinib gavage. Muscle fiber size and function were assessed. RNAseq, mitochondrial morphology, respirometry, and mass spectrometry were determined. Results: CMV significantly reduced diaphragm size and specific force by 45% (p < 0.01), associated with a two-fold P-STAT3 upregulation (p < 0.001). CMV disrupted mitochondrial content and reduced the oxygen consumption rate (p < 0.01). Expression of the motor protein myosin was unaffected, however CMV alters myosin function via post-translational modifications (PTMs). Daily administration of JI increased animal survival (40% vs. 87%; p < 0.05), restricted P-STAT3 (p < 0.001), and preserved diaphragm size and specific force. JI was associated with preserved mitochondrial content and respiratory function (p < 0.01), and the reversal or augmentation of myosin deamidation PTMs of the rod and head region. Conclusion: JI preserved diaphragm function, leading to increased survival in an experimental model of VIDD. Functional enhancement was associated with maintenance of mitochondrial content and respiration and the reversal of ventilator-induced PTMs of myosin. These results demonstrate the potential of repurposing Ruxolitinib for treatment of VIDD.
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| 2. |
- Addinsall, Alex B., et al.
(author)
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Ruxolitinib Prevents Ventilator Induced Diaphragm Dysfunction
- 2022
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In: The FASEB Journal. - : John Wiley & Sons. - 0892-6638 .- 1530-6860. ; 36:S1
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Journal article (peer-reviewed)abstract
- Mechanical ventilation (MV), however brief results in the loss of diaphragm muscle mass and strength, termed ventilator induced diaphragm dysfunction (VIDD). VIDD increases dependence, complicates and prolongs weaning and significantly increases discharge mortality rate and health care costs worldwide. The Janus kinase (JAK)/Signal Transducer and Activator of Transcription (STAT) pathway was recently identified as an important signalling pathway implicated in VIDD, upregulated in the diaphragm following MV and limb muslces during critical care. Regulation of STAT3 is imperritve to skeletal muscle mass and function, as STAT3 is required in proper muscle growth and regeneration, while chronic activation of STAT3 is implicated in muscle dysfunction. As JAK/STAT pathway inhibition can restrict the development of chronic muscle wasting conditons, this study aimed to explore the therapeutic potential of Ruxolitinib, an approved JAK1/2 inhibitor for myelofibrosis, for treatment of CIM. We hypothesised Ruxolitinib would reduce loss of muscle mass and function associated with VIDD. Here, rats were subjected to five days controlled MV (CMV) with and without daily Ruxolitinib gavage. Five-days CMV significantly reduced diaphragm muscle size and impaired specific force, which was associated with 2-fold upregulation of P-STAT3, disrupted mitochondrial structure and respiratory function. Expression of the motor protein myosin was not affected, however CMV may alter myosin function through deamidation post translational modification. Ruxolitinib increases five-day survival rate, restored P-STAT3 expression and preserved diaphragm muscle size and specific force. These functional improvements were associated with improved mitochondrial structure, augmented mitochondrial respiratory function and reversal or augmentation of myosin deamidations. These results provide evidence of the preclinical potential of repurposing Ruxolitinib for the treatment of VIDD.
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| 3. |
- Bader, Erik, et al.
(author)
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Identification of proliferative and mature beta-cells in the islets of Langerhans
- 2016
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In: Nature. - : Springer Science and Business Media LLC. - 0028-0836 .- 1476-4687. ; 535:7612, s. 430-
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Journal article (peer-reviewed)abstract
- Insulin-dependent diabetes is a complex multifactorial disorder characterized by loss or dysfunction of beta-cells. Pancreatic beta-cells differ in size, glucose responsiveness, insulin secretion and precursor cell potential(1-5); understanding the mechanisms that underlie this functional heterogeneity might make it possible to develop new regenerative approaches. Here we show that Fltp (also known as Flattop and Cfap126), a Wnt/planar cell polarity (PCP) effector and reporter gene(6), acts as a marker gene that subdivides endocrine cells into two subpopulations and distinguishes proliferation-competent from mature beta-cells with distinct molecular, physiological and ultrastructural features. Genetic lineage tracing revealed that endocrine subpopulations from Fltp-negative and -positive lineages react differently to physiological and pathological changes. The expression of Fltp increases when endocrine cells cluster together to form polarized and mature 3D islet mini-organs(7-9). We show that 3D architecture and Wnt/PCP ligands are sufficient to trigger beta-cell maturation. By contrast, the Wnt/PCP effector Fltp is not necessary for beta-cell development, proliferation or maturation. We conclude that 3D architecture and Wnt/PCP signalling underlie functional beta-cell heterogeneity and induce beta-cell maturation. The identification of Fltp as a marker for endocrine subpopulations sheds light on the molecular underpinnings of islet cell heterogeneity and plasticity and might enable targeting of endocrine subpopulations for the regeneration of functional beta-cell mass in diabetic patients.
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| 4. |
- Johansson, Alina, et al.
(author)
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miR-31 regulates energy metabolism and is suppressed in Tcells from patients with Sjögren's syndrome
- 2019
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In: European Journal of Immunology. - : Wiley. - 0014-2980 .- 1521-4141. ; 49:2, s. 313-322
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Journal article (peer-reviewed)abstract
- Systemic autoimmune diseases are characterized by the overexpression of type I IFN stimulated genes, and accumulating evidence indicate a role for type I IFNs in these diseases. However, the underlying mechanisms for this are still poorly understood. To explore the role of type I IFN regulated miRNAs in systemic autoimmune disease, we characterized cellular expression of miRNAs during both acute and chronic type I IFN responses. We identified a Tcell-specific reduction of miR-31-5p levels, both after intramuscular injection of IFN and in patients with Sjogren's syndrome (SjS). To interrogate the role of miR-31-51p in Tcells we transfected human CD4(+) Tcells with a miR-31-5p inhibitor and performed metabolic measurements. This identified an increase in basal levels of glucose metabolism after inhibition of miR-31-5p. Furthermore, treatment with IFN- also increased the basal levels of human CD4(+) T-cell metabolism. In all, our results suggest that reduced levels of miR-31-5p in Tcells of SjS patients support autoimmune T-cell responses during chronic type I IFN exposure.
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| 5. |
- Kabra, Uma D., et al.
(author)
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Drp1 Overexpression Decreases Insulin Content in Pancreatic MIN6 Cells
- 2022
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In: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 23:20
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Journal article (peer-reviewed)abstract
- Mitochondrial dynamics and bioenergetics are central to glucose-stimulated insulin secretion by pancreatic beta cells. Previously, we demonstrated that a disturbance in glucose-invoked fission impairs insulin secretion by compromising glucose catabolism. Here, we investigated whether the overexpression of mitochondrial fission regulator Drp1 in MIN6 cells can improve or rescue insulin secretion. Although Drp1 overexpression slightly improves the triggering mechanism of insulin secretion of the Drp1-knockdown cells and has no adverse effects on mitochondrial metabolism in wildtype MIN6 cells, the constitutive presence of Drp1 unexpectedly impairs insulin content, which leads to a reduction in the absolute values of secreted insulin. Coherent with previous studies in Drp1-overexpressing muscle cells, we found that the upregulation of ER stress-related genes (BiP, Chop, and Hsp60) possibly impacts insulin production in MIN6 cells. Collectively, we confirm the important role of Drp1 for the energy-coupling of insulin secretion but unravel off-targets effects by Drp1 overexpression on insulin content that warrant caution when manipulating Drp1 in disease therapy.
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| 6. |
- Moruzzi, Noah
(author)
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Interplay between mitochondria, primary cilium, diabetes and its complications
- 2017
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Doctoral thesis (other academic/artistic)abstract
- Diabetes is one of the major health problems of the 21st century. Dysfunction of the insulin secreting pancreatic β-cell together with insulin resistance is central to the pathogenesis of type 2 diabetes mellitus (T2DM). The cause of the disease and the underlying mechanisms linking hyperglycemia to diabetes complications are still unclear. This thesis is focused on two cellular organelles, the mitochondrion and the primary cilium, and their role in the pathophysiological mechanisms of diabetes and its complications. In the first paper, we studied the effect of hyperglycemia on cell biology and energy metabolism in human primary fibroblasts and endothelial cells. Acute hyperglycemia triggered a metabolic switch from mitochondrial respiration to aerobic glycolysis, which was persistent after prolonged exposure together with reduced ATP/ADP ratio without increase in reactive oxygen species (ROS). An acute decrease in mitochondrial transmembrane potential and cellular proliferation with changes in cytoskeletal reorganization was linked to the increased osmotic pressure induced by hyperglycemia. In the second and third papers we investigated the effect of hypoxia, a common feature in diabetes, and hyperoxia in pancreatic islets. Here, we found deleterious effects on mitochondrial content, respiration and glucose-stimulated insulin secretion. Preconditioning with the K+ATP channel opener diazoxide enhanced insulin release, HIF-1α and AMPK activation and improved β-cell survival in response to hypoxia. In the fourth paper, a role for the β-cell primary cilium in diabetes was reported. We found reduced first phase insulin secretion in ciliary defective cells and islets, and impaired glucose tolerance in a ciliopathy mouse model. These results were linked to impaired recruitment of insulin receptor A to the cilium, necessary for proper insulin signaling. Mitochondrial respiration and glucose uptake was unaffected by cilia impairment. Additionally, in vivo evidence of ciliary morphology alteration in the GK rat, a model of T2DM, supported a relationship between ciliary defect and T2DM. Preliminary results show that decreasing intracellular ATP and increasing mitochondrial ROS production impaired cilia morphology and/or number in two different cell types. Further, cilia were decreased in number with altered morphology in the kidneys of a mouse model of T2DM with diabetic nephropathy, characterized by increased ROS and altered mitochondrial metabolism. Finally, a reduction of 60-80% in mtDNA content (reported in diabetes) did not affect mitochondrial metabolism, respiration and energy production in two different cell types. In summary, mitochondrial dysfunctions during diabetes and its complications are most probably due to a combination of hyperglycemia and other factors such as hypoxia, depending on the cells and tissues involved. A proper ciliary/basal body function is necessary for insulin release and signaling in β-cell. Cilia morphology and number can be affected by mitochondrial dysfunction/ROS and thus related to diabetic complications such as diabetic nephropathy.
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| 7. |
- Olesen, Kim, et al.
(author)
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Diversity of respiratory parameters and metabolic adaptation to low oxygen tension in mesenchymal stromal cells
- 2022
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In: Metabolism Open. - : Elsevier. - 2589-9368. ; 13:March
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Journal article (peer-reviewed)abstract
- ObjectiveCell metabolism has been shown to play an active role in regulation of stemness and fate decision. In order to identify favorable culture conditions for mesenchymal stromal cells (MSCs) prior to transplantation, this study aimed to characterize the metabolic function of MSCs from different developmental stages in response to different oxygen tension during expansion.Materials and methodsWe cultured human fetal cardiac MSCs and human adult bone-marrow MSCs for a week under hypoxia (3% O2) and normoxia (20% O2). We performed mitochondrial characterization and assessed oxygen consumption- and extracellular acidification-rates (OCR and ECAR) in addition to oxygen-sensitive respiration and mitochondrial complex activities, using both the Seahorse and Oroboros systems.ResultsAdult and fetal MSCs displayed similar basal respiration and mitochondrial amount, however fetal MSCs had lower spare respiratory capacity and apparent coupling efficiency. Fetal MSCs expanded in either hypoxia or normoxia demonstrated similar acidification rates, while adult MSCs downregulated their aerobic glycolysis in normoxia. Acute decrease in oxygen tension caused a higher respiratory inhibition in adult compared to fetal MSCs. In both sources of MSCs, minor changes in complex activities in normoxic and hypoxic cultures were found.ConclusionsIn contrast to adult MSCs, fetal MSCs displayed similar respiration and aerobic glycolysis at different O2 culture concentrations during expansion. Adult MSCs adjusted their respiration to glycolytic activities, depending on the culture conditions thus displaying a more mature metabolic function. These findings are relevant for establishing optimal in vitro culturing conditions, with the aim to maximize engraftment and therapeutic outcome.
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| 8. |
- Sabatier, Pierre, et al.
(author)
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An integrative proteomics method identifies a regulator of translation during stem cell maintenance and differentiation
- 2021
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In: Nature Communications. - : Springer Nature. - 2041-1723. ; 12
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Journal article (peer-reviewed)abstract
- To characterize molecular changes during cell type transitions, the authors develop a method to simultaneously measure protein expression and thermal stability changes. They apply this approach to study differences between human pluripotent stem cells, their progenies, parental and allogeneic cells. Detailed characterization of cell type transitions is essential for cell biology in general and particularly for the development of stem cell-based therapies in regenerative medicine. To systematically study such transitions, we introduce a method that simultaneously measures protein expression and thermal stability changes in cells and provide the web-based visualization tool ProteoTracker. We apply our method to study differences between human pluripotent stem cells and several cell types including their parental cell line and differentiated progeny. We detect alterations of protein properties in numerous cellular pathways and components including ribosome biogenesis and demonstrate that modulation of ribosome maturation through SBDS protein can be helpful for manipulating cell stemness in vitro. Using our integrative proteomics approach and the web-based tool, we uncover a molecular basis for the uncoupling of robust transcription from parsimonious translation in stem cells and propose a method for maintaining pluripotency in vitro.
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| 9. |
- Saghaleyni, Rasool, 1987, et al.
(author)
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Enhanced metabolism and negative regulation of ER stress support higher erythropoietin production in HEK293 cells
- 2022
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In: Cell Reports. - : Elsevier BV. - 2211-1247. ; 39:11
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Journal article (peer-reviewed)abstract
- Recombinant protein production can cause severe stress on cellular metabolism, resulting in limited titer and product quality. To investigate cellular and metabolic characteristics associated with these limitations, we compare HEK293 clones producing either erythropoietin (EPO) (secretory) or GFP (non-secretory) protein at different rates. Transcriptomic and functional analyses indicate significantly higher metabolism and oxidative phosphorylation in EPO producers compared with parental and GFP cells. In addition, ribosomal genes exhibit specific expression patterns depending on the recombinant protein and the production rate. In a clone displaying a dramatically increased EPO secretion, we detect higher gene expression related to negative regulation of endoplasmic reticulum (ER) stress, including upregulation of ATF6B, which aids EPO production in a subset of clones by overexpression or small interfering RNA (siRNA) knockdown. Our results offer potential target pathways and genes for further development of the secretory power in mammalian cell factories.
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| 10. |
- Valladolid-Acebes, Ismael, et al.
(author)
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Lowering apolipoprotein CIII protects against high-fat diet-induced metabolic derangements
- 2021
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In: Science Advances. - : American Association for the Advancement of Science. - 2375-2548. ; 7:11
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Journal article (peer-reviewed)abstract
- Increased levels of apolipoprotein CIII (apoCIII), a key regulator of lipid metabolism, result in obesity-related metabolic derangements. We investigated mechanistically whether lowering or preventing high-fat diet (HFD)-induced increase in apoCIII protects against the detrimental metabolic consequences. Mice, first fed HFD for 10 weeks and thereafter also given an antisense (ASO) to lower apoCIII, already showed reduced levels of apoCIII and metabolic improvements after 4 weeks, despite maintained obesity. Prolonged ASO treatment reversed the metabolic phenotype due to increased lipase activity and receptor-mediated hepatic uptake of lipids. Fatty acids were transferred to the ketogenic pathway, and ketones were used in brown adipose tissue (BAT). This resulted in no fat accumulation and preserved morphology and function of liver and BAT. If ASO treatment started simultaneously with the HFD, mice remained lean and metabolically healthy. Thus, lowering apoCIII protects against and reverses the HFD-induced metabolic phenotype by promoting physiological insulin sensitivity.
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