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Oscillating glucose induces microRNA-185 and impairs an efficient antioxidant response in human endothelial cells
BACKGROUND: Intracellular antioxidant response to high glucose is mediated by Cu/Mn-superoxide dismutases (SOD-1/SOD-2), catalase (CAT) and glutathione peroxidases (GPx), particularly glutathione peroxidase-1 (GPx-1). Although oscillating glucose can induce a more deleterious effect than high glucos...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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BioMed Central
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4852407/ https://www.ncbi.nlm.nih.gov/pubmed/27137793 http://dx.doi.org/10.1186/s12933-016-0390-9 |
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author | La Sala, Lucia Cattaneo, Monica De Nigris, Valeria Pujadas, Gemma Testa, Roberto Bonfigli, Anna R. Genovese, Stefano Ceriello, Antonio |
author_facet | La Sala, Lucia Cattaneo, Monica De Nigris, Valeria Pujadas, Gemma Testa, Roberto Bonfigli, Anna R. Genovese, Stefano Ceriello, Antonio |
author_sort | La Sala, Lucia |
collection | PubMed |
description | BACKGROUND: Intracellular antioxidant response to high glucose is mediated by Cu/Mn-superoxide dismutases (SOD-1/SOD-2), catalase (CAT) and glutathione peroxidases (GPx), particularly glutathione peroxidase-1 (GPx-1). Although oscillating glucose can induce a more deleterious effect than high glucose on endothelial cells, the mechanism by which oscillating glucose exerts its dangerous effects is incompletely understood; however, the involvement of oxidative damage has been generally accepted. In this study we sought to determine whether oscillating glucose differentially modulates antioxidant response, and to elucidate the potential regulatory mechanisms exerted by the microRNA-185 (miR-185). METHODS: Human endothelial cells were exposed for 1 week to constant and oscillating high glucose. SOD-1, SOD-2, CAT and GPx-1, as well as two markers of oxidative stress [8-hydroxy-2′-deoxyguanosine (8-OHdG) and the phosphorylated form of H2AX (γ-H2AX)] were measured at the end of the experiment. Intracellular miR-185 was measured and loss-of function assays were performed in HUVEC. Bioinformatic tool was used to predict the link between miR-185 on 3′UTR of GPx-1 gene. Luciferase assay was performed to confirm the binding on HUVEC. RESULTS: After exposure to constant high glucose SOD-1 and GPx-1 increased, while in oscillating glucose SOD-1 increased and GPx-1 did not. SOD-2 and CAT remained unchanged under both conditions. A critical involvement of oscillating glucose-induced miR-185 in the dysregulation of endogenous GPx-1 was found. Computational analyses predict GPx-1 as miR-185′s target. HUVEC cultures were used to confirm glucose’s causal role on the expression of miR-185, its target mRNA and protein and finally the activation of antioxidant response. In vitro luciferase assays confirmed computational predictions targeting of miR-185 on 3′-UTR of GPx-1 mRNA. Knockdown of miR-185, using anti-miR-185 inhibitor, was accompanied by a significant upregulation of GPx-1 in oscillating glucose. 8-OHdG and γ-H2AX increased more in oscillating glucose than in constant high glucose. CONCLUSIONS: Glucose oscillations may exert more deleterious effects on the endothelium than high glucose, likely due to an impaired response of GPx-1, coupled by the upregulation of miR-185. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12933-016-0390-9) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-4852407 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-48524072016-05-03 Oscillating glucose induces microRNA-185 and impairs an efficient antioxidant response in human endothelial cells La Sala, Lucia Cattaneo, Monica De Nigris, Valeria Pujadas, Gemma Testa, Roberto Bonfigli, Anna R. Genovese, Stefano Ceriello, Antonio Cardiovasc Diabetol Original Investigation BACKGROUND: Intracellular antioxidant response to high glucose is mediated by Cu/Mn-superoxide dismutases (SOD-1/SOD-2), catalase (CAT) and glutathione peroxidases (GPx), particularly glutathione peroxidase-1 (GPx-1). Although oscillating glucose can induce a more deleterious effect than high glucose on endothelial cells, the mechanism by which oscillating glucose exerts its dangerous effects is incompletely understood; however, the involvement of oxidative damage has been generally accepted. In this study we sought to determine whether oscillating glucose differentially modulates antioxidant response, and to elucidate the potential regulatory mechanisms exerted by the microRNA-185 (miR-185). METHODS: Human endothelial cells were exposed for 1 week to constant and oscillating high glucose. SOD-1, SOD-2, CAT and GPx-1, as well as two markers of oxidative stress [8-hydroxy-2′-deoxyguanosine (8-OHdG) and the phosphorylated form of H2AX (γ-H2AX)] were measured at the end of the experiment. Intracellular miR-185 was measured and loss-of function assays were performed in HUVEC. Bioinformatic tool was used to predict the link between miR-185 on 3′UTR of GPx-1 gene. Luciferase assay was performed to confirm the binding on HUVEC. RESULTS: After exposure to constant high glucose SOD-1 and GPx-1 increased, while in oscillating glucose SOD-1 increased and GPx-1 did not. SOD-2 and CAT remained unchanged under both conditions. A critical involvement of oscillating glucose-induced miR-185 in the dysregulation of endogenous GPx-1 was found. Computational analyses predict GPx-1 as miR-185′s target. HUVEC cultures were used to confirm glucose’s causal role on the expression of miR-185, its target mRNA and protein and finally the activation of antioxidant response. In vitro luciferase assays confirmed computational predictions targeting of miR-185 on 3′-UTR of GPx-1 mRNA. Knockdown of miR-185, using anti-miR-185 inhibitor, was accompanied by a significant upregulation of GPx-1 in oscillating glucose. 8-OHdG and γ-H2AX increased more in oscillating glucose than in constant high glucose. CONCLUSIONS: Glucose oscillations may exert more deleterious effects on the endothelium than high glucose, likely due to an impaired response of GPx-1, coupled by the upregulation of miR-185. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12933-016-0390-9) contains supplementary material, which is available to authorized users. BioMed Central 2016-04-30 /pmc/articles/PMC4852407/ /pubmed/27137793 http://dx.doi.org/10.1186/s12933-016-0390-9 Text en © La Sala et al. 2016 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Original Investigation La Sala, Lucia Cattaneo, Monica De Nigris, Valeria Pujadas, Gemma Testa, Roberto Bonfigli, Anna R. Genovese, Stefano Ceriello, Antonio Oscillating glucose induces microRNA-185 and impairs an efficient antioxidant response in human endothelial cells |
title | Oscillating glucose induces microRNA-185 and impairs an efficient antioxidant response in human endothelial cells |
title_full | Oscillating glucose induces microRNA-185 and impairs an efficient antioxidant response in human endothelial cells |
title_fullStr | Oscillating glucose induces microRNA-185 and impairs an efficient antioxidant response in human endothelial cells |
title_full_unstemmed | Oscillating glucose induces microRNA-185 and impairs an efficient antioxidant response in human endothelial cells |
title_short | Oscillating glucose induces microRNA-185 and impairs an efficient antioxidant response in human endothelial cells |
title_sort | oscillating glucose induces microrna-185 and impairs an efficient antioxidant response in human endothelial cells |
topic | Original Investigation |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4852407/ https://www.ncbi.nlm.nih.gov/pubmed/27137793 http://dx.doi.org/10.1186/s12933-016-0390-9 |
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