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Inhibition of MicroRNA‐155 Supports Endothelial Tight Junction Integrity Following Oxygen‐Glucose Deprivation
BACKGROUND: Brain microvascular endothelial cells form a highly selective blood brain barrier regulated by the endothelial tight junctions. Cerebral ischemia selectively targets tight junction protein complexes, which leads to significant damage to cerebral microvasculature. Short noncoding molecule...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
John Wiley and Sons Inc.
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6064884/ https://www.ncbi.nlm.nih.gov/pubmed/29945912 http://dx.doi.org/10.1161/JAHA.118.009244 |
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author | Pena‐Philippides, Juan Carlos Gardiner, Amy Sabrina Caballero‐Garrido, Ernesto Pan, Rong Zhu, Yiliang Roitbak, Tamara |
author_facet | Pena‐Philippides, Juan Carlos Gardiner, Amy Sabrina Caballero‐Garrido, Ernesto Pan, Rong Zhu, Yiliang Roitbak, Tamara |
author_sort | Pena‐Philippides, Juan Carlos |
collection | PubMed |
description | BACKGROUND: Brain microvascular endothelial cells form a highly selective blood brain barrier regulated by the endothelial tight junctions. Cerebral ischemia selectively targets tight junction protein complexes, which leads to significant damage to cerebral microvasculature. Short noncoding molecules called microRNAs are implicated in the regulation of various pathological states, including endothelial barrier dysfunction. In the present study, we investigated the influence of microRNA‐155 (miR‐155) on the barrier characteristics of human primary brain microvascular endothelial cells (HBMECs). METHODS AND RESULTS: Oxygen‐glucose deprivation was used as an in vitro model of ischemic stroke. HBMECs were subjected to 3 hours of oxygen‐glucose deprivation, followed by transfections with miR‐155 inhibitor, mimic, or appropriate control oligonucleotides. Intact normoxia control HBMECs and 4 oxygen‐glucose deprivation–treated groups of cells transfected with appropriate nucleotide were subjected to endothelial monolayer electrical resistance and permeability assays, cell viability assay, assessment of NO and human cytokine/chemokine release, immunofluorescence microscopy, Western blot, and polymerase chain reaction analyses. Assessment of endothelial resistance and permeability demonstrated that miR‐155 inhibition improved HBMECs monolayer integrity. In addition, miR‐155 inhibition significantly increased the levels of major tight junction proteins claudin‐1 and zonula occludens protein‐1, while its overexpression reduced these levels. Immunoprecipitation and colocalization analyses detected that miR‐155 inhibition supported the association between zonula occludens protein‐1 and claudin‐1 and their stabilization at the HBMEC membrane. Luciferase reporter assay verified that claudin‐1 is directly targeted by miR‐155. CONCLUSIONS: Based on these results, we conclude that miR‐155 inhibition–induced strengthening of endothelial tight junctions after oxygen‐glucose deprivation is mediated via its direct target protein claudin‐1. |
format | Online Article Text |
id | pubmed-6064884 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | John Wiley and Sons Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-60648842018-08-09 Inhibition of MicroRNA‐155 Supports Endothelial Tight Junction Integrity Following Oxygen‐Glucose Deprivation Pena‐Philippides, Juan Carlos Gardiner, Amy Sabrina Caballero‐Garrido, Ernesto Pan, Rong Zhu, Yiliang Roitbak, Tamara J Am Heart Assoc Original Research BACKGROUND: Brain microvascular endothelial cells form a highly selective blood brain barrier regulated by the endothelial tight junctions. Cerebral ischemia selectively targets tight junction protein complexes, which leads to significant damage to cerebral microvasculature. Short noncoding molecules called microRNAs are implicated in the regulation of various pathological states, including endothelial barrier dysfunction. In the present study, we investigated the influence of microRNA‐155 (miR‐155) on the barrier characteristics of human primary brain microvascular endothelial cells (HBMECs). METHODS AND RESULTS: Oxygen‐glucose deprivation was used as an in vitro model of ischemic stroke. HBMECs were subjected to 3 hours of oxygen‐glucose deprivation, followed by transfections with miR‐155 inhibitor, mimic, or appropriate control oligonucleotides. Intact normoxia control HBMECs and 4 oxygen‐glucose deprivation–treated groups of cells transfected with appropriate nucleotide were subjected to endothelial monolayer electrical resistance and permeability assays, cell viability assay, assessment of NO and human cytokine/chemokine release, immunofluorescence microscopy, Western blot, and polymerase chain reaction analyses. Assessment of endothelial resistance and permeability demonstrated that miR‐155 inhibition improved HBMECs monolayer integrity. In addition, miR‐155 inhibition significantly increased the levels of major tight junction proteins claudin‐1 and zonula occludens protein‐1, while its overexpression reduced these levels. Immunoprecipitation and colocalization analyses detected that miR‐155 inhibition supported the association between zonula occludens protein‐1 and claudin‐1 and their stabilization at the HBMEC membrane. Luciferase reporter assay verified that claudin‐1 is directly targeted by miR‐155. CONCLUSIONS: Based on these results, we conclude that miR‐155 inhibition–induced strengthening of endothelial tight junctions after oxygen‐glucose deprivation is mediated via its direct target protein claudin‐1. John Wiley and Sons Inc. 2018-06-26 /pmc/articles/PMC6064884/ /pubmed/29945912 http://dx.doi.org/10.1161/JAHA.118.009244 Text en © 2018 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley. This is an open access article under the terms of the http://creativecommons.org/licenses/by-nc/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes. |
spellingShingle | Original Research Pena‐Philippides, Juan Carlos Gardiner, Amy Sabrina Caballero‐Garrido, Ernesto Pan, Rong Zhu, Yiliang Roitbak, Tamara Inhibition of MicroRNA‐155 Supports Endothelial Tight Junction Integrity Following Oxygen‐Glucose Deprivation |
title | Inhibition of MicroRNA‐155 Supports Endothelial Tight Junction Integrity Following Oxygen‐Glucose Deprivation |
title_full | Inhibition of MicroRNA‐155 Supports Endothelial Tight Junction Integrity Following Oxygen‐Glucose Deprivation |
title_fullStr | Inhibition of MicroRNA‐155 Supports Endothelial Tight Junction Integrity Following Oxygen‐Glucose Deprivation |
title_full_unstemmed | Inhibition of MicroRNA‐155 Supports Endothelial Tight Junction Integrity Following Oxygen‐Glucose Deprivation |
title_short | Inhibition of MicroRNA‐155 Supports Endothelial Tight Junction Integrity Following Oxygen‐Glucose Deprivation |
title_sort | inhibition of microrna‐155 supports endothelial tight junction integrity following oxygen‐glucose deprivation |
topic | Original Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6064884/ https://www.ncbi.nlm.nih.gov/pubmed/29945912 http://dx.doi.org/10.1161/JAHA.118.009244 |
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