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Toll-like receptor 3-mediated inflammation by p38 is enhanced by endothelial nitric oxide synthase knockdown
BACKGROUND: Vascular dysfunction is commonly seen during severe viral infections. Endothelial nitric oxide synthase (eNOS), has been postulated to play an important role in regulating vascular homeostasis as well as propagation of the inflammatory reaction. We hypothesized that the loss of eNOS woul...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2019
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6466662/ https://www.ncbi.nlm.nih.gov/pubmed/30987646 http://dx.doi.org/10.1186/s12964-019-0345-3 |
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author | Koch, Stephen R. Choi, Hyehun Mace, Eric H. Stark, Ryan J. |
author_facet | Koch, Stephen R. Choi, Hyehun Mace, Eric H. Stark, Ryan J. |
author_sort | Koch, Stephen R. |
collection | PubMed |
description | BACKGROUND: Vascular dysfunction is commonly seen during severe viral infections. Endothelial nitric oxide synthase (eNOS), has been postulated to play an important role in regulating vascular homeostasis as well as propagation of the inflammatory reaction. We hypothesized that the loss of eNOS would negatively impact toll-like receptor 3 (TLR3) signaling and worsen vascular function to viral challenge. METHODS: Human microvascular endothelial cells (HMVECs) were exposed to either control or eNOS siRNA and then treated with Poly I:C, a TLR3 agonist and mimicker of dsRNA viruses. Cells were assessed for protein-protein associations, cytokine and chemokine analysis as well as transendothelial electrical resistance (TEER) as a surrogate of permeability. RESULTS: HMVECs that had reduced eNOS expression had a significantly elevated increase in IL-6, IL-8 and IP-10 production after Poly I:C. In addition, the knockdown of eNOS enhanced the change in TEER after Poly I:C stimulation. Western blot analysis showed enhanced phosphorylation of p38 in sieNOS treated cells with Poly I:C compared to siControl cells. Proximity ligation assays further demonstrated direct eNOS-p38 protein-protein interactions. The addition of the p38 inhibitor, SB203580, in eNOS knockdown cells reduced both cytokine production after Poly I:C, and as well as mitigated the reduction in TEER, suggesting a direct link between eNOS and p38 in TLR3 signaling. CONCLUSIONS: These results suggest that reduction of eNOS increases TLR3-mediated inflammation in human endothelial cells in a p38-dependent manner. This finding has important implications for understanding the pathogenesis of severe viral infections and the associated vascular dysfunction. [Image: see text] ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12964-019-0345-3) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-6466662 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-64666622019-04-22 Toll-like receptor 3-mediated inflammation by p38 is enhanced by endothelial nitric oxide synthase knockdown Koch, Stephen R. Choi, Hyehun Mace, Eric H. Stark, Ryan J. Cell Commun Signal Research BACKGROUND: Vascular dysfunction is commonly seen during severe viral infections. Endothelial nitric oxide synthase (eNOS), has been postulated to play an important role in regulating vascular homeostasis as well as propagation of the inflammatory reaction. We hypothesized that the loss of eNOS would negatively impact toll-like receptor 3 (TLR3) signaling and worsen vascular function to viral challenge. METHODS: Human microvascular endothelial cells (HMVECs) were exposed to either control or eNOS siRNA and then treated with Poly I:C, a TLR3 agonist and mimicker of dsRNA viruses. Cells were assessed for protein-protein associations, cytokine and chemokine analysis as well as transendothelial electrical resistance (TEER) as a surrogate of permeability. RESULTS: HMVECs that had reduced eNOS expression had a significantly elevated increase in IL-6, IL-8 and IP-10 production after Poly I:C. In addition, the knockdown of eNOS enhanced the change in TEER after Poly I:C stimulation. Western blot analysis showed enhanced phosphorylation of p38 in sieNOS treated cells with Poly I:C compared to siControl cells. Proximity ligation assays further demonstrated direct eNOS-p38 protein-protein interactions. The addition of the p38 inhibitor, SB203580, in eNOS knockdown cells reduced both cytokine production after Poly I:C, and as well as mitigated the reduction in TEER, suggesting a direct link between eNOS and p38 in TLR3 signaling. CONCLUSIONS: These results suggest that reduction of eNOS increases TLR3-mediated inflammation in human endothelial cells in a p38-dependent manner. This finding has important implications for understanding the pathogenesis of severe viral infections and the associated vascular dysfunction. [Image: see text] ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12964-019-0345-3) contains supplementary material, which is available to authorized users. BioMed Central 2019-04-15 /pmc/articles/PMC6466662/ /pubmed/30987646 http://dx.doi.org/10.1186/s12964-019-0345-3 Text en © The Author(s). 2019 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 | Research Koch, Stephen R. Choi, Hyehun Mace, Eric H. Stark, Ryan J. Toll-like receptor 3-mediated inflammation by p38 is enhanced by endothelial nitric oxide synthase knockdown |
title | Toll-like receptor 3-mediated inflammation by p38 is enhanced by endothelial nitric oxide synthase knockdown |
title_full | Toll-like receptor 3-mediated inflammation by p38 is enhanced by endothelial nitric oxide synthase knockdown |
title_fullStr | Toll-like receptor 3-mediated inflammation by p38 is enhanced by endothelial nitric oxide synthase knockdown |
title_full_unstemmed | Toll-like receptor 3-mediated inflammation by p38 is enhanced by endothelial nitric oxide synthase knockdown |
title_short | Toll-like receptor 3-mediated inflammation by p38 is enhanced by endothelial nitric oxide synthase knockdown |
title_sort | toll-like receptor 3-mediated inflammation by p38 is enhanced by endothelial nitric oxide synthase knockdown |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6466662/ https://www.ncbi.nlm.nih.gov/pubmed/30987646 http://dx.doi.org/10.1186/s12964-019-0345-3 |
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