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Extracellular vesicles derived from endothelial cells modulate macrophage phenotype in vitro
Extracellular vesicles (EVs) mediate cell-to-cell communication by horizontally transferring biological materials from host cells to target cells. During exposure to pathogens, pathogen-associated molecular patterns (e.g., lipopolysaccharide, LPS) get in contact with endothelial cells and stimulate...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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BioMed Central
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10634087/ https://www.ncbi.nlm.nih.gov/pubmed/37946271 http://dx.doi.org/10.1186/s40001-023-01427-6 |
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author | He, Zhizhen Greven, Johannes Shi, Yulong Qin, Kang Zhao, Qun Zhang, Xing Buhl, Eva Miriam Eschweiler, Jörg Hildebrand, Frank Balmayor, Elizabeth Rosado |
author_facet | He, Zhizhen Greven, Johannes Shi, Yulong Qin, Kang Zhao, Qun Zhang, Xing Buhl, Eva Miriam Eschweiler, Jörg Hildebrand, Frank Balmayor, Elizabeth Rosado |
author_sort | He, Zhizhen |
collection | PubMed |
description | Extracellular vesicles (EVs) mediate cell-to-cell communication by horizontally transferring biological materials from host cells to target cells. During exposure to pathogens, pathogen-associated molecular patterns (e.g., lipopolysaccharide, LPS) get in contact with endothelial cells and stimulate the secretion of endothelial cell-derived EVs (E-EVs). The triggered EVs secretion is known to have a modulating influence on the EVs-receiving cells. Macrophages, a major component of innate immunity, are polarized upon receiving external inflammatory stimuli, in which toll-like receptor4 (TLR4)—nuclear factor kappa B (NFκB) pathway plays a key role. However, the functions of LPS-induced E-EVs (E(LPS)-EVs) in modulating macrophage phenotype and activation remain elusive. We collected the EVs from quiescent endothelial cells (E(Nor)-EVs) and E(LPS)-EVs to detect their stimulatory role on NR8383 macrophages. Isolated EVs were characterized by transmission electron microscopy (TEM), western blot assay, and nanoparticle tracking analysis (NTA). NR8383 macrophages were stimulated with ELPS-EVs, ENor-EVs, or PBS for 24 h. Hereafter, the uptake of EVs by the macrophages was investigated. Upon EVs stimulation, cellular viability was determined by MTT assay, while macrophage phenotype was analyzed by flow cytometry and immunofluorescence analysis. Furthermore, a western blot assay was conducted to evaluate the potentially involved TLR4–NFκB pathway. Interestingly, upon exposure to LPS, endothelial cells secreted significantly higher amounts of EVs (i.e., E(LPS)-EVs) when compared to quiescent cells or cells in PBS. The E(LPS)-EVs were also better internalized by NR8383 macrophages than E(Nor)-EVs. The cellular viability of E(LPS)-EVs-treated macrophages was 1.2 times higher than those in the E(Nor)-EVs and PBS groups. In addition, E(LPS)-EVs modulated NR8383 macrophages towards a proinflammatory macrophage M1-like phenotype. This was indicated by the significantly upregulated expressions of proinflammatory macrophage biomarkers CD86 and inducible nitric oxide synthase (iNOS) observed in E(LPS)-EVs-treated macrophages. The TLR4–NFκB signaling pathway was substantially activated in E(LPS)-EVs-treated macrophages, indicated by the elevated expressions of makers TLR4 and phosphorylated form of nuclear factor kappa B p65 subunit (p-NFκBp65). Overall, our results indicate that E-EVs play a crucial role in macrophage phenotype modulation under inflammatory conditions. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s40001-023-01427-6. |
format | Online Article Text |
id | pubmed-10634087 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-106340872023-11-10 Extracellular vesicles derived from endothelial cells modulate macrophage phenotype in vitro He, Zhizhen Greven, Johannes Shi, Yulong Qin, Kang Zhao, Qun Zhang, Xing Buhl, Eva Miriam Eschweiler, Jörg Hildebrand, Frank Balmayor, Elizabeth Rosado Eur J Med Res Research Extracellular vesicles (EVs) mediate cell-to-cell communication by horizontally transferring biological materials from host cells to target cells. During exposure to pathogens, pathogen-associated molecular patterns (e.g., lipopolysaccharide, LPS) get in contact with endothelial cells and stimulate the secretion of endothelial cell-derived EVs (E-EVs). The triggered EVs secretion is known to have a modulating influence on the EVs-receiving cells. Macrophages, a major component of innate immunity, are polarized upon receiving external inflammatory stimuli, in which toll-like receptor4 (TLR4)—nuclear factor kappa B (NFκB) pathway plays a key role. However, the functions of LPS-induced E-EVs (E(LPS)-EVs) in modulating macrophage phenotype and activation remain elusive. We collected the EVs from quiescent endothelial cells (E(Nor)-EVs) and E(LPS)-EVs to detect their stimulatory role on NR8383 macrophages. Isolated EVs were characterized by transmission electron microscopy (TEM), western blot assay, and nanoparticle tracking analysis (NTA). NR8383 macrophages were stimulated with ELPS-EVs, ENor-EVs, or PBS for 24 h. Hereafter, the uptake of EVs by the macrophages was investigated. Upon EVs stimulation, cellular viability was determined by MTT assay, while macrophage phenotype was analyzed by flow cytometry and immunofluorescence analysis. Furthermore, a western blot assay was conducted to evaluate the potentially involved TLR4–NFκB pathway. Interestingly, upon exposure to LPS, endothelial cells secreted significantly higher amounts of EVs (i.e., E(LPS)-EVs) when compared to quiescent cells or cells in PBS. The E(LPS)-EVs were also better internalized by NR8383 macrophages than E(Nor)-EVs. The cellular viability of E(LPS)-EVs-treated macrophages was 1.2 times higher than those in the E(Nor)-EVs and PBS groups. In addition, E(LPS)-EVs modulated NR8383 macrophages towards a proinflammatory macrophage M1-like phenotype. This was indicated by the significantly upregulated expressions of proinflammatory macrophage biomarkers CD86 and inducible nitric oxide synthase (iNOS) observed in E(LPS)-EVs-treated macrophages. The TLR4–NFκB signaling pathway was substantially activated in E(LPS)-EVs-treated macrophages, indicated by the elevated expressions of makers TLR4 and phosphorylated form of nuclear factor kappa B p65 subunit (p-NFκBp65). Overall, our results indicate that E-EVs play a crucial role in macrophage phenotype modulation under inflammatory conditions. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s40001-023-01427-6. BioMed Central 2023-11-09 /pmc/articles/PMC10634087/ /pubmed/37946271 http://dx.doi.org/10.1186/s40001-023-01427-6 Text en © The Author(s) 2023 https://creativecommons.org/licenses/by/4.0/Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/ (https://creativecommons.org/publicdomain/zero/1.0/) ) applies to the data made available in this article, unless otherwise stated in a credit line to the data. |
spellingShingle | Research He, Zhizhen Greven, Johannes Shi, Yulong Qin, Kang Zhao, Qun Zhang, Xing Buhl, Eva Miriam Eschweiler, Jörg Hildebrand, Frank Balmayor, Elizabeth Rosado Extracellular vesicles derived from endothelial cells modulate macrophage phenotype in vitro |
title | Extracellular vesicles derived from endothelial cells modulate macrophage phenotype in vitro |
title_full | Extracellular vesicles derived from endothelial cells modulate macrophage phenotype in vitro |
title_fullStr | Extracellular vesicles derived from endothelial cells modulate macrophage phenotype in vitro |
title_full_unstemmed | Extracellular vesicles derived from endothelial cells modulate macrophage phenotype in vitro |
title_short | Extracellular vesicles derived from endothelial cells modulate macrophage phenotype in vitro |
title_sort | extracellular vesicles derived from endothelial cells modulate macrophage phenotype in vitro |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10634087/ https://www.ncbi.nlm.nih.gov/pubmed/37946271 http://dx.doi.org/10.1186/s40001-023-01427-6 |
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