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GM-CSF increases LPS-induced production of proinflammatory mediators via upregulation of TLR4 and CD14 in murine microglia
BACKGROUND: Microglia are resident macrophage-like cells in the central nervous system (CNS) and cause innate immune responses via the LPS receptors, Toll-like receptor (TLR) 4 and CD14, in a variety of neuroinflammatory disorders including bacterial infection, Alzheimer’s disease, and amyotrophic l...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2012
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3565988/ https://www.ncbi.nlm.nih.gov/pubmed/23234315 http://dx.doi.org/10.1186/1742-2094-9-268 |
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author | Parajuli, Bijay Sonobe, Yoshifumi Kawanokuchi, Jun Doi, Yukiko Noda, Mariko Takeuchi, Hideyuki Mizuno, Tetsuya Suzumura, Akio |
author_facet | Parajuli, Bijay Sonobe, Yoshifumi Kawanokuchi, Jun Doi, Yukiko Noda, Mariko Takeuchi, Hideyuki Mizuno, Tetsuya Suzumura, Akio |
author_sort | Parajuli, Bijay |
collection | PubMed |
description | BACKGROUND: Microglia are resident macrophage-like cells in the central nervous system (CNS) and cause innate immune responses via the LPS receptors, Toll-like receptor (TLR) 4 and CD14, in a variety of neuroinflammatory disorders including bacterial infection, Alzheimer’s disease, and amyotrophic lateral sclerosis. Granulocyte macrophage-colony stimulating factor (GM-CSF) activates microglia and induces inflammatory responses via binding to GM-CSF receptor complex composed of two different subunit GM-CSF receptor α (GM-CSFRα) and common β chain (βc). GM-CSF has been shown to be associated with neuroinflammatory responses in multiple sclerosis and Alzheimer’s disease. However, the mechanisms how GM-CSF promotes neuroinflammation still remain unclear. METHODS: Microglia were stimulated with 20 ng/ml GM-CSF and the levels of TLR4 and CD14 expression were evaluated by RT-PCR and flowcytometry. LPS binding was analyzed by flowcytometry. GM-CSF receptor complex was analyzed by immunocytechemistry. The levels of IL-1β, IL-6 and TNF-α in culture supernatant of GM-CSF-stimulated microglia and NF-κB nuclear translocation were determined by ELISA. Production of nitric oxide (NO) was measured by the Griess method. The levels of p-ERK1/2, ERK1/2, p-p38 and p38 were assessed by Western blotting. Statistically significant differences between experimental groups were determined by one-way ANOVA followed by Tukey test for multiple comparisons. RESULTS: GM-CSF receptor complex was expressed in microglia. GM-CSF enhanced TLR4 and CD14 expressions in microglia and subsequent LPS-binding to the cell surface. In addition, GM-CSF priming increased LPS-induced NF-κB nuclear translocation and production of IL-1β, IL-6, TNF-α and NO by microglia. GM-CSF upregulated the levels of p-ERK1/2 and p-p38, suggesting that induction of TLR4 and CD14 expression by GM-CSF was mediated through ERK1/2 and p38, respectively. CONCLUSIONS: These results suggest that GM-CSF upregulates TLR4 and CD14 expression in microglia through ERK1/2 and p38, respectively, and thus promotes the LPS receptor-mediated inflammation in the CNS. |
format | Online Article Text |
id | pubmed-3565988 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2012 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-35659882013-02-11 GM-CSF increases LPS-induced production of proinflammatory mediators via upregulation of TLR4 and CD14 in murine microglia Parajuli, Bijay Sonobe, Yoshifumi Kawanokuchi, Jun Doi, Yukiko Noda, Mariko Takeuchi, Hideyuki Mizuno, Tetsuya Suzumura, Akio J Neuroinflammation Research BACKGROUND: Microglia are resident macrophage-like cells in the central nervous system (CNS) and cause innate immune responses via the LPS receptors, Toll-like receptor (TLR) 4 and CD14, in a variety of neuroinflammatory disorders including bacterial infection, Alzheimer’s disease, and amyotrophic lateral sclerosis. Granulocyte macrophage-colony stimulating factor (GM-CSF) activates microglia and induces inflammatory responses via binding to GM-CSF receptor complex composed of two different subunit GM-CSF receptor α (GM-CSFRα) and common β chain (βc). GM-CSF has been shown to be associated with neuroinflammatory responses in multiple sclerosis and Alzheimer’s disease. However, the mechanisms how GM-CSF promotes neuroinflammation still remain unclear. METHODS: Microglia were stimulated with 20 ng/ml GM-CSF and the levels of TLR4 and CD14 expression were evaluated by RT-PCR and flowcytometry. LPS binding was analyzed by flowcytometry. GM-CSF receptor complex was analyzed by immunocytechemistry. The levels of IL-1β, IL-6 and TNF-α in culture supernatant of GM-CSF-stimulated microglia and NF-κB nuclear translocation were determined by ELISA. Production of nitric oxide (NO) was measured by the Griess method. The levels of p-ERK1/2, ERK1/2, p-p38 and p38 were assessed by Western blotting. Statistically significant differences between experimental groups were determined by one-way ANOVA followed by Tukey test for multiple comparisons. RESULTS: GM-CSF receptor complex was expressed in microglia. GM-CSF enhanced TLR4 and CD14 expressions in microglia and subsequent LPS-binding to the cell surface. In addition, GM-CSF priming increased LPS-induced NF-κB nuclear translocation and production of IL-1β, IL-6, TNF-α and NO by microglia. GM-CSF upregulated the levels of p-ERK1/2 and p-p38, suggesting that induction of TLR4 and CD14 expression by GM-CSF was mediated through ERK1/2 and p38, respectively. CONCLUSIONS: These results suggest that GM-CSF upregulates TLR4 and CD14 expression in microglia through ERK1/2 and p38, respectively, and thus promotes the LPS receptor-mediated inflammation in the CNS. BioMed Central 2012-12-13 /pmc/articles/PMC3565988/ /pubmed/23234315 http://dx.doi.org/10.1186/1742-2094-9-268 Text en Copyright ©2012 Parajuli et al.; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Parajuli, Bijay Sonobe, Yoshifumi Kawanokuchi, Jun Doi, Yukiko Noda, Mariko Takeuchi, Hideyuki Mizuno, Tetsuya Suzumura, Akio GM-CSF increases LPS-induced production of proinflammatory mediators via upregulation of TLR4 and CD14 in murine microglia |
title | GM-CSF increases LPS-induced production of proinflammatory mediators via upregulation of TLR4 and CD14 in murine microglia |
title_full | GM-CSF increases LPS-induced production of proinflammatory mediators via upregulation of TLR4 and CD14 in murine microglia |
title_fullStr | GM-CSF increases LPS-induced production of proinflammatory mediators via upregulation of TLR4 and CD14 in murine microglia |
title_full_unstemmed | GM-CSF increases LPS-induced production of proinflammatory mediators via upregulation of TLR4 and CD14 in murine microglia |
title_short | GM-CSF increases LPS-induced production of proinflammatory mediators via upregulation of TLR4 and CD14 in murine microglia |
title_sort | gm-csf increases lps-induced production of proinflammatory mediators via upregulation of tlr4 and cd14 in murine microglia |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3565988/ https://www.ncbi.nlm.nih.gov/pubmed/23234315 http://dx.doi.org/10.1186/1742-2094-9-268 |
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