期刊
JOURNAL OF NEUROINFLAMMATION
卷 9, 期 -, 页码 -出版社
BIOMED CENTRAL LTD
DOI: 10.1186/1742-2094-9-268
关键词
Microglia; TLR4; CD14; GM-CSF; NF-kappa B
资金
- National Institute of Biomedical Innovation (NIBIO)
- Ministry of Education, Culture, Sports, Science and Technology of Japan
- Ministry of Health, Labour and Welfare of Japan
- Grants-in-Aid for Scientific Research [24111520, 24659430] Funding Source: KAKEN
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 alpha (GM-CSFR alpha) and common beta chain (beta c). 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-1 beta, IL-6 and TNF-alpha in culture supernatant of GM-CSF-stimulated microglia and NF-kappa B 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-kappa B nuclear translocation and production of IL-1 beta, IL-6, TNF-alpha 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.
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