Abstract
Purpose:
To investigate the effect of the complement activation product C5a on toll-like receptor (TLR) 4-induced responses in RPE cells.
Methods:
Confluent cultures of human RPE cells (ARPE-19) were stimulated with C5a, lipopolysaccharide (LPS), or a combination of the two. The expression of TLR4 was determined by real-time PCR and flow cytometry. Cytokine profiles were determined by real-time PCR and enzyme-linked immunosorbent assay (ELISA). The phosphorylation of p38, ERK 1/2, and JNK was measured by flow cytometry.
Results:
C5a stimulation enhanced the expression of TLR4 in a dose- and time-dependent manner. C5a was able to stimulate the production of TLR4-induced IL-6 and IL-8 by ARPE-19 cells. Blocking experiments showed that the effect of C5a on cytokine production was mediated via C5aR. ERK1/2, but not JNK or p38, were involved in the production of IL-6 and IL-8.
Conclusions:
The results indicate that C5a can induce the TLR4 expression and enhance the production of TLR4-induced IL-6 and IL-8 by ARPE-19. The effect of C5a on cytokine production was mediated by C5aR and the phosphorylation of ERK1/2.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Cell Line
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Complement C5a / metabolism
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Complement C5a / pharmacology*
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Dose-Response Relationship, Drug
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Drug Combinations
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Epithelial Cells / cytology
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Epithelial Cells / drug effects*
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Epithelial Cells / immunology
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Gene Expression Regulation
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Humans
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Interleukin-6 / genetics
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Interleukin-6 / immunology
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Interleukin-8 / genetics
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Interleukin-8 / immunology
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Lipopolysaccharides / pharmacology*
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MAP Kinase Kinase 4 / genetics
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MAP Kinase Kinase 4 / immunology
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Mitogen-Activated Protein Kinase 1 / genetics
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Mitogen-Activated Protein Kinase 1 / immunology
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Mitogen-Activated Protein Kinase 3 / genetics
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Mitogen-Activated Protein Kinase 3 / immunology
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Phosphorylation
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Receptor, Anaphylatoxin C5a / genetics
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Receptor, Anaphylatoxin C5a / immunology
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Retinal Pigment Epithelium / cytology
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Retinal Pigment Epithelium / drug effects*
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Retinal Pigment Epithelium / immunology
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Signal Transduction
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Toll-Like Receptor 4 / genetics*
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Toll-Like Receptor 4 / immunology
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p38 Mitogen-Activated Protein Kinases / genetics
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p38 Mitogen-Activated Protein Kinases / immunology
Substances
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C5AR1 protein, human
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Drug Combinations
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IL6 protein, human
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Interleukin-6
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Interleukin-8
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Lipopolysaccharides
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Receptor, Anaphylatoxin C5a
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TLR4 protein, human
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Toll-Like Receptor 4
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Complement C5a
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MAPK1 protein, human
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Mitogen-Activated Protein Kinase 1
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Mitogen-Activated Protein Kinase 3
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p38 Mitogen-Activated Protein Kinases
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MAP Kinase Kinase 4