Abstract
The modulation of cell signaling by free radicals is important for the pathogenesis of inflammatory diseases. Recently, we have shown that NO reduces IL-1beta-induced matrix metalloproteinase (MMP-9) expression in glomerular mesangial cells (MC). Here we report that exogenously administrated superoxide, generated by the hypoxanthine/xanthine oxidase system (HXXO) or by the redox cycler 2, 3-dimethoxy-1,4-naphtoquinone, caused a marked amplification of IL-1beta-primed, steady state, MMP-9 mRNA level and an increase in gelatinolytic activity in the conditioned medium. Superoxide generators alone were ineffective. Cytokine-induced steady state mRNA levels of TIMP-1, an endogenous inhibitor of MMP-9, were affected similarly by HXXO. Transient transfection of rat mesangial cells with 0.6 kb of the 5'-flanking region of the rat MMP-9 gene proved a transcriptional regulation of MMP-9 expression by superoxide. HXXO augmented the IL-1beta-triggered nuclear translocation of p65 and c-Jun and, in parallel, increased DNA binding activities of NF-kappaB and AP-1. Mutation of either response element completely prevented MMP-9 promoter activation by IL-1beta. Moreover, specific inhibitors of the classical extracellular signal-regulated kinase (ERK) pathway and p38 mitogen-activated protein kinase (MAPK) cascade, partially reversed the HXXO-mediated effects on MMP-9 mRNA levels, thus demonstrating involvement of ERKs and p38 MAPKs in MMP-9 expression. Furthermore, IL-1beta-triggered phosphorylation of all three MAPKs, including p38-MAPK, c-Jun N-terminal kinase, and ERK, was substantially enhanced by superoxide. Our data identify superoxide as a costimulatory factor amplifying cytokine-induced MMP-9 expression by interfering with the signaling cascades leading to the activation of AP-1 and NF-kappaB.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adjuvants, Immunologic / pharmacology
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Animals
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Base Sequence
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Binding Sites / genetics
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Binding Sites / immunology
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Biological Transport / immunology
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Cell Nucleus / immunology
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Cell Nucleus / metabolism
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DNA-Binding Proteins / metabolism
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Enzyme Activation / drug effects
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Enzyme Activation / immunology
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Enzyme Inhibitors / pharmacology
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Flavonoids / pharmacology
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Gene Amplification / immunology
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Gene Expression Regulation / immunology
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Glomerular Mesangium / cytology
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Glomerular Mesangium / enzymology*
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Glomerular Mesangium / immunology*
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Glomerular Mesangium / metabolism
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Hydrolysis
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I-kappa B Proteins*
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Imidazoles / pharmacology
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Interleukin-1 / physiology*
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JNK Mitogen-Activated Protein Kinases
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MAP Kinase Signaling System / drug effects
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MAP Kinase Signaling System / immunology*
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Matrix Metalloproteinase 9 / biosynthesis*
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Matrix Metalloproteinase 9 / genetics
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Matrix Metalloproteinase 9 / metabolism
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Matrix Metalloproteinase Inhibitors
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Mitogen-Activated Protein Kinases / metabolism
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Molecular Sequence Data
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NF-KappaB Inhibitor alpha
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NF-kappa B / antagonists & inhibitors
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NF-kappa B / metabolism
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NF-kappa B / physiology*
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Phosphorylation
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Promoter Regions, Genetic / immunology
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Pyridines / pharmacology
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RNA, Messenger / antagonists & inhibitors
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RNA, Messenger / biosynthesis
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RNA, Messenger / metabolism
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Rats
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Superoxides / pharmacology*
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Tissue Inhibitor of Metalloproteinase-1 / biosynthesis
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Tissue Inhibitor of Metalloproteinase-1 / genetics
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Transcription Factor AP-1 / metabolism
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Transcription Factor AP-1 / physiology*
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Transcription Factor RelA
Substances
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Adjuvants, Immunologic
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DNA-Binding Proteins
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Enzyme Inhibitors
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Flavonoids
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I-kappa B Proteins
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Imidazoles
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Interleukin-1
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Matrix Metalloproteinase Inhibitors
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NF-kappa B
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Nfkbia protein, rat
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Pyridines
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RNA, Messenger
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Tissue Inhibitor of Metalloproteinase-1
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Transcription Factor AP-1
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Transcription Factor RelA
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Superoxides
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NF-KappaB Inhibitor alpha
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JNK Mitogen-Activated Protein Kinases
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Mitogen-Activated Protein Kinases
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Matrix Metalloproteinase 9
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SB 203580
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2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one