Abstract
Mitogen-activated protein (MAP) kinases play a pivotal role in the macrophages in the production of proinflammatory cytokines triggered by lipopolysaccharides. However, their function in the responses of macrophages to Gram-positive bacteria is poorly understood. Even less is known about the attenuation of MAP kinase signaling in macrophages exposed to Gram-positive bacteria. In the present study, we have investigated the regulation of MAP kinases and the role of MAP kinase phosphatase (MKP)-1 in the production of pro-inflammatory cytokines using murine RAW264.7 and primary peritoneal macrophages after peptidoglycan stimulation. Treatment of macrophages with peptidoglycan resulted in a transient activation of JNK, p38, and extracellular signal-regulated kinase. Most interestingly, MKP-1 expression was potently induced by peptidoglycan, and this induction was concurrent with MAP kinase dephosphorylation. Triptolide, a diterpenoid triepoxide, potently blocked the induction of MKP-1 by peptidoglycan and prolonged the activation of JNK and p38. Overexpression of MKP-1 substantially attenuated the production of tumor necrosis factor (TNF)-alpha induced by peptidoglycan, whereas knockdown of MKP-1 by small interfering RNA substantially increased the production of both TNF-alpha and interleukin-1 beta. Finally, we found that in primary murine peritoneal macrophages, MKP-1 induction following peptidoglycan stimulation also coincided with inactivation of JNK and p38. Blockade of MKP-1 induction resulted in a sustained activation of both JNK and p38 in primary macrophages. Our results reveal that MKP-1 critically regulates the expression of TNF-alpha and interleukin-1 beta in RAW264.7 cells and further suggest a central role for this phosphatase in controlling the inflammatory responses of primary macrophages to Gram-positive bacterial infection.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Animals
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Anti-Inflammatory Agents, Non-Steroidal / pharmacology
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / physiology*
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Cell Line
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Diterpenes / pharmacology
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Dual Specificity Phosphatase 1
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Enzyme Activation / drug effects
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Epoxy Compounds
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Extracellular Signal-Regulated MAP Kinases / metabolism
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Female
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Gene Expression
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Immediate-Early Proteins / genetics
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Immediate-Early Proteins / physiology*
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Inflammation
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Interleukin-1 / biosynthesis
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JNK Mitogen-Activated Protein Kinases / antagonists & inhibitors
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JNK Mitogen-Activated Protein Kinases / metabolism
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MAP Kinase Kinase 4
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Macrophages / drug effects
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Macrophages / enzymology*
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Macrophages / physiology*
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Macrophages, Peritoneal / drug effects
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Macrophages, Peritoneal / enzymology
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Macrophages, Peritoneal / physiology
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Mice
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Mice, Inbred C3H
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Mice, Inbred C57BL
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Mitogen-Activated Protein Kinase Kinases / antagonists & inhibitors
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Mitogen-Activated Protein Kinase Kinases / metabolism
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Peptidoglycan / pharmacology*
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Phenanthrenes / pharmacology
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Phosphoprotein Phosphatases / genetics
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Phosphoprotein Phosphatases / physiology*
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Phosphorylation
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Protein Phosphatase 1
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Protein Tyrosine Phosphatases / genetics
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Protein Tyrosine Phosphatases / physiology*
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Transfection
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Tumor Necrosis Factor-alpha / biosynthesis
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p38 Mitogen-Activated Protein Kinases / antagonists & inhibitors
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p38 Mitogen-Activated Protein Kinases / metabolism
Substances
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Anti-Inflammatory Agents, Non-Steroidal
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Cell Cycle Proteins
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Diterpenes
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Epoxy Compounds
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Immediate-Early Proteins
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Interleukin-1
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Peptidoglycan
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Phenanthrenes
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Tumor Necrosis Factor-alpha
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triptolide
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Extracellular Signal-Regulated MAP Kinases
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JNK Mitogen-Activated Protein Kinases
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p38 Mitogen-Activated Protein Kinases
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MAP Kinase Kinase 4
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Mitogen-Activated Protein Kinase Kinases
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Phosphoprotein Phosphatases
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Protein Phosphatase 1
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Dual Specificity Phosphatase 1
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Dusp1 protein, mouse
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Protein Tyrosine Phosphatases