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p38 mitogen-activated protein kinase regulates cyclooxygenase-2 mRNA stability and transcription in
1Kennedy Institute of Rheumatology, 1 Aspenlea Road, Hammersmith, London W6 8LH, United Kingdom.
Abstract:
p38 mitogen-activated protein kinase (MAPK) is activated by inflammatory stimuli such as bacterial lipopolysaccharide (LPS), interleukin-1, and tumor necrosis factor. We have previously shown that the pyridinyl imidazole SB 203580, which inhibits it, blocks the interleukin-1 induction of cyclooxygenase-2 (COX-2) and matrix metalloproteinase 1 and 3 mRNAs in fibroblasts. Here we explore the role of p38 MAPK in the response of human monocytes to LPS. 0.1 microM SB 203580 significantly inhibited the LPS induction of COX-2 and tumor necrosis factor protein and mRNAs. The activity of MAPK-activated protein kinase-2 (a substrate of p38 MAPK) in the cells was commensurately reduced. Some isoforms of c-jun N-terminal kinase (which is also activated by LPS) are sensitive to SB 203580; the inhibitor had little effect on monocyte c-jun N-terminal kinases up to 2 microM. We investigated the mechanism of inhibition of COX-2 induction. Transcription (measured by a nuclear run-on assay) was 60% inhibited by SB 203580 (2 microM). Importantly, we found that p38 MAPK was essential for stabilizing COX-2 mRNA: when cells stimulated for 4 h with LPS were treated with actinomycin D, COX-2 mRNA decayed slowly. Treatment of stimulated cells with 2 microM SB 203580 caused a rapid disappearance of COX-2 mRNA, even with actinomycin D present. We conclude p38 MAPK plays a role in the transcription and stabilization of COX-2 mRNA.
Insights
p38 mitogen-activated protein kinase (MAPK) regulates inflammatory responses. This study shows p38 MAPK is crucial for stabilizing cyclooxygenase-2 (COX-2) mRNA in human monocytes stimulated by lipopolysaccharide (LPS).
Area of Science:
- Molecular Biology
- Immunology
- Cell Signaling
Background:
- p38 mitogen-activated protein kinase (MAPK) is activated by inflammatory stimuli like lipopolysaccharide (LPS).
- p38 MAPK inhibition blocks interleukin-1 induction of cyclooxygenase-2 (COX-2) and matrix metalloproteinase mRNAs in fibroblasts.
Purpose of the Study:
- To investigate the role of p38 MAPK in human monocyte responses to LPS.
- To elucidate the mechanism by which p38 MAPK affects COX-2 expression.
Main Methods:
- Human monocytes were stimulated with LPS and treated with the p38 MAPK inhibitor SB 203580.
- COX-2 and tumor necrosis factor (TNF) protein and mRNA levels were measured.
- MAPK-activated protein kinase-2 activity was assessed.
- Nuclear run-on assays were used to measure transcription.
- Actinomycin D was used to assess mRNA stability.
Main Results:
- SB 203580 significantly inhibited LPS-induced COX-2 and TNF protein and mRNA.
- p38 MAPK activity was reduced, while c-jun N-terminal kinase activity was largely unaffected.
- SB 203580 inhibited COX-2 mRNA transcription by 60%.
- p38 MAPK was essential for COX-2 mRNA stabilization; SB 203580 caused rapid COX-2 mRNA decay.
Conclusions:
- p38 MAPK plays a significant role in both the transcription and stabilization of COX-2 mRNA in human monocytes stimulated by LPS.
- Inhibition of p38 MAPK leads to decreased expression of inflammatory mediators like COX-2.