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Cytokine mRNA decay is accelerated by an inhibitor of p38-mitogen-activated protein kinase
S W Wang1, J Pawlowski, S T Wathen
1Department of Biology, Amgen Inc., Boulder, CO 80301, USA.
Objective:
To identify the site(s) in tumor necrosis factor (TNFalpha), interleukin-6 (IL-6), and macrophage inflammatory protein-1alpha (MIP-1alpha) biosynthesis that is blocked by SB202190, a selective inhibitor of p38-mitogen activated protein kinase (p38).
Materials:
Human blood monocytes isolated by centrifugal elutriation.
Methods:
Monocytes were stimulated with lipopolysaccharide in the presence of 0, 0.3, 1 and 3 microM SB202190. Induced TNFalpha, IL-6, and MIP-1alpha protein and mRNA were measured by ELISA and quantitative RT-PCR, respectively. The half-lives of cytokine mRNA levels were determined following treatment of cells with actinomycin D or SB202190.
Results:
SB202190 suppressed >60% of lipopolysaccharide-induced TNFalpha, IL-6, and MIP-1alpha protein and mRNA expression. Suppressed mRNA levels could be attributed to a >2 to 7-fold reduction in cytokine mRNA half-lives. In contrast, SB202190 did not destabilize mRNAs encoding interferon-induced gene 15 protein and glyceraldehyde-3-phosphate dehydrogenase.
Conclusions:
Specific mRNA destabilization represents an important and novel site of action for the cytokine suppressive effects of p38 inhibitors.
Insights
SB202190, a p38 inhibitor, blocks tumor necrosis factor-alpha, interleukin-6, and macrophage inflammatory protein-1alpha production by reducing cytokine mRNA stability. This reveals a novel mechanism for p38 inhibitors in suppressing inflammation.
Area of Science:
- Immunology
- Molecular Biology
- Pharmacology
Background:
- Tumor necrosis factor-alpha (TNFα), interleukin-6 (IL-6), and macrophage inflammatory protein-1alpha (MIP-1α) are key inflammatory cytokines.
- p38-mitogen activated protein kinase (p38) is a critical regulator of inflammatory responses.
- SB202190 is a selective inhibitor of p38 kinase.
Purpose of the Study:
- To determine the mechanism by which SB202190 inhibits the biosynthesis of TNFα, IL-6, and MIP-1α.
- To investigate the role of p38 kinase inhibition in regulating cytokine production at the post-transcriptional level.
Main Methods:
- Human blood monocytes were isolated and stimulated with lipopolysaccharide (LPS) in the presence of varying concentrations of SB202190.
- TNFα, IL-6, and MIP-1α protein and mRNA levels were quantified using ELISA and RT-PCR.
- Cytokine mRNA half-lives were assessed after treatment with actinomycin D or SB202190.
Main Results:
- SB202190 significantly suppressed LPS-induced TNFα, IL-6, and MIP-1α protein and mRNA expression by over 60%.
- This suppression was linked to a 2- to 7-fold reduction in the half-lives of these cytokine mRNAs.
- SB202190 did not affect the mRNA stability of control genes, such as interferon-induced gene 15 and glyceraldehyde-3-phosphate dehydrogenase.
Conclusions:
- Specific mRNA destabilization is a significant and novel mechanism of action for p38 inhibitors.
- Inhibition of p38 kinase leads to the destabilization of cytokine mRNAs, thereby reducing inflammatory cytokine production.
- This finding provides a deeper understanding of the anti-inflammatory effects of p38 inhibitors and potential therapeutic strategies.