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Fish oil modulates macrophage P44/P42 mitogen-activated protein kinase activity induced by lipopolysaccharide
1Department of Surgery, University of California, Los Angeles, USA. clo@mednet.ucla.edu
Background:
Mitogen-activated protein kinase (MAPK) cascades represent a major signal system to transduce extracellular signals into cellular responses. Overactivity of MAPK has been implicated in the development of many diseases, including cancer and sepsis. This study investigated the hypothesis that fish oil altered the membrane phospholipid composition and modulated MAPK activity.
Methods:
RAW 264.7 cells, a mouse macrophage (Mphi) cell line, were grown in eicosapentaenoic acid (EPA)-rich media (114 micromol/L) for 48 hours. Mphi were washed and exposed to Escherichia coli lipopolysaccharide (LPS; 1 microg/mL) for 10 minutes. Both total and activated (phosphorylated) portions of MAPK (P44 and P42) were determined by Western blot assays. AP-1 transcription factor activity was determined by electrophoretic mobility gel shift assays (EMSA). Mphi tumor necrosis factor (TNF) mRNA expression was measured by Northern blot assays.
Results:
LPS stimulation induced RAW cell phosphorylation of P44/P42. In contrast, RAW cells grown in EPA-rich media had less P44/P42 activation in the presence of LPS. Total P44/P42 were not affected by EPA or LPS. Similarly, EPA also inhibited AP-1 activity. Inhibition of P44/P42 activity with PD98059 reduced both AP-1 activity and TNF mRNA expression of LPS-stimulated Mphi.
Conclusions:
Our data suggest that fish oil regulates macrophage proinflammatory gene activation, at least in part, by modulating the MAPK activity.
Insights
Fish oil, rich in eicosapentaenoic acid (EPA), reduces mitogen-activated protein kinase (MAPK) activation in macrophages. This modulation of MAPK signaling by EPA impacts inflammatory gene expression, suggesting a role for fish oil in regulating immune responses.
Area of Science:
- Cellular Biology
- Immunology
- Molecular Biology
Background:
- Mitogen-activated protein kinase (MAPK) cascades are crucial for signal transduction.
- Dysregulated MAPK activity is linked to diseases like cancer and sepsis.
- This study explored fish oil's effect on macrophage MAPK activity and membrane composition.
Purpose of the Study:
- To investigate if fish oil alters macrophage membrane phospholipids.
- To determine the effect of fish oil on MAPK pathway activation.
- To assess the impact of fish oil on inflammatory gene expression.
Main Methods:
- RAW 264.7 macrophage cell line cultured in eicosapentaenoic acid (EPA)-rich media.
- Stimulation with lipopolysaccharide (LPS) to induce inflammatory response.
- Western blot to quantify MAPK (P44/P42) phosphorylation.
- Electrophoretic mobility gel shift assays (EMSA) for AP-1 activity.
- Northern blot to measure TNF mRNA expression.
Main Results:
- EPA-rich media significantly reduced LPS-induced phosphorylation of P44/P42 MAPK.
- EPA inhibited the activity of the AP-1 transcription factor.
- EPA did not alter total P44/P42 levels.
- Inhibition of MAPK with PD98059 mimicked EPA's effect on AP-1 and TNF mRNA.
Conclusions:
- Fish oil, specifically EPA, modulates MAPK activity in macrophages.
- This modulation of MAPK signaling is a mechanism by which fish oil regulates inflammatory gene activation.
- Findings suggest fish oil's potential role in managing inflammatory diseases.