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Prostaglandin E2 stimulates AP-1-mediated CD14 expression in mouse macrophages via cyclic AMP-dependent protein
H Iwahashi1, A Takeshita, S Hanazawa
1Department of Oral Microbiology, Meikai University School of Dentistry, Keyakidai, Sakado City, Saitama, Japan.
Abstract:
PGs play a functional role in the early stage of Gram-negative bacterial infections, because this prostanoid is produced rapidly by epithelial cells after a bacterial infection. CD14, one of the LPS receptors, is a key molecule in triggering the response to bacterial LPS in association with a Toll-like molecule. Therefore, in this study, we investigated the effect of PG on CD14 expression in mouse macrophages. PGE1, PGE2, and PGA1 among the PGs tested strongly stimulated the expression of the CD14 gene in the cells. The stimulatory action also was observed by Western blot analysis. cAMP-elevating agents stimulated expression of CD14 gene as well. Protein kinase A inhibitor, N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide (H-89), but not protein kinase C inhibitor 3-(1-[3-(dimethylamino)propyl]-1H-indol-3-yl)-4-(1H-indol-3-yl)-1H-py rrole-2,5-dione (GF109203X), abolished the stimulated expression of CD14. A run-on assay showed that PGE2 stimulated the CD14 gene expression at the transcriptional level via protein kinase A. PGE2 also stimulated activation of AP-1, a heterodimer of c-Jun and c-Fos, because the prostanoid increased specific binding of nuclear proteins to the AP-1 consensus sequence and stimulated AP-1-promoted luciferase activity. PGE2-stimulated expression of CD14 was inhibited by antisense c-fos and c-jun oligonucleotides, but not by their sense oligonucleotides. Finally, PGE2 pretreatment synergistically stimulated LPS-induced expression of IL-1beta and IL-6 genes in mouse macrophages. Therefore, the present study demonstrates that PGE2 has the ability to stimulate AP-1-mediated expression of CD14 in mouse macrophages via cAMP-dependent protein kinase A.
Insights
Prostaglandin E2 (PGE2) significantly boosts CD14 gene expression in mouse macrophages by activating the AP-1 pathway through protein kinase A. This enhances the cells' response to bacterial lipopolysaccharides (LPS).
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- Prostaglandins (PGs) are rapidly produced by epithelial cells during Gram-negative bacterial infections.
- CD14 is a key receptor for bacterial lipopolysaccharide (LPS), initiating inflammatory responses.
- The role of PGs in modulating CD14 expression during bacterial infections requires further investigation.
Purpose of the Study:
- To investigate the effect of Prostaglandin E2 (PGE2) on CD14 gene expression in mouse macrophages.
- To elucidate the signaling pathways involved in PGE2-mediated CD14 regulation.
- To determine the functional consequence of PGE2 on macrophage response to LPS.
Main Methods:
- Primary mouse macrophages were treated with various PGs (PGE1, PGE2, PGA1) and cAMP-elevating agents.
- CD14 gene expression was analyzed using quantitative PCR and Western blotting.
- Involvement of protein kinase A (PKA) and protein kinase C (PKC) was assessed using specific inhibitors (H-89 and GF109203X).
- Transcriptional activity was measured via run-on assays.
- AP-1 transcription factor activity was evaluated using DNA-binding assays and luciferase reporter assays.
- The role of AP-1 components (c-Jun, c-Fos) was studied using antisense oligonucleotides.
- The effect of PGE2 on LPS-induced cytokine production (IL-1beta, IL-6) was examined.
Main Results:
- PGE1, PGE2, and PGA1 significantly stimulated CD14 gene expression at both mRNA and protein levels.
- cAMP-elevating agents also increased CD14 expression.
- PGE2-induced CD14 expression was mediated by PKA, not PKC.
- PGE2 enhanced CD14 gene expression at the transcriptional level.
- PGE2 stimulated the transcriptional activity of AP-1.
- Inhibition of c-Jun and c-Fos by antisense oligonucleotides blocked PGE2-induced CD14 expression.
- PGE2 pretreatment synergistically enhanced LPS-induced expression of IL-1beta and IL-6.
Conclusions:
- PGE2 stimulates CD14 gene expression in mouse macrophages via a cAMP-dependent PKA pathway.
- PGE2-induced CD14 expression is regulated at the transcriptional level and involves the activation of AP-1 transcription factors (c-Jun and c-Fos).
- PGE2 enhances the inflammatory response of macrophages to LPS by upregulating CD14 expression and subsequently cytokine production.