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Biochim Biophys Acta. 2005 Feb 21;1687(1-3):64-75.

Molecular characterization of the lipopolysaccharide/platelet activating factor- and zymosan-induced pathways leading to prostaglandin production in P388D1 macrophages.

Author information

1
Department of Pharmacology, School of Medicine, University of California at San Diego, La Jolla, CA 92093-0601, USA. rschalos@ucsd.edu

Abstract

P388D1 cells release free arachidonic acid (AA) and prostaglandin E2 (PGE2) upon stimulation with platelet-activating factor (PAF) and zymosan. The response to PAF is dependent on priming of the cells with bacterial lipopolysaccharide (LPS). In the LPS/PAF pathway, both AA and PGE2 release are dependent on transcription and translation, whereas in the zymosan pathway the release of these compounds appears to be largely independent of these processes. Using quantitative real-time PCR, we analyzed the expression of mRNAs that encode proteins potentially responsible for the dependency of the LPS/PAF pathway on gene expression. These include all the phospholipases A2 (PLA2) that we detected in P388D1 cells, cyclooxygenases (COX), COX-1 and COX-2, the membrane-associated prostaglandin E synthase-1 (mPGES-1), the lipocalin-type prostaglandin D2 synthase (PGDS), hematopoietic PGDS and the subunit G(alpha i2) of heterotrimeric G-proteins. None of the mRNAs encoding PLA2s, PGDSs, or G(alpha i2) are substantially altered during LPS priming. However, cyclooxygenase-2 is up-regulated during LPS priming and after stimulation of the cells with zymosan. A modest but significant increase of mPGES-1 mRNA was also detected upon stimulation with zymosan. Thus, the dependency of the LPS/PAF-induced PGE2 production on gene expression can be attributed to the production of cyclooxygenase-2. The dependency of AA release on gene expression is not due to altered expression of any of the PLA2s. We suggest that an accessory regulatory protein affecting the release of AA must be responsible. Using HPLC we separated lipids that are secreted upon stimulation with LPS/PAF and zymosan and found that in both pathways PGD2 is the dominant prostaglandin produced and also detected PGE2, PGF(2alpha) and AA besides several unidentified compounds.

PMID:
15708354
DOI:
10.1016/j.bbalip.2004.11.005
[Indexed for MEDLINE]

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