JPET

Home Help [Feedback] [For Subscribers] [Archive] [Search] --
QUICK SEARCH:   [advanced]


     

Journal of Pharmacology And Experimental Therapeutics Fast Forward
First published on September 16, 2005; DOI: 10.1124/jpet.105.093922

This Article
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
jpet.105.093922v1
jpet.105.093922v2
316/1/371    most recent
Right arrow Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Du, L.
Right arrow Articles by Keeney, D. S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Du, L.
Right arrow Articles by Keeney, D. S.


Received for publication August 8, 2005.
Revised September 7, 2005.
Accepted for publication September 14, 2005.

A Biosynthetic Pathway Generating 12-Hydroxy-5,8,14-eicosatrienoic Acid from Arachidonic Acid is Active in Mouse Skin Microsomes

Liping Du 1, Valery Yermalitsky 1, David L. Hachey 1, Setti J. Jagadeesh 2, John R. Falck 2, Diane S. Keeney 1*

1 Vanderbilt University 2 University of Texas Southwestern Medical Center

* Address correspondence to: E-mail: diane.keeney{at}vanderbilt.edu

Abstract

The epidermis expresses cyclooxygenases, lipoxygenases and cytochromes P450, which utilize arachidonic acid to generate a diverse array of lipid mediators affecting epidermal differentiation and cellular functions. Recent studies show that mouse epidermis expresses CYP2B19, a keratinocyte-specific epoxygenase that generates 11,12- and 14,15-epoxyeicosatrienoic (EET) acids from arachidonate. We studied CYP2B19-dependent metabolism in mouse epidermal microsomes, reconstituted in the presence of [1-14C]-arachidonic acid. The majority of the [14C]-products were formed independently of NADPH, indicative of robust epidermal cyclooxygenase and lipoxygenase activities. We studied two NADPH-dependent products generated in a highly reproducible manner from arachidonate. One of these (product I) co-eluted with the CYP2B19 product 14,15-EET on a reversed-phase HPLC system; there was no evidence for other regioisomeric EET products. Further analyses proved that product I was not an epoxy fatty acid, based on different retention times on a normal phase HPLC system and failure of product I to undergo hydrolysis in acidic solution. We analyzed purified epidermal [14C]-products by liquid chromatography negative electrospray ionization mass spectrometry. Structures of the NADPH-dependent products were confirmed to be 12-oxo-5,8,14-eicosatrienoic acid (I) and 12-hydroxy-5,8,14-eicosatrienoic acid (II). This was the first evidence for a 12-hydroxy-5,8,14-eicosatrienoic acid biosynthetic pathway in mouse epidermis. Epidermal microsomes also generated 12-hydroperoxy, 12-hydroxy, and 12-oxo eicosatetraenoic acids from arachidonate, possible intermediates in the 12-hydroxy-5,8,14-eicosatrienoic acid biosynthetic pathway. These results predict hydroxyeicosatrienoic acids are synthesized from arachidonate in human epidermis. This would have important implications for human skin diseases given the known pro- and anti-inflammatory activities of stereo- and regioisomeric hydroxyeicosatrienoic acids.


Key words: CYP, cytochrome P450, eicosanoids, epidermis, keratinocyte, skin




Home Help [Feedback] [For Subscribers] [Archive] [Search] --
All ASPET Journals Molecular Pharmacology Pharmacological Reviews
Molecular Interventions Drug Metabolism and Disposition

Copyright © 2005 by the American Society for Pharmacology and Experimental Therapeutics.