title: Lipoxygenases mediate the effect of essential fatty acid in skin barrier formation: a proposed role in releasing omega-hydroxyceramide for construction of the corneocyte lipid envelope.
abstract: A barrier to water loss is vital to maintaining life on dry land. Formation of the mammalian skin barrier requires both the essential fatty acid linoleate and the two lipoxygenases 12R-lipoxygenase (12R-LOX) and epidermal lipoxygenase-3 (eLOX3), although their roles are poorly understood. Linoleate occurs in O-linoleoyl-ω-hydroxyceramide, which, after hydrolysis of the linoleate moiety, is covalently attached to protein via the free ω-hydroxyl of the ceramide, forming the corneocyte lipid envelope, a scaffold between lipid and protein that helps seal the barrier. Here we show using HPLC-UV, LC-MS, GC-MS, and (1)H NMR that O-linoleoyl-ω-hydroxyceramide is oxygenated in a regio- and stereospecific fashion by the consecutive actions of 12R-LOX and eLOX3 and that these products occur naturally in pig and mouse epidermis. 12R-LOX forms 9R-hydroperoxy-linoleoyl-ω-hydroxyceramide, further converted by eLOX3 to specific epoxyalcohol (9R,10R-trans-epoxy-11E-13R-hydroxy) and 9-keto-10E,12Z esters of the ceramide; an epoxy-ketone derivative (9R,10R-trans-epoxy-11E-13-keto) is the most prominent oxidized ceramide in mouse skin. These products are absent in 12R-LOX-deficient mice, which crucially display a near total absence of protein-bound ω-hydroxyceramides and of the corneocyte lipid envelope and die shortly after birth from transepidermal water loss. We conclude that oxygenation of O-linoleoyl-ω-hydroxyceramide is required to facilitate the ester hydrolysis and allow bonding of the ω-hydroxyceramide to protein, providing a coherent explanation for the roles of multiple components in epidermal barrier function. Our study uncovers a hitherto unknown biochemical pathway in which the enzymic oxygenation of ceramides is involved in building a crucial structure of the epidermal barrier.
authors.affiliation: Department of Pharmacology and the Vanderbilt Institute of Chemical Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
authors.forename: Yuxiang
authors.initials: Y
authors.lastname: Zheng
authors.forename: Huiyong
authors.initials: H
authors.lastname: Yin
authors.forename: William E
authors.initials: WE
authors.lastname: Boeglin
authors.forename: Peter M
authors.initials: PM
authors.lastname: Elias
authors.forename: Debra
authors.initials: D
authors.lastname: Crumrine
authors.forename: David R
authors.initials: DR
authors.lastname: Beier
authors.forename: Alan R
authors.initials: AR
authors.lastname: Brash
mesh_terms: D000818:Animals; D001092:Arachidonate 12-Lipoxygenase / Q000235:genetics / Q000378:metabolism*; D002518:Ceramides / Q000235:genetics / Q000378:metabolism*; D004817:Epidermis / Q000378:metabolism*; D005228:Fatty Acids, Essential / Q000235:genetics / Q000378:metabolism*; D008084:Lipoxygenase / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D010084:Oxidation-Reduction; D013552:Swine
chemical_list: D002518:Ceramides; D005228:Fatty Acids, Essential; D008084:Lipoxygenase; C518403:eLOX3 protein, mouse; C520168:Alox12b protein, mouse; D001092:Arachidonate 12-Lipoxygenase