The principal investigator has found that diets high in linoleic acid (LA) inhibit mouse skin carcinogenesis in the two stage DMBA - initiation, TPA - promotion model, and proposes studies to determine if a lipoxygenase product of linoleic acid, 13-HODE (13-hydroxyoctadecadienoic acid) can, in part, mediate this effect. The hypothesis to be tested is that the pro-tumor promoting 12-HETE (12-hydroxyeicosatetraenoic acid), a product of lipoxygenase action on arachidonic acid (AA), exerts its TPA-like effects by binding to a unique receptor and activating protein kinase C (PKC) delta, and that 13-HODE counteracts the effects of 12-HETE by competing with 12-HETE for binding. To test this hypothesis, the following specific aims will be carried out.
Aim 1. The specific lipoxygenase metabolites of AA and LA that mimic or inhibit TPA-elicited changes in specific markers of keratinocyte differentiation/proliferation and adhesion will be assessed.
Aim 2. Using whole cell and subcellular binding assays, attempts will be made to determine if 12-HETE binds to its own receptor, and if 13-HODE competes for this binding.
Aim 3 A. To assess possible mechanisms by which 12-HETE may act as a promoter, and 13-HODE as an inhibitor, a determination will be made if one or more PKC isoenzymes is activated by 12-HETE. The functional significance of this activation will be evaluated using antisense transient transfections.
Aim 3 B. The possibility that PKC delta or another PKC isoform may be the receptor for 12-HETE will be explored in PKC-downregulated and antisense and sense transfected cells, and co-immunoprecipitation studies (PKC delta with radiolabeled HETEs and HODEs).
