25-Hydroxycholesterol enhances eicosanoid production in cultured bovine coronary artery endothelial cells by increasing prostaglandin G/H synthase-2.
Oxidized derivatives of cholesterol, the oxysterols, have been implicated in the development of atherosclerosis. We have found that the oxysterol, 25-hydroxycholesterol (25OHC), is a potent stimulator of eicosanoid production in endothelial cells. Confluent monolayers of bovine coronary artery endothelial cells (BCAECs) were treated with 25OHC (10 micrograms/ml) or interleukin-1 beta (IL-1 beta, 1 ng/ml), a known prostaglandin G/H synthase-2 (PGHS-2) inducer, for 48 h at 37 degrees C. Following incubation with [14C]arachidonic acid, the 14C-labeled metabolites of arachidonic acid were extracted and resolved using both normal and reverse phase high-performance liquid chromatography (HPLC) systems. 25OHC-treated cells had a five-fold increase in their prostaglandin production when compared to untreated cells. Eicosanoid production in IL-1 beta treated cells was not as pronounced. The major component in both sets of cells comigrated with 6-ketoPGF1 alpha. Other PGHS metabolites, 15- and 11-hydroxyeicosatetraenoic acids (HETEs) and 12-hydroxyheptadecatrienoic acid (HHT) were also identified and increased following 25OHC treatment. Epoxyeicosatrienoic acids, metabolites of cytochrome P450, were not effected. Enhanced production of 6-ketoPGF1 alpha, 15-HETE, 11-HETE and HHT were inhibited by indomethacin. Thus, all eicosanoids induced by 25OHC treatment were PGHS products. Western immunoblot analysis of lysates from 25OHC, IL-1 beta, or vehicle treated cells using anti-PGHS-1 and -2 antibodies revealed a significant increase in PGHS-2, but not PGHS-1, in both 25OHC and IL-1 beta treated cells. The notion that oxysterols can modulate vascular eicosanoid production through enzyme induction may be important in ultimately understanding their role in the pathogenesis of atherosclerosis.
Wohlfeil
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Department of Anesthesiology, Medical College of Wisconsin, Milwaukee 53226, USA. wohlfeil@post.its.mcw.edu
Campbell
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WB
D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D003331:Coronary Vessels / Q000187:drug effects; D015777:Eicosanoids / Q000096:biosynthesis*; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000201:enzymology; D004790:Enzyme Induction; D006888:Hydroxycholesterols / Q000494:pharmacology*; D006893:Hydroxyeicosatetraenoic Acids / Q000096:biosynthesis; D007375:Interleukin-1 / Q000494:pharmacology; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis*
D015777:Eicosanoids; D006888:Hydroxycholesterols; D006893:Hydroxyeicosatetraenoic Acids; D007375:Interleukin-1; C007997:25-hydroxycholesterol; D011451:Prostaglandin-Endoperoxide Synthases