title: Extra-hepatic metabolism of 7-ketocholesterol occurs by esterification to fatty acids via cPLA2α and SOAT1 followed by selective efflux to HDL.
abstract: Accumulation of 7-ketocholesterol (7KCh) in tissues has been previously associated with various chronic aging diseases. Orally ingested 7KCh is readily metabolized by the liver and does not pose a toxicity threat. However, 7KCh formed in situ, usually associated with lipoprotein deposits, can adversely affect surrounding tissues by causing inflammation and cytotoxicity. In this study we have investigated various mechanisms for extra-hepatic metabolism of 7KCh (e.g. hydroxylation, sulfation) and found only esterification to fatty acids. The esterification of 7KCh to fatty acids involves the combined action of cytosolic phospholipase A2 alpha (cPLA2α) and sterol O-acyltransferase (SOAT1). Inhibition of either one of these enzymes ablates 7KCh-fatty acid ester (7KFAE) formation. The 7KFAEs are not toxic and do not induce inflammatory responses. However, they can be unstable and re-release 7KCh. The higher the degree of unsaturation, the more unstable the 7KFAE (e.g. 18:0>18:1>18:2>18:3≫20:4). Biochemical inhibition and siRNA knockdown of SOAT1 and cPLA2α ablated the 7KFAE synthesis in cultured ARPE19 cells, but had little effect on the 7KCh-induced inflammatory response. Overexpression of SOAT1 reduced the 7KCh-induced inflammatory response and provided some protection from cell death. This effect is likely due to the increased conversion of 7KCh to 7KFAEs, which reduced the intracellular 7KCh levels. Addition of HDL selectively increased the efflux of 7KFAEs and enhanced the effect of SOAT1 overexpression. Our data suggests an additional function for HDL in aiding extra-hepatic tissues to eliminate 7KCh by returning 7KFAEs to the liver for bile acid formation.
authors.affiliation: Mechanisms of Retinal Diseases Section, Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD, USA. Electronic address: leej@nei.nih.gov.
authors.forename: Jung Wha
authors.initials: JW
authors.lastname: Lee
authors.affiliation: Mechanisms of Retinal Diseases Section, Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Jiahn-Dar
authors.initials: JD
authors.lastname: Huang
authors.affiliation: Mechanisms of Retinal Diseases Section, Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Ignacio R
authors.initials: IR
authors.lastname: Rodriguez
mesh_terms: D000818:Animals; D001692:Biological Transport; D001711:Biotransformation; D016923:Cell Death; D002460:Cell Line; D008076:Cholesterol, HDL / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004951:Esterification; D005227:Fatty Acids / Q000378:metabolism*; D054513:Group IV Phospholipases A2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007653:Ketocholesterols / Q000378:metabolism*; D008253:Macaca mulatta; D008297:Male; D013058:Mass Spectrometry; D034622:RNA Interference; D055213:Retinal Pigment Epithelium / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D002785:Sterol O-Acyltransferase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection
chemical_list: D008076:Cholesterol, HDL; D004791:Enzyme Inhibitors; D005227:Fatty Acids; D007653:Ketocholesterols; D002785:Sterol O-Acyltransferase; C493492:sterol O-acyltransferase 1; D054513:Group IV Phospholipases A2; C516389:PLA2G4A protein, human; C003001:7-ketocholesterol
keywords: 7-Ketocholesterol; 7-Ketocholesterol fatty acid ester; Cultured RPE cell; HDL; SOAT1; cPLA2α