Conversion of 7-ketocholesterol to oxysterol metabolites by recombinant CYP27A1 and retinal pigment epithelial cells

Published

June 1, 2011

Author(s)

Illarion V. Turko, Gun-Young Heo, Ilya Bederman, Natalia Mast, Wei-Li Liao, Irina A. Pikuleva

Abstract

Of different oxygenated cholesterol metabolites, 7-ketocholesterol (7KCh) is considered noxious oxysterol implicated in the development of certain pathologies including those in the neural retina and the underlying retinal pigment epithelium (RPE). It is currently unknown how 7KCh is eliminated from the retina. To elucidate whether cytochrome P450 27A1 (CYP27A1) is involved, we used purified recombinant enzyme and compared the binding and catalytic properties of 7KCh against the endogenous substrate cholesterol. The affinities of CYP27A1 for the two sterols were similar but 7KCh was metabolized at a 4-fold higher rate than cholesterol in the reconstituted system in vitro. Lipid extracts from bovine neural retina and RPE were then analyzed by isotope dilution gas chromatography-mass spectrometry with single ion monitoring for the presence of the CYP27A1 products of 7K hydroxylation. Two metabolites of 7KCh, 3β,27-dihydroxy-5-cholesten-7-one and 3β-hydroxy-5-cholesten-7-one-26-oic acid, were detected in the RPE but not in the neural retina. 3β,27-Dihydroxy-5-cholesten-7-one was also formed when RPE homogenates were supplemented with NADPH and the mitochondrial redox system. The data obtained represents strong evidence for the role of CYP27A1 in retinal metabolism of 7K and suggests that in addition to cholesterol removal the functions of this enzyme could also include elimination of toxic cholesterol metabolites.

Citation

Journal of Lipid Research

Volume

52

Issue

6

Pub Type

Journals

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DOI Link

Keywords

cholesterol, 3β, 27-dihydroxy-5-cholesten-7-one, 3β-hydroxy-5-cholesten-7-one-26-oic acid, isotope dilution gas chromatography-mass spectrometry, age-related macular degeneration

Citation

Turko, I. , Heo, G. , Bederman, I. , Mast, N. , Liao, W. and Pikuleva, I. (2011), Conversion of 7-ketocholesterol to oxysterol metabolites by recombinant CYP27A1 and retinal pigment epithelial cells, Journal of Lipid Research, [online], https://doi.org/10.1194/jlr.M014217 (Accessed December 15, 2024)