An oxysterol biomarker for 7-dehydrocholesterol oxidation in cell/mouse models for Smith-Lemli-Opitz syndrome[S]

• Published in: Journal of lipid research Vol. 52; no. 6; pp. 1222 - 1233
• Main Authors: Xu, Libin, Korade, Zeljka, Rosado, Jr. Dale A., Liu, Wei, Lamberson, Connor R., Porter, Ned A.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.06.2011; The American Society for Biochemistry and Molecular Biology; Elsevier
• Subjects: Animals; antioxidant; Antioxidants - pharmacology; Biomarkers - analysis; Biomarkers - chemistry; Brain - embryology; Brain - metabolism; Brain - pathology; Cell Line, Tumor; Cholestenones - analysis; Cholestenones - chemistry; Chromatography, High Pressure Liquid; Chromatography, Liquid - methods; Dehydrocholesterols - isolation & purification; Dehydrocholesterols - metabolism; Disease Models, Animal; Embryo, Mammalian - metabolism; Embryo, Mammalian - pathology; Female; Fibroblasts - cytology; Fibroblasts - metabolism; free radical oxidation; Humans; Isotope Labeling; lipid peroxidation; Mass Spectrometry - methods; metabolism; Mice; Mice, Knockout; Oxidation-Reduction - drug effects; oxidative stress; Oxidoreductases Acting on CH-CH Group Donors - deficiency; Oxidoreductases Acting on CH-CH Group Donors - genetics; Pregnancy; Reference Standards; Smith-Lemli-Opitz Syndrome - embryology; Smith-Lemli-Opitz Syndrome - genetics; Smith-Lemli-Opitz Syndrome - metabolism; Smith-Lemli-Opitz Syndrome - pathology; antioxidant; metabolism; lipid peroxidation; free radical oxidation; oxidative stress
• ISSN: 0022-2275; 1539-7262
• DOI: 10.1194/jlr.M014498

The level of 7-dehydrocholesterol (7-DHC) is elevated in tissues and fluids of Smith-Lemli-Opitz syndrome (SLOS) patients due to defective 7-DHC reductase. Although over a dozen oxysterols have been identified from 7-DHC free radical oxidation in solution, oxysterol profiles in SLOS cells and tissues have never been studied. We report here the identification and complete characterization of a novel oxysterol, 3β,5α-dihydroxycholest-7-en-6-one (DHCEO), as a biomarker for 7-DHC oxidation in fibroblasts from SLOS patients and brain tissue from a SLOS mouse model. Deuterated (d7)-standards of 7-DHC and DHCEO were synthesized from d7-cholesterol. The presence of DHCEO in SLOS samples was supported by chemical derivatization in the presence of d7-DHCEO standard followed by HPLC-MS or GC-MS analysis. Quantification of cholesterol, 7-DHC, and DHCEO was carried out by isotope dilution MS with the d7-standards. The level of DHCEO was high and correlated well with the level of 7-DHC in all samples examined (R = 0.9851). Based on our in vitro studies in two different cell lines, the mechanism of formation of DHCEO that involves 5α,6α-epoxycholest-7-en-3β-ol, a primary free radical oxidation product of 7-DHC, and 7-cholesten-3β,5α,6β-triol is proposed. In a preliminary test, a pyrimidinol antioxidant was found to effectively suppress the formation of DHCEO in SLOS fibroblasts.