title: An oxysterol biomarker for 7-dehydrocholesterol oxidation in cell/mouse models for Smith-Lemli-Opitz syndrome.
abstract: 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.
authors.affiliation: Department of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235.
authors.forename: Libin
authors.initials: L
authors.lastname: Xu
authors.affiliation: Department of Psychiatry and Vanderbilt Kennedy Center for Research on Human Development, Vanderbilt University, Nashville, TN 37235.
authors.forename: Zeljka
authors.initials: Z
authors.lastname: Korade
authors.affiliation: Department of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235.
authors.forename: Jr Dale A
authors.initials: JDA
authors.lastname: Rosado
authors.affiliation: Department of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235.
authors.forename: Wei
authors.initials: W
authors.lastname: Liu
authors.affiliation: Department of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235.
authors.forename: Connor R
authors.initials: CR
authors.lastname: Lamberson
authors.affiliation: Department of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235. Electronic address: n.porter@vanderbilt.edu.
authors.forename: Ned A
authors.initials: NA
authors.lastname: Porter
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D015415:Biomarkers / Q000032:analysis* / Q000737:chemistry; D001921:Brain / Q000196:embryology / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D002783:Cholestenones / Q000032:analysis* / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid / Q000379:methods*; D003684:Dehydrocholesterols / Q000302:isolation & purification / Q000378:metabolism; D004195:Disease Models, Animal; D004622:Embryo, Mammalian / Q000378:metabolism* / Q000473:pathology; D005260:Female; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D007553:Isotope Labeling; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D018345:Mice, Knockout; D010084:Oxidation-Reduction / Q000187:drug effects; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000172:deficiency* / Q000235:genetics; D011247:Pregnancy; D012015:Reference Standards; D019082:Smith-Lemli-Opitz Syndrome / Q000196:embryology / Q000235:genetics / Q000378:metabolism* / Q000473:pathology
chemical_list: C561089:3,5-dihydroxycholest-7-en-6-one; D000975:Antioxidants; D015415:Biomarkers; D002783:Cholestenones; D003684:Dehydrocholesterols; C016705:7-dehydrocholesterol; D044925:Oxidoreductases Acting on CH-CH Group Donors; C019995:7-dehydrocholesterol reductase