title: Accurate detection of Smith-Lemli-Opitz syndrome carriers by measurement of the rate of reduction of the ergosterol C-7 double bond in cultured skin fibroblasts.
abstract: The activity of ergosterol delta 7-reductase (3 beta-hydroxysteroid delta 7-reductase) was measured in cultured skin fibroblasts from 7 controls, 10 Smith-Lemli-Opitz syndrome (SLOS) patients, and 10 parents (obligate carriers). The fibroblasts were exposed to delipidated medium supplemented with lovastatin for 24 h and the enzyme activity was determined by incubating cell-free homogenate with ergosterol (ergosta-5,7,22-trien-3 beta-ol) and measuring the mass of brassicasterol (ergosta-5,22-dien-3 beta-ol) formed by gas chromatography-mass spectrometry with selected-ion monitoring. In carriers, the activity was significantly lower than in controls (22 +/- 2 vs 65 +/- 10 pmol/min per mg protein, p < 0.0005), and no overlap was observed. The mean activity in carriers' fibroblasts was more than 100 times higher than in patients' cells (0.2 pmol/min per mg protein). The use of ergosterol avoids the many problems caused by the instability and lack of availability of radiolabelled 7-dehydrocholesterol. The present method makes it possible to discriminate SLOS carriers from both controls and patients using a commercially available substrate and common analytical equipment.
authors.affiliation: Department of Medicine, University of Medicine and Dentistry of New Jersey, New Jersey Medical School, Newark, USA.
authors.forename: M
authors.initials: M
authors.lastname: Honda
authors.forename: G S
authors.initials: GS
authors.lastname: Tint
authors.forename: S
authors.initials: S
authors.lastname: Shefer
authors.forename: A
authors.initials: A
authors.lastname: Honda
authors.forename: A K
authors.initials: AK
authors.lastname: Batta
authors.forename: G
authors.initials: G
authors.lastname: Xu
authors.forename: T S
authors.initials: TS
authors.lastname: Chen
authors.forename: G
authors.initials: G
authors.lastname: Salen
mesh_terms: D002478:Cells, Cultured; D002775:Cholestadienols / Q000032:analysis*; D004875:Ergosterol / Q000032:analysis; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D006579:Heterozygote; D006801:Humans; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000378:metabolism; D044925:Oxidoreductases Acting on CH-CH Group Donors; D010840:Phytosterols; D012867:Skin / Q000166:cytology; D019082:Smith-Lemli-Opitz Syndrome / Q000201:enzymology / Q000378:metabolism*; D013265:Stigmasterol / Q000032:analysis
chemical_list: D002775:Cholestadienols; D010840:Phytosterols; C003532:brassicasterol; D013265:Stigmasterol; D010088:Oxidoreductases; D044925:Oxidoreductases Acting on CH-CH Group Donors; C019995:7-dehydrocholesterol reductase; D004875:Ergosterol