Department of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN, USA.
J Lipid Res. 2011 Oct;52(10):1810-20. doi: 10.1194/jlr.M018366. Epub 2011 Aug 4.
Treatment of Sprague-Dawley rats with AY9944, an inhibitor of 3β-hydroxysterol-Δ(7)-reductase (Dhcr7), leads to elevated levels of 7-dehydrocholesterol (7-DHC) and reduced levels of cholesterol in all biological tissues, mimicking the key biochemical hallmark of Smith-Lemli-Opitz syndrome (SLOS). Fourteen 7-DHC-derived oxysterols previously have been identified as products of free radical oxidation in vitro; one of these oxysterols, 3β,5α-dihydroxycholest-7-en-6-one (DHCEO), was recently identified in Dhcr7-deficient cells and in brain tissues of Dhcr7-null mouse. We report here the isolation and characterization of three novel 7-DHC-derived oxysterols (4α- and 4β-hydroxy-7-DHC and 24-hydroxy-7-DHC) in addition to DHCEO and 7-ketocholesterol (7-kChol) from the brain tissues of AY9944-treated rats. The identities of these five oxysterols were elucidated by HPLC-ultraviolet (UV), HPLC-MS, and 1D- and 2D-NMR. Quantification of 4α- and 4β-hydroxy-7-DHC, DHCEO, and 7-kChol in rat brain, liver, and serum were carried out by HPLC-MS using d(7)-DHCEO as an internal standard. With the exception of 7-kChol, these oxysterols were present only in tissues of AY9944-treated, but not control rats, and 7-kChol levels were markedly (>10-fold) higher in treated versus control rats. These findings are discussed in the context of the potential involvement of 7-DHC-derived oxysterols in the pathogenesis of SLOS.
用 3β-羟甾醇-Δ(7)-还原酶（Dhcr7）抑制剂 AY9944 处理 Sprague-Dawley 大鼠，会导致所有生物组织中的 7-脱氢胆固醇（7-DHC）水平升高，胆固醇水平降低，这模拟了 Smith-Lemli-Opitz 综合征（SLOS）的关键生化特征。此前已有 14 种 7-DHC 衍生的氧化固醇在体外被鉴定为自由基氧化产物；其中一种氧化固醇，3β,5α-二羟基胆甾-7-烯-6-酮（DHCEO），最近在 Dhcr7 缺陷细胞和 Dhcr7 基因敲除小鼠的脑组织中被鉴定出来。我们在此报告，除了 Dhceo 和 7-酮胆固醇（7-kChol）之外，还从 AY9944 处理的大鼠脑组织中分离并鉴定出三种新的 7-DHC 衍生的氧化固醇（4α-和 4β-羟基-7-DHC 和 24-羟基-7-DHC）。这些五种氧化固醇的身份通过 HPLC-紫外（UV）、HPLC-MS 以及 1D 和 2D-NMR 得以阐明。通过 HPLC-MS 使用 d(7)-DHCEO 作为内标，对大鼠脑、肝和血清中的 4α-和 4β-羟基-7-DHC、DHCEO 和 7-kChol 进行定量。除了 7-kChol 之外，这些氧化固醇仅存在于 AY9944 处理的大鼠组织中，而不存在于对照组大鼠中，并且处理组大鼠的 7-kChol 水平明显（>10 倍）高于对照组。这些发现与 7-DHC 衍生的氧化固醇在 SLOS 发病机制中的潜在作用有关。
