SLOS and Neuronal Oxidative Stress

说明(申请人提供)：7-脱氢胆固醇(7-DHC)是人体生物学中一种重要的脂类。它是胆固醇的直接生物合成前体，也是维生素D3的前体。最近的发现表明，7-DHC很容易与分子氧发生自由基链氧化，即脂质过氧化。事实上，7-DHC的反应性使其比几乎任何其他已知的化合物更容易发生脂质过氧化。脂质过氧化与许多人类疾病有关，包括帕金森氏症、肌萎缩侧索硬化症、阿尔茨海默氏症和亨廷顿病等神经退行性疾病。许多在脂质过氧化过程中形成的化合物具有很强的生物活性，神经退行性变可能与这些有毒的过氧化产物有关。每10-30,000个人中就有一个人患有Smith-Lemli-Opitz综合征(SLOS)，它是由促进胆固醇生物合成最后一步的酶(Dhcr-7)缺陷引起的。这一缺陷导致患有这种综合征的个体的7-DHC浓度增加了10,000倍。SLOS导致一系列大脑异常，这些患者还表现出智力低下和自闭症样症状。这项提案的重点是7-DHC在可持续土地利用系统中积累的后果。一个指导性假设是7-DHC及其过氧化代谢产物对神经元功能有害。该假说还指出，7-DHC及其过氧化副产物的积累会导致神经元的生长和功能发生变化，结果是在这些人身上观察到毁灭性的异常。对7-DHC的过氧化产物进行了分离、纯化和充分表征。将开发方法来确定这些过氧化产物或其代谢产物是否被观察到作为7-DHC水平升高的组织和细胞的生物标记物。所研究的系统将包括三个SLOS小鼠模型、经过基因工程改造为具有高水平7-DHC的神经细胞、来自七名SLOS患者的皮肤细胞(成纤维细胞)以及来自NIH/NICHD的福布斯·波特博士从患者样本中提取的血浆、尿液和脑脊液。这项研究的另一个主要主题是评估细胞或动物暴露于7-DHC过氧化产物所造成的生物后果。将评估过氧化产物对神经细胞活性、形态和基因表达的影响，最后，将努力寻找保护细胞并逆转SLOS啮齿动物模型表型特征的脂质过氧化小分子抑制剂(抗氧化剂)。脂质过氧化经常与神经退行性疾病有关，这项研究与公共健康的相关性是过氧化的化学和生物学基础研究及其在此提出的对神经退行性疾病的抑制与神经退行性疾病的联系，包括一种毁灭性的综合征SLOS。 公共卫生相关性：Smith-Lemli-Opitz综合征(SLOS)是一种毁灭性的神经发育代谢疾病，由胆固醇生物合成缺陷导致有毒氧固醇化合物积聚引起。建立对人体体液中有毒化合物的检测并了解它们的基本神经生物学将有助于揭示SLOS，并导致治疗这种疾病和其他改变胆固醇代谢的疾病。

DESCRIPTION (provided by applicant): 7-Dehydrocholesterol (7-DHC) is an important lipid in human biology. It is the immediate biosynthetic precursor of cholesterol and it is also a precursor to vitamin D3. Very recent discoveries indicate that 7-DHC is very prone to undergo free radical chain oxidation with molecular oxygen, i.e. lipid peroxidation. Indeed, 7- DHC's reactivity makes it more susceptible to lipid peroxidation than nearly any other known compound. Lipid peroxidation is associated with many human diseases, including neurodegenerative disorders such as Parkinson's, ALS, Alzheimer's and Huntington's diseases. Many of the compounds formed during lipid peroxidation have potent biological activities and neurodegeneration may be associated with these toxic peroxidation products. A human syndrome affecting 1 in 10-30,000 individuals, Smith-Lemli-Opitz Syndrome (SLOS), is caused by a defect in the enzyme (Dhcr-7) that promotes the last step of cholesterol biosynthesis. This defect results in an increase by up to 10,000-fold of 7-DHC concentrations in individuals suffering from this syndrome. SLOS causes a range of brain abnormalities and these patients also exhibit mental retardation and autism-like symptoms. This proposal is focused on the consequences of the accumulation of 7-DHC in SLOS. A guiding hypothesis is that 7-DHC and its peroxidation metabolites are detrimental to neuronal function. The hypothesis also states that accumulation of 7-DHC and its peroxidation-derived byproducts leads to changes in the growth and function of neurons, the consequences being the devastating abnormalities observed in these individuals. The peroxidation products of 7-DHC have been isolated, purified and fully characterized. Methods will be developed to determine if these peroxidation products or their metabolites are observed as biomarkers in tissues and cells having elevated levels of 7-DHC. The systems studied will include three SLOS mouse models, neuronal cells that are genetically engineered to have high levels of 7-DHC, skin-cells (fibroblasts) from seven SLOS patients and plasma, urine and cerebral spinal fluid from patient samples by Dr. Forbes Porter at the NIH/NICHD. Another major theme of the research is to assess the biological consequences resulting from exposure of a cell or animal to 7-DHC peroxidation products. The effect of peroxidation products on neuronal cell viability, morphology and gene expression will be assessed and finally, an effort will be initiated to find small molecule inhibitors of lipid peroxidation (antioxidants) that protect cells and reverse the phenotypic characteristics of SLOS rodent models. Lipid peroxidation is frequently linked to neurodegenerative disorders and the relevance of this research to public health is the linkage of the fundamental studies in the chemistry and biology of peroxidation and its inhibition proposed here to neurodegenerative disorders, including a devastating syndrome, SLOS. PUBLIC HEALTH RELEVANCE: Smith-Lemli-Opitz syndrome (SLOS) is a devastating neurodevelopmental metabolic disorder caused by a defect in cholesterol biosynthesis that leads to a build up of toxic oxysterol compounds. Establishing assays for the toxic compounds in human fluids and understanding their fundamental neurobiology will shed light on SLOS and lead to therapies for this disorder and others that have altered cholesterol metabolism.