Significance of CYP46A1 and other P450s in retinal function

CYP46A1 和其他 P450 在视网膜功能中的意义

• 批准号: 8290853
• 负责人: Irina A Pikuleva
• 金额: $ 39.25万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8290853
• 关键词: Acids; Address; Affect; Age; Age related macular degeneration; Aging; Aldehydes; Alzheimer' s Disease; Anastomosis - action; Animals; Arachidonic Acids; Biochemical; Blood flow; Bruch' s basal membrane structure; CYP46A1 gene; Cardiovascular Diseases; Catabolism; Cell physiology; Cholesterol; Cholesterol Homeostasis; Chronic; Complex; Cytochrome P450; Data; Defect; Deposition; Development; Diagnostic; Diet; Disease; Electroretinography; Enzymes; Event; Excision; Exudative age-related macular degeneration; Eye; Eye diseases; Fluorescein Angiography; Funding; Genes; Genetic; Goals; Grant; High Density Lipoproteins; Histocytochemistry; Human; Individual; Inflammation; Intervention; Knowledge; Label; Lead; Life; Link; Lipids; Lipoproteins; Maintenance; Mammals; Mass Spectrum Analysis; Measures; Mediating; Methodology; Methods; Modification; Molecular Biology; Mus; Optical Coherence Tomography; Oxidative Stress; Pathologic; Pathology; Phenotype; Play; Polyunsaturated Fatty Acids; Post-Translational Protein Processing; Prevention; Procedures; Protein Isoforms; Proteins; Protocols documentation; Pyridoxamine; Research; Retina; Retinal; Retinal Neovascularization; Risk Factors; Role; Specimen; Sterols; Structure; Structure of retinal pigment epithelium; Testing; Tissues; Vascularization; Vision; Visual; Visual impairment; Voriconazole; base; behavior test; cholesterol biosynthesis; density; in vivo; insight; instrumentation; mRNA Expression; multiple reaction monitoring; novel therapeutics; oxidation; premature; prevent; protein expression; protein function; senescence; therapeutic target

DESCRIPTION (provided by applicant): Cholesterol is essential for life in mammals. Yet, chronic excess of cholesterol is a risk factor for cardiovascular and Alzheimer's diseases and likely age-related macular degeneration (AMD). Cholesterol and other lipids accumulate with age between the retinal pigment epithelium and Bruch's membrane, but in people with AMD this accumulation is more prominent suggesting defects in cholesterol removal. Cholesterol removal always involves two mechanisms, lipoprotein-mediated transport and catabolism to oxysterols by cytochrome P450 enzymes (CYP) including CYPs 27A1 and 46A1. While recent genetic studies revealed an association between AMD and genes affecting the formation of high density lipoproteins, the significance of the enzymatic cholesterol removal from the retina is still unknown. We only know that in humans, ocular manifestations of CYP27A1 deficiency include premature retinal senescence and cholesterol deposition, and that inhibition of CYP46A1 with voriconazole causes visual disturbances. To begin to gain insight into cholesterol catabolism in the retina, we analyzed cadaveric human eyes and established that retinal cholesterol is mainly metabolized by CYP27A1 while the other P450 isoform, CYP46A1, plays a lesser role. We also found that retinal levels of the CYP27A1 protein do not correlate with the levels of its metabolite 5-cholestenoic acid and ascertained the basis for this discrepancy. We demonstrated that CYP27A1 is subjected to a deleterious post-translational modification by isolevuglandins generated from arachidonic acid. We also found that mice lacking one (CYP27A1) or two (CYPs 27A1 and 46A1) cholesterol-catabolizing enzymes have structural retinal pathologies, altered blood flow and pathologic retinal vascularization. These findings confirm that enzymatic cholesterol elimination is an important contributor to the normal function of the retina. In this competing renewal, we propose to further explore our findings. The Specific Aims of this application are: 1) to characterize the ocular phenotype of mice with impaired cholesterol catabolism; 2) to identify the major mechanisms regulating retinal CHO levels; and 3) to examine the isoLG-adduction in the retina. Advanced diagnostic instrumentation, state-of-the-art mass spectrometry methodologies, biochemical and molecular biology analyses of human and mouse specimens, as well as dietary and pharmacological interventions in mice will be used to address the goals of the project. The data obtained will significantly advance our understanding of the role of cholesterol in vision and the mechanisms for pathologic vascularization in the retina. These studies may lead to the identification of therapeutic targets for prevention and treatment of neovascular AMD. PUBLIC HEALTH RELEVANCE: This research will be critical for understanding the role of cholesterol in development of certain eye diseases that reduce vision.

描述（申请人提供）：胆固醇对哺乳动物的生命至关重要。然而，慢性过量胆固醇是心血管和阿尔茨海默氏病的危险因素，可能是与年龄有关的黄斑变性（AMD）。胆固醇和其他脂质随着视网膜色素上皮和Bruch膜的年龄而积聚，但是在AMD的人中，这种积累更为突出，这表明去除胆固醇的缺陷。去除胆固醇始终涉及两种机制：脂蛋白介导的转运和分解代谢，包括细胞色素P450酶（CYP），包括CYPS 27A1和46A1。虽然最近的遗传研究揭示了AMD与影响高密度脂蛋白形成的基因之间的关联，但从视网膜中去除酶促胆固醇的重要性仍然未知。我们只知道在人类中，CYP27A1缺乏的眼部表现包括过早的视网膜衰老和胆固醇沉积，并且用伏立康唑抑制CYP46A1会引起视觉障碍。为了开始深入了解视网膜中胆固醇的分解代谢，我们分析了尸体人眼睛，并确定视网膜胆固醇主要由CYP27A1代谢，而其他P450同型CYOForm，CYP46A1扮演的角色较小。我们还发现，CYP27A1蛋白的视网膜水平与其代谢物的水平无关 5-胆固醇酸并确定了这种差异的基础。我们证明了CYP27A1受到蛛网膜酸产生的异甲抗物素的经过有害的翻译后修饰。我们还发现，缺乏一只（CYP27A1）或两只（CYPS 27A1和46A1）胆固醇促胆固醇酶的小鼠具有结构性视网膜病理，血液流动和病理性视网膜血管化的改变。这些发现证实，酶促胆固醇消除是视网膜正常功能的重要促进者。在这种竞争的续约中，我们建议进一步探索我们的发现。该应用的具体目的是：1）表征胆固醇分解代谢受损的小鼠的眼部表型； 2）确定调节视网膜CHO水平的主要机制； 3）检查视网膜中的隔离。先进的诊断仪器，最先进的质谱方法，人和小鼠标本的生化和分子生物学分析，以及小鼠的饮食和药理干预措施，以解决项目的目标。获得的数据将大大提高我们对视觉中胆固醇作用以及视网膜病理血管形成机制的理解。这些研究可能导致鉴定用于预防和治疗新血管AMD的治疗靶标。 公共卫生相关性：这项研究对于理解胆固醇在某些减少视力的眼病发展中的作用至关重要。