Genetic susceptibility and common multi-factorial retinal/macular diseases

遗传易感性和常见的多因素视网膜/黄斑疾病

• 批准号: 7968423
• 负责人: ANAND SWAROOP
• 金额: $ 176.09万
• 依托单位: NATIONAL EYE INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7968423
• 关键词: Aging; Antibodies; Cell Culture Techniques; Cell physiology; Cells; Cholesterol Homeostasis; Chromosomes, Human, Pair 22; Diabetic Retinopathy; Elderly; Electroporation; Electroretinography; Etiology; Gene Transfer; Genes; Genetic Polymorphism; Genetic Predisposition to Disease; Genomics; Genotype; Genus Hippocampus; Goals; Homeostasis; Human; In Vitro; Lead; Maintenance; Measures; Mediating; Meta-Analysis; Metabolic; Methods; Microarray Analysis; Mitochondria; Mitochondrial Proteins; Monkeys; Mus; Photoreceptors; Predisposition; Proteins; Protocols documentation; RNA Interference; Retina; Retinal; Retinal Degeneration; Role; Sorting - Cell Movement; Structure of retinal pigment epithelium; Techniques; Variant; base; case control; cohort; disease phenotype; disorder of macula of retina; follow-up; genetic variant; genome wide association study; in vivo; insertion/deletion mutation; mouse model; novel; retinal rods

We have recently completed a whole-genome scan of a large cohort of case-controls for finding new AMD-associated variants. Replication studies suggest that a new region on chromosome 22 is associated with susceptibility to AMD. Based on the GWAS study, we performed follow-up genotyping to narrow down a susceptibility region. Furthermore, we established a protocol for gene transfer into the developing mouse retinal pigment epithelium (RPE) using in vivo electroporation. Utilizing this technique, we have studied the roles of genes implicated in AMD identified through GWAS. Genes involved in cholesterol metabolism and transport are being evaluated by RNAi-mediated knockdown studies. Information about the functional contribution of these genes in RPE cell physiology and maintenance could lead to further elucidation of the etiology of AMD. We recently identified a key AMD-susceptibility variant in ARMS2 gene that encodes a mitochondrial protein based on in vitro cell culture studies. We have generated a number of ARMS2 antibodies to perform in vivo localization of ARMS2 protein in human and monkey retina. We are also examining how a specific insertion-deletion polymorphism can lead to AMD susceptibility. Finally, we are determining cellular/functional changes associated with aging of the retina in order to understand their contribution to the onset of retinal degeneration. We have combined microarray analysis of flow-sorted rod photoreceptors from young and aging mice, validated with qPCR and IHC, with electroretinograms and a novel method (Seahorse XF24 Analyzer) to measure mitochondrial function in enzymatically-dissociated retina cells.

我们最近完成了对大量病例对照的全基因组扫描，以寻找新的 AMD 相关变异。复制研究表明，22 号染色体上的一个新区域与 AMD 易感性相关。基于 GWAS 研究，我们进行了后续基因分型以缩小易感区域。此外，我们建立了使用体内电穿孔将基因转移到发育中的小鼠视网膜色素上皮（RPE）的方案。 利用这项技术，我们研究了通过 GWAS 鉴定的与 AMD 相关的基因的作用。 参与胆固醇代谢和运输的基因正在通过 RNAi 介导的敲低研究进行评估。有关这些基因在 RPE 细胞生理学和维护中的功能贡献的信息可能有助于进一步阐明 AMD 的病因。 我们最近根据体外细胞培养研究，在编码线粒体蛋白的 ARMS2 基因中发现了一个关键的 AMD 易感性变异。 我们已经生成了许多 ARMS2 抗体，用于在人和猴子视网膜中进行 ARMS2 蛋白的体内定位。我们还在研究特定的插入缺失多态性如何导致 AMD 易感性。 最后，我们正在确定与视网膜衰老相关的细胞/功能变化，以了解它们对视网膜变性发生的贡献。我们对年轻和衰老小鼠的流式分选视杆光感受器进行微阵列分析，并通过 qPCR 和 IHC 进行验证，并结合视网膜电图和一种新方法（Seahorse XF24 分析仪）来测量酶解视网膜细胞中的线粒体功能。