Functional Analysis of Novel Genes in Eye Development and Vision

眼睛发育和视力新基因的功能分析

• 批准号: 9555682
• 负责人: graeme j wistow
• 金额: $ 116.04万
• 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/9555682
• 关键词: Afferent Neurons; Age related macular degeneration; Aging; Alpha Cell; Animal Model; Apical; Auditory; Bioinformatics; Biophysics; Blindness; Bruch' s basal membrane structure; Cataract; Cell Culture Techniques; Cells; Cellular biology; Centrosome; Cholesterol; Cilia; Complement Factor H; Complex; Defect; Deposition; Disease; Employee Strikes; Epithelium; Eye; Eye Development; Genes; Genomics; Glaucoma; Growth; Hair Cells; Human; Hybrids; Knock-out; Knockout Mice; Labyrinth; Lens Fiber; Mass Spectrum Analysis; Methods; MicroRNAs; Modeling; Pathway interactions; Photoreceptors; Process; Proteins; Publishing; Recombinant Proteins; Retina; Retinal Photoreceptors; Ribonucleases; Role; S1-5 protein; Serum; Surgical sutures; Techniques; Tissues; Up-Regulation; Vision; Visual; Work; Yeasts; Zinc; age related; capillary bed; ciliopathy; deprivation; gene product; lens; mouse model; novel; protein biomarkers; response; retbindin; zinc-binding protein

Age-related macular degeneration (AMD) is a major cause of vision loss. We have shown that dry AMD is associated with specific deposits of complement factor H and fibulin 3 within domains rich in cholesterol basal to the RPE. Mass spectroscopy has been used to identify further components of complexes that are implicated in formation of protein depositions in AMD. We have developed a cell-culture model for serum-deprivation AMD using RPE-derived cells. In AMD, changes at Bruch's membrane and in the capillary bed are likely to restrict access of serum components to the RPE. We have shown that serum deprivation of RPE cells leads to a marked upregulation of cholesterol synthesis and transport and the accumulation of cholesterol in the RPE. This is strongly reminiscent of the accumulation of cholesterol RPE that we and others have seen in human AMD. Furthermore, serum-deprivation leads to depletion of intracellular zinc, while many zinc-binding proteins are induced. AREDS studies have shown that zinc is protective against AMD. We have extended this work to further define the process in the starved cells and have identified other pathways relevant to AMD. This work has used cell biology and RNAsed methods. Retbindin is a novel protein of retinal photoreceptors. A knockout mouse model shows progressive deficits in visual response and age-related defects in the outer retina that have some striking similarities to some forms of age-related macular degeneration and to glaucoma. Characterization of this model shows many marked similarities to AMD, including deposition of key proteins that are markers for human AMD. In the lens, we have two mouse models for age-related cortical cataract in knockouts for the lens-specific proteins KLPH and Lengsin. We show that KLPH is specific to lens epithelium and is required for normal lens suture formation. Deletion of KLPH leads to complete loss of expression of Clic5 in the lens. In normal lens, Clic5 is localized to the cilium/centrosome complex at the apical tip of the lens fibers. Clic5 is known to be associated with a ciliopathy in the inner ear. We have now published the results of an extensive collaborative study of a targeted deletion model for the miR183C miRNA cluster that has an important role in maturation of sensory neurons, including photoreceptors and auditory hair cells. The deletion has aspects of ciliopathies and provides a possible mouse model for Ushers-like disease.

与年龄相关的黄斑变性（AMD）是视力丧失的主要原因。我们已经表明，干燥的AMD与富含RPE的胆固醇的结构域内的补体H和Fibulin 3的特定沉积有关。质谱学已用于鉴定与AMD中蛋白质沉积形成有关的复合物的进一步成分。我们已经开发了一种使用RPE衍生的细胞的细胞培养模型，用于血清剥夺AMD。在AMD中，BRUCH的膜和毛细管床的变化可能会限制血清成分进入RPE。我们已经表明，RPE细胞的血清剥夺导致胆固醇合成和转运的明显上调以及胆固醇在RPE中的积累。这强烈让人联想到我们和其他人在人类AMD中看到的胆固醇RPE的积累。此外，血清剥夺会导致细胞内锌的消耗，而许多锌结合蛋白是诱导的。 AREDS研究表明，锌具有保护性AMD。我们已经扩展了这项工作，以进一步定义饥饿的细胞中的过程，并确定了与AMD相关的其他途径。这项工作使用了细胞生物学和基于RN的方法。 视毒素是视网膜感光体的一种新型蛋白质。敲除小鼠模型显示出视觉反应的逐渐缺陷和外部视网膜中与年龄相关的缺陷，这些缺陷与某些形式的与年龄相关的黄斑变性和青光眼具有明显的相似性。该模型的表征显示了与AMD的许多明显相似性，包括沉积是人类AMD标记的关键蛋白。 在镜头中，我们有两个小鼠模型用于与年龄相关的皮质性白内障，用于敲除晶状体特异性蛋白KLPH和Lengsin的敲除。我们表明，KLPH是特定于晶状体上皮的，是正常透镜缝合形成所必需的。 KLPH的缺失导致镜头中Clic5表达的完全丧失。在正常晶状体中，Clic5位于晶状体纤维顶端的纤毛/中心体络合物中。已知Clic5与内耳中的纤毛病有关。 现在，我们已经发布了对MiR183C miRNA群集的靶向缺失模型的广泛协作研究的结果，该模型在感觉神经元的成熟（包括光感受器和听觉毛细胞）中具有重要作用。缺失具有纤毛病的各个方面，并为引诱者样疾病提供了可能的小鼠模型。