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.

视网膜相关性黄斑变性（AMD）是视力丧失的主要原因。我们已经证明干性AMD与RPE基底富含胆固醇的区域内补体因子H和纤蛋白3的特异性沉积有关。质谱已被用于鉴定涉及AMD中蛋白质沉积形成的复合物的其他组分。我们已经开发了一个细胞培养模型，用于血清剥夺AMD使用RPE衍生的细胞。在AMD中，布鲁赫膜和毛细血管床的变化可能限制血清组分进入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显著上调以及胆固醇在RPE中的积累。这强烈地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，血清剥夺导致细胞内锌耗尽，同时诱导许多锌结合蛋白。 AREDS研究表明，锌对AMD有保护作用。我们已经扩展了这项工作，以进一步定义饥饿细胞中的过程，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RNAsed方法。 Retbindin是一种新的视网膜光感受器蛋白。基因敲除小鼠模型显示视觉反应的进行性缺陷和外层视网膜中与年龄相关的缺陷，这些缺陷与某些形式的年龄相关性黄斑变性和青光眼有一些惊人的相似之处。该模型的表征显示出与AMD的许多显著相似性，包括作为人类AMD标志物的关键蛋白质的沉积。 在透镜中，我们有两个年龄相关性皮质性白内障的小鼠模型，敲除晶状体特异性蛋白KLPH和Lengsin。我们发现，KLPH是特定的透镜上皮细胞和所需的正常透镜缝合形成。KLPH的缺失导致透镜中Clic 5表达的完全丧失。在正常透镜中，Clic 5定位于透镜纤维顶端的纤毛/中心体复合体。已知Clic 5与内耳纤毛病变有关。 我们现在已经发表了一项广泛合作研究的结果，该研究针对miR 183 C miRNA簇的靶向缺失模型，该模型在感觉神经元（包括光感受器和听觉毛细胞）的成熟中发挥着重要作用。该缺失具有纤毛病的特征，并为Ushers样疾病提供了可能的小鼠模型。