Molecular insights into the pathogenesis of Sorsby Fundus Dystrophy

索斯比眼底营养不良发病机制的分子见解

基本信息

批准号：
8518343
负责人：
Jian Hua Qi
金额：
$ 18.64万
依托单位：
CLEVELAND CLINIC LERNER COM-CWRU
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-08-01 至 2014-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8518343
关键词：
Accounting Address Age related macular degeneration Angiogenesis Inhibitors Biochemical Blindness Blood Vessels Bruch&apos s basal membrane structure Characteristics Choroid Choroidal Neovascularization Coupled Degenerative Disorder Deposition Development Diabetic Retinopathy Disease Employee Strikes Endothelial Cells Epithelial Event Extracellular Matrix Eye Functional disorder Genes In Vitro Induced Mutation Inherited Knock-in Mouse Macular degeneration Matrix Metalloproteinases Mediating Modeling Molecular Mutagenesis Mutation N-Glycosylation Site Pathogenesis Pathway interactions Patients Phenotype Play Process Property Protein Glycosylation Proteins Rare Diseases Research Resistance Retinal Degeneration Retinal Pigments Role Sorsby&apos s fundus dystrophy Structure Structure of retinal pigment epithelium Testing Therapeutic Tissue Inhibitor of Metalloproteinase-3 Tissues Vascular Endothelial Growth Factor Receptor-2 Work angiogenesis base design dimer disease phenotype early onset glycosylation in vivo innovation insight mouse model mutant neovascularization novel novel therapeutic intervention prevent therapeutic target

项目摘要

DESCRIPTION (provided by applicant): Tissue Inhibitors of Metalloproteinase-3 (TIMP3) is a regulator of matrix metalloproteinase (MMPs) and a potent angiogenesis inhibitor. Sorsby fundus dystrophy (SFD), a rare, dominantly inherited, early onset macular degenerative disease that is caused by mutations in the TIMP3 gene has marked similarities to age- related macular degeneration (ARMD) disease phenotype including an increased accumulation of TIMP3 in Bruch's membrane (BM) and the development of choroidal neovasculaization (CNV), a major cause of severe vision loss in patients. Our studies using in vitro and in vivo SFD models indicate that SFD-related S156C- TIMP3 promotes CNV via vascular endothelial growth factor receptor-2 (VEGFR-2)- and retinal pigment epithelial (RPE) cell-dependent angiogenic pathways. We also found that increased mutant protein glycosylation is a characteristic of vascular endothelial and RPE cells. Furthermore, we have evidence of increased mutant TIMP3 protein glycosylation with a concomitant increase in mutant TIMP3 levels in RPE- choroid tissue of S156C-TIMP3 knock-in mice. We hypothesize that that S156C-TIMP3 mutation induces N- glycosylation at position184, the only potential N-glycosylation site of TIMP3, resulting in increased accumulation in BM and/or CNV development via VEGFR-2-and/or RPE cell-dependent angiogenic pathways. Using in vitro and in vivo SFD models coupled with pharmacological, mutagenesis, protein over-expression and biochemical characterization approaches our hypothesis will be tested with the following Specific Aims: 1. To determine if inhibition of TIMP-3 glycosylation prevents the increased accumulation of mutant protein in BM and CNV development in S156C-Timp3 knock-in mice. 2. To determine if mutation of Asn184 in S156C-TIMP3 inhibits VEGFR-2- and RPE cell-mediated angiogenesis. The long-term objectives of the proposed research is to identify and dissect the consequences of SFD mutations on TIMP3 structure and functions with particular emphasis on TIMP3 glycosylation as a potential molecular event mediating its accumulation in BM and the subsequent angiogenic or CNV phenotype. Undoubtedly, this would provide novel insights into the pathogenesis of SFD and ARMD that share similar disease phenotype, and thereby establish a potential therapeutic target for these ocular diseases.

描述（由申请人提供）：金属蛋白酶组织抑制剂-3（TIMP3）是基质金属蛋白酶（MMP）的调节剂和有效的血管生成抑制剂。索斯比眼底营养不良 (SFD) 是一种罕见的显性遗传性早发性黄斑变性疾病，由 TIMP3 基因突变引起，与年龄相关性黄斑变性 (ARMD) 疾病表型有显着相似之处，包括 TIMP3 在 Bruch 膜中积累增加（BM）和脉络膜新生血管（CNV）的发展，这是患者严重视力丧失的主要原因。我们使用体外和体内 SFD 模型的研究表明，SFD 相关的 S156C-TIMP3 通过血管内皮生长因子受体 2 (VEGFR-2) 和视网膜色素上皮 (RPE) 细胞依赖性血管生成途径促进 CNV。我们还发现突变蛋白糖基化增加是血管内皮细胞和 RPE 细胞的一个特征。此外，我们有证据表明，S156C-TIMP3 敲入小鼠的 RPE 脉络膜组织中突变型 TIMP3 蛋白糖基化增加，同时突变型 TIMP3 水平也随之增加。我们假设 S156C-TIMP3 突变在 184 位（TIMP3 唯一潜在的 N-糖基化位点）诱导 N-糖基化，从而通过 VEGFR-2 和/或 RPE 细胞依赖性血管生成途径导致 BM 和/或 CNV 发育中积累增加。使用体外和体内 SFD 模型，结合药理学、诱变、蛋白质过表达和生化表征方法，我们的假设将通过以下具体目标进行测试： 1. 确定 TIMP-3 糖基化的抑制是否会阻止突变体积累的增加S156C-Timp3 敲入小鼠的 BM 和 CNV 发育中的蛋白质。 2.确定S156C-TIMP3中Asn184的突变是否抑制VEGFR-2和RPE细胞介导的血管生成。拟议研究的长期目标是识别和剖析 SFD 突变对 TIMP3 结构和功能的影响，特别强调 TIMP3 糖基化作为介导其在 BM 中积累和随后的血管生成或 CNV 表型的潜在分子事件。毫无疑问，这将为具有相似疾病表型的 SFD 和 ARMD 的发病机制提供新的见解，从而为这些眼部疾病建立潜在的治疗靶点。