Mechanisms of Retinal Vascular Permeability in Diabetes

糖尿病视网膜血管通透性的机制

基本信息

批准号：
8462825
负责人：
David Antonetti
金额：
$ 6.11万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
依托单位国家：
美国
项目类别：
财政年份：
1998
资助国家：
美国
起止时间：
1998-09-01 至 2014-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8462825
关键词：
Advanced Glycosylation End Products Alanine Biochemical Blindness Blood Vessels Blood-Retinal Barrier CCL2 gene Cattle Cell Death Cell Proliferation Cell division Cell membrane Cells Complex Cytoplasm Data Development Diabetes Mellitus Diabetic Retinopathy Disease Dose Elements Endocytosis Endosomes Endothelial Cells Epithelial FDA approved Figs - dietary Functional disorder Funding Gene Deletion Gene Expression Goals Golgi Apparatus Growth Factor Health Hyperglycemia Hyperlipidemia Hyperplasia Immigration Inflammatory Integral Membrane Protein Interleukin-1 Laboratories Lead Life Link Literature Mass Spectrum Analysis Measures Medical Metabolic Molecular Mus Mutate Mutation Oxygen Pathogenesis Pathway interactions Permeability Phenotype Phosphorylation Phosphorylation Site Proliferating Protein Isoforms Protein Kinase C Proteins Publishing Rattus Receptor Gene Regulation Research Retina Retinal Retinal Diseases Role Signal Pathway Signal Transduction Site Small Interfering RNA Stomach Structure of retinal pigment epithelium Testing Therapeutic Therapeutic Intervention Tight Junctions Time Tube Tumor Necrosis Factor Receptor Tumor Necrosis Factor-alpha Ubiquitination United States Vascular Endothelial Cell Vascular Endothelial Growth Factors Vascular Permeabilities angiogenesis atypical protein kinase C base cell growth cellular imaging comparative cytokine epsin immunocytochemistry in vivo inhibitor/antagonist macular edema mutant new therapeutic target novel novel therapeutics occludin overexpression prevent protein kinase C beta research study response therapeutic target trafficking

项目摘要

DESCRIPTION (provided by applicant): Diabetic retinopathy remains a leading cause of blindness in the United States with no FDA approved medical therapy. The goal of this proposal is to understand the molecular mechanisms that lead to increased vascular permeability and macular edema and to elucidate the relationship of vascular permeability to angiogenesis so that novel therapies to treat or prevent diabetic retinopathy may be developed. Vascular dysfunction in diabetic retinopathy may result from both the direct effect of hyperglycemia, advanced glycation end products, and hyperlipidemia on vascular endothelial cells and the indirect effect of these metabolites through induction of growth factors such as vascular endothelial growth factor (VEGF) and inflammatory cytokines such as tumor necrosis factor (TNF). In the current proposal, it is hypothesized that VEGF and TNF alter the tight junction complex leading to increased endothelial permeability. Research over the previous funding period has demonstrated that VEGF induces phosphorylation of the tight junction protein occludin in a protein kinase C dependent manner that is associated with vascular permeability. Further, VEGF induces redistribution of occludin and other tight junction proteins from the plasma membrane to the cell cytoplasm and over time, leads to the degradation of occludin. Analysis of occludin phosphorylation sites by mass spectrometry has identified VEGF responsive phospho-sites. In this proposal, data is presented demonstrating that mutation of occludin to prevent phosphorylation, blocks VEGF-induced permeability and occludin endocytosis. Furthermore, preliminary data demonstrate that TNF alters the tight junction complex by reducing the tight junction proteins claudin 5 and zonula occludens 1 but does not decrease occludin content suggesting at least partially divergent mechanisms. Occludin content is closely associated with proliferation of cells that possess tight junctions. Silencing occludin expression in retinal pigment epithelium cells induces a two-fold increase in cell proliferation. Therefore, we will examine the mechanisms by which VEGF and TNF alter the tight junction complex to induce endothelial permeability and the relationship of VEGF-induced occludin phosphorylation and degradation to angiogenesis. While VEGF and TNF diverge in control of occludin, preliminary data demonstrates both factors utilize the atypical PKC pathway to alter the junctional complex and induce endothelial permeability. Therapies targeting atypical PKC pathway alone or in conjunction with PKC inhibitors may provide an effective means to control vascular permeability in diabetic retinopathy and other retinal diseases involving VEGF and inflammatory cytokines. PUBLIC HEALTH RELEVANCE: The experiments in this proposal will elucidate the mechanisms by which growth factors and inflammatory cytokines alter the tight junction complex of the blood-retinal barrier contributing to macular edema in diabetic retinopathy. Further, the proposal will explore the link between vascular permeability and angiogenesis and the role of the tight junction protein occludin in endothelial cell growth control.

描述（由申请人提供）：糖尿病性视网膜病仍然是美国失明的主要原因，没有FDA批准的医疗疗法。该建议的目的是了解导致血管通透性和黄斑水肿增加的分子机制，并阐明血管通透性与血管生成的关系，以便可以开发出新的治疗或预防糖尿病性视网膜病的疗法。糖尿病性视网膜病中的血管功能障碍可能是由于高血糖，晚期糖基化最终产物的直接作用以及高脂血症对血管内皮细胞的直接作用，以及这些代谢产物通过诱导生长因子（例如血管内皮生长因子（VEGGF））和炎症性Cytokine（TUMOR）诸如TUMOR NECR的诱导因素而造成这些代谢物的间接作用。在当前的提案中，假设VEGF和TNF改变了紧密的连接络合物，导致内皮通透性增加。在上一个资金期间的研究表明，VEGF诱导与血管通透性相关的蛋白激酶C依赖性方式的紧密连接蛋白闭塞蛋白的磷酸化。此外，VEGF诱导叶解蛋白和其他紧密连接蛋白从质膜重新分布，随着时间的流逝，会导致occludin降解。通过质谱法分析occludin磷酸化位点已鉴定出VEGF反应性磷酸化位置。在此提案中，介绍了数据表明，闭塞蛋白的突变可防止磷酸化，阻断VEGF诱导的渗透性和occludin内吞作用。此外，初步数据表明，TNF通过降低紧密连接蛋白Claudin 5和Zonula occludens 1来改变紧密的连接络合物，但不会降低闭合蛋白含量至少表明至少部分发散机制。 occludin含量与具有紧密连接的细胞的增殖密切相关。视网膜色素上皮细胞中的沉默咬合蛋白表达诱导细胞增殖增加了两倍。因此，我们将研究VEGF和TNF改变紧密连接复合物以诱导内皮渗透性以及VEGF诱导的闭塞磷酸化和降解对血管生成的机制。虽然VEGF和TNF在控制occludin方面存在分歧，但初步数据证明了这两个因素都利用非典型的PKC途径来改变连接式复合物并诱导内皮渗透性。仅针对非典型PKC途径的疗法或与PKC抑制剂结合使用，可以提供有效的手段，以控制糖尿病性视网膜病变和其他涉及VEGF和炎性细胞因子的视网膜疾病中的血管通透性。公共卫生相关性：该提案中的实验将阐明生长因子和炎症细胞因子改变血液 - 视网膜屏障的紧密连接络合物的机制，导致糖尿病性视网膜病中黄斑水肿。此外，该提案将探讨血管通透性与血管生成之间的联系，以及紧密连接蛋白闭塞蛋白在内皮细胞生长控制中的作用。