Mechanisms of Retinal Vascular Permeability in Diabetes

糖尿病视网膜血管通透性的机制

• 批准号: 8460893
• 负责人: David Antonetti
• 金额: $ 40.64万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-01 至 2014-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8460893
• 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

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.

描述（由申请人提供）：糖尿病视网膜病变仍然是美国失明的主要原因，没有FDA批准的药物治疗。本提案的目的是了解导致血管通透性增加和黄斑水肿的分子机制，并阐明血管通透性与血管生成的关系，以便开发治疗或预防糖尿病视网膜病变的新疗法。糖尿病视网膜病变中的血管功能障碍可能由高血糖、晚期糖基化终产物和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如血管内皮生长因子（VEGF）和炎性细胞因子如肿瘤坏死因子（TNF）的间接作用引起。在目前的提议中，假设VEGF和TNF改变了紧密连接复合物，导致内皮通透性增加。在之前的资助期间的研究已经证明，VEGF以蛋白激酶C依赖的方式诱导紧密连接蛋白occludin的磷酸化，这与血管通透性相关。此外，VEGF诱导闭合蛋白和其他紧密连接蛋白从质膜到细胞质的再分布，并且随着时间的推移，导致闭合蛋白的降解。通过质谱分析闭合蛋白磷酸化位点已经鉴定了VEGF响应性磷酸化位点。在这个建议中，数据表明，occludin的突变，以防止磷酸化，阻断VEGF诱导的渗透性和occludin的内吞作用。此外，初步数据表明，TNF改变紧密连接复合物，通过减少紧密连接蛋白claudin 5和封闭小带1，但不减少occludin含量，这表明至少部分不同的机制。封闭蛋白含量与具有紧密连接的细胞的增殖密切相关。在视网膜色素上皮细胞中沉默occludin表达诱导细胞增殖增加两倍。因此，我们将研究VEGF和TNF改变紧密连接复合物诱导内皮通透性的机制，以及VEGF诱导的occludin磷酸化和降解与血管生成的关系。虽然VEGF和TNF在occludin的控制方面存在分歧，但初步数据表明这两种因素都利用非典型PKC途径来改变连接复合物并诱导内皮渗透性。针对非典型PKC通路单独或与PKC抑制剂联合的治疗可能提供控制糖尿病视网膜病变和其他涉及VEGF和炎性细胞因子的视网膜疾病的血管通透性的有效手段。