Study of the RPE barrier

RPE屏障的研究

• 批准号: 7949302
• 负责人: YUN Zheng LE
• 金额: $ 37万
• 依托单位: UNIVERSITY OF OKLAHOMA HLTH SCIENCES CTR
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7949302
• 关键词: Age related macular degeneration; Albumins; Animal Model; Biochemical; Biochemical Pathway; Biological Assay; Biological Markers; Blindness; Blood; Blood Circulation; Blood Vessels; Cell Culture Techniques; Cells; Data; Diabetes Mellitus; Diabetic Retinopathy; Disease; Endothelial Cells; Epithelial; Epithelial Cells; Experimental Models; Extravasation; Functional disorder; Human; Imagery; Immunoblotting; In Vitro; Inflammation; Inflammatory; Intestines; Ischemia; Kidney; Knockout Mice; Knowledge; Literature; Measures; Mediating; Methods; Microscopic; Molecular; Muller' s cell; Mus; Neuroglia; Nose; Pathogenesis; Pathology; Patients; Permeability; Phosphorylation; Physiological; Play; Positioning Attribute; Protein Isoforms; Protein Kinase C; Proteins; Public Health; Regulation; Retina; Retinal; Retinal Detachment; Retinal Neovascularization; Retinal Pigments; Retinopathy of Prematurity; Role; Severities; Signal Transduction; Structure of retinal pigment epithelium; System; Testing; Therapeutic; Tight Junctions; Uveitis; Vascular Diseases; Vascular Endothelial Growth Factor Receptor; Vascular Endothelial Growth Factor Receptor-2; Vascular Endothelial Growth Factors; Vascular Permeabilities; autocrine; autoimmune uveitis; base; bevacizumab; cellular targeting; design; diabetic; inhibitor/antagonist; knockout gene; macular edema; monolayer; occludin; public health relevance; receptor; research study; success; therapeutic target

DESCRIPTION (provided by applicant): The inner and outer blood-retina barriers (BRBs) are formed by tight junctions between adjacent endothelial or retinal pigment epithelial (RPE) cells. Breakdown of BRBs is a major pathological change in age-related macular degeneration, diabetic retinopathy, retinopathy of prematurity, and uveitis. While previous studies have yielded a better understanding about the roles of the inner BRB under physiological and pathological conditions, surprisingly little is known about the regulation and pathophysiology of the outer BRB. Since the outer BRB is responsible for ~85% of blood circulation to the retina, it is unimaginable that ischemia-induced pathological change in the outer BRB plays an insignificant role in the overall pathology of retinochoroidal vascular diseases. To investigate the mechanisms of outer BRB breakdown and to test the concept of inhibiting outer BRB permeability as a therapeutic strategy for retinochoroidal vascular diseases, we have prepared gene knockout systems for the mouse RPE and Muller glia, cells that regulate the function of both inner and outer BRBs through vascular endothelial growth factor (VEGF-A). Using these conditional gene knockout systems, we have disrupted VEGF and its receptor (VEGFR2) in the mouse RPE and have generated mice with VEGF disruption in the Muller cells. In Specific Aim 1, we will test our hypothesis that outer BRB breakdown is a significant contributor to diabetes/ischemia-induced overall retinal "vascular leakage" through autocrine VEGF/VEGF-R2 signaling in the RPE by measuring the total retinal vascular leakage, the number of significant breakpoints in the outer BRB, and the quantity of outer BRB-specific leakage in the RPE-specific VEGF and VEGFR2 knockout mice after inducing ischemia or diabetes. In Specific Aim 2, we will test the concept of inhibiting outer BRB permeability as a therapeutic strategy for uveitis by examining the total retinal vascular leakage, the number and severity of retinal detachment, the quantity of outer BRB-specific leakage, and the expression of inflammatory biomarkers in the RPE-specific VEGFR2 knockout mice after inducing uveitis. As a control for inner BRB breakdown, we will also measure the same parameters in uveitic Muller cell- specific VEGF knockout mice. In Specific Aim 3, we will determine the molecular mechanism of diabetes/ischemia-induced outer BRB breakdown by investigating the biochemical pathway governing the regulation of tight-junction proteins. PUBLIC HEALTH RELEVANCE: This application is relevant to an important public health issue: the pathogenic mechanism of leading causes of blindness: diabetic retinopathy, retinopathy of prematurity, and uveitis. Our study will focus on the regulatory mechanism and the pathophysiology of blood-retina barriers.

描述（由申请人提供）：内部和外部血视网膜屏障（BRB）由相邻内皮或视网膜色素上皮（RPE）细胞之间的紧密连接形成。 BRBs的分解是年龄相关性黄斑变性、糖尿病性视网膜病变、早产儿视网膜病变和葡萄膜炎的主要病理变化。虽然之前的研究对内部 BRB 在生理和病理条件下的作用有了更好的了解，但令人惊讶的是，人们对外部 BRB 的调节和病理生理学知之甚少。由于外层 BRB 负责约 85% 的视网膜血液循环，因此难以想象外层 BRB 缺血引起的病理变化在视网膜脉络膜血管疾病的整体病理学中起着微不足道的作用。为了研究外层 BRB 分解的机制并测试抑制外层 BRB 通透性作为视网膜脉络膜血管疾病治疗策略的概念，我们为小鼠 RPE 和 Muller 胶质细胞制备了基因敲除系统，这些细胞通过血管内皮生长因子 (VEGF-A) 调节内层和外层 BRB 的功能。使用这些条件基因敲除系统，我们破坏了小鼠 RPE 中的 VEGF 及其受体 (VEGFR2)，并产生了 Muller 细胞中 VEGF 被破坏的小鼠。 在具体目标 1 中，我们将通过测量总视网膜血管渗漏、外层 BRB 中显着断点的数量以及 RPE 特异性 VEGF 和外层 BRB 特异性渗漏的数量来测试我们的假设，即外层 BRB 破裂是通过 RPE 中自分泌 VEGF/VEGF-R2 信号传导导致糖尿病/缺血引起的总体视网膜“血管渗漏”的重要因素。 诱导缺血或糖尿病后的 VEGFR2 敲除小鼠。在具体目标 2 中，我们将通过检查诱导葡萄膜炎后 RPE 特异性 VEGFR2 敲除小鼠的总视网膜血管渗漏、视网膜脱离的数量和严重程度、外层 BRB 特异性渗漏的数量以及炎症生物标志物的表达，来测试抑制外层 BRB 通透性作为葡萄膜炎治疗策略的概念。作为内部 BRB 分解的对照，我们还将在葡萄膜炎 Muller 细胞特异性 VEGF 敲除小鼠中测量相同的参数。在具体目标 3 中，我们将通过研究控制紧密连接蛋白调节的生化途径来确定糖尿病/缺血引起的外部 BRB 分解的分子机制。 公共健康相关性：本申请与一个重要的公共健康问题相关：导致失明的主要原因：糖尿病性视网膜病变、早产儿视网膜病变和葡萄膜炎。我们的研究将重点关注血视网膜屏障的调节机制和病理生理学。