Molecular Basis of Retinal Angiogenesis

项目摘要 过氧化物酶体增殖物激活受体（PPARs）是核受体超家族的成员， 配体激活的转录因子。三种亚型，PPAR-<$，PPAR-<$/<$和PPAR-<$，每一种都由一个 单个基因，已经被识别。PPARs可以被多种配体激活，包括天然的 存在的脂肪酸、贝特类和甘草素。在上一个项目期间，我们对 视网膜血管生成中的胰高血糖素确定了PPARs，特别是PPARs-β/β，作为潜在的治疗靶点。 所有三种PPARs均在血管内皮中表达。在这种情况下，PPAR-<$和PPAR-<$通常 显示出抗炎、抗增殖和抗血管生成作用。相比之下， 关于过氧化物酶体增殖物激活受体，虽然它已被证明可以刺激血管内皮细胞的增殖， 在某些癌细胞系中，活化导致VEGF诱导，但在其它细胞系中则不然。先前的研究清楚地表明 这三种PPARs通过组织和细胞的高度多效性机制影响血管功能， 特定.因此，为了理解视网膜血管生成背景下的这些机制， 表达和活化必须在所涉及的特定视网膜细胞中表征。这些细胞 产生血管生成因子的M？ller细胞和视网膜微血管内皮细胞（RMEC）， 对血管生成因子有反应。使用这两种细胞类型的原代培养物， 可以研究过氧化物酶体增殖物激活受体（PPAR）、生长因子产生和血管生成细胞行为之间的相互作用 系统地，导致识别适当的PPARs治疗操作。我们的初步 数据表明，在与早期和晚期炎症相关的条件下， 晚期糖尿病视网膜病变（DR）。因此，我们建议使用工具的组合来表征的作用， 视网膜血管疾病中的PPAR-/：1）从野生型和PPAR-/-/-小鼠中分离的细胞的比较; 2） 从野生型小鼠分离的细胞中的高选择性激动剂和拮抗剂; 3）野生型和 在早期和晚期DR模型中的PPAR-/-/-小鼠;和4）在早期和晚期DR模型中的高度特异性激动剂和拮抗剂。 野生型小鼠和大鼠中的DR。我们的体内模型已被选择为与早期（STZ-1） 诱发糖尿病）和晚期（氧诱导视网膜病变）DR。我们认为这种策略是重点， 全面，它将产生重要的信息，我们对DR的基本理解和 制定合理的治疗方法。

PROJECT SUMMARY Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear receptor superfamily of ligand-activated transcription factors. Three subtypes, PPAR-¿, PPAR-¿/¿ and PPAR-¿, each encoded by a single gene, have been identified. PPARs can be activated by a variety of ligands that include naturally occurring fatty acids, fibrates and prostaglandins. During the last project period, our studies of the roles of prostaglandins in retinal angiogenesis identified PPARs, specifically PPAR-¿/¿, as potential therapeutic targets. All three PPARs are expressed in vascular endothelium. In this context, PPAR-¿ and PPAR-¿ generally demonstrate anti-inflammatory, anti-proliferative, and anti-angiogenic effects. In contrast, much less is known regarding PPAR-¿/¿, although it has been shown to stimulate proliferation of vascular endothelial cells, and its activation leads to VEGF induction in some cancer cell lines, but not others. Previous work clearly suggests that all three PPARs influence vascular function by highly pleiotropic mechanisms that are tissue- and cell- specific. Thus, in order to understand these mechanisms in the context of retinal angiogenesis, PPAR expression and activation must be characterized in the specific retinal cells that are involved. These cells are the M¿ller cells, which produce angiogenic factors, and the retinal microvascular endothelial cells (RMEC), which respond to the angiogenic factors. Using primary cultures of these two cell types, the complicated interactions between PPAR-¿/¿, growth factor production and angiogenic cell behaviors can be studied systematically, leading to the identification of appropriate therapeutic manipulations of PPARs. Our preliminary data indicate that PPAR-¿/¿ is activated in M¿ller cells and RMEC under conditions relevant to both early and late diabetic retinopathy (DR). Thus, we propose to use a combination of tools to characterize the role of PPAR-¿/¿ in retinal vascular disease: 1) comparisons of cells isolated from wild type and PPAR-¿/¿-/- mice; 2) highly selective agonists and antagonists in cells isolated from wild type mice; 3) comparisons of wild type and PPAR-¿/¿-/- mice in models of early and late DR; and 4) highly specific agonists and antagonists in models of DR in wild type mice and in rats. Our in vivo models have been selected for their relevance to early (STZ- induced diabetes) and late (oxygen-induced retinopathy) DR. We believe this strategy is focused and comprehensive, and it will yield information of importance to our fundamental understanding of DR and to the development of rational therapeutic approaches.