Novel Genes Expressed in Proliferating Endothelial Cells

眼睛新生血管是许多疾病的中心组成部分。导致严重视力损害的疾病。增殖性糖尿病视网膜病变是20-74岁美国人失明的主要原因。尽管已经取得了重大进展，但我们预防和治疗新生血管疾病的能力显然将受益于在细胞和分子水平上对血管生成的更彻底的了解。我们研究项目的长期目标是了解当正常静止的眼内皮细胞（EC）去分化为新血管形成所需的增殖性血管生成表型时发生的事件序列。本研究的重点是我们实验室最近发现的一组在增殖EC中表达的新基因。这些基因对应的mrna通过亲和层析分离（随后克隆），使用一种必需的RNA结合蛋白作为诱饵，其本身在增殖EC中选择性表达。我们提出的项目的目的是表征这些新的基因产物，并测试它们是否对血管生成表型至关重要。Specific Aim #1旨在告诉我们哪些在增殖EC中被上调，因此更有可能在促进或维持血管生成表型中起作用。在静息和血管生成条件下培养的视网膜EC将通过免疫化学和杂交技术进行检查，在体内诱导进行新生血管形成的眼部组织也将进行检查。在Aim #2中，我们将生成全长cDNA克隆，以促进结构分析和功能探针（分子和免疫学）的开发。在目标3中，我们将验证这些分子对于维持血管生成表型至关重要的假设。这将通过将拮抗剂引入细胞并评估其抑制增殖和其他表型特异性性状表达的能力来实现。这项研究将增加我们对眼睛血管EC增殖的基因表达的一般知识，但也有可能揭示眼部血管生成和其他涉及细胞去分化和增殖的疾病过程中的特定调控点。

Neovascularization in the eye is a central component of many. disorders leading to severe vision impairment. Proliferative diabetic retinopathy alone is the leading cause of blindness in Americans aged 20-74. Although significant progress has been made, our ability to prevent and treat neovascular disorders would clearly benefit from a more thorough knowledge of angiogenesis at the cellular and molecular levels. The long term goal of our research program has been to understand the sequence of events that occur when normally quiescent ocular endothelial cells (EC) dedifferentiate into the proliferative, angiogenic phenotype required for new blood vessel formation. This study focuses on a set of novel genes recently identified in our laboratory that are expressed in proliferating EC. mRNAs corresponding to these genes were isolated by affinity chromatography (and subsequently cloned) using an essential RNA binding protein as bait that is itself expressed selectively in proliferating EC. Our aims for the proposed project are to characterize these novel gene products and test whether they are essential to the angiogenic phenotype. Specific Aim #1 is designed to tell us which are upregulated in proliferating EC, and therefore more likely to function in promoting or maintaining the angiogenic phenotype. Cultured retinal EC under quiescent and angiogenic conditions will be examined by immunochemical and hybridization techniques, as will be ocular tissues induced to undergo neovascularization in vivo. In Aim #2, we will generate full length cDNA clones to facilitate structural analysis and development of functional probes (molecular and immunological). In Aim #3 we will test the hypothesis that these molecules are essential for maintaining the angiogenic phenotype. This will be accomplished by introducing antagonists into cells and assessing their ability to inhibit proliferation and expression of other phenotype-specific traits. This study will add to our general knowledge of gene expression in proliferating vascular EC of the eye, but also has the potential for revealing specific regulatory points in ocular angiogenesis and other disease processes involving cell dedifferentiation and proliferation.