Extracellular Proteinases in Ocular Neovascularization

描述(申请人提供)：视网膜新生血管是美国工作年龄成人(糖尿病视网膜病变)和婴儿(早产儿视网膜病变)失明的主要原因。血管生成级联的一个重要事件是毛细血管基底膜的破裂和微血管内皮细胞通过细胞外基质的侵袭和迁移。血管生成的这一步骤部分依赖于蛋白水解酶的表达和活性，其中包括尿激酶(UPA)及其受体。该酶的活性受特定的内源性抑制物、纤溶酶原激活物抑制物PAI-1的调节。PAI-1在新血管形成期间在视网膜中升高，似乎是新血管反应所必需的。血管生成的启动以及对蛋白水解酶和抑制物表达的刺激受特定生长因子的活性调节，其中包括众所周知的血管内皮生长因子(VEGF)。其他因素可能在这一过程中发挥重要作用，包括肝细胞生长因子(HGF)，它在眼血管生成中的作用尚未引起足够的重视。了解蛋白水解酶、蛋白水解酶抑制物和生长因子的表达机制，并确定其功能的特异性抑制物，可能为开发新的视网膜新生血管治疗干预措施提供独特的机会。这一提议将检验一种假说，即视网膜新生血管是由特定的生长因子、细胞外蛋白水解酶及其抑制物的表达促进的，抑制它们可能导致新的有用的治疗方法。本研究的目的是：(1)研究PAI-1在视网膜新生血管中的表达和作用。我们将使用生化、组织学和分子技术，包括使用基因敲除动物来解决这一目标。(2)探讨肝细胞生长因子(HGF)在视网膜新生血管中的表达及其在调节蛋白水解酶表达和视网膜毛细血管内皮细胞行为中的作用。利用野生型和转基因动物，研究HGF在调节蛋白酶表达和内皮细胞行为中的作用。(3)直接或间接抑制PAI-1或HGF功能的药物能否抑制视网膜新生血管的形成。这些研究的结果将使我们能够进一步确定视网膜中新血管形成的机制，这可能导致严重的视觉并发症，以及血管生成过程的特定阶段是否适合进行治疗干预。这种方法将为开发潜在有效的治疗眼部新生血管的方法提供合理的基础。

DESCRIPTION (provided by applicant): Retinal neovascularization is the leading cause of blindness in the United States among working-age adults (diabetic retinopathy) and infants (retinopathy of prematurity). An important event of the angiogenesis cascade is the breakdown of the capillary basement membrane and the invasion and migration of microvascular endothelial cells through the extracellular matrix. This step of angiogenesis is partially dependent upon the expression and activity of proteinases, including urokinase (uPA) and its receptor. The activity of this enzyme is regulated by specific endogenous inhibitor, plasminogen activator inhibitor PAI-1. PAI-1 is elevated in the retina during new vessel formation and appears to be required for the neovascular response. The initiation of angiogenesis and the stimulation of proteinase and inhibitor expression are regulated by the activity of specific growth factors including the well known vascular endothelial growth factor (VEGF). Other factors may play important roles during this process including the hepatocyte growth factor (HGF) which has received little attention in terms of its function in ocular angiogenesis. Understanding the mechanisms of proteinase, proteinase inhibitor and growth factor expression, and identifying specific inhibitors of their functions may provide unique opportunities for the development of new therapeutic interventions for retinal neovascularization. This proposal will test the hypothesis that retinal neovascularization is facilitated by the expression of specific growth factors, extracelluar proteinases and their inhibitors, the inhibition of which may lead to new and useful therapies. The aims of this study which will address the hypothesis are: (1) To characterize the expression and role of PAI-1 in retinal neovascularization. We will use biochemical, histological and molecular techniques, including the use of knockout animals to address this aim. (2) To determine the expression and role of hepatocyte growth factor (HGF) in the regulation of proteinase expression and retinal capillary endothelial cell behavior in retinal neovascularization. Studies will focus on the role of HGF in regulating proteinase expression and endothelial cell behavior using wildtype and transgenic animals. (3) To determine if the extent of retinal neovascularization can be suppressed through the use of agents which directly or indirectly inhibit the function of PAI-1 or HGF. The results of these studies will allow us to further define the mechanisms involved in the formation of new vessels in the retina which can lead to serious visual complications, and whether a specific stage of the angiogenic process is an appropriate target for therapeutic intervention. Such an approach will provide a rational basis for the development of potentially powerful and effective therapeutics for ocular neovascularization.