Birth and Death of Choriocapillaris.

描述(申请人提供)：视网膜色素上皮细胞(RPE)和脉络膜毛细血管(CC)在眼睛中有密切的关系。作为最近的邻居，只有布鲁赫膜隔开，它们相互依赖，相互帮助，完成维持健康光感受器细胞的关键角色。本研究探讨了两种不同类型的老年性黄斑变性(AMD)、地理萎缩(GA)和渗出性AMD中，从脉络膜毛细血管的发育或出生(妊娠5-23周)到两种不同细胞类型的功能障碍或死亡之间的关系。应用酶免疫组织化学(IH)和TUNEL标记技术对CC的代谢状态和活性进行评价。用免疫组织化学和原位末端标记法检测RPE细胞的活性和功能。两种类型的细胞为自身(自分泌)或其邻近细胞(旁分泌)及其同源受体产生和使用的生长因子(成纤维细胞生长因子、血管内皮生长因子、PIGF、血管生成素、促红细胞生成素、干细胞因子、基质衍生因子-1)及其同源受体将通过IH和组织漂白进行研究。在胎儿RPE存在或不存在的情况下，胎儿人内皮细胞在体外形成的管道将使用中和抗体和siRNA来评估生长因子的旁分泌和自分泌效应。IH将研究白细胞和补体在AMD患者CC死亡和功能障碍以及CNV中的作用，以确定细胞类型、白细胞数量(非特异性酯酶、CD45、CD68)、白细胞黏附分子(ICAM-1、VCAM-1、P-选择素)、促炎性细胞因子(TNFa、IL6、IL8、MCP-1)和血管生成细胞因子(成纤维细胞生长因子、血管内皮生长因子、PIGF、血管生成素)。还将研究补体因子H、攻击复合体、白细胞和CC活性之间的空间关系。利用我们的碱性磷酸酶平面包埋技术，我们已经建立并将继续研究CC和RPE之间的互惠共生关系。这一分析将使我们能够确定这种关系在GA和渗出性AMD中是如何被破坏的。这项建议的目的是为了更好地了解这两种相邻细胞类型之间的关系，并阐明可能有针对性地维持或改善这种健康关系的重要因素。

DESCRIPTION (provided by applicant): The retinal pigment epithelial cells (RPE) and the choriocapillaris (CC) have an intimate relationship in the eye. As nearest neighbors, separated only by Bruch's membrane, they rely on each other and assist each other in accomplishing their critical role of sustaining healthy photoreceptor cells. This proposal explores the relationship between these two cell types from the development or birth of choriocapillaris (5-23 weeks gestation) to the dysfunction or death of either cell type in two different forms of age-related macular degeneration (AMD), geographic atrophy (GA) and exudative AMD. The metabolic status and viability of CC will be evaluated using enzyme and immunohistochemistry (IH) and TUNEL labeling. RPE cell viability and function will be investigated with IH and TUNEL labeling. Growth factors (FGF's, VEGF, PIGF, angiopoietins, erythropoietin, stem cell factor, stromal derived factor-1) that the two cells types produce and use for themselves (autocrine) or their neighbor (paracrine), and their cognate receptors will be investigated by IH and bleaching of the tissue. Tube formation in vitro by fetal human endothelial cells in the presence or absence of fetal RPE will be assessed for paracrine and autocrine effects of growth factors using neutralizing antibodies and siRNA. The contribution of leukocytes and complement to CC death and dysfunction and to CNV in AMD will be investigated by IH to identify cell types, leukocyte numbers (nonspecific esterase, CD45, CD68), leukocyte adhesion molecules (ICAM-1, VCAM-1, P-selectin), and pro-inflammatory (TNFa, IL6, IL8, MCP-1) and angiogenic (FGF's, VEGF, PIGF, angiopoietins) cytokines. The spatial relationship between complement factor H, attack complex, leukocytes and CC viability will also be investigated. Using our alkaline phosphatase flat-embedding technique in which CC vascular density and percent of Bruch's membrane covered with RPE can be quantified, we have established and will continue to investigate the mutualistic symbiotic relationship between CC and RPE. This analysis will permit us to establish how this relationship is disrupted in GA and exudative AMD. The goal of this proposal is to better understand the relationship between these two neighboring cell types and elucidate important factors that could be targeted to maintain or improve this healthy relationship.