The Role of PECAM in Models of Lung Injury

描述(申请人提供)：血小板-内皮细胞黏附分子-1(PECAM-1，CD31)是一种黏附和信号分子，表达于内皮细胞(EC)、白细胞和血小板上。它是调节白细胞跨内皮细胞迁移的关键分子，尽管它在肺循环中有高水平的表达，但对其涉及的机制或在肺部疾病中的作用知之甚少。利用PECAM-1基因敲除(KO)小鼠，研究人员发现，免疫复合体沉积疾病中的肺中性粒细胞浸润高度依赖于PECAM。这项资助的目标是利用体内和体外模型确定PECAM调节转归的机制，并确定PECAM-1在其他肺部疾病模型中的作用。有待检验的假设是，PECAM通过PECAM胞浆结构域的局部区域介导的内皮和中性粒细胞信号事件调节转运，并且PECAM-1仅在特定类型的肺损伤中发挥重要作用。提出了三个独立但相关的特定目标：在目标1中，将使用野生型和KO动物之间产生的骨髓嵌合体，并通过将特定的PECAM-1亚型重组为具有细胞质区域突变的PECAM-1亚型，来确定PECAM-1调节中性粒细胞向肺内迁移的机制。在目标2中，PECAM-1调节跨内皮细胞迁移的机制将使用包括人肺内皮细胞迁移模型和实验迁移模型的体外系统来确定，在该模型中，突变形式的PECAM-1将被转染到PECAM阴性的人内皮样细胞系或从KO小鼠获得的PECAM阴性的小鼠EC中。在目标3中，将使用抗PECAM-1抗体和PECAM基因敲除小鼠来测试PECAM-1在“内皮驱动”和“白细胞驱动”的肺炎症、氧化剂诱导的肺损伤模型和其他免疫介导的肺损伤模型中调节白细胞向肺内迁移的重要性。这些研究将填补对肺和细胞黏附生物学的基本了解的空白，并提供潜在有用的治疗信息。

DESCRIPTION (provided by applicant): Platelet-Endothelial Cell Adhesion Molecule-1 (PECAM-1, CD31) is an adhesion and signaling molecule that is expressed on endothelial cells (EC), leukocytes, and platelets. It is a key molecule in regulating transendothelial migration of leukocytes, however little in known about the mechanisms involved or its role in lung disease, despite high levels of expression in the pulmonary circulation. Using PECAM-1 knockout (KO) mice, the investigators have found that lung neutrophil infiltration in immune complex deposition disease is highly PECAM-dependent. The goals of this grant are to define the mechanisms by which PECAM regulates transmigration using in vivo and in vitro models and to define the role of PECAM-1 in other models of lung disease. The hypotheses to be tested are that PECAM regulates transmigration through endothelial and neutrophil signaling events mediated by localized regions of the PECAM cytoplasmic domain and that PECAM-1 will play an important role in only specific types of lung injury. Three independent, but related specific aims are proposed: In Aim 1, the mechanisms by which PECAM-1 regulates migration of neutrophils into the lung will be determined using bone marrow chimeras generated between wild type and KO animals and by reconstituting PECAM KO mice with specific PECAM-1 isoforms that have mutations in their cytoplasmic domains. In Aim 2, the mechanisms by which PECAM-1 regulates transendothelial migration will be defined using in vitro systems that include a human lung EC transmigration model and experimental transmigration models in which mutant forms of PECAM-1 will be transfected into a PECAM-negative human endothelial-like cell line or into PECAM-negative murine EC's derived from the KO mice. In Aim 3, anti-PECAM-1 antibodies and PECAM knockout mice will be used to test the importance of PECAM-1 in regulating leukocyte transmigration into the lung in "endothelial-driven" vs. "leukocyte-driven" lung inflammation, in models of oxidant-induced lung injury, and in other immune-mediated lung injury models. These studies will fill a gap in the basic understanding of lung and cell adhesion biology, as well as provide potentially useful therapeutic information.