The Rac-2-induced macrophage proteome and vascular pathology

描述（由申请人提供）：众所周知，巨噬细胞介导的炎症在血管病理生理学中起关键作用，包括导致动脉粥样硬化和血管生成。整合素粘附受体在单核细胞/巨噬细胞募集、定位、分化和效应子功能中起关键作用。基于我们实验室的初步数据，即白细胞（T淋巴细胞）2整合素参与导致Rho家族GT3 Rac-2的激活，并且这种激活导致基因表达的调节，该提议集中于鉴定巨噬细胞中的Rac-2靶标和确定Rac-2在血管病理模型中的重要性。假设Rac-2通过对基因表达和/或蛋白质磷酸化的影响，是需要巨噬细胞定位和活化的那些血管病理学（或生理学）状态（例如动脉粥样硬化和炎性血管生成）的关键介质。这个为期三年的项目的具体目标是：1。使用Rac-GTP下拉测定和磷酸-MAP激酶免疫印迹证明白细胞整联蛋白接合导致鼠巨噬细胞中Rac-2和Rac-2效应子活化; 2.使用蛋白质组学和磷酸化蛋白质组学分析，通过生物信息学努力鉴定新的Rac-2靶标，鉴定其水平和/或磷酸化状态在用表达组成型活性Rac-2（Q61L）的构建体转染的鼠巨噬细胞中增加的蛋白质;和3.在动脉粥样硬化（ApoE-/-）和后肢缺血模型中使用Rac-2基因缺失小鼠，记录Rac-2在血管病理学和生理学中的作用。拟议工作的长期目标是定义巨噬细胞特异性炎症生物标志物和/或血管病理学中的分子成像靶点，包括急性和慢性动脉粥样硬化综合征以及血管生成。我的目标是利用这个项目和这些发现作为我的医生科学家职业生涯的基础。包括炎症在内的免疫系统对于防御感染至关重要。它还在改变血管健康，导致冠状动脉疾病和血液供应缺乏时新血管形成方面发挥作用。这项工作将定义炎症蛋白，表达在免疫细胞中，可以作为进行性血管疾病的标志物，并作为成像或治疗靶点。

DESCRIPTION (provided by applicant): It is well recognized that macrophage-mediated inflammation plays a key role in vascular pathophysiology, including that leading to atherosclerosis and angiogenesis. Integrin adhesion receptors are critically involved in monocyte/macrophage recruitment, localization, differentiation, and effector function. Based on the preliminary data in our laboratory that leukocyte (T lymphocyte) ¿2 integrin engagement leads to activation of the Rho family GTPase Rac-2, and that this activation results in modulation of gene expression, this proposal is focused on identifying Rac-2 targets in macrophages and on determining the importance of Rac-2 in vascular pathology models. The hypothesis is that Rac-2, through effects on gene expression and/or protein phosphorylation, is a critical mediator of those vascular pathology (or physiology) states requiring macrophage localization and activation, such as atherosclerosis and inflammatory angiogenesis. The specific aims of this 3 year project are to: 1. Demonstrate that leukocyte integrin engagement results in Rac-2 and Rac-2 effector activation in murine macrophages, using Rac-GTP pull-down assays and phospho-MAP kinase immunoblots; 2. Identify proteins whose levels and/or phosphorylation status are increased in murine macrophages transfected with a construct expressing a constitutively active Rac-2 (Q61L), using proteomic and phosphoproteomic analyses, with bioinformatic efforts to identify novel Rac-2 targets; and 3. Document a role for Rac-2 in vascular pathology and physiology, using Rac-2 gene-deleted mice in atherosclerosis (ApoE-/-) and hindlimb ischemia models. The long term objective of the proposed work is defining macrophage-specific inflammatory biomarkers and/or molecular imaging targets in vascular pathology, including acute and chronic atherosclerotic syndromes, as well as angiogenesis. My goal is to utilize this project and these findings as the foundation for my physician-scientist career. The immune system, which includes inflammation, is critical to defense against infections. It also plays a role in altering blood vessel health, leading to coronary artery disease and to new vessel formation when blood supply is lacking. This work will define inflammation proteins, expressed in immune cells that could serve as markers of progressive, vascular disease, and as imaging or therapeutic targets.