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中起着关键作用，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、分化和效应功能中起关键作用。基于我们实验室的初步数据，白细胞（T淋巴细胞）¿2整合素参与导致Rho家族GTPase Rac-2的激活，并且这种激活导致基因表达的调节，本提案的重点是识别巨噬细胞中的Rac-2靶点，并确定Rac-2在血管病理模型中的重要性。我们的假设是，通过影响基因表达和/或蛋白磷酸化，Rac-2是需要巨噬细胞定位和激活的血管病理（或生理）状态的关键介质，如动脉粥样硬化和炎症性血管生成。这个为期三年的项目的具体目标是：1。利用Rac-GTP拉下实验和磷酸map激酶免疫印迹，证明白细胞整合素参与导致小鼠巨噬细胞中Rac-2和Rac-2效应激活；2. 通过蛋白质组学和磷酸化蛋白质组学分析，利用生物信息学方法鉴定新的Rac-2靶标，在转染了表达组成型活性Rac-2 （Q61L）的构建体的小鼠巨噬细胞中，鉴定其水平和/或磷酸化状态增加的蛋白；和3。通过在动脉粥样硬化（ApoE-/-）和后肢缺血模型中使用Rac-2基因缺失小鼠，证明Rac-2在血管病理和生理中的作用。这项工作的长期目标是确定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抵御感染至关重要。它还在改变血管健康方面发挥作用，导致冠状动脉疾病，并在血液供应不足时形成新的血管。这项工作将定义炎症蛋白，在免疫细胞中表达，可作为进展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.