Monocyte Differentiation and Mac-1-Regulated Forkhead

描述（申请人提供）：炎症在动脉粥样硬化和再狭窄的血管损伤中起重要作用。血管细胞间的粘附相互作用协调了这种炎症反应。我们专注于白细胞整合素Mac-1 (CD11b/CD18)，确定这种粘附受体是动脉损伤后新内膜增厚的分子决定因素。这让我们假设Mac-1的“由外而内”信号可能启动了一个促进血管炎症的基因程序。我们利用差分显示PCR克隆了一个新的叉头转录因子（Mac-1调控叉头，MFH），该叉头转录因子在Mac-1聚集的单核细胞中与非聚集的单核细胞相比下调。MFH作为已知的调节单核细胞分化的基因的转录抑制因子，包括PU.1和c-fms，并且在体外过度表达MFH可阻止巨噬细胞样分化。该建议的中心假设是，Mac-1参与通过调节一种新型叉头转录抑制因子的表达来协调单核细胞分化信号，从而调节血管炎症，最终在损伤后新内膜增厚。我们的具体目标是：(1)确定MFH调节转录活性的分子机制，并表征Mac-1调控MFH的性质；(2)确定调控MFH表达对体外和体内单核细胞分化的影响；(3)观察逆转录病毒骨髓干细胞过表达MFH对动脉损伤后巨噬细胞积累和新生内膜增厚的影响。本提案中概述的实验应阐明Mac-1信号在单核细胞分化中的作用及其在血管损伤中的重要性。了解单核细胞分化的分子机制将为开发抗炎策略以调节动脉粥样硬化、再狭窄和移植动脉病变中的血管损伤提供必要的见解。

DESCRIPTION (provided by applicant): Inflammation plays an essential role in vascular injury in atherosclerosis and restenosis. Adhesive interactions between vascular cells orchestrate this inflammatory response. We have focused on the leukocyte integrin Mac-1 (CD11b/CD18), identifying this adhesive receptor as a molecular determinant of neointimal thickening after arterial injury. This led us to hypothesize that "outside-in" signaling by Mac-1 may initiate a gene program that promotes vascular inflammation. We have cloned a novel forkhead transcription factor (Mac-l-regulated forkhead, MFH) using differential display PCR that is down-regulated in Mac-1 clustered compared to non-clustered monocytic cells. MFH functions as a transcriptional repressor of genes known to regulate monocyte differentiation, including PU.1 and c-fms, and over-expressing MFH prevents macrophage-like differentiation in vitro. The central hypothesis of this proposal is that Mac-1 engagement orchestrates monocyte differentiation signals by regulating the expression of a novel forkhead transcription repressor, thereby modulating vascular inflammation and ultimately neointimal thickening after injury. Our specific aims are: (1) to define the molecular mechanism of MFH's ability to modulate transcriptional activity and to characterize the nature of MFH regulation by Mac-1; (2) to determine the effect of manipulating MFH expression on monocyte differentiation in vitro and in vivo; and (3) to examine the effect of retroviral bone marrow stem cell over-expression of MFH on macrophage accumulation and neointimal thickening after arterial injury. The experiments outlined in this proposal should clarify the role of Mac-1 signaling in monocyte differentiation and its importance in vascular injury. Understanding the molecular mechanisms of monocyte differentiation will provide insights necessary to develop anti-inflammatory strategies for modulating vascular injury in atherosclerosis, restenosis, and transplant arteriopathy.