Functions and Mechanisms of Ref-1 in the Endothlium

描述（由申请人提供）：由活性氧（ROS）和内皮一氧化氮（NO）的产生和消除决定的内皮还原-氧化（氧化还原）状态的扰动有助于许多血管疾病的发病机制，如动脉粥样硬化、再狭窄和高血压。氧化还原因子-1（ref-1）是一种普遍存在的DNA修复酶/转录调节因子。ref-1是否以及如何影响内皮细胞氧化还原状态和功能尚不清楚。基于初步证据，该提议提出了ref-i调节ROS和NO的内皮产生的新概念，因此是内皮氧化还原状态的关键决定因素。建议的实验将测试的作用和机制的ref-1在调节内皮细胞的ROS和NO的生产。ref-1的eNOS，Akt激酶，NAD（P）H氧化酶的活性，和热休克蛋白的表达，作为可能的机制，其对内皮细胞的NO和ROS的产生的影响将进行探讨。领域的参考-1，是重要的，在调节这些介质的活动/表达的特点。除了确定ref-1在培养的内皮细胞中的功能外，在全血管中的腺病毒基因转移实验还将阐明其在调节内皮衍生的生物可利用的NO和内皮依赖性血管张力中的作用。最后，eNOS衍生的NO和来自rac 1调节的NAD（P）H氧化酶的ROS在调节内皮中ref-1的功能、表达和亚细胞定位中的作用将被定义。Ref-1作为转录的主要调节因子和DNA修复酶，参与基本的细胞过程，如增殖、凋亡和分化。ref-1在心血管系统中的重要性才刚刚开始被认识。通过研究ref-1在内皮中的调节，其在内皮生理学和病理生理学中的作用，并通过研究ref-1的转录和新的非转录作用机制，该提案有望推进我们目前对其在血管生物学和疾病中的功能的理解。

DESCRIPTION (provided by the applicant): Perturbations in the reduction-oxidation (redox) status of the endothelium, determined by the production and elimination of reactive oxygen species (ROS) and endothelial nitric oxide (NO), contributes to the pathogenesis of many vascular disorders such as atherosclerosis, restenosis, and hypertension.Redox factor-1 (ref-1) is a ubiquitous DNA repair enzyme/transcriptional regulator. If and how ref-1 affects endothelial redox status and function is not known. Based on preliminary evidence, this proposal advances the novel concept that ref-i regulates endothelial production of ROS and NO, and therefore is a crucial determinant of endothelial redox state. Proposed experiments will test the role and mechanisms of ref-1 in regulating the production of endothelial ROS and NO. The effect of ref-1 on eNOS, Akt kinase, and NAD(P)H oxidase activities, and heat shock protein expression, as possible mechanisms for its effects on endothelial NO and ROS generation will be explored. The domains of ref-1 that are important in regulating the activities/expression of these mediators will be characterized. In addition to defining the function(s) of ref-1 in cultured endothelial cells, adenoviral gene transfer experiments in whole vessels will also elucidate its role in regulating endothelium-derived bioavailable NO, and endothelium-dependent vascular tone. Finally, the roles of eNOS-derived NO, and ROS derived from the rac1 -regulated NAD (P) H oxidase in regulating the function, expression, and sub-cellular localization of ref-1 in the endothelium will be defined.Ref-1, as a master regulator of transcription and a DNA repair enzyme, participates in fundamental cellular processes such as proliferation, apoptosis, and differentiation. The importance of ref-1 in the cardiovascular system is only beginning to be appreciated. By investigating the regulation of ref-1 in the endothelium, its role in endothelial physiology and pathophysiology, and by examining both transcriptional and novel non-transcriptional mechanisms of action of ref-1, this proposal promises to advance our current understanding of its functions in vascular biology and disease.