Copper transport protein and inflammatory angiogenesis

铜转运蛋白与炎症血管生成

• 批准号: 9484073
• 负责人: TOHRU FUKAI
• 金额: $ 23.39万
• 依托单位: AUGUSTA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-12 至 2019-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9484073
• 关键词: Copper; transport; protein; inflammatory; angiogenesis

DESCRIPTION (provided by applicant): The overall aim of this grant is to elucidate the novel linkage between copper transport protein "Antioxidant1 (Atox1)" and "NADPH oxidase" involved in inflammatory angiogenesis. Ischemic disease is a leading cause of morbidity and mortality in worldwide. Neovascularization is an important repair process in response to ischemia, which depends on angiogenesis, inflammation and reactive oxygen species (ROS). Copper (Cu), an essential micronutrient, is involved in physiological repair processes such as wound healing and angiogenesis as well as in various pathophysiologies including tumor growth, atherosclerosis and inflammatory diseases. Since excess Cu is toxic, bioavailability of intracellular Cu is tightly controlled by Cu transport proteins such as Cu chaperone Atox1. Our laboratories provided the first evidence that Atox1 functions as a Cu-dependent transcription factor to regulate Cu-induced cell growth. Furthermore, we are one of the first to demonstrate that ROS derived from NADPH oxidase (Nox) play an important role in angiogenic signaling in endothelial cells (ECs) as well as postnatal angiogenesis in response to ischemic injury. However, the role of Cu transport proteins in inflammatory angiogenesis and its linkage with Nox are entirely unknown. Our preliminary data suggest that Atox1 deficient mice have impaired angiogenesis and inflammatory cell recruitment due to decrease in endothelial ROS production in ischemic tissues. Bone marrow (BM) reconstitution indicates that Atox1 in ECs, but not BM cells, is required for post-ischemic revascularization. Based on new preliminary data, we hypothesize that Atox1 functions as a novel regulator for Nox by transcriptional regulation of p47phox as well as activating Rac1; both are critical cytosolic components of Nox, in a Cu-dependent manner. This in turn promotes ROS-dependent signaling linked to inflammatory and angiogenic responses in ECs, which contributes to neovascularization in response to ischemic injury. Aim1 will focus on establishing a role of Atox1 in regulating NADPH oxidase and ROS-dependent inflammatory and angiogenic signaling and function in ECs in a Cu-dependent manner. Aim 2 will focus on identifying molecular mechanisms of how Atox1 is involved in activation of NADPH oxidase through transcriptional regulation of p47phox and activating Rac1 via binding to a Rac1-binding scaffold protein IQGAP1 in ECs in a Cu-dependent manner. Aim 3 will focus on determining the functional role of Atox1 in neovascularization in vivo by regulating ROS production, angiogenesis and inflammatory cell recruitment in injured tissues in a Cu-dependent manner using hindlimb ischemia model with Atox1-/- mice. Bone marrow transplantation, in vivo intravital microscopy and bioluminescence imaging, highly innovative Cu imaging analysis in vitro and in vivo will be performed. Our study will provide novel insight into Cu transport protein and their regulators as potential therapeutic targets for treatment of angiogenesis- and inflammation-dependent ischemic cardiovascular diseases.

描述（由申请人提供）：该赠款的总体目的是阐明铜转运蛋白“抗氧化剂1（ATOX1）”和与炎症血管生成有关的新型联系。缺血性疾病是全球发病率和死亡率的主要原因。新血管形成是响应缺血的重要修复过程，它取决于血管生成，炎症和活性氧（ROS）。铜（CU）是一种必需的微量营养素，参与了生理修复过程，例如伤口愈合和血管生成以及各种病理生理，包括肿瘤生长，动脉粥样硬化和炎症性疾病。由于过量的Cu有毒，因此细胞内Cu的生物利用度紧密 由Cu转运蛋白（例如Cu Chaperone Atox1）控制。我们的实验室提供了第一个证据，表明ATOX1起着CU依赖性转录因子的作用，以调节CU诱导的细胞生长。此外，我们是第一个证明源自NADPH氧化酶（NOX）的ROS在内皮细胞（ECS）的血管生成信号传导以及响应缺血性损伤的产后血管生成中起重要作用的人之一。但是，Cu转运蛋白在炎症性血管生成中的作用及其与NOX的联系完全未知。我们的初步数据表明，由于缺血组织内皮ROS的产生减少，ATOX1缺乏小鼠的血管生成和炎症细胞募集受损。骨髓（BM）的重构表明EC中的ATOX1（而不是BM细胞）是缺血后血运重建所必需的。基于新的初步数据，我们假设Atox1通过P47Phox的转录调节和激活Rac1作为NOX的新调节剂。两者都是以Cu依赖性方式的NOX的关键胞质成分。反过来，这促进了与EC中炎症和血管生成反应有关的ROS依赖性信号传导，这有助于响应缺血性损伤的新血管形成。 AIM1将专注于以Cu依赖性方式在EC中建立ATOX1在调节NADPH氧化酶以及ROS依赖性炎症和血管生成信号和功能中的作用。 AIM 2将集中在识别ATOX1如何通过p47phox的转录调节和通过与cu依赖性方式中EC中的Rac1结合脚手架蛋白IQGAP1结合而激活RAC1的分子机制。 AIM 3将专注于通过使用ATOX1 - / - 小鼠的后肢缺血模型来以CU依赖性方式调节损伤组织中的ROS产生，血管生成和炎症细胞募集，以确定ATOX1在体内新血管化的功能作用。将进行骨髓移植，体内插入式显微镜和生物发光成像，体外和体内高度创新的CU成像分析。我们的研究将提供有关CU转运蛋白及其调节剂的新见解，作为治疗血管生成和炎症依赖性缺血性心血管疾病的潜在治疗靶标。