The role of endothelial connexins in vascular wound repair

内皮连接蛋白在血管伤口修复中的作用

基本信息

批准号：
10751730
负责人：
Meghan Sedovy
金额：
$ 4.25万
依托单位：
VIRGINIA POLYTECHNIC INST AND ST UNIV
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-08-10 至 2026-02-09
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10751730
关键词：
Alanine Antibodies Aorta Area Arteries Biological Biology Biomedical Research Blood Vessels C-terminal Carotid Arteries Cell Communication Cell Culture Techniques Cell Line Cell Proliferation Cell Separation Cells Cessation of life Communication Confocal Microscopy Connexin 43 Connexins Coronary Thrombosis Coronary artery Data Data Analyses Defect Deoxyuridine Disease Endothelial Cells Endothelium Ensure Environment Equipment Failure Fellowship Female Gap Junctions Genetic Goals Health Heart Home Hour Human Immunofluorescence Immunologic Impaired healing Impairment Injections Injury Investigation Knock-out Knockout Mice Legal patent Life Ligation Link Measures Mediating Mentorship Microscope Mission Modeling Modernization Modification Mus Mutation Myocardial Infarction Operative Surgical Procedures Pathway interactions Patients Pharmaceutical Preparations Phosphorylation Phosphorylation Inhibition Phosphorylation Site Phosphotransferases Plasmids Post-Translational Protein Processing Preparation Process Proliferating Proliferation Marker Proto-Oncogene Proteins c-akt Research Research Institute Research Personnel Role Saphenous Vein Serine Signal Pathway Signal Transduction Site Stents Survival Rate Technical Expertise Techniques Testing Therapeutic Thrombus Tissues Training Transfection Travel United States National Institutes of Health Universities Up-Regulation Vascular Diseases Vascular Endothelial Cell Vascular Graft Virginia Western Blotting Work animal facility burden of illness cell growth cell injury cell motility design endothelial regeneration endothelial repair experience experimental study healing improved in vivo in vivo Model injured male migration mimetics novel pharmacologic protein expression repaired response restenosis sex stroke risk therapeutic target transcriptome sequencing vascular injury wound closure wound healing

项目摘要

|| Project Summary . Impaired endothelial wound healing in blood vessels like the coronary artery and saphenous vein is a primary contributor to vascular stent failure and deadly coronary thrombosis. Identifying the mechanism of delayed endothelial repair is essential for improving vessel patency and patient survival rates. As such, this proposal focuses on identifying regulators of normal endothelial healing and understanding how these factors are dysregulated when healing is impaired. Previous studies have shown connexin 43 (Cx43) gap junctions may regulate wound healing. However, the specific role of connexin 43 in endothelial cell (EC) healing is unknown. Preliminary data in this proposal indicates that ligation-induced vascular injury promotes increases in the expression of EC Cx43 surrounding the damaged area in the aorta and carotid arteries in mice. Cx43-mediated gap junction intracellular communication is controlled by phosphorylation at the Cx43 C-terminus. My preliminary data demonstrate that specific Cx43 phosphorylation at its serine (s) 368, associated with gap junction closure, is present during the final stages of EC wound healing in vivo. Cx43-s368 also reduced the rate of wound closure in cultured human EC. These findings inform the hypothesis that Cx43 expression and channel functions are critical for EC wound healing in large arteries. The aims of this proposal are to define the role of connexin 43 in endothelial wound healing (Aim 1), investigate if a loss of Cx43 limits healing in vivo (Aim 1), and test if channel functions, regulated by posttranslational modifications, improve/delay vascular repair (Aim 2). This investigation will be completed using a novel mouse carotid EC injury survival surgery in mice I developed, which will allow for the assessment of Cx43 expression in carotid EC during the healing process. Both genetic and pharmacological strategies will be used to alter Cx43 expression and phosphorylation in mouse injury models and in cultured human EC, and the impact of these modifications on the rate and quality of EC healing will be quantified. Additionally, RNAseq approaches will be used to identify Cx43 gap junction-dependent signaling in the regenerating endothelium. These aims will be accomplished under the mentorship of researchers with extensive experience in vascular biology, Dr. Scott Johnstone, Dr. Robert Gourdie, and Dr. Brant Isakson, at Virginia Tech’s Fralin Biomedical Research Institute (FBRI). The FBRI is home to research equipment including but not limited to animal facilities, confocal microscopes, flow cytometers, and cell culture equipment. This state- of-the-art research environment will allow for a detailed and mechanistic investigation. Ultimately, this study will characterize a novel role for Cx43 in vascular EC. This project is designed to identify new potential therapeutic targets in vascular disease that will promote the mission of the NIH to enhance patient health and to promote longer life free of illness.

|| 项目摘要。冠状动脉和隐静脉等血管的内皮伤口愈合受损是主要的原因确定延迟性冠状动脉血栓形成的机制。内皮修复对于改善血管通畅和患者生存率至关重要。专注于识别正常内皮愈合的调节因子并了解这些因子是如何发挥作用的先前的研究表明，当愈合受损时，连接蛋白 43 (Cx43) 间隙连接可能会失调。然而，连接蛋白 43 在内皮细胞 (EC) 愈合中的具体作用尚不清楚。该提案中的初步数据表明，结扎诱导的血管损伤会促进小鼠主动脉和颈动脉受损区域周围 EC Cx43 的表达。间隙连接细胞内通讯由 Cx43 C 末端的磷酸化控制。数据表明，Cx43 丝氨酸 368 处的特异性磷酸化与间隙连接闭合相关， Cx43-s368 存在于体内 EC 伤口愈合的最后阶段，也降低了伤口闭合率。这些发现为 Cx43 表达和通道功能的假设提供了依据。对于大动脉 EC 伤口愈合至关重要该提案的目的是确定连接蛋白 43 在大动脉中的作用。内皮伤口愈合（目标 1），研究 Cx43 的缺失是否会限制体内愈合（目标 1），并测试通道是否功能，由翻译后修饰调节，改善/延迟血管修复（目标 2）。将使用我开发的小鼠颈动脉 EC 损伤生存手术来完成，这将允许用于评估愈合过程中颈动脉 EC 中的 Cx43 表达。药理学策略将用于改变小鼠损伤模型中的 Cx43 表达和磷酸化在培养的人类 EC 中，这些修饰对 EC 愈合速度和质量的影响将是此外，RNAseq 方法将用于识别 Cx43 间隙连接依赖性信号传导。这些目标将在研究人员的指导下实现。 Scott Johnstone 博士、Robert Gourdie 博士和 Brant Isakson 博士在血管生物学方面拥有丰富的经验弗吉尼亚理工大学弗拉林生物医学研究所 (FBRI) 拥有包括以下在内的研究设备。但不限于动物设施、共聚焦显微镜、流式细胞仪和细胞培养设备。最先进的研究环境将允许进行详细和机械的调查。表征 Cx43 在血管 EC 中的新作用该项目旨在确定新的潜在治疗方法。血管疾病的目标将促进 NIH 的使命，即增强患者健康并促进无病长寿。