Inflammation and Cerebral Aneurysm Formation: Targets for Modulation and Vascular

项目摘要/摘要 候选人：我的长期职业目标是成为脑动脉瘤研究领域的领先临床医生和科学家。自从斯坦福大学的一名本科生在爱德华·穆卡西博士的实验室从事一个研究人类巨细胞病毒B2.7启动子调控的项目以来，我一直致力于生物医学研究。作为哥伦比亚大学的一名医学生，我在索尔·西尔弗斯坦博士手下工作，绘制了单纯疱疹病毒IE0基因的功能域。然后，我在E·桑德·康诺利博士和大卫·平斯基博士的指导下，在小鼠模型中研究中风的病理生理机制。AS 作为一名麻省总医院的住院医生和研究员，我在约瑟夫·瓦康蒂博士的实验室里参与了一个项目，将内皮细胞植入聚合物中，创造出一种治疗脑动脉瘤的血管移植物。我未来五年的近期目标是成为一名独立调查员。我将启动一项严格的教学计划，重点是获得实现这一目标所需的知识和技能。我将在获奖期间获得科学技能，包括动物外科、组织学、免疫组织化学、活体显微镜、干细胞生物学技术和高级数据统计分析。为了补充我的实验室经验，我将参加佛罗里达大学跨学科项目的研究生课程 生物医学科学。我制定了一个全面的职业发展计划，其中包括实验室培训、密切指导、研究生水平的课程工作、研究会议和口头陈述。我的目标是在奖项结束时获得NIH R01奖项。 环境：我将在爱德华·斯科特博士的实验室工作，他是一位杰出的导师，他成功地培训了几名临床医生、科学家和之前的K08奖获得者。我将聘请材料科学与工程系的克里斯托弗·巴蒂奇博士作为顾问，他是一位出色的合作者。我将与干细胞生物学和再生医学项目、麦克奈特脑研究所和材料科学与工程系的合作者一起，在一个杰出的科学和创造性环境中工作，指导我的研究进展。我得到了我的主席、神经外科和佛罗里达大学的全力支持，将至少50%的时间和精力投入到我的研究事业发展中。所有的组成部分都已就位，以促进我的研究提案和我整个研究事业发展的成功结果。 研究计划：该项目的长期目标是更好地确定脑动脉瘤形成的病理生理学，从而开发一种治疗脑动脉瘤的转化性疗法。总体目标是回答关于炎症是导致脑动脉瘤形成还是只是一种附带现象的认识上的根本差距；识别可能成为调节动脉瘤炎症反应的靶点的信号机制；以及识别能够修复血管炎症的退化机制的血管修复细胞。中心假说是：脑动脉瘤的形成是一个炎症过程。调节炎症反应将改变脑动脉瘤的发展。血管修复细胞可以被鉴定为修复血管炎症引起的退变机制。这项拟议研究的基本原理是，由于脑动脉瘤可以在破裂之前被诊断出来，因此修复动脉瘤炎症过程并防止动脉瘤进展到破裂的治疗方法，将对公众健康产生重大有益影响。在强大的初步数据的指导下，这一假说将通过追求三个具体目标来检验：1)证明脑动脉瘤的形成是由炎症过程引起的；2)确定参与炎症过程的信号机制作为调节目标，以改变动脉瘤的发展；以及3)确定修复血管炎症引起的退化机制的血管修复细胞。在第一个目标下，将进行两个实验：1)通过激活炎症(将激活的巨噬细胞和中性粒细胞注射到 正常小鼠颈动脉)；以及2)在免疫缺陷小鼠中建立不能引起炎症反应的弹力酶囊状动脉瘤模型。在第二个目标下，白细胞募集级联反应的每一步的信号机制：1)趋化，2)滚动，3)黏附，4)移行；将在弹力酶囊状动脉瘤模型中被阻断。在第三个目标下，CD133+内皮祖细胞(EPC)、造血干细胞和间充质干细胞的细胞特异性群体将被种植到我们的合作者Christopher Batich博士开发的Matrigel和其他聚合物中，并被注射到小鼠弹性蛋白酶囊状动脉瘤中，稍后将对其进行评估，以寻找再内皮化和抑制血管炎症反应的证据。

PROJECT SUMMARY/ABSTRACT Candidate: My long-term career goal is to become a leading clinician-scientist in the field of cerebral aneurysm research. I have had a long commitment to biomedical research since an undergraduate at Stanford University working in the laboratory of Dr. Edward Mocarksi on a project studying regulation of the B2.7 promoter of human cytomegalovirus. As a medical student at Columbia University I worked under Dr. Saul Silverstein mapping functional domains of the IE0 gene of herpes simplex virus. I then worked under Dr. E. Sander Connolly and Dr. David Pinsky studying pathophysiologic mechanisms of stroke in a mouse model. As a resident and fellow at the Massachusetts General Hospital, I worked in the laboratory of Dr. Joseph Vacanti on a project seeding polymers with endothelial cells to create a vascular graft for treating cerebral aneurysms. My immediate objective for the next five years is to become an independent investigator. I will start a rigorous didactic program focused on gaining the knowledge and skills necessary to achieve this objective. Scientific skills I will acquire during the award period, include animal surgery, histology, immunohistochemistry, intravital microscopy, stem cell biology techniques, and advanced statistical analysis of data. To complement my laboratory experience, I will take graduate coursework in the University of Florida Interdisciplinary Program in Biomedical Sciences. I have developed a comprehensive career development program which includes laboratory training, close mentorship, graduate level coursework, research conferences, and oral presentations. My goal is to obtain an NIH R01 award by the time the award finishes. Environment: I will be working in the laboratory of Dr. Edward Scott, an outstanding mentor, who has successfully trained several clinician-scientists and previous recipients of the K08 award. I will have as a consultant, Dr. Christopher Batich in the Department of Materials Science and Engineering, a superb collaborator. I will be working in an outstanding scientific and creative environment with collaborators in the Program in Stem Cell Biology and Regenerative Medicine, the McKnight Brain Institute, and the Department of Materials Science and Engineering to guide my research progress. I have the full committed support of my chairman, the Department of Neurological Surgery, and the University of Florida to devote at least 50% of my time and effort to my research career development. All of the components are in-place to promote a successful outcome in my research proposal and my overall research career development. Research Plan: The long-term goal of this project is to better define the pathophysiology of cerebral aneurysm formation, and, thereby, develop a translational therapy that would treat cerebral aneurysms. The overall objective is to answer the fundamental gap in knowledge of whether inflammation causes cerebral aneurysm formation, or is an epiphenomenon; identifying signaling mechanisms which could be targets for modulation of the aneurysm inflammatory response; and identifying vascular repair cells that can repair the degenerative mechanisms of vascular inflammation. The central hypothesis is that: Cerebral aneurysm formation is an inflammatory process. Modulating the inflammatory response would alter the development of cerebral aneurysms. Vascular repair cells can be identified that repair the degenerative mechanisms caused by vascular inflammation. The rationale for the proposed research is that because cerebral aneurysms can be diagnosed before they rupture, a therapy that would repair the aneurysm inflammatory process and prevent aneurysms from progressing to rupture, would have a significant beneficial impact for the public health. Guided by strong preliminary data, this hypothesis will be tested by pursuing three specific aims: 1) Demonstrate that cerebral aneurysm formation is caused by an inflammatory process; 2) Identify signaling mechanisms involved in the inflammatory process as targets for modulation to alter the development of aneurysms; and 3) Identify vascular repair cells that repair the degenerative mechanisms caused by vascular inflammation. Under the first aim, two lines of experiments will be performed: 1) aneurysms will be produced in mice by activating inflammation (injecting activated macrophages and neutrophils into normal mouse carotid artery); and 2) performing an elastase saccular aneurysm model in immune-deficient mice that can not mount an inflammatory response. Under the second aim, signaling mechanisms at each step of the leukocyte recruitment cascade: 1) chemotaxis, 2) rolling, 3) adhesion, 4) transmigration; will be blocked in an elastase saccular aneurysm model. Under the third aim, cell-specific populations of CD133+ endothelial progenitor cells (EPCs), hematopoietic stem cells, and mesenchymal stem cells will be seeded into Matrigel and other polymers developed by our collaborator, Dr. Christopher Batich, and injected into murine elastase saccular aneurysms which will later be evaluated for evidence of re-endothelialization and inhibition of the vascular inflammatory response.