The role of the Wnt/planar cell polarity pathway in pulmonary angiogenesis

描述（由申请人提供）：这里提出的建议分为两部分：第一部分包含我提出的职业规划，第二部分描述了拟议项目的研究策略。我对血管生物学研究的兴趣始于我第一次临床接触患有特发性肺动脉高压（IPAH）的患者。对这种疾病的病理生物学的兴趣使我继续在科罗拉多大学接受肺病学家的培训，但由于家庭原因，我在第一年的奖学金后从丹佛搬到了斯坦福大学，在血管研究领域的领导者Marlene Rabinovitch博士的指导下继续我的研究培训。在过去的四年里，我 已经完成了两个项目，处理无翼（Wnt）和骨形态发生蛋白（BMP）信号在肺动脉内皮细胞（PAEC）和平滑肌细胞中的相互作用。此外，在同一时期，我被授予K12和哈罗德·阿莫斯职业发展奖，以继续我目前作为一名医生科学家的职业生涯。从最初提交以来，我已经成为斯坦福大学的医学助理教授，这一职位使我能够将80%的时间投入到与我的研究项目相关的活动中，例如参加科学会议，学习新的实验技术和数据分析方法，并向我的顾问和同行提交我的研究进展报告。除了定期与我的导师拉宾诺维奇博士进行一对一的会谈外，我每季度还将与一个顾问小组会面，他们将提供批评和帮助 讨论与拟议研究有关的想法。为了跟上最新的思想， 我将参加由斯坦福大学教授的生物学、遗传学和生物统计学的研究生课程，学习负责任的研究行为的基本原则。最后，我的剩余时间（20%）将用于教学活动，旨在培训住院医师和研究员在住院和门诊环境中管理PAH患者。我的长期目标是有足够的发表和初步数据，以支持我作为一个独立的研究者，并提交R 01赠款，将重点应用在这些拟议的研究中获得的数据，以调查异常Wnt信号如何可能有助于IPAH的发展。第二部分是研究计划。IPAH是一种罕见的进行性疾病，与肺动脉压异常升高相关，如果不治疗，会导致充血性心力衰竭和过早死亡。PAH患者表现出进行性远端小肺动脉狭窄和丢失，无显著代偿性血管再生，表明存在肺血管生成缺陷。我们小组和其他人最近的研究表明，骨形态发生蛋白受体（BMPR）II的突变可能通过减少肺动脉内皮细胞（PAEC）的存活和增殖而导致血管损失，但BMPRI调节这些血管生成反应的机制尚未得到很好的表征。除了PAEC，周细胞募集对于新形成的血管的稳定是至关重要的，但它们对IPAH发病机制的贡献尚未得到很好的确定。我们最近发现BMP信号通过募集PAEC中的两条Wnt信号通路促进肺血管生成：“经典的”Wnt/β信号通路。连环蛋白（？C）和“非经典”Wnt/平面细胞极性（PCP）途径。基于我们的初步研究和文献综述，我们提出Wnt/PCP通路的激活是肺血管生成所需的，通过1）诱导PAEC组织成正确排列的血管管和2）促进周细胞募集到血管管以形成功能性血管。在使用从健康供体和IPAH患者的肺获得的PAEC和周细胞的一系列实验中，我们将证明Wnt/PCP信号传导是血管生成期间指导PAEC（Aim 1）和周细胞（Aim 2）行为所必需的，并且活性的丧失与减少的肺血管生成相关。此外，我们将通过对两种功能失调的Wnt/PCP信号传导动物模型的研究来补充这些细胞研究，这将有助于深入了解该途径与肺切除术后补偿性血管生成的生物学相关性（目标3）。未来的研究将集中在减少Wnt/PCP信号传导的突变如何与BMPRII突变发生遗传相互作用以促进IPAH的发展，以及旨在使PAEC和周细胞中的Wnt/PCP通路活性正常化的治疗干预如何通过恢复IPAH患者再生失去的血管的能力来帮助预防进展并提高其生存率。 公共卫生相关性：我们的目标是了解Wnt/PCP通路如何调节肺血管生成，并证明破坏通路激活的突变可导致PAH。新基因的鉴定可能会导致新的治疗方法，可以防止进展和/或逆转已建立的疾病。

DESCRIPTION (provided by applicant): The proposal presented here is divided into two parts: the first contains my proposed career plan and the second describes the research strategy for the proposed project. My interest in vascular biology research began with my first clinical encounters with patients suffering from idiopathic pulmonary hypertension (IPAH). Interest in the pathobiology of the disease led me to continue training as a pulmonologist at the University of Colorado but, due to family related reasons, I moved from Denver to Stanford University after my first year of fellowship to continue my research training under the mentorship of Dr. Marlene Rabinovitch, a leader in the field of vascular research. Over the last four years, I have completed two projects dealing with the interaction between Wingless (Wnt) and bone morphogenetic protein (BMP) signaling in pulmonary artery endothelial (PAEC) and smooth muscle cells. Furthermore, during the same time period, I have been awarded both a K12 and, a Harold Amos career development award to continue my current career track as a physician scientist. Since the initial submission, I have become an assistant professor of medicine at Stanford University, a position that allows me to devote 80% of my time to activities related to my research project such as participating in scientific conferences, learning new experimental techniques and methods of data analysis and presenting progress reports of my research to my advisors and peers. In addition to regular one-on-one meetings with my mentor Dr. Rabinovitch, I will meet quarterly with a panel of advisors and consultants that will provide criticism and help discuss ideas related to the proposed research. In an effort to keep abreast of the latest ideas in my field of research, I will enroll in a series of graduate courses in biology, genetics and biostatistics taught by Stanford University faculty and learn the basic principles for responsible conduct in research. Finally, the remainder of my time (20%) will be spent in teaching activities aimed at training residents and fellows in the management of PAH patients in both inpatient and outpatient settings. My long term goals are to have sufficient published and preliminary data to support my independence as an investigator and to submit an R01 grant that will focus on applying the data obtained in these proposed studies to investigating how abnormal Wnt signaling may contribute to IPAH development. The second part of the proposal contains the research plan. IPAH is a rare and progressive disorder associated with abnormally elevated pulmonary pressures that, if untreated, lead to congestive heart failure and premature death. Patients with PAH demonstrate progressive narrowing and loss of small distal pulmonary arteries without significant compensatory vascular regeneration indicating that a defect in pulmonary angiogenesis is present. Recent studies by our group and others have shown that mutations in the bone morphogenetic protein receptor (BMPR) II could contribute to vessel loss by reducing pulmonary artery endothelial cell (PAEC) survival and proliferation in response to injury but the mechanisms by which BMPRI can regulate these angiogenic responses have not been well characterized. In addition to PAECs, pericyte recruitment is critical for stabilization o newly formed vessels but their contribution to the pathogenesis of IPAH has not been well established. We have recently shown that BMP signaling promotes pulmonary angiogenesis by recruitment of two Wnt signaling pathways in PAECs: the "canonical" Wnt/?-catenin (? C) and the "non-canonical" Wnt/planar cell polarity (PCP) pathways. Based on our preliminary studies and our review of the literature, we propose that activation of the Wnt/PCP pathway is required for pulmonary angiogenesis by 1) inducing PAEC to organize into properly aligned vascular tubes and 2) by promoting pericyte recruitment to vascular tubes to form functional blood vessels. In a series of experiments using PAECs and pericytes obtained from the lungs of healthy donors and IPAH patients, we will demonstrate that Wnt/PCP signaling is required to direct PAEC (Aim 1) and pericyte (Aim 2) behavior during angiogenesis and that loss of activity correlates with reduced pulmonary angiogenesis. Furthermore, we will complement these cell studies with work on two animal models of dysfunctional Wnt/PCP signaling that will help provide insight into the biological relevance of this pathway to compensatory angiogenesis following pneumonectomy (Aim 3). Future studies will center on how mutations that reduce Wnt/PCP signaling can genetically interact with BMPRII mutations to promote development of IPAH and how therapeutic interventions aimed at normalizing Wnt/PCP pathway activity in PAEC and pericytes could help prevent progression and improve survival in IPAH patients by restoring their ability to regenerate lost vessels. PUBLIC HEALTH RELEVANCE: Our goal is to understand how the Wnt/PCP pathway regulates pulmonary angiogenesis and to demonstrate that mutations that disrupt pathway activation can result in PAH. Identification of novel genes could lead to novel therapies that could prevent progression and/or reverse established disease.