Angiogenesis in hyperoxic lung fibrosis

描述（由申请人提供）： 这个为期5年的培训计划提出了职业发展计划，作为一个独立的生物医学研究人员在肺损伤和血管生成领域。 主要研究者通过ABIM研究途径完成了肺部和重症监护奖学金培训。 与赞助商和经验丰富的合作者，他将扩大他的科学技能，为职业发展作为一个独立的医生，科学家做准备。 该计划将强调技能的分子生物学血管生成和巨噬细胞调节纤维化使用小鼠高氧模型。 为了提高他在计算生物学方面的知识，他将参加应用统计学课程。 史蒂文·格林伯格医学博士一位巨噬细胞生物学家，和保罗·罗斯曼医学博士，一位在细胞因子信号分子调节领域享有盛誉的免疫学家将提供赞助。 该计划将受益于合作的专业知识，扬kitaubrski博士，在血管生成和帕蒂李，医学博士的专家。世卫组织将为高氧研究提供咨询支持。 此外，世界著名的研究者乔治扬科普洛斯将每8周与赞助商、Kitaubrski博士和Lee博士在一个咨询委员会中合作和服务。 该委员会将审查进展情况，并提供密切的科学支持和职业建议。 长期高氧导致人类和小鼠肺纤维化。 伴随的血管重构导致肺动脉高压、右心衰竭和过早死亡。 该项目解决了以下问题：巨噬细胞衍生的血管生成调节因子，特别是血管生成素-2（Ang-2），是否有助于长时间亚致死性高氧引起的血管重塑。 这些血管改变是否独立地促进了肺纤维化的发展？ 巨噬细胞源性血管紧张素-2是否直接抑制内皮细胞对高氧反应的存活？ 巨噬细胞源性介质在高氧肺损伤中的作用尚不清楚。 我们在初步实验中表明，增加血管紧张素-2 mRNA和蛋白质在高氧暴露RAW 264.7和亚致死高氧肺纤维化的小鼠模型。 这些小鼠发生了巨噬细胞为主的细胞浸润、胶原沉积和肺血管退化，其概括了人类亚急性肺纤维化的特征。 1）通过免疫组织化学、Western和北方印迹和ELISA来表征高氧暴露小鼠中的纤维化和血管重塑反应，以量化肺纤维化、血管生成和血管生成调节剂及其受体的表达的标志物的变化。 将通过共聚焦显微镜评价对肺血管的影响。2)通过血管生成测定，对巨噬细胞衍生的血管生成调节因子表达改变的功能变化作出反应。 巨噬细胞耗竭小鼠将用于评估肺巨噬细胞的贡献。3)在高氧之前通过腺病毒转染或阻断抗体肺靶向过表达Ang-1或2，以确定Ang-2在长期亚致死性高氧期间对肺血管重塑和纤维化的贡献。4)确定Ang-2介导的内皮细胞存活抑制是否是PI 3 K/Akt依赖性的。 哥伦比亚大学在为K 08职业获奖者准备独立职业生涯方面有着成功的指导和培训记录。

DESCRIPTION (provided by applicant): This 5-year training program proposes the development plan for a career as an independent biomedical researcher in the area of lung injury and angiogenesis. The principal investigator, has completed Pulmonary and Critical Care Fellowship training through the ABIM Research Pathway. With the sponsors and experienced collaborators he will expand on his scientific skills in preparation for career progression as an independent physician-scientist. The program will emphasize skills in molecular biology of angiogenesis and macrophage regulation of fibrosis using a murine hyperoxia model. To advance his knowledge in computational biology he will attend courses in applied statistics. Steven Greenberg M.D. a macrophage biologist, and Paul Rothman M.D., a renowned immunologist in the area of molecular regulation of cytokine signaling, will provide sponsorship. The program will benefit from collaborative expertise of Jan Kitajewski PhD, an expert in angiogenesis and Patty Lee, M.D. who will provide consultative support for the hyperoxia studies. Additionally, George Yancopoulos, a world renowned investigator, will collaborate and serve with the sponsors, Drs. Kitajewski and Lee on an advisory committee every 8 weeks. This committee will review progress and provide close scientific support and career advice. Prolonged hyperoxia results in lung fibrosis in humans and mice. The accompanying vascular remodeling contributes to pulmonary hypertension, right heart failure, and premature death. This program addresses the questions: Do macrophage-derived angiogenic regulatory factors, particularly angiopoietin-2 (Ang-2), contribute to vascular remodeling in response to prolonged sublethal hyperoxia. Do these vascular alterations contribute, independently, to the development of lung fibrosis? And does macrophage-derived Ang-2 directly inhibit endothelial cell survival in response to hyperoxia? The role of macrophage-derived mediators in hyperoxic lung injury remains unclear. We demonstrate in preliminary experiments, increased Ang-2 mRNA and protein in response to hyperoxic exposure in RAW 264.7 and in a mouse model of sublethal hyperoxic lung fibrosis. These mice develop macrophage-predominant cellular infiltration, collagen deposition and pulmonary vascular regression that recapitulates features of subacute lung fibrosis in humans. The following specific aims will be accomplished: 1) The characterization of fibrotic and vascular remodeling responses in hyperoxia-exposed mice by immuno-histochemistry, Western and Northern blotting and ELISA to quantify changes in markers of lung fibrosis, angiogenesis and expression of angiogenic regulators and their receptors. The effects on pulmonary vasculature will be evaluated by confocal microscopy. 2) Functional changes in response to altered expression of macrophage-derived angiogenic regulators by angiogenesis assays. Macrophage-depleted mice will be used to assess the contribution of lung macrophages. 3) Lung targeted overexpression of Ang-1 or 2 by adenovirus transfection or blocking antibodies prior to hyperoxia to determine the contribution of Ang-2 to pulmonary vascular remodeling and fibrosis during prolonged sublethal hyperoxia. 4) Determine if Ang-2 mediated endothelial survival inhibition is PI3K/Akt dependent. Columbia University has an established record of successful mentorship and training for K08 Career awardees in their preparation for careers as independent.