Interleukin-13-mediated mechanisms of pulmonary vascular remodeling

白细胞介素13介导的肺血管重塑机制

• 批准号: 8996962
• 负责人: Won-Kyung Cho
• 金额: $ 13.35万
• 依托单位: BROWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2016-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8996962
• 关键词: Address; Advisory Committees; Animals; Arginine; Biological Assay; Blood Vessels; Cell Proliferation; Chronic; Clinical; Committee Members; Complex; Development; Disease; Environment; Future; Gene Silencing; Generations; Genes; Glycolysis; Grant; Human; Hypoxia; Hypoxia Inducible Factor; Immune; Immunobiology; In Vitro; Inflammation Mediators; Interleukin-13; Isoenzymes; K-Series Research Career Programs; Link; Lung; Lung Inflammation; Medial; Mediating; Mentors; Mentorship; Metabolic; Metabolism; Mitochondria; Mus; Mutant Strains Mice; Nitric Oxide Pathway; Outcome; Oxidative Phosphorylation; Oxygen Consumption; Pathogenesis; Pathway interactions; Patients; Pharmacology; Phenotype; Physicians; Play; Process; Proliferating; Pulmonary Hypertension; Pulmonary Vascular Resistance; Pulmonary artery structure; Receptor Signaling; Regulation; Regulatory Pathway; Reporting; Research; Research Personnel; Role; Scientist; Signal Transduction; Small Interfering RNA; Smooth Muscle Myocytes; Testing; Time; Transgenic Mice; Transgenic Organisms; Universities; Vascular Smooth Muscle; Vascular remodeling; Vascular resistance; Vocational Guidance; Warburg Effect; Western Blotting; Work; aerobic glycolysis; arginase; cancer cell; career; improved; in vivo; insight; interleukin-13 receptor; mutant; novel; null mutation; programs; pulmonary arterial hypertension; receptor; research study; vascular smooth muscle cell proliferation

PROJECT SUMMARY Pulmonary Arterial Hypertension (PAH) is a fatal disease caused by elevated vascular resistance due to lumen-obliterating processes in small pulmonary arteries. To improve clinical outcome, a better understanding regarding the pathogenesis of PAH is desperately needed. Recently we reported that chronic exposure of interleukin-13 (IL-13) in murine lungs, using a transgenic approach (IL-13 Tg), induced impressive smooth muscle cell-driven pulmonary vascular remodeling (PVR) leading to pulmonary hypertension (PH). We further showed that IL-13-induced proliferation of vascular smooth muscle cells in vivo and in vitro is mediated through Arginase2 (Arg2) via IL-13 receptor alpha2 (IL-13Ra2). Also, we observed that IL-13 increased the expression and activity of hypoxic inducible factor1a (HIF1a) via IL-13Ra2, as well, and that IL-13-induced proliferation of human pulmonary artery smooth muscle cell (hPASMC) is significantly decreased with knocking HIF1a gene down. Furthermore, HIF1a regulates Arg2 expression at transcriptional level. These findings implicate the important role of HIF1a in IL-13-induced proliferation of vascular smooth muscle cells. Given that HIF1a is a metabolic switch from oxidative phosphorylation to glycolysis, these findings suggest that IL-13 might induce the Warburg type effect (increased aerobic glycolysis and decreased oxidative phosphorylation in cancer cells leading to proliferation) to stimulate proliferation of vascular smooth muscle cells. Specific Aims We propose the following hypotheses: 1. IL-13 stimulates the proliferation of hPASMC through IL-13Ra2. 2. HIF1a plays a critical role in IL-13-stimulated proliferation of hPASMC via IL-13Ra2. 3. IL-13 regulates the metabolic shift from oxidative phosphorylation to glycolysis via IL-13Ra2, HIF1a and Arg2, thereby inducing PASMC proliferation and PH. Our Aims are to: AIM 1. Define the role of IL-13Ra2 in the development of IL-13-induced PH in vitro and in vivo. AIM 2. Define the role of HIF1a in the generation of the IL-13-IL-13Ra2-induced proliferative phenotypes in PASMC in vitro and in vivo. AIM 3. Define the role of IL-13 in the development of the metabolic shift from oxidative phosphorylation to glycolysis in PASMC in vitro and in vivo. Experimental Approach We will use both in vivo and in vitro approaches using hPASMC and IL-13 Tg mice. To address AIM 1, the pulmonary vascular phenotype will be assessed in IL-13 Tg mice crossed with IL-13Ra2 null mutant compared to IL-13 Tg. Further hPASMC transfected with the specific siRNA of IL-13Ra2 will be treated with IL-13 and proliferation assays will be done. The results will be compared to those from hPASMC transfected with control siRNA. To address AIM 2, HIF1a expression will be examined and compared using Western blot in IL-13- treated hPSAMC with or without knocking the IL-13Ra2 gene down, and IL-13 Tg and IL-13 Tg crossed with IL-13Ra2 null mutant mice. To address AIM 3, lactate levels and oxygen consumption rates in IL-13-stimulated hPASMC will be assessed. A metabolic modifier will be given to IL-13 Tg mice to see whether the metabolic shift in IL-13 Tg through HIF1a alters the pulmonary vascular phenotype in these animals. This work will provide us with a novel insight into the pathogenesis of pulmonary hypertension induced by a complex interaction between immune and metabolic processed in addition to a novel regulatory pathway of HIF1a. This grant proposes a research mentorship program at Yale University under the primary sponsorship of Dr. Jack Elias, a world-renowned investigator in lung inflammation and immunobiology, and Dr. Hyung Joon Chun, an expert in vascular signaling and pharmacology, as a co-mentor. We have also enlisted the expertise of Yale investigators Drs. Gerald Shadel, Patty Lee, William Sessa, Frank Giordano and Chun Geun Lee as advisory committee members to provide scientific and career counseling. The proposed career and research program as outlined will provide an extraordinary scientific environment wherein Dr. Cho can launch her future independent career as a physician-scientist.

项目摘要 肺动脉高压（PAH）是一种致命的疾病，由血管阻力升高引起， 小肺动脉的管腔闭塞过程。为了改善临床结果，更好地了解 关于PAH发病机制的研究是迫切需要的。最近，我们报告说，长期接触 白细胞介素-13（IL-13）在小鼠肺中，使用转基因方法（IL-13 Tg），诱导了令人印象深刻的平滑 肌细胞驱动的肺血管重构（PVR）导致肺动脉高压（PH）。我们进一步 表明IL-13诱导的血管平滑肌细胞在体内和体外的增殖是通过 精氨酸酶2（Arg 2）通过IL-13受体α 2（IL-13 Ra 2）。此外，我们观察到IL-13增加了表达 和缺氧诱导因子1a（HIF 1a）的活性，以及IL-13诱导的 敲除HIF 1a基因后，人肺动脉平滑肌细胞（hPASMC）数量明显减少 下来此外，HIF 1a在转录水平上调节Arg 2的表达。这些发现表明， HIF 1a在IL-13诱导血管平滑肌细胞增殖中的重要作用HIF 1a是一种 从氧化磷酸化到糖酵解的代谢转换，这些发现表明IL-13可能诱导 瓦尔堡型效应（癌细胞中有氧糖酵解增加和氧化磷酸化减少 导致增殖）以刺激血管平滑肌细胞的增殖。 具体目标 我们提出以下假设： 1. IL-13通过IL-13 Ra 2刺激hPASMC增殖。 2. HIF 1a通过IL-13 Ra 2在IL-13刺激的hPASMC增殖中起关键作用。 3. IL-13通过IL-13 Ra 2、HIF 1a调节从氧化磷酸化到糖酵解的代谢转变 和Arg 2，从而诱导PASMC增殖和PH。 我们的目标是： AIM 1.定义IL-13 Ra 2在体外和体内IL-13诱导的PH发展中的作用。 AIM 2.定义HIF 1a在IL-13-IL-13 Ra 2诱导的增殖表型的产生中的作用， 体外和体内PASMC。 AIM 3.确定IL-13在从氧化磷酸化到磷酸化的代谢转变中的作用。 在体外和体内PASMC的糖酵解。 实验方法 我们将使用hPASMC和IL-13 Tg小鼠的体内和体外方法。为了解决AIM 1问题， 肺血管表型将在与IL-13 Ra 2无效突变体杂交的IL-13 Tg小鼠中进行评估， 至IL-13 Tg.用IL-13 Ra 2的特异性siRNA转染的另外的hPASMC将用IL-13和IL-14处理。 进行增殖测定。将结果与用对照转染的hPASMC的结果进行比较 siRNA。为了解决AIM 2，将使用IL-13- 1中的蛋白质印迹法检查并比较HIF 1a表达。 用或不用敲低IL-13 Ra 2基因处理hPSAMC，IL-13 Tg和IL-13 Tg与 IL-13 Ra 2无效突变小鼠。为了解决AIM 3，在IL-13刺激的小鼠中， 将评估hPASMC。将代谢调节剂给予IL-13 Tg小鼠，以观察代谢调节剂是否能够抑制IL-13 Tg小鼠的生长。 IL-13 Tg通过HIF 1a的转移改变了这些动物的肺血管表型。 这项工作将为我们提供一个新的见解肺动脉高压的发病机制， 免疫和代谢过程之间的复杂相互作用，以及一种新的调节途径， HIF 1a。 这项拨款建议在耶鲁大学的研究导师计划的主要赞助下，博士。 世界著名的肺部炎症和免疫生物学研究者Jack Elias和Hyung Joon Chun博士 血管信号和药理学方面的专家，作为共同导师。我们还邀请了耶鲁大学的专家 研究人员Gerald Shadel博士，Patty Lee，William Sessa，Frank Giordano和Chun Geun Lee担任顾问 委员会成员提供科学和职业咨询。拟议的职业和研究计划 将提供一个非凡的科学环境，赵博士可以在那里启动她的未来， 作为一名独立的物理学家和科学家。