ALVEOLO-CAPILLARY SIGNALING IN LUNG IMMUNOLOGY

肺免疫学中的肺泡毛细血管信号转导

• 批准号: 7643372
• 负责人: Jahar Bhattacharya
• 金额: $ 38.16万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-30 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7643372
• 关键词: Abbreviations; Address; Alveolar; Alveolar Cell; Alveolus; Blood; Blood capillaries; Bypass; Capillary Endothelial Cell; Cell membrane; Confocal Microscopy; Cytosolic Phospholipase A2; Detection; Dyes; Endoplasmic Reticulum; Endothelial Cells; Enzymes; Epithelial; Epithelial Cells; Funding; Hand; Hydrogen Peroxide; Image; Immunology; Investigation; Leukocytes; Lung; Lung Inflammation; Mediating; Membrane; Methods; Mitochondria; Mus; Oxidases; Pathway interactions; Play; Preparation; Procedures; Production; Rattus; Reactive Oxygen Species; Regulation; Reporting; Research; Role; Signal Transduction; Stimulus; Testing; Time; Translating; arachidonate; capillary; chemokine; fluorophore; novel; optical imaging; receptor; response

DESCRIPTION (provided by applicant): Aims: Our overall aim is to understand the role of alveoli in the initiation of lung inflammation. Previously, we reported that alveolar TNFalpha induces vectorial crosstalk between epithelial and endothelial cells of intact alveolo-capillary membranes, and that increases in the AEC cytosolic Ca2+ (cytCa2+) drive the crosstalk. Here, we will test the hypothesis that the crosstalk messenger is H2O2 originating from mitochondria or NAD(P)H oxidase of AEC. Our Specific Aims are to define alveolo-capillary signaling in the context of the regulation of cytCa2+ and H2O2 production in AEC by receptor-mediated (Specific Aims 1 and 2) and receptor-independent (Specific Aim 3) mechanisms. Procedures: We will use the isolated, blood-perfused lung preparation for mouse and rat. Alveoli will be loaded with specific fluorophores for the detection of cytosolic, mitochondrial and endosomal store Ca2+, and for ROS production. Concomitantly, capillaries will be loaded with similar dyes to determine crosstalk responses. Single AEC and EC in the alveolo-capillary region will be optically imaged in real-time by of wide-angle and confocal microscopy. For proinflammatory activation, alveoli capillaries will be challenged with TNFalpha, LPS, arachidonate, concentrated HCI and photolytic uncaging of Ca2+. EC responses will be determined in terms of cytCa2+ and ROS production and the recruitment of leukocytes. The hypotheses will also be tested in mice containing genetically defective in NAD(P)H oxidase (gp91phox(-/-)), and in cPLA2. Significance: Although pathological stimuli in the alveolus induce lung inflammation, the role of the alveolar epithelial barrier in generating this response is not understood. Our research will reveal the importance of different modes of Ca2+ mobilization in AEC, namely those that are receptor-mediated versus receptor-independent, in establishing the messenger that translates the alveolar response to proinflammatory activation of EC in capillaries. This research is novel and it will advance our understanding of fundamental mechanisms regulating lung inflammation.

描述（由申请人提供）：目的：我们的总体目标是了解肺泡在肺部炎症发生中的作用。以前，我们报告说，肺泡TNF α诱导完整的肺泡毛细血管膜的上皮细胞和内皮细胞之间的矢量串扰，并在AEC胞浆Ca 2+（cytCa 2+）驱动串扰的增加。在这里，我们将测试的假设，串扰信使是H2 O2来源于线粒体或NAD（P）H氧化酶的AEC。我们的具体目标是通过受体介导（具体目标1和2）和受体非依赖性（具体目标3）机制，在AEC中调节细胞内Ca 2+和H2 O2产生的背景下定义肺泡-毛细血管信号传导。 程序：我们将使用小鼠和大鼠的离体血液灌注肺制剂。肺泡将装载特异性荧光团，用于检测胞质、线粒体和内体储存Ca 2+，以及ROS产生。同时，毛细管将装载类似的染料以确定串扰响应。单个AEC和肺泡毛细血管区的EC将通过广角和共聚焦显微镜进行实时光学成像。对于促炎活化，将用TNF α、LPS、花生四烯酸、浓HCl和Ca 2+的光解释放来激发肺泡毛细血管。EC反应将根据cytCa 2+和ROS的产生以及白细胞的募集来确定。还将在含有NAD（P）H氧化酶（gp 91 phox（-/-））和cPLA 2遗传缺陷的小鼠中检验这些假设。 重要性：虽然肺泡中的病理刺激诱导肺部炎症，但肺泡上皮屏障在产生这种反应中的作用尚不清楚。我们的研究将揭示AEC中不同模式的Ca 2+动员的重要性，即那些受体介导的与受体无关的，在建立将肺泡反应转化为毛细血管中EC的促炎激活的信使中。这项研究是新颖的，它将促进我们对调节肺部炎症的基本机制的理解。