Cyclic nucleotide gradients regulate the balance of mechanical forces underlying pulmonary endothelial barrier integrity

环核苷酸梯度调节肺内皮屏障完整性的机械力平衡

• 批准号: 10402898
• 负责人: THOMAS C RICH
• 金额: $ 35.02万
• 依托单位: UNIVERSITY OF SOUTH ALABAMA
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-12-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10402898
• 关键词: 3-Dimensional; A kinase anchoring protein; Acute Respiratory Distress Syndrome; Adenylate Cyclase; Adhesions; Affect; Agonist; Alprostadil; Anisotropy; Apical; Bacterial Infections; Bicarbonates; Blood; Blood Circulation; Cell membrane; Cell-Matrix Junction; Cells; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cyclic GMP; Cyclic Nucleotides; Data; Development; Dimensions; Distant; Endothelial Cells; Endothelium; Equilibrium; Extracellular Matrix; Face; Feedback; Filtration; Fingerprint; Forskolin; Frequencies; Future; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; Generations; Goals; Heterogeneity; Imaging technology; Individual; Inflammation; Inflammation Mediators; Intercellular Junctions; Investigation; Isometric Exercise; Isoproterenol; Kinetics; Liquid substance; Location; Lung; Maps; Measurement; Measures; Mechanics; Mediating; Membrane; Modulus; Molecular; Monolayer Stress Microscopy; Movement; Oxygen; Permeability; Phenotype; Physiological; Prostaglandins; Proteins; Pulmonary Edema; Regulation; Second Messenger Systems; Signal Pathway; Signal Transduction; Site; Stabilizing Agents; Syndrome; Testing; Time; Work; base; cell motility; experimental study; imaging approach; insight; interstitial; lung microvascular endothelial cells; mechanical force; mechanical properties; monolayer; novel; receptor; response; spatiotemporal; temporal measurement

PROJECT SUMMARY Pulmonary microvascular endothelial cells (PMVECs) form contiguous, semi-permeable barriers between the bloodstream and the interstitial space. cAMP generated by plasma membrane-localized adenylyl cyclases (ACs) enhances PMVEC barriers. In contrast, cytosolic cAMP, cGMP, and cUMP generated by exogenous and endogenous soluble cyclases disrupt PMVEC barriers. These observations suggest that cyclic nucleotide signals are highly localized, or compartmentalized, and that near-membrane and cytosolic cAMP, cGMP, and perhaps cUMP signals have opposing effects on endothelial function in the lung microvasculature. The concept of compartmentalized signals implies that feedback networks localized to specific subcellular domains control the kinetics of second messenger signals. However, our understanding of the physiological and pathophysiological implications of localized feedback networks within pulmonary endothelial cells is at best rudimentary. Thus, the overall goal of this project is to determine the spatial and temporal relationships between compartmentalized cAMP signals, PKA-mediated feedback networks, and regulation of mechanical forces in pulmonary endothelial cells. Experiments described in this proposal will for the first time identify where cAMP signals occur in the 3D space of PMVECs, identify important temporal components of cAMP signals, and chart feedback mechanisms contributing to signal localization and kinetics of these signals. In other words, we will provide roadmaps identifying the spatial locations of cAMP signals that are critical for controlling the dynamics of cellular forces. We will then overlay these responses onto PKA activity maps and underlying distributions of A kinase anchoring proteins (AKAPs). As such, successful completion of the studies proposed in this application will identify the spatial and temporal fingerprints of specific cAMP signalosomes that regulate mechanical forces within pulmonary endothelial cells, and thus control endothelial barrier integrity. The spatial and temporal fingerprints will direct future studies aimed at identifying target proteins within these signalosomes, leading to both a better understanding of the molecular mechanisms underlying localized signal transduction and identifying translational targets within signalosomes.

项目摘要 肺微血管内皮细胞（PMVEC）形成连续的，半渗透的屏障 血液和间质空间。由质膜 - 定位腺苷循环酶产生的cAMP （ACS）增强PMVEC屏障。相比之下，外源性和 内源性可溶性循环酶破坏PMVEC屏障。这些观察结果表明环状核苷酸 信号是高度局部或分隔的，并且近膜和胞质营，CGMP和 也许Cump信号对肺微脉管系统中的内皮功能具有相反的影响。这个概念 隔室化信号意味着反馈网络位于特定的亚细胞域控制 第二信号信号的动力学。但是，我们对生理学和 肺内皮细胞内局部反馈网络的病理生理意义充其量是 基本。因此，该项目的总体目标是确定空间和时间关系 在分隔的营地信号，PKA介导的反馈网络和机械调节之间 肺内皮细胞中的力。该提案中描述的实验将首次确定 在PMVEC的3D空间中出现营地信号的地方，确定cAMP的重要时间成分 信号和图表反馈机制有助于信号定位和这些信号的动力学。在 其他词语，我们将提供路线图，以识别营地信号的空间位置 控制细胞力的动力学。然后，我们将这些响应叠加到PKA活动图上，并 激酶锚定蛋白（AKAP）的基础分布。因此，成功完成研究 在此应用中提出的建议将确定特定cAMP信号体的空间和时间指纹 调节肺内皮细胞中的机械力，从而控制内皮屏障完整性。 空间和时间指纹将指导未来的研究，旨在识别其中的靶蛋白 信号体，导致对局部信号的分子机制有更好的理解 转导和识别信号体内的翻译目标。