Alvarez Roger A, Miller Megan P, Hahn Scott A, Galley Joseph C, Bauer Eileen, Bachman Timothy, Hu Jian, Sembrat John, Goncharov Dmitry, Mora Ana L, Rojas Mauricio, Goncharova Elena, Straub Adam C
1 Division of Pulmonary, Allergy, Critical Care, and Sleep Medicine, Miller School of Medicine, University of Miami, Miami, Florida; and.
2 Heart, Lung, Blood, and Vascular Medicine Institute.
Am J Respir Cell Mol Biol. 2017 Dec;57(6):733-744. doi: 10.1165/rcmb.2016-0418OC.
Pulmonary hypertension is characterized by pulmonary endothelial dysfunction. Previous work showed that systemic artery endothelial cells (ECs) express hemoglobin (Hb) α to control nitric oxide (NO) diffusion, but the role of this system in pulmonary circulation has not been evaluated. We hypothesized that up-regulation of Hb α in pulmonary ECs contributes to NO depletion and pulmonary vascular dysfunction in pulmonary hypertension. Primary distal pulmonary arterial vascular smooth muscle cells, lung tissue sections from unused donor (control) and idiopathic pulmonary artery (PA) hypertension lungs, and rat and mouse models of SU5416/hypoxia-induced pulmonary hypertension (PH) were used. Immunohistochemical, immunocytochemical, and immunoblot analyses and transfection, infection, DNA synthesis, apoptosis, migration, cell count, and protein activity assays were performed in this study. Cocultures of human pulmonary microvascular ECs and distal pulmonary arterial vascular smooth muscle cells, lung tissue from control and pulmonary hypertensive lungs, and a mouse model of chronic hypoxia-induced PH were used. Immunohistochemical, immunoblot analyses, spectrophotometry, and blood vessel myography experiments were performed in this study. We find increased expression of Hb α in pulmonary endothelium from humans and mice with PH compared with controls. In addition, we show up-regulation of Hb α in human pulmonary ECs cocultured with PA smooth muscle cells in hypoxia. We treated pulmonary ECs with a Hb α mimetic peptide that disrupts the association of Hb α with endothelial NO synthase, and found that cells treated with the peptide exhibited increased NO signaling compared with a scrambled peptide. Myography experiments using pulmonary arteries from hypoxic mice show that the Hb α mimetic peptide enhanced vasodilation in response to acetylcholine. Our findings reveal that endothelial Hb α functions as an endogenous scavenger of NO in the pulmonary endothelium. Targeting this pathway may offer a novel therapeutic target to increase endogenous levels of NO in PH.
肺动脉高压的特征是肺内皮功能障碍。先前的研究表明，全身动脉内皮细胞（ECs）表达血红蛋白（Hb）α以控制一氧化氮（NO）的扩散，但该系统在肺循环中的作用尚未得到评估。我们假设肺ECs中Hbα的上调导致肺动脉高压中NO的消耗和肺血管功能障碍。使用了原发性远端肺动脉血管平滑肌细胞、来自未使用供体（对照）和特发性肺动脉（PA）高压肺的肺组织切片，以及SU5416/低氧诱导的肺动脉高压（PH）的大鼠和小鼠模型。本研究进行了免疫组织化学、免疫细胞化学和免疫印迹分析以及转染、感染、DNA合成、凋亡、迁移、细胞计数和蛋白质活性测定。使用了人肺微血管ECs和远端肺动脉血管平滑肌细胞的共培养物、对照和肺动脉高压肺的肺组织以及慢性低氧诱导的PH小鼠模型。本研究进行了免疫组织化学、免疫印迹分析、分光光度法和血管肌动描记术实验。我们发现，与对照组相比，患有PH的人和小鼠的肺内皮中Hbα的表达增加。此外，我们显示在低氧条件下与PA平滑肌细胞共培养的人肺ECs中Hbα上调。我们用一种破坏Hbα与内皮型NO合酶结合的Hbα模拟肽处理肺ECs，发现与乱序肽相比，用该肽处理的细胞表现出增强的NO信号。使用低氧小鼠肺动脉进行的肌动描记术实验表明，Hbα模拟肽增强了对乙酰胆碱的血管舒张反应。我们的研究结果表明，内皮Hbα在肺内皮中作为NO的内源性清除剂发挥作用。靶向该途径可能为增加PH中内源性NO水平提供一个新的治疗靶点。
