Nikitopoulou Ioanna, Orfanos Stylianos E, Kotanidou Anastasia, Maltabe Violetta, Manitsopoulos Nikolaos, Karras Panagiotis, Kouklis Panos, Armaganidis Apostolos, Maniatis Nikolaos A
First Department of Critical Care Medicine & Pulmonary Services, GP Livanos and M Simou Laboratories, University of Athens Medical School, Evangelismos Hospital, Athens, Greece;
First Department of Critical Care Medicine & Pulmonary Services, GP Livanos and M Simou Laboratories, University of Athens Medical School, Evangelismos Hospital, Athens, Greece; Second Department of Critical Care, University of Athens Medical School, Attikon Hospital, Haidari, Greece; and.
Am J Physiol Lung Cell Mol Physiol. 2016 Aug 1;311(2):L352-63. doi: 10.1152/ajplung.00156.2014. Epub 2016 May 27.
Increased pulmonary vascular resistance in pulmonary hypertension (PH) is caused by vasoconstriction and obstruction of small pulmonary arteries by proliferating vascular cells. In analogy to cancer, subsets of proliferating cells may be derived from endothelial cells transitioning into a mesenchymal phenotype. To understand phenotypic shifts transpiring within endothelial cells in PH, we injected rats with alkaloid monocrotaline to induce PH and measured lung tissue levels of endothelial-specific protein and critical differentiation marker vascular endothelial (VE)-cadherin. VE-cadherin expression by immonoblotting declined significantly 24 h and 15 days postinjection to rebound to baseline at 30 days. There was a concomitant increase in transcriptional repressors Snail and Slug, along with a reduction in VE-cadherin mRNA. Mesenchymal markers α-smooth muscle actin and vimentin were upregulated by immunohistochemistry and immunoblotting, and α-smooth muscle actin was colocalized with endothelial marker platelet endothelial cell adhesion molecule-1 by confocal microscopy. Apoptosis was limited in this model, especially in the 24-h time point. In addition, monocrotaline resulted in activation of protein kinase B/Akt, endothelial nitric oxide synthase (eNOS), nuclear factor (NF)-κB, and increased lung tissue nitrotyrosine staining. To understand the etiological relationship between nitrosative stress and VE-cadherin suppression, we incubated cultured rat lung endothelial cells with endothelin-1, a vasoconstrictor and pro-proliferative agent in pulmonary arterial hypertension. This resulted in activation of eNOS, NF-κB, and Akt, in addition to induction of Snail, downregulation of VE-cadherin, and synthesis of vimentin. These effects were blocked by eNOS inhibitor N(ω)-nitro-l-arginine methyl ester. We propose that transcriptional repression of VE-cadherin by nitrosative stress is involved in endothelial-mesenchymal transdifferentiation in experimental PH.
肺动脉高压（PH）中肺血管阻力增加是由血管收缩以及增殖的血管细胞阻塞小肺动脉所致。与癌症类似，增殖细胞亚群可能源自内皮细胞向间充质表型的转变。为了解PH中内皮细胞内发生的表型变化，我们给大鼠注射生物碱野百合碱以诱导PH，并测量肺组织中内皮特异性蛋白和关键分化标志物血管内皮（VE）-钙黏蛋白的水平。注射后24小时和15天，通过免疫印迹法检测到的VE-钙黏蛋白表达显著下降，至30天时反弹至基线水平。转录抑制因子Snail和Slug随之增加，同时VE-钙黏蛋白mRNA减少。免疫组织化学和免疫印迹法显示间充质标志物α-平滑肌肌动蛋白和波形蛋白上调，共聚焦显微镜观察发现α-平滑肌肌动蛋白与内皮标志物血小板内皮细胞黏附分子-1共定位。在该模型中细胞凋亡有限，尤其是在24小时时间点。此外，野百合碱导致蛋白激酶B/Akt、内皮型一氧化氮合酶（eNOS）、核因子（NF）-κB激活，肺组织硝基酪氨酸染色增加。为了解亚硝化应激与VE-钙黏蛋白抑制之间的病因学关系，我们用内皮素-1（一种肺动脉高压中的血管收缩剂和促增殖剂）培养大鼠肺内皮细胞。这除了诱导Snail、下调VE-钙黏蛋白和合成波形蛋白外，还导致eNOS、NF-κB和Akt激活。这些作用被eNOS抑制剂N（ω）-硝基-L-精氨酸甲酯阻断。我们认为亚硝化应激对VE-钙黏蛋白的转录抑制参与了实验性PH中的内皮-间充质转分化。
