Pulmonary arterial hypertension (PAH) is a fatal cardiopulmonary disease characterized by increased vascular cell proliferation with resistance to apoptosis and occlusive remodeling of the small pulmonary arteries in humans. The Notch family of proteins are proximal signaling mediators of an evolutionarily conserved pathway that effect cell proliferation, fate determination, and development. In endothelial cells (ECs), Notch receptor 2 (Notch2) has been shown to promote endothelial apoptosis. However, a pro- or anti-proliferative role for Notch2 in pulmonary endothelial proliferation and ensuing PAH is unknown. Herein, we postulated that suppressed Notch2 signaling drives pulmonary endothelial proliferation in the setting of PAH. We observed that levels of Notch2 are ablated in lung and PA tissue samples from PAH patients compared to non-PAH controls. Interestingly, Notch2 expression was attenuated in human pulmonary artery endothelial cells (hPAECs) exposed to vasoactive factors including hypoxia, TGFβ, ET-1, and IGF-1. Gene silencing of Notch2 increased EC proliferation and reduced apoptosis. At the molecular level, Notch2-deficient hPAECs activated Akt, Erk1/2 and anti-apoptotic protein Bcl-2, and reduced levels of p21^cip and Bax. Intriguingly, loss of Notch2 elicits a paradoxical activation of Notch1 and transcriptional upregulation of canonical Notch target genes Hes1, Hey1 and Hey2. Further, reduction in Rb and increased E2F1 binding to the Notch1 promoter appear to explain the upregulation of Notch1. In aggregate, our results demonstrate that loss of Notch2 derepresses Notch1 and elicits aberrant EC hallmarks of PAH. The data underscore a novel role for Notch in the maintenance of endothelial cell homeostasis.

中文翻译：

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Notch2 抑制模拟人类肺动脉高压的变化调节 Notch1 并促进内皮细胞增殖

肺动脉高压 (PAH) 是一种致命的心肺疾病，其特征是血管细胞增殖增加，具有抗细胞凋亡和人类小肺动脉闭塞性重塑的特点。Notch 蛋白家族是影响细胞增殖、命运决定和发育的进化保守途径的近端信号传导介质。在内皮细胞 (EC) 中，Notch 受体 2 (Notch2) 已被证明可促进内皮细胞凋亡。然而，Notch2 在肺内皮增殖和随后的 PAH 中的促增殖或抗增殖作用尚不清楚。在此，我们假设在 PAH 的情况下，抑制的 Notch2 信号传导会驱动肺内皮细胞增殖。我们观察到，与非 PAH 对照相比，来自 PAH 患者的肺和 PA 组织样本中 Notch2 的水平被消融。有趣的是，在暴露于血管活性因子（包括缺氧、TGFβ、ET-1 和 IGF-1）的人肺动脉内皮细胞 (hPAEC) 中，Notch2 的表达减弱。Notch2的基因沉默增加了EC增殖并减少了细胞凋亡。在分子水平上，Notch2 缺陷型 hPAECs 激活 Akt、Erk1/2 和抗凋亡蛋白 Bcl-2，并降低 p21 水平^cip和 Bax。有趣的是，Notch2 的缺失引起了 Notch1 的矛盾激活和典型 Notch 靶基因 Hes1、Hey1 和 Hey2 的转录上调。此外，Rb 的减少和 E2F1 与 Notch1 启动子的结合增加似乎可以解释 Notch1 的上调。总之，我们的结果表明，Notch2 的缺失会抑制 Notch1 并引发 PAH 的异常 EC 标志。这些数据强调了 Notch 在维持内皮细胞稳态中的新作用。