Department of Pulmonary and Critical Care Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, 1095 Jiefang Avenue, Wuhan, 430030, China.
Key Laboratory of Respiratory Diseases, National Ministry of Health of the People's Republic of China and National Clinical Research Center for Respiratory Disease, Wuhan, China.
Respir Res. 2022 Jan 11;23(1):6. doi: 10.1186/s12931-022-01927-9.
Hypoxic pulmonary hypertension (HPH) is a chronic progressive advanced disorder pathologically characterized by pulmonary vascular remodeling. Notch4 as a cell surface receptor is critical for vascular development. However, little is known about the role and mechanism of Notch4 in the development of hypoxic vascular remodeling.
Lung tissue samples were collected to detect the expression of Notch4 from patients with HPH and matched controls. Human pulmonary artery smooth muscle cells (HPASMCs) were cultured in hypoxic and normoxic conditions. Real-time quantitative PCR and western blotting were used to examine the mRNA and protein levels of Notch4. HPASMCs were transfected with small interference RNA (siRNA) against Notch4 or Notch4 overexpression plasmid, respectively. Cell viability, cell proliferation, apoptosis, and migration were assessed using Cell Counting Kit-8, Edu, Annexin-V/PI, and Transwell assay. The interaction between Notch4 and ERK, JNK, P38 MAPK were analyzed by co-immunoprecipitation. Adeno-associated virus 1-mediated siRNA against Notch4 (AAV1-si-Notch4) was injected into the airways of hypoxic rats. Right ventricular systolic pressure (RVSP), right ventricular hypertrophy and pulmonary vascular remodeling were evaluated.
In this study, we demonstrate that Notch4 is highly expressed in the media of pulmonary vascular and is upregulated in lung tissues from patients with HPH and HPH rats compared with control groups. In vitro, hypoxia induces the high expression of Delta-4 and Notch4 in HPASMCs. The increased expression of Notch4 promotes HPASMCs proliferation and migration and inhibits cells apoptosis via ERK, JNK, P38 signaling pathways. Furthermore, co-immunoprecipitation result elucidates the interaction between Notch4 and ERK/JNK/P38. In vivo, silencing Notch4 partly abolished the increase in RVSP and pulmonary vascular remodeling caused by hypoxia in HPH rats.
These findings reveal an important role of the Notch4-ERK/JNK/P38 MAPK axis in hypoxic pulmonary remodeling and provide a potential therapeutic target for patients with HPH.
低氧性肺动脉高压（HPH）是一种慢性进行性进展性疾病，其病理特征为肺血管重构。Notch4 作为细胞表面受体，对血管发育至关重要。然而，Notch4 在低氧性血管重构发展中的作用和机制知之甚少。
收集 HPH 患者和匹配对照的肺组织样本，检测 Notch4 的表达。在低氧和常氧条件下培养人肺动脉平滑肌细胞（HPASMCs）。实时定量 PCR 和 Western blot 检测 Notch4 的 mRNA 和蛋白水平。分别用 Notch4 小干扰 RNA（siRNA）或 Notch4 过表达质粒转染 HPASMCs。用细胞计数试剂盒-8（Cell Counting Kit-8）、Edu、Annexin-V/PI 和 Transwell 检测细胞活力、增殖、凋亡和迁移。通过共免疫沉淀分析 Notch4 与 ERK、JNK、P38 MAPK 的相互作用。将腺相关病毒 1 介导的 Notch4 siRNA（AAV1-si-Notch4）注入低氧大鼠气道。评估右心室收缩压（RVSP）、右心室肥厚和肺血管重构。
本研究表明，Notch4 在肺血管中层高度表达，并在 HPH 患者和 HPH 大鼠的肺组织中上调，高于对照组。体外，低氧诱导 HPASMCs 中 Delta-4 和 Notch4 的高表达。Notch4 的高表达促进 HPASMCs 增殖和迁移，通过 ERK、JNK、P38 信号通路抑制细胞凋亡。此外，共免疫沉淀结果阐明了 Notch4 与 ERK/JNK/P38 的相互作用。在体内，沉默 Notch4 部分阻断了 HPH 大鼠低氧引起的 RVSP 升高和肺血管重构。
这些发现揭示了 Notch4-ERK/JNK/P38 MAPK 轴在低氧性肺重构中的重要作用，为 HPH 患者提供了一个潜在的治疗靶点。
