Teaching and Research Office of Clinical Pharmacology, College of Pharmacy, Xinxiang Medical University, Xinxiang, China.
Department of Pharmacy, The Second Affiliated Hospital, Nanchang University, Nanchang, China.
J Cell Physiol. 2019 Jun;234(6):9535-9550. doi: 10.1002/jcp.27642. Epub 2018 Oct 26.
Pulmonary arterial hypertension (PAH) is characterized by the apoptosis resistance and hyperproliferation of pulmonary artery smooth muscle cells (PASMCs). Its pathogenesis has not been revealed. Here, we carried out experiments to investigate the functions of miR-140-5p and tumor necrosis factor-α (TNF-α).
We selected GSE703 from Gene Expression Omnibus (GEO) Database to conduct microarray analysis using R software and Gene Set Enrichment Analysis (GSEA). Combing bioinformatics results, the upregulation of miR-140-5p inhibited PAH progression through targeting TNF-α. RNA expression was measured by quantitative real-time polymerase chain reaction (RT-qPCR) and protein level was measured by western blot analysis and enzyme-linked immunosorbent assays (ELISA). We conducted monocrotaline (MCT) injection to rats to form PAH animal models. The lung tissues were observed by hematoxylin-eosin (HE) staining and Sirius red-picric acid staining. Right ventricular systolic pressure (RVSP) and the ratio of right ventricle (RV)-to-left ventricle (LV) plus septum (S) weight (RV/[LV + S]) were measured in MCT-induced animal models. Overexpression of miR-140-5p and TNF-α were utilized to research the proliferation, migration, and phenotypic variation of hypoxia-mediated PASMCs. The binding between miR-140-5p and TNF-α 3'-untranslated region (3'-UTR) was confirmed via luciferase reporter assay.
Downregulation of miR-140-5p and upregulation of TNF-α were observed in PAH rat model and hypoxia-mediated PASMCs. And we proved that overexpression of miR-140-5p could suppress the proliferation, migration, and phenotypic variation of PASMCs, therefore inhibiting PAH pathogenesis. Luciferase assay verified that miR-140-5p targeted TNF-α directly. A converse correlation was also shown between miR-140-5p and TNF-α in PASMCs.
miR-140-5p and TNF-α are important regulators in PAH pathology and may serve as a therapeutic target for PAH.
肺动脉高压（PAH）的特征是肺动脉平滑肌细胞（PASMC）的凋亡抵抗和过度增殖。其发病机制尚未阐明。在这里，我们进行了实验来研究 miR-140-5p 和肿瘤坏死因子-α（TNF-α）的功能。
我们从基因表达综合数据库（GEO）中选择 GSE703，使用 R 软件和基因集富集分析（GSEA）进行微阵列分析。结合生物信息学结果，上调的 miR-140-5p 通过靶向 TNF-α抑制 PAH 的进展。通过实时定量聚合酶链反应（RT-qPCR）测量 RNA 表达，通过 Western blot 分析和酶联免疫吸附测定（ELISA）测量蛋白质水平。我们对大鼠进行了单硝酸异山梨酯（MCT）注射以形成 PAH 动物模型。通过苏木精-伊红（HE）染色和天狼星红苦味酸染色观察肺组织。在 MCT 诱导的动物模型中测量右心室收缩压（RVSP）和右心室（RV）-左心室（LV）加室间隔（S）重量比（RV/[LV+S]）。利用 miR-140-5p 和 TNF-α 的过表达来研究缺氧介导的 PASMC 增殖、迁移和表型变化。通过荧光素酶报告基因测定证实 miR-140-5p 与 TNF-α 3'-非翻译区（3'-UTR）的结合。
在 PAH 大鼠模型和缺氧介导的 PASMC 中观察到 miR-140-5p 的下调和 TNF-α 的上调。并且我们证明过表达 miR-140-5p 可以抑制 PASMC 的增殖、迁移和表型变化，从而抑制 PAH 发病机制。荧光素酶测定证实 miR-140-5p 直接靶向 TNF-α。在 PASMC 中还显示出 miR-140-5p 与 TNF-α 之间存在相反的相关性。
miR-140-5p 和 TNF-α 是 PAH 病理中的重要调节剂，可能成为 PAH 的治疗靶点。
