PDGF通过下调miR-1281诱发肺动脉高压形成的分子机制

Pulmonary arterial hypertension (PAH), a complex vascular disease with a dreadful survival rate, is characterized by abnormal proliferation of pulmonary artery smooth muscle cells (PASMCs) and progressive structural remolding of pulmonary arteries. Platelet-derived growth factor (PDGF), a potent mitogen, has been implicated in these processes. Inhibition of the PDGF signaling pathway is consideraed as a potential target for the treatment of pulmonary hypertension. Our preliminary data showed that PDGF-BB down-regulated miR-1281 and EP300 expression. HDAC4, a functional antagonism of EP300, is a direct target of miR-1281. Functionally, miR-1281 inhibits the proliferation of PASMCs. Based on the results above, we propose that PDGF-reduced miR-1281 might contribute to the development of PAH. In this proposal, we will explore the molecular mechenism of miR-1281 downregulation induced by PDGF-BB and the potential biological function of miR-1281. The impact of synthetic miR-1281 agomir on pulmonary vascular remodeling and PAH symtoms will be evaluated by using hypoxia induced PAH rat model. Finally, the clinical relationship between miR-1281 level and PAH disease will be evaluated. A new molecular mechanism of PDGF-induced PAH will be elucidated by epigenetic studies including miRNA, methylation, histone acetylation and deacetylation. This project will also provide a new clue for the development of miRNA-based treatment and early diagnosis means for PAH.

肺动脉高压（PAH）发病机制复杂，死亡率高。肺动脉平滑肌细胞(PASMC)增殖和肺血管结构重建是PAH的重要特征，PDGF是参与这一过程强有力的有丝分裂原，阻断PDGF通路已成为PAH治疗的有效途径。前期研究发现，PDGF-BB下调miR-1281及EP300；miR-1281靶基因则是与EP300功能拮抗的HDAC4；过表达miR-1281能抑制PASMC增殖。推测miR-1281参与调控PDGF-BB诱发的PAH。本项目拟从分子水平揭示PDGF-BB如何调控miR-1281；研究miR-1281下调的功能及机制；以PAH大鼠为模型，探讨导入miR-1281对PAH症状的改善作用；明确miR-1281与PAH在临床上的相关性。研究将从表观遗传学角度，包括miRNA、甲基化、组蛋白乙酰化和去乙酰化等改变阐述PDGF-BB诱发PAH的新机制，也为开发基于miRNA的治疗和早期诊断提供新思路。

背景：肺动脉平滑肌细胞（PASMC）异常增殖、迁移引发肺动脉血管重构是PAH的重要病理基础。血小板衍生生长因子（PDGF）是PASMC的一种重要的促有丝分裂原，与肺动脉血管重构密切相关。microRNA（miRNA）是一类非编码单链小RNA，通过与靶基因3`非翻译区相互作用，在转录后水平抑制目的基因的表达。近期的研究表明，PDGF调控的miRNAs在调控PASMC的细胞功能和介导PDGF刺激的细胞高增殖状态中起着关键作用。.方法和结果：前期的miRNA表达谱分析发现，miR-1281是在PDGF-BB处理的PASMC中下调最显著的miRNA。进一步检测证实其在PDGF刺激的人和大鼠PASMC中均下调，其表达在物种中具有保守性。通过过表达或干扰表达的细胞功能研究发现，miR-1281抑制PASMC的增殖和迁移。生物信息学预测和3'-非翻译区报告基因分析证实组蛋白去乙酰化酶4（HDAC4）是miR-1281的靶基因。增殖和迁移实验证明HDAC4的细胞功能与miR-1281呈负相关。机制上，PDGF-BB激活磷脂酰肌醇3激酶（PI3K）途径，然后诱导DNA甲基转移酶1（DNMT1）的表达，导致启动子上游CpG岛甲基化增强，使得miR-1281的表达被抑制。此外，在低氧诱导的PASMC、野百合碱（MCT）诱导的PAH大鼠的肺动脉，以及冠心病合并肺动脉高压（CHD-PAH）患者血清中都发现miR-1281的表达水平显著降低。.结论：我们阐释了一种全新的、整合了多种表观遗传调节因子的调控途径，即PI3K-DNMT1-miR-1281-HDAC4，证明其参与调节PDGF-BB诱导的PASMC的增殖和迁移以及肺血管重塑。这些数据提示，miR-1281可能作为一种潜在的生物标志物和小分子药物应用于PAH的诊断和治疗。