Role of SIN3a in the epigenetic regulation of the Bone Morphogenetic Protein Receptor Type 2 in pulmonary arterial hypertension

项目概要 PAH 是一种致命性疾病，其特征是远端肺血管进行性重塑， 导致肺血管阻力和肺动脉压升高，可能导致严重的并发症 并发症，例如右心衰竭，并最终死亡。越来越多的证据表明内皮细胞 功能障碍是 PAH 的首要触发因素之一，导致血管细胞不受控制的增殖、血管 肺血管的重塑和闭塞。 PAH 最常见的病理机制之一 是血管细胞中骨形态发生蛋白 2 型受体 (BMPR2) 信号的改变。的 由突变或表达缺失引起的 BMPR2 功能丧失与严重的血流动力学相关 PAH 患者的概况和不良预后。多项研究指出肺内皮是细胞类型 PAH 中 BMPR2 缺失的影响最为严重。然而，其背后的分子机制 BMPR2 表达的调控及其相关的 PAH 样表型仍然很大程度上未知。我们组有 最近表明，SIN3a 在 BMPR2 启动子的 DNA 和组蛋白甲基化中起着核心作用 肺动脉平滑肌细胞与PAH的发病机制。然而，表观遗传和转录 SIN3a 在 PAEC 中调节 BMPR2 基因的机制仍有待阐明。 建议揭示 SIN3a 在肺内皮功能障碍中的作用 我们在这方面的目标 在 PAH 中，确定 PAEC 中 BMPR2 表达调节的下游机制，并评估治疗效果 编码 SIN3a 的修饰 mRNA 对 PAH 动物模型的影响。我们的初步数据表明 SIN3a 是 从 PAH 患者中分离出的 hPAEC 显着下调。在体外，我们观察到 SIN3a 沉默 下调 BMPR2 表达和信号传导，同时增强 PAEC 增殖和迁移。 从机制上讲，我们发现了 SIN3a 调节 hPAEC 中 BMPR2 水平的新分子途径。 我们的数据表明，SIN3a 过表达通过抑制 Zeste 同源物 2 的增强子来上调 FOXK2 FOXK2 启动子中 (EZH2) 介导的组蛋白甲基化。最终，SIN3a 过度表达增加 FOXK2 与 BMPR2 启动子结合并上调 hPAEC 中的 BMPR2 水平。根据这些发现， 我们假设 SIN3a 的缺失会损害 PAEC 中 BMPR2 的表达，触发内皮细胞 PAH 的功能障碍和血管重塑。在本提案中，我们的假设将通过追求 以下三个具体目标： 目标 1) 研究 SIN3a 在内皮细胞功能障碍中的作用并破译 PAH-hPAEC 中 BMPR2 调节的分子机制。目标 2) 阐明效果 使用双重方法研究 SIN3a 缺陷对平滑肌细胞和内皮细胞 PAH 发病的影响。目标 3) 评价SIN3a modRNA在PAH临床前模型中的治疗效果。定义监管 PAH 中 BMPR2 表达缺失的机制具有重要意义。最后，恢复 使用 SIN3a 修饰的 RNA 在肺部表达 SIN3a 可能是治疗 PAH 的一种新的有前景的策略。

Project Summary PAH is a fatal disease characterized by the progressive remodeling of the distal pulmonary vascular arteries, resulting in elevated pulmonary vascular resistance and pulmonary artery pressure, which may lead to serious complications, such as right heart failure, and ultimately death. Accumulating evidence indicates that endothelial dysfunction is one of the first triggers in PAH that leads to uncontrolled proliferation of vascular cells, vascular remodeling, and occlusion of the pulmonary blood vessels. One of the most common pathomechanisms in PAH is the alteration of the Bone Morphogenetic Protein Type 2 Receptor (BMPR2) signaling in vascular cells. The loss of BMPR2 function, induced by mutation or a loss of expression, is associated with a severe hemodynamic profile and poor outcomes in PAH patients. Multiple studies point to the pulmonary endothelium as the cell type that is most critically impacted by BMPR2 loss in PAH. However, the molecular mechanisms underlying the regulation of BMPR2 expression and its associated PAH-like phenotype remain largely unknown. Our group has recently shown that SIN3a plays a central role in the DNA and histone methylation of the BMPR2 promoter in pulmonary artery smooth muscle cells and the pathogenesis of PAH. However, the epigenetic and transcriptional mechanisms by which SIN3a regulates the BMPR2 gene in PAEC remain to be elucidated. proposal are to uncover the role of SIN3a in the pulmonary endothelial dysfunction Our objectives in this in PAH, identify the downstream mechanisms underlying the regulation of BMPR2 expression in PAEC, and evaluate the therapeutic effects of modified mRNA encoding SIN3a in animal models of PAH. Our preliminary data showed that SIN3a is significantly downregulated in hPAEC isolated from PAH patients. In vitro, we observed that SIN3a silencing downregulates BMPR2 expression and signaling while potentiating PAEC proliferation and migration. Mechanistically, we discovered a novel molecular pathway by which SIN3a modulates BMPR2 levels in hPAECs. Our data showed that SIN3a overexpression upregulates FOXK2 by repressing Enhancer of Zeste Homolog 2 (EZH2)-mediated histone methylation in the FOXK2 promoter. Ultimately, SIN3a overexpression increases FOXK2 binding to the BMPR2 promoter and upregulates BMPR2 levels in hPAECs. Based upon these findings, we hypothesize that the loss of SIN3a impairs BMPR2 expression in PAEC, triggers endothelial dysfunction and vascular remodeling in PAH. In this proposal, our hypothesis will be tested by pursuing the following three specific aims: Aim 1) To investigate the role of SIN3a in endothelial cell dysfunction and decipher the molecular mechanism underlying the regulation of BMPR2 in PAH-hPAEC. Aim 2) To elucidate the effects of SIN3a deficiency in the onset of PAH in smooth muscle cells and endothelial cells using a dual approach. Aim 3) To evaluate the therapeutic efficacy of SIN3a modRNA in preclinical models of PAH. Defining the regulatory mechanisms underlying the loss of BMPR2 expression in PAH will be of great relevance. Finally, restoring the expression of SIN3a in the lungs using SIN3a modified RNA might be a new promising strategy for treating PAH.