Novel roles of RNA modifications in the pathogenesis of pulmonary vascular remodeling and PAH

肺动脉高压(PAH)的特征是肺血管进行性增加。 阻力和闭塞性血管重塑，导致右心衰竭和过早死亡。vt.给出 闭塞性血管重塑的潜在分子机制仍然是个谜，目前的治疗方法 没有针对基本的疾病修改机制，因此只导致了适度的 发病率和死亡率的改善。而几个转录因子和转录协同- 激活剂已经在PAH的背景下进行了研究，PAH是mRNAs的转录后调节，可以影响 关键蛋白的表达在很大程度上仍未被探索。包括N6-甲基腺苷在内的RNA修饰 (M6A)最近被发现是基因表达的重要调节因子。M6A改装 控制着RNA的稳定性、运输和翻译，并与人类疾病有关，如肥胖和 癌症。尽管M6A在各种基本生物过程中具有重要的功能，但对M6A修饰的研究 在PAH中缺少mRNAs。我们的支持数据表明，脂肪质量的表达与肥胖相关 蛋白质(FTO)是一种具有良好特性的RNA去甲基酶，在肺血管内皮细胞中显著升高 特发性肺高压患者的细胞(ECs)。Tie2Cre介导的内皮细胞FTO缺失(FtoTie2Cre)抑制PH 慢性低氧所致。我们的机制研究提供了证据表明，几个导致PAH的基因 人肺内皮细胞中潜在的FTO靶点。此外，野百合碱对FTO的药理抑制作用-- 应激组大鼠肺血管重塑和肺高压均受到抑制。因此，我们假设内皮细胞 FTO作为一种主要的m6A擦除剂，对mRNA的稳定性和导致PAH的关键因素的积累起着关键的调节作用 内皮细胞中调节肺血管收缩和血管重塑的基因是一种新的治疗方法 多环芳烃的目标。我们提出了以下三个具体目标。在目标1中，我们将定义 内皮细胞FTO在PAH发病机制中的作用在目标2中，我们将描述潜在的分子机制 FTO对导致肺血管收缩和血管重构的内皮功能障碍的调节。 我们将确定ECs中的关键FTO靶点，并探讨挽救FtoTieCre表型的可能性 通过新型纳米颗粒介导的体内EC特异性基因转移。在目标3中，我们将探索治疗 包括FDA批准的改变用途的药物在内的FTO抑制剂在治疗PAH方面的潜力 赠送PAH动物模型。基于我们新发现的明确的临床相关性，我们预计 通过描绘分子和细胞，拟议的研究具有显著的翻译潜力 内皮功能障碍导致肺血管收缩和血管重塑的机制 确定可用药的靶点，并探索可以在药理上 抑制/逆转血管重塑和抑制血管收缩防治PAH 在病人身上。因此，我们相信拟议的研究具有很大的翻译潜力。

Pulmonary arterial hypertension (PAH) is characterized by progressive increase of pulmonary vascular resistance and obliterative vascular remodeling that causes right heart failure and premature death. Given the underlying molecular mechanisms of obliterative vascular remodeling remain enigmatic, current therapies have not targeted the fundamental disease modifying mechanisms and hence only resulted in a modest improvement in the morbidity and mortality. While several transcription factors and transcriptional co- activators have been studied in the context of PAH, post-transcriptional regulations of mRNAs that can affect expression of key proteins remain largely unexplored. RNA modifications including N6-methyladenosine (m6A) have recently been discovered as essential regulators of gene expression. The m6A modification controls RNA stability, transport, and translation and has been linked to human diseases such as obesity and cancers. Despite its functional importance in various fundamental bioprocesses, studies of m6A modification of mRNAs in PAH are lacking. Our Supporting Data show that expression of fat mass and obesity-associated protein (FTO), a well-characterized RNA demethylase, is markedly elevated in pulmonary vascular endothelial cells (ECs) of idiopathic PAH patients. Tie2Cre-mediated deletion of Fto in ECs (FtoTie2Cre) inhibited PH induced by chronic hypoxia. Our mechanistic studies provide evidence that several PAH-causing genes are potential FTO targets in human lung ECs. Furthermore, pharmacological inhibition of FTO in monocrotaline- challenged rats inhibited pulmonary vascular remodeling and PH. Thus, we hypothesize that endothelial FTO, as a major m6A eraser, acts as a key regulator of mRNA stability and accumulation of key PAH-causing genes in ECs to regulate pulmonary vasoconstriction and vascular remodeling and thus is a novel therapeutic target for PAH. We propose the following 3 Specific Aims. In Aim 1, we will define the fundamental role of endothelial FTO in the pathogenesis of PAH. In Aim 2, we will delineate the molecular mechanisms underlying FTO regulation of endothelial dysfunction leading to pulmonary vasoconstriction and vascular remodeling. We will identify the key FTO targets in ECs and address the possibility of rescuing the phenotype of FtoTieCre by novel nanoparticle-mediated in vivo EC-specific gene transfer. In Aim 3, we will explore the therapeutic potential of FTO inhibitors including a repurposed FDA-approved drug in the treatment of PAH employing 3 complimentary animal models of PAH. Based on the clear clinical relevance of our novel findings, we expect that the proposed studies have significant translational potential by delineating the molecular and cellular mechanisms of endothelial dysfunction leading to pulmonary vasoconstriction and vascular remodeling, identifying druggable targets, and exploring pharmacological agents that can pharmacologically inhibit/reverse vascular remodeling and also inhibit vasoconstriction for the prevention and treatment of PAH in patients. Thus, we believe the proposed studies have great translational potential.