血管外膜氧应激参与PAH血管重塑的表观遗传学机制

Pulmonary arterial hypertension (PAH) is best characterized by progressive inward hypertrophic remodeling and prominent restructuring and activation of the adventitia, especially fibroblast. The mechanism underlying this phenomenon and its significance to the PAH development remains elusive. The therapeutic strategy to lower pulmonary vascular resistance by reducing vascular tone is only temporarily effective. In the preliminary data, we found that both Nox4 mRNA and protein expression are robustly upregulated in the adventitia of pulmonary arteries (PA) in 3 different experimental models of PAH (MCT, Sugen/hypoxia and FHR) in rats. By the administration of different novel and selective Nox4 inhibitors in vivo, we also found selective inhibition of Nox4 prevents and reverses PA remodeling and right ventricle hypotrophy, as well as improve the hemodynamic profile of PA in PAH rat model. HDAC3, 4 and 5 are significantly elevated in PA of MCT treated rats, and both HDAC inhibitor and siRNA potently suppressed the ability of TGF-β1 to increase Nox4 in human fibroblast. Collectively, these data support a significant and novel role for Nox4 and HDACs in the pathogenesis of PAH. This application will address the deficits in our knowledge by: 1) indentifying the functional significance of Nox4 in PAH, 2) understand the role of Nox4 in PA adventitia cell behavior and vascular remodeling in PAH, 3) define the epigenetic mechanisms of PAH, 4) explore the reciprocal relationship of HDACs and Nox4 and their functional significance in PAH. The proposed studies will move the field forward by identifying the vascular significance of oxidative stress and epigenetics and its translational potential in PAH.

肺血管重塑和血管周围组织激活是肺动脉高压（PAH）的主要特点，其发病机制不清。我们的初步研究表明，Nox4主要分布在血管外膜Fibroblast中，其在PAH中的基因表达水平明显增高，Nox4选择性抑制剂明显阻止了肺血管重塑、改善了肺及右心功能。 同时，在PAH肺动脉中，HDAC3-5的蛋白表达显著上调，HDAC 抑制剂和siRNA在肺Fibroblast中显著了降低Nox4在基础和TGF-β1刺激下的基因表达。本课题将探讨：1）Nox4在PAH血管外膜中的作用；2）血管外膜Nox4参与血管重塑调节的分子机制及对管周细胞生物学行为的影响；3）阐明参与PAH的表观遗传学机制；4）明确Nox4与HDACs交互调节的分子机制及其在PAH中的意义。本项目的完成将明确氧应激和表观遗传在PAH中的作用及分子调控机制，为进一步寻找具有鉴别诊断价值的分子标记及临床治疗的靶标分子提供重要的理论依据。

肺血管重塑和血管周围组织激活是肺动脉高压（PAH）的主要特点，其发病机制不清。本项目在细胞分子水平、整体动物和临床样本中开展研究工作，深入解析了NADPH氧化酶及信号通路分子在PAH发生发展中的作用，以及组蛋白乙酰化介导的表观遗传学修饰参与血管外膜氧化应激之间的调控作用，及其在PAH血管重塑发生发展中的分子机制。主要完成了：1）阐明了Nox4在PAH中肺动脉中的细胞分布、表达及重要的功能意义，为靶点研究及转化医学研究提供有价值的资料；2）发现了Nox4主要分布在血管外膜Fibroblasts中，其在PAH中的基因表达水平明显增高，Nox4选择性抑制剂明显阻止了肺血管重塑、改善了肺及右心功能；3）在PAH肺动脉中HDAC3-5的蛋白表达显著上调，而HDAC 抑制剂和siRNA在肺Fibroblasts中显著了降低Nox4在基础和TGF-β1刺激下基因表达；4）血管外膜Nox4参与血管重塑调节的分子机制及对管周细胞生物学行为的影响；5）阐明参与Nox4转录调控的表观遗传学机制及其在PAH中的作用；6）在PAH临床样本中，利用备用候选基因关联研究的遗传学策略了解Nox4，KIRs等氧化应激相关基因内部及上下游调控区多态性与PAH的关联，发现了Nox4多态性rs11821838 tag SNP与PAH显著相关，在华东汉族人群中证实KIR3DL1在PAH发病机制中起保护作用，KIR2DS4发挥与KIR3DL1相似的生物学功能。本研究为探索氧化应激和表观遗传在PAH中的作用及机制奠定了坚实的工作基础，并为临床上防治肺动脉高压提供了新的分子靶点。