特异性 ssDNA 适配子靶向调控 NADPH 氧化酶-1在主动脉夹层中膜退行性病变中的作用机制

Aortic dissection (AD) is a catastrophic vascular disease with an incredibly high mortality, but its exact pathogenesis has not been fully defined. Our experiment found an enhanced expression of NADPH oxidase-1 (NOX1) in tunica media of the dissected aorta. NOX1 could not only inhibit the synthesis of elastic fibre but also promote the apoptosis of vascular smooth muscle cell (SMC). All these evidences have proven that NOX1 is involved in aortic media degeneration (AMD), which is the morphologic evidence of aortic dissection. Therefore, NOX1 may play an important role in AD..Our study innovatively uses Systematic Evolution of Ligands by Exponential Enrichment (SELEX) technique to obtain the specific ssDNA aptamers targeted NOX1. The specific ssDNA aptamers will then be used to interfere the activity of NOX1 in vitro and vivo. Next, the effects of NOX1 and its specific ssDNA aptamers on downstream signaling pathway (ROS/p38 MAPK) and Sp1/Sp3 transcription factors will be investigated. Furthermore, their regulatory mechanism of elastic fiber synthesis will also be studied. This study will hopefully elucidate the mechanism of specific ssDNA aptamer on AMD in process of aortic dissection through targeted-regulating NOX1. Consequently, it may provide not only a new perspective for the pathogenesis of AD but also novel targets for aorta remodeling and better prognosis of AD.

主动脉夹层（AD）是灾难性的血管疾病，死亡率高，但发病机制不明。我们研究发现 NADPH 氧化酶-1（NOX1）在人 AD 血管中膜内表达明显增高，其能抑制弹性纤维的合成并促进平滑肌细胞（SMC）凋亡，可能参与了中膜退行性病变（AMD）。而 AMD 是 AD 发生的形态学基础，推测 NOX1 在 AD 中可能起到了重要作用。本项目以 NOX1 为靶点，首次运用 SELEX 技术筛选出特异性的 ssDNA 适配子。通过 SMC 培养及小鼠 AD 模型的建立，利用ssDNA 适配子靶向干预 NOX1，观察其对下游信号通路（ROS/p38 MAPK）、Sp1/Sp3转录因子、 Nogo-B、Fibulin-5、以及弹性纤维合成的影响。探讨 ssDNA 适配子靶向调控 NOX1 在 AMD 中的作用及机制，以期为 AD 发病机制研究提供新线索，以及为促进主动脉修复，改善 AD 预后提供新靶点。

主动脉夹层（AD）是灾难性的血管疾病，死亡率高，但发病机制不明。主动脉中膜退行性病变（Aortic media degeneration, AMD）是 AD 发生的病理学基础。AMD 特征为：弹性纤维形态和排列不规则、断裂、损耗，血管平滑肌细胞（vascular smooth muscle cells，VSMCs）缺失，以及胶原蛋白在间质的积聚等。我们成功建立了AD动物模型，免疫组织化学检测亦证实了上述改变。通过免疫组化化学法、蛋白印迹等技术对AD及正常主动脉临床标本检测发现：NOX1在 AMD 中高表达，而在AMD中低表达，随后在血管平滑肌细胞实验中，我们通过过表达及抑制NOX1，发现它能反向调控Fibulin 5表达及纤维蛋白形成。同时我们采用SELEX技术，体外筛选成功获得与NOX1功能基团RAC1的ssDNA适配子，Rac1 适配子能与靶目标高亲和力结合，而抑制NOX1活性。最后，我们获得如下结论：1、初步阐明了 NOX1 在 AMD 中的作用及机制；2、上调NOX1可通过抑制Fibμlin 5合成，影响弹力纤维合成，从而诱发AMD；抑制NOX1，Fibμlin 5表达上升，弹力纤维合成增多，降低AMD发生；3、NOX1可作为防治ADM的潜在靶点。