肌动蛋白解聚因子在脑动脉瘤发病过程中的作用和机制

Cerebral aneurysm (CA) causes catastrophic subarachnoid hemorrhage, which is characterized by a high mortality and morbidity rate. Despite its clinical and social importance, the detailed molecular mechanism of CA formation remains to be elusive, resulting in the absence of effective medical treatment against CA. Our preliminary experiment demonstrated that actin-depolymerizing factor (ADF) was downregualted in CA. In addition, modeled high wall shear stress and high wall shear stress gradients suppressed ADF expression in human carotid artery endothelial cells (HCAECs); simulation with IL-1β suppressed ADF expression in human carotid artery smooth muscle cells (HCASMCs). The aim of this study is to elucidate the role of ADF in the development of CA. In vitro, ADF will be overexpressed by transfecting with recombinant adenovirus containing ADF-overexpressing plasmid or suppressed by transfecting with ADF shRNA lentiviral particles in both HCAECs and HCASMCs. Then, we will evaluate the role of ADF in modeled high wall shear stress and high wall shear stress gradients-induced injury in HCAECs and IL-1β-induced injury in HCASMCs. In vivo, after ligation of the left carotid artery and posterior branches of the bilateral renal arteries in ADF knockout mice, they will be fed with a high-salt diet containing 8% sodium chloride and 0.12% b-aminopropionitrile. Five months after aneurysm induction, the incidence and size of CA will be evaluated to elucidate the role of ADF in the formation of CA. To elucidate the underlying molecular mechanism of ADF in CA formation, inflammation, oxidative stress, nitric oxide formation, degradation of extracellular matrix in CA will be evaluated. This study will elucidate the potential role of ADF in CA and will provide a novel therapeutic target for CA.

目前，临床上尚无有效的预防和治疗脑动脉瘤的特异性药物。为了开发该类药物，对脑动脉瘤发生的分子机制的研究就显得尤为重要和迫切。课题预实验证实脑动脉瘤组织中肌动蛋白解聚因子（actin-depolymerizing factor， ADF）蛋白表达降低。本课题首先通过转染含有ADF高表达质粒或RNAi质粒的重组病毒，观察ADF表达调控对流体剪切力刺激损伤颈动脉内皮细胞和IL-1β损伤颈动脉平滑肌细胞这两个过程的影响。使用ADF基因敲除小鼠，采用左颈总动脉和双侧肾动脉后支结扎手术，并以喂食高盐饮食的方法来诱导形成脑动脉瘤，观察ADF基因敲除对脑动脉瘤形成的影响。通过观察脑动脉瘤组织中炎症反应、氧化应激、一氧化氮合成、细胞外基质降解，探讨ADF蛋白影响脑动脉瘤形成的具体机制。本课题将对脑动脉瘤的发病机制提出新的理论，并为脑动脉瘤的药物防治提供一个全新的蛋白靶点。

目前，临床上尚无有效的预防和治疗脑动脉瘤的特异性药物。为了开发该类药物，对脑动脉瘤发生的分子机制的研究就显得尤为重要和迫切。本课题以肌动蛋白解聚因子（actin-depolymerizing factor, ADF）为核心，研究其在脑动脉瘤发生发展中的作用。首先在细胞水平通过转染含有ADF高表达质粒或RNAi质粒的重组病毒，发现高表达ADF可以减轻流体剪切力刺激诱导血管内皮细胞凋亡，增加内皮细胞的屏障功能，减少炎症因子的生成，增加一氧化氮的合成；相反低表达ADF可以恶化流体剪切力刺激诱导的血管内皮细胞损伤。高表达ADF可以减轻氧化性低密度脂蛋白诱导的内皮细胞凋亡，减少炎症因子的合成。高表达ADF可以减轻高级糖化终末产物诱导的内皮细胞凋亡，增加内皮细胞的屏障功能，减少炎症因子的生成，增加一氧化氮的合成。高表达ADF可以通过抑制金属蛋白酶活性，减轻IL-1β诱导的颈动脉平滑肌细胞胶原合成下调。在整体水平，通过小鼠颅内注射弹性蛋白酶合并给予血管经张素II诱导颅内动脉瘤，发现ADF基因敲除可以减轻脑动脉瘤的发生和破裂。最终证明ADF对脑动脉瘤形成和发展具有重要作用。本课题为脑动脉瘤的发病机制提出新的假说，并为脑动脉瘤的药物防治提供一个全新的蛋白靶点。