Slit/Robo通路在异常增大的血流动力学应力诱导脑动脉瘤形成中的作用机制研究

Cerebral aneurysms (CA) are a great threat to human health because of its high mortality and morbidity，however, the molecular mechanism of CA formation has not clear. Endothelial cell injure induced by hemodynamic abnornmalities was the earlist change in aneurysm wall. Slit2/Robo4 is specifically expressed in vessel endothelial cells (ECs). In the present study, we firstly revealed that Slit2/Robo4 was highly expressed in aneurysms, and then functional studies showed that Slit/Robo pathway mediates ECs apoptosis. However, the effect of Slit/Robo pathway on aneurysm initiation and formation remain to be established. Our previous studies showed that most CAs were deviated to smaller lateral angles, and the Slit2/Robo4 mRNA expression levels and hemodynamic stresses in the daughter branch forming a smaller angle with the parent vessel were significantly greater than the contralateral branch. We thus hypothesized that the high hemodynamic stresses site in the branch forming a smaller lateral angle with the parent artery is the aneurysm initiation site. Furthermore, a stent that was deployed via the parent vessel into the branch vessel with aneurysm initiation site could decrease the hemodynamic stresses, ECs apoptosis induced by the Slit/Robo pathway, and aneurysm recurrence rate after stent treatment. To test this hypothesis, a parallel-plate flow chamber model in vitro was used to investigate the effect of Slit/Robo pathway on high wall shear stress induced ECs apoptosis. A bilateral common carotid artery ligation model was used to examine the effect of Slit/Robo pathway on vessel wall remodeling induced by high hemodynamic stresses. Three-dimensional angiography of pre/post stent treatment and latest follow-up was used to analyze the impact of stent on hemodynamic stresses, Slit/Robo signal pathway and recurernce rate alteration at the aneurysm initiation site. Moreover, This study will reveal the molecular mechanism of CA formation and optimize the method of stent implantation to decrease aneurysm recurrence rate, which will thus provide us with theoretical basis for the therapy of cerebral aneurysm and therapy target selection.

脑动脉瘤（CA）破裂导致高致死/残率，CA形成机制尚未阐明，异常血流动力学应力（Hs）诱导内皮细胞（ECs）功能损伤是CA形成的第一步。Slit2/Robo4特异表达于ECs，本课题组首次发现其在CA中高表达且调控ECs凋亡。但是，Slit/Robo通路在CA形成中的作用尚无报道。在前期研究中，CA多朝向小外侧角，且该侧Hs和Slit/Robo表达显著大于对侧分支血管，我们推测，该侧高Hs位点是动脉瘤的起始位点，支架植入该位点可降低Hs和Slit/Robo介导的ECs凋亡，降低术后复发率。为证明该假说，平行平板流动腔系统和双侧颈总动脉结扎成瘤动物模型研究Slit/Robo通路在高Hs诱导ECs凋亡及管壁重构中的作用；影像学资料研究支架植入动脉瘤起始位点对降低Hs、抑制Slit/Robo通路和术后复发率的作用。本课题对阐述CA发生的分子机制和降低复发率提供新思路，并为CA的防治提供新靶点。

血流动力学应力（Hemodynamic stress）诱导的内皮功能障碍和炎性因子表达上调为颅内动脉瘤（Cerebral aneurysm）形成的起始关键步骤，而血流动力学应力的大小取决于动脉分叉的形态学结构。本研究根据颅内高发病率动脉瘤及其动脉分叉的形态学特点，构建血管模型分析血流动力学应力特点，并构建高曲度动物模型研究动脉瘤形成的相关分子机制。回顾性收集大脑前动脉（ACA，N=58）、大脑中动脉（MCA，N=64）、基底动脉（BA，N=19）和颈内动脉-大脑后交通动脉（ICA-PComA，N=57）分叉动脉瘤患者，研究发现190例（96.0%）动脉瘤朝向小外侧角，多因素分析结果显示该侧的高曲度为MCA动脉瘤形成的独立危险因素，ROC曲线分析证明该曲度用于预测动脉瘤存在的曲线下面积为0.932，但是血管高曲度诱发的血流动力学改变和炎性反应尚不清晰。利用不同曲度的血管模型研究在一个心动周期内的血流动力学应力变化及其与血管曲度间的相关性，证明总压力、剪切力、动态压力和涡流强度均与血管曲度显著正相关。利用具有生物相容性的模具垫高腹主动脉，增大血管曲度，术后15个月检测到炎性因子、MMPs及Slit/Robo家族成员的表达显著增加。进一步利用乙醇（EtOH）诱导体外培养人脐静脉血管内皮细胞（Human Umbilical Vein Endothelial Cells，HUVECs）的炎性反应，同时伴随着Slit/Robo通路的激活、氧化应激反应和细胞迁移率的下降，而苦参根部提取的抗炎药物氧化苦参碱（Oxymatrine，OMT）缓解了HUVECs的炎性反应和Slit/Robo通路的激活，同时协同抑制细胞迁移。本研究揭示了血管高曲度诱导的血流动力学改变及炎性反应可能与管壁重构及动脉瘤的形成有关，而Slit/Robo通路参与了这一过程。

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