EphrinB2及其介导的血管重塑在静脉高压诱导硬脑膜动静脉瘘形成中的作用及激活机制

Complex dural arteriovenous fistula (DAVF) is associate with high morbidity and mortality rates, and difficult to cure. Venous hypertension is regarded as a major cause of it's formation because of the tissue hypoxia and elevated shear stress in the venous system, while the mechanism is unknown. High level expression of the hypoxia-inducible factor (HIF)-1α and the downstream angiogenesis-related factors in rats with intracranial venous hypertension has been observed in our previous study, suggesting that HIF-1α and subsequent angiogenesis take the important part in the formation of DAVF, While the inducing rate of DAVF has not been increased by promoting angiogenesis. Overexpression of EphrinB2 has been reported in the specimen of DAVF, which has been thought to play a key role in the vessel remodeling. We hypothesize that EphrinB2 and vessle remodeling midiated by it have played key roles in the formation of DAVF, and that the elevated shear stress on the wall of venous sinus and accessory veins account for the upregulation and activation of EphrinB2. The study will be carried out in vivo. Ephrin B2 will be detected in the brain and dura of venous hypertension rats and the micro vessel density and the induced ratio of DAVF will be detached after lentivirus vectors are administrated to alter the expression level of EphrinB2 in rats. In vitro study will also be performed. Human umbilical vein endothelial cells will be cultured in deferent shear stress level and HIF expression level to reveal the relationship between EprhinB2 and shear stress. The research project   will clarify the role and mechanism of the upregulation of EphrinB2 and vessle remodeling in the formation of DAVF induced by venous hypertension and will provide new ideas for the treatment of DAVF and targets for drug therapy.

复杂硬脑膜动静脉瘘（DAVF）致残致死率高，难以治愈。前期研究显示静脉高压可通过组织缺氧和静脉应力增高诱发DAVF，缺氧诱导因子（HIF）及其诱导的血管新生是其重要环节，但单纯诱导血管新生并不能提高DAVF的诱导率。EphrinB2介导血管重塑，在DAVF标本中亦高表达。我们假设，EphrinB2及其介导的血管重塑在DAVF形成过程中发挥了重要作用，静脉内剪应力升高可能通过HIF激活EphrinB2。本研究拟建立静脉高压大鼠模型，并利用慢病毒载体干预EphrinB2的表达水平，检测其在脑和硬膜中EphrinB2的表达水平，以及微血管和DAVF诱发情况，以证实EphrinB2的重要作用；另外研究不同剪应力和HIF水平下人脐静脉内皮细胞EphrinB2的表达水平，揭示EprinB2的激活机制。通过研究，可望阐明EphrinB2在静脉高压诱导DAVF形成中的作用和激活机制，为DAVF的药物治疗提供依据。

硬脑膜动静脉瘘（dural arteriovenous fistula，DAVF）致死致残率高，尤其是复杂型DAVF，更是难以治愈。静脉高压为DAVF公认的初始诱发因素，可通过组织缺氧和改变血管应力环境诱发DAVF，缺氧诱导因子(hypoxia-inducible factor, HIF）启动血管内皮生长因子（vascular endothelial growth factor，VEGF）介导的血管新生是其重要环节，但是该通道并不是诱导DAVF发生的唯一通路。有研究发现，人的DAVF标本中VEGF和EphrinB2表达均增加，而二者同时参与新血管的形成过程。因此本研究通过建立不同静脉端压力的大鼠模型，并利用慢病毒载体干预EphrinB2表达水平，检测颅内各组织中EphrinB2的表达水平及对新血管形成的影响，并通过研究不同剪应力环境下培养的脐静脉内皮细胞中，HIF-1α和EphrinB2的关系，初步探讨启动EphrinB2的上游分子通路。结果发现静脉高压窦汇及硬膜组织中EphrinB2、VEGFR2表达明显增加，窦汇表达部位主要在血管内皮层。静脉高压并静脉窦内EphrinB2 shRNA慢病毒载体感染后，窦汇及硬膜组织的EphrinB2的表达明显受到抑制，VEGFR2表达显著减少，VIII因子微血管增生受到抑制，脑组织内无显著变化。静脉窦闭塞并静脉窦内重组EphrinB2慢病毒感染组中，窦汇及硬膜组织EphrinB2的表达显著增加，VEGFR2表达显著增加，VIII因子微血管增生活跃，脑组织内无显著变化。免疫共沉淀反应中，EphrinB2可以在胞内与VEGFR2发生相互作用。高剪应力环境下脐静脉内皮细胞EphrinB2/EphB4表达均显著增加，高剪应力环境下沉默HIF-1α后， EphrinB2/EphB4表达无明显差异。结果提示1.EphrinB2能与VEGFR2发生特异性相互作用，静脉高压环境下，硬膜组织中的EphrinB2表达增加，并上调VEGFR2介导的血管新生，使血管新生活跃。2.EphrinB2/EphB4通路激活与静脉高压造成的静脉内高剪应力环境有关，静脉高压激活EphrinB2/EphB4通路与高剪应力介导内皮细胞HIF-1α表达无关。