DKK3介导血管壁干细胞迁移对腹主动脉瘤形成的影响

Abdominal Aortic Aneurysm (AAA) is a permanent abnormal bulge as a result of a localized weak area in the abdominal aortic wall. Injuries of major cellular components such as smooth muscle cells and endothelial cells in the aortic wall play a vital role in the pathogenesis of AAA. Our previous studies have demonstrated that Sca-1+ vascular progenitor cells (VPCs) have the capability to differentiate into both smooth muscle cells and endothelial cells and participate into repair of injured vessels. Dickkopf-related protein 3 (DKK3) promotes reendothelialization and stabilizes atherosclerotic plaque by either inducing endothelial cells migration or increasing VPC-derived SMC numbers. Together with our preliminary data showed that DKK3 provided protection against AAA and promotion of VPC migration via MEK/ERK and PI3K/Akt signaling pathways. Thus, our hypothesis is that DKK3 inhibits the formation of AAA via inducing VPC migration which benefits the repair of injured vascular cells on the vessel wall. This project aims to assess the effects of DKK3 on AAA formation in a DKK3 knockout mice model and confirm the therapeutic potential of DKK3 by injections of DKK3 adenovirus in vivo. Then, on a model of VPCs transplantation in the adventitia of abdominal aortic wall, we plan to study the role of DKK3-induced VPC migration in the process of AAA development. Furthermore, we will uncover the molecular mechanisms of DKK3-mediated VPC migration in vitro. The results of the present project would provide new theoretical bases and therapeutic agents in prevention and treatment of AAA, suggesting a potential value of DKK3 in clinical application in future.

腹主动脉瘤(AAA)是由腹主动脉血管壁局部结构薄弱引起的永久性异常扩张，平滑肌细胞和内皮细胞受损是其主要发病机制之一。我们前期已证实，血管壁干细胞(VPC)具有向平滑肌细胞和内皮细胞分化并参与修复损伤血管的功能。Dickkopf相关蛋白3(DKK3)可促进内皮细胞迁移和VPC向平滑肌细胞分化从而修复损伤血管内皮及稳定动脉粥样硬化斑块。结合本项目预实验结果，DKK3抑制AAA形成并经MEK/ERK与PI3K/Akt通路促进VPC迁移。我们提出假设：DKK3通过促进VPC迁移修复损伤的血管壁细胞抑制AAA形成。本项目拟在DKK3敲除小鼠模型上，探讨DKK3对AAA形成的影响及DKK3腺病毒潜在的治疗作用；并采用血管外膜移植VPC模型，阐明DKK3介导VPC迁移对AAA形成的影响及机制；进一步揭示DKK3在VPC迁移中的作用及分子机制。本项目将为诊治腹主动脉瘤提供新的理论基础和潜在临床治疗药物。

腹主动脉瘤（AAA）是腹主动脉的永久性病理性扩张，患者通常无症状，一旦形成主动脉夹层或动脉瘤瘤体破裂，死亡率极高，目前临床仍缺乏有效的早期诊断及逆转发病进程的药物。AAA患者主动脉血管平滑肌细胞（VSMCs）大量凋亡坏死、细胞外基质降解等是AAA的主要病理特征，但其具体分子机制尚不明确。近年来，有研究报道Dickkopf相关蛋白3（DKK3）可通过介导细胞的迁移与分化参与多种心血管疾病的发生发展。而DKK3对AAA发生发展的影响及潜在机制并不清楚。本课题从临床现象出发，通过在DKK3敲除的Apoe-/-小鼠上构建血管紧张素II（Ang II) 诱导的AAA模型，探究DKK3对腹主动脉发生发展的影响及潜在分子机制。本课题结果显示：与正常对照组相比，AAA患者血清中的DKK3水平显著升高，且与AAA的发生率呈正相关。免疫荧光染色及GEO数据挖掘分析发现患者主动脉瘤组织DKK3表达水平也比正常组织高。在小鼠腹主动脉瘤模型中，DKK3敲除能显著降低AAA的发生率和瘤体破裂率，提高小鼠生存率。形态学结果显示，DKK3敲除可降低主动脉弹力板断裂程度。机制上，DKK3敲除显著降低主动脉血管壁趋化因子CCL12、CXCL16和CX3CL1的基因及蛋白表达水平。不同基因型小鼠骨髓移植结果显示，非骨髓来源的DKK3对AAA的发生发展起到更加重要的促进作用。免疫荧光染色发现DKK3主要表达于主动脉瘤组织血管中、外膜，且可与主动脉VSMCs共定位。DKK3敲除能明显抑制VSMCs由收缩型向合成型表型的转换，并通过降低基质金属蛋白酶的分泌，减少细胞外基质降解，非经典WNT信号通路中JNK，ERK磷酸化水平均有降低。此外，DKK3敲除对于Ang II诱导的原代骨髓和腹腔巨噬细胞趋化因子的表达水平改变，以及Sca-1+血管干细胞的迁移能力变化均无显著影响。综上所述，人血清DKK3与AAA 发生率呈正相关，DKK3可能通过激活非经典WNT信号通路促进VSMCs由收缩表型向合成表型转化，并促进VSMCs分泌基质金属蛋白酶，促进主动脉弹力板断裂，从而促进 AAA 的发生发展。DKK3可能成为AAA疾病临床早期诊断的标志物及潜在的治疗靶点。

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