内皮祖细胞ALK1基因在脑动静脉畸形修复中的作用及其机制研究

Cerebral arteriovenous malformation (bAVM) is an important cause of intracranial spontaneous hemorrhage, which has high mortality and disability rate. The overall efficacy of clinical intervention for bAVM remains unsatisfactory due to late diagnosis. Alk1 is one of the TGF - receptor superfamily, and its mutation is one of the genetic susceptibility factors of bAVM. Both clinical specimens and animal models indicated that Alk1 gene is closely correlated with the appearance of bAVM, however, its exact mechanism remains unclear. The latest studies showed that Alk1 is involved in the migration and repair of vascular endothelial cells in the arteries, which is closely related to the biological function of endothelial progenitor cells. Furthermore, endothelial progenitor cells are crucial for vascular development. Our preliminary results also demonstrated that the specific knockout of Alk1 gene in endothelial progenitor cells could lead to the occurrence of bAVM. In addition, endothelial progenitor cells replenishment to mice that underwent radiotherapy was found to restore the function of peripheral blood cell. Therefore, we propose that “Alk1 deletion in endothelial progenitor cells is sufficient to induce bAVMs in the adult mice, normal endothelial progenitor cells replenishment could reinforce the weak vasculature of bAVM and mitigate bAVM". This study is designed to investigate the therapeutic effect of bone marrow derived normal endothelial progenitor cells on bAVM by changing bAVM blood vessel structure using gene knockout mice as the bAVM model. Our study will provide new insights for the clinical treatment of bAVM.

脑动静脉畸形（bAVM）是颅内自发性出血的主要原因之一，有较高的致死致残率。临床干预多在严重症状出现后，总体疗效仍不满意。ALK1属TGFβ受体超家族，其突变是bAVM的遗传易感因素之一。临床标本和动物模型均提示Alk1基因与bAVM的形成密切相关，其机制仍不清楚。研究表明Alk1参与血管内皮在动脉的迁移和修复，且与内皮祖细胞的生物学功能密切相关，而后者对血管发育至关重要。我们前期数据提示特异性敲除内皮祖细胞Alk1基因足以形成bAVM，且补充内皮祖细胞可恢复放疗后小鼠的外周血细胞功能。据此我们提出“Alk1缺失的内皮细胞可以导致bAVM的发生，补充正常内皮祖细胞可以修复bAVM中异常的血管结构，从而达到治疗bAVM的作用”这一假说。本项目将在现有转基因小鼠和bAVM的模型上，采用骨髓移植的方式阐明正常内皮祖细胞可通过改变bAVM的血管结构达到修复bAVM的作用，从而为临床治疗提供新思路。

目的：探讨 Alk1基因缺失的内皮细胞（ECs）在脑动静脉畸形（bAVM）形成中的作用及移植正常内皮祖细胞（EPCs）是否可以修复bAVM中异常的血管结构，从而为bAVM的防治提供新思路。方法：在小鼠实验性 bAVM 模型中，通过转基因动物实验、干细胞移植和分子生物学实验，观察内皮细胞Alk1基因在bAVM形成发展中的作用。结果：在bAVM形成过程中，需要外源性VEGF局部的高压刺激和Alk1基因缺失的ECs。在已形成bAVMs后，移植正常的骨髓内皮祖细胞（BM-EPCS）可缓解bAVM的严重程度。在已形成的bAVMs脑区，Alk1基因缺失的ECs基本来源于供体pdgfb-iCreER/Alk12f/2f小鼠的EPCs。bAVMs脑区Alk1阳性细胞及蛋白表达量都显著减少，且在bAVM发育早期，主要是Alk1− ECs的局部增殖。结论：Alk1基因缺失的ECs在局部VEGF的刺激下可以导致bAVM的发生；补充正常EPCS，主要是Alk1− EPCs可以修复bAVM中异常的血管结构，从而达到治疗bAVM的作用。

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