神经细胞来源的DAPK1是脑缺血造成神经元死亡的关键分子，新近报道DAPK1还表达于脑血管内皮细胞，其在脑缺血损伤中的作用尚不清楚。申请人的预实验显示，敲减DAPK1可降低缺血再灌注（I/R）后血脑屏障（BBB）的高通透性，减轻脑组织损伤，其机制可能涉及对脑血管内皮细胞cPLA2磷酸化的调节。基于此，我们提出科学假说：脑I/R后，脑血管内皮细胞的DAPK1通过激活cPLA2破坏脑血管内皮完整性，导致BBB损伤。本项目拟采用内皮细胞特异性敲除、过表达和敲减DAPK1的方法和体外与动物脑I/R模型，研究：（1）脑I/R后DAPK1对于脑血管内皮完整性和连接结构的作用如何；（2）脑I/R后DAPK1是否是通过cPLA2调控脑血管内皮完整性和连接结构；（3）干预内皮DAPK1对脑I/R的保护作用如何。本项目预期将揭示脑血管内皮细胞DAPK1对于BBB完整性的调控，为脑卒中治疗提供新的潜在靶点。

The DAPK1 from neuronal cells is critical for neuronal death in cerebral ischemia. Recently DAPK1 has been reported to express not only in neuronal cells but also in cerebral endothelial cells; however, its role in cerebral ischemic injury is still unclear. Our pre-experiments show that DAPK1 knockdown decreased blood-brain barrier (BBB) hyperpermeability and brain injury after ischemia-reperfusion (I/R), the mechanisms of which could involve the regulation of cPLA2 phosphorylation in cerebral endothelial cells. Therefore, we hypothesize that after cerebral I/R the endothelial DAPK1 impairs cerebrovascular integrity via cPLA2 activation, resulting in BBB breakdown. By using the endothelial specific knockout, overexpression and knockdown of DAPK1 and the in vitro and in vivo models of cerebral I/R, this project will explore: (1) how DAPK1 regulates endothelial integrity and junction structure after cerebral I/R; (2) whether DAPK1 regulates endothelial integrity and junction structure by activating cPLA2 after cerebral I/R; and (3) whether endothelial DAPK1 intervention protects cerebral I/R injury. We expect to reveal the regulation of cerebrovascular endothelial DAPK1 on BBB integrity and provide a new therapeutic target for stroke treatment.