芪参益气滴丸抑制颈动脉血栓形成后tPA溶栓引起的脑血管性水肿和出血的作用及机理

Ischemic stroke is a common disease of brain tissue injury caused by insufficient cerebral blood supply. It has brought a heavy burden to families and society for its high disability rate and high fatality rate. Tissue plasminogen activator (tPA) is the only drug that included in the guidelines for the treatment of ischemic stroke. However, tPA is associated with risk of cerebrovascular edema and hemorrhage. There is no effective drug to this complication. Damage of the blood-brain barrier is the key pathological basis that leading to cerebrovascular edema and hemorrhage caused by tPA. Ischemia reperfusion injury is the main cause of blood-brain barrier damage. Previous studies have shown that QiShen YiQi Pills (QSYQ) can improve myocardial ischemia reperfusion injury and maintain vascular permeability by tonifying Qi. The main component of QSYQ, T541, has been proved to inhibit cerebrovascular edema and hemorrhage caused by tPA thrombolysis. Since T541 is still in the early stage of new drug development, we selected QSYQ, a listed drug containing its active ingredient, for research. We speculate that QSYQ can also inhibit cerebral edema and hemorrhage due to tPA thrombolysis by improving ischemia reperfusion injury. This project intends to reveal the scientific principle of QSYQ for the treatment of cerebrovascular edema and hemorrhage caused by tPA thrombolysis. This study will be researched through the detection of cerebral edema and cerebral hemorrhage, the integrity of related structures of blood-brain barrier, energy metabolism and oxidative stress related molecules.

缺血性脑卒中是一种由于脑供血不足而引起脑组织损伤的常见疾病，具有其高致死率和高致残率。组织型纤溶酶原激活剂（tPA）是治疗该病唯一纳入指南的药物，但tPA伴有脑血管性水肿和出血的风险。目前临床上尚无针对该种并发症的药物。血脑屏障受损是tPA引起的脑水肿和出血的关键病理环节。而缺血再灌注损伤又是造成血脑屏障受损的主要原因。既往研究表明芪参益气滴丸（QSYQ）可通过补气固摄改善心肌缺血再灌注损伤。QSYQ的主要组分配伍T541已被证实能够抑制tPA引起的脑水肿和出血。鉴于T541尚处于新药研发早期，我们选择了包含其有效成分的上市药物QSYQ进行研究。我们推测QSYQ也可通过改善缺血再灌注损伤而抑制tPA溶栓引起的脑水肿和出血。本项目拟通过检测脑水肿和脑出血、血脑屏障相关结构的完整性、能量代谢和氧化应激相关分子等揭示QSYQ补气固摄治疗tPA溶栓引起的脑血管性水肿和出血的科学原理。

背景：脑卒中已成为我国居民首位致死原因，其中缺血性卒中占60%～80%。组织型纤溶酶原激活剂（tPA）是目前唯一被批准用于治疗急性缺血性脑卒中的药物。而临床上使用tPA静脉溶栓治疗可能继发脑血管水肿和脑出血等严重并发症。芪参益气滴丸是由黄芪、丹参、三七和降香组成的中成药制剂，具有益气通脉、活血止痛的作用。本团队近期研究发现芪参益气滴丸的主要活性成分T541能改善卒中后tPA所致的脑水肿和出血，但是芪参益气滴丸是否具有类似作用尚不清楚。.目的：观察芪参益气滴丸能否缓解小鼠缺血性卒中后tPA静脉溶栓引起的脑水肿和脑出血，并探讨其可能的作用机制。.方法：采用氯化铁化学刺激法制备缺血性卒中小鼠模型，术后2.5小时芪参益气滴丸灌胃给药，术后4.5小时tPA静脉给药，术后28.5小时检测相关指标并取材。使用激光多普勒血流仪观察脑血流量变化，TTC染色分析脑梗死情况，神经行为学评分评估神经功能缺损，伊文思蓝染色和脑细静脉白蛋白漏出评价血脑屏障的通透性，脑干湿重比分析脑水肿状况，比色法检测脑出血水平，免疫荧光法和蛋白质印迹法检测大脑皮层紧密连接、黏附连接、基底膜、基质金属蛋白酶（MMPs）、质膜微囊和能量代谢相关蛋白的表达情况。观察并记录术后7天小鼠的存活状况。.结果：芪参益气滴丸抑制了tPA所加重的脑梗死、神经行为学缺损、死亡率、血脑屏障损伤、脑水肿和脑出血；芪参益气滴丸抑制了tPA所致的质膜微囊相关蛋白Caveolin-1的高表达和Src蛋白的磷酸化水平升高；芪参益气滴丸逆转了tPA引起的能量代谢相关蛋白ATP 5D的减少；芪参益气滴丸抑制了tPA所致的紧密连接蛋白ZO-1、JAM-1、Occludin、Claudin-5，黏附连接蛋白VE-cadherin、α-catenin、β-catenin，和基底膜主要成分Collagen IV、Laminin 的低表达和排列不连续；芪参益气滴丸逆转了tPA所致MMP-9的高表达，脑细静脉黏附白细胞的增多和浸润白细胞数的增多。.结论：芪参益气滴丸能抑制卒中后tPA所加重的血脑屏障损伤，改善脑水肿和脑出血，有望作为一种提高tPA安全性的辅助药物应用于临床。