冰片配伍黄芪甲苷和三七总皂苷通过Notch信号通路对大鼠脑缺血再灌注损伤模型的神经保护作用

目的 从Notch信号通路探讨冰片配伍黄芪甲苷（astragaloside IV，AST IV）和三七总皂苷（Panax notoginseng saponins，PNS）对大鼠脑缺血再灌注损伤（cerebral ischemia reperfusion injury，CIRI）模型的神经保护作用。方法 SD大鼠随机分为假手术组、模型组、冰片（7.5 mg/kg）组、PNS（25 mg/kg）组、AST IV（10 mg/kg）组、AST IV（10 mg/kg）+PNS（25 mg/kg）组、冰片（7.5 mg/kg）+AST IV（10 mg/kg）+PNS（25 mg/kg）低剂量组、冰片（15 mg/kg）+AST IV（20 mg/kg）+PNS（50 mg/kg）高剂量组、依达拉奉（4 mg/kg）组。假手术组、模型组ig 0.5% CMC-Na，依达拉奉组ip相应药物，其余各组ig相应药物，2次/d，间隔12 h。末次给药后2 h采用线栓法阻断大鼠右侧大脑中动脉，建立CIRI模型。缺血2 h，再灌注22 h后，进行神经功能缺损评分；HE染色法观察缺血皮质区病理变化；免疫组化法检测神经元特异性核蛋白（neuron specific nuclear，NeuN）、内皮屏障抗原蛋白（endothelial barrier antigen，EBA）表达；Western blotting法检测血管内皮生长因子（vascular endothelial growth factor，VEGF）、Notch1、Notch胞内域（intracellular domain of Notch，NICD）蛋白表达。结果 模型组大鼠神经功能缺损评分和细胞损伤率显著增加（P<0.01）；各给药组大鼠神经功能缺损评分和细胞损伤率显著降低（P<0.05、0.01），冰片+AST IV+PNS组的效应强于各药物单用及AST IV+PNS组（P<0.05、0.01）。模型组大鼠缺血皮质区NeuN、EBA蛋白表达均显著降低（P<0.01）；各给药组NeuN、EBA蛋白表达显著增加（P<0.05、0.01），冰片+AST IV+PNS组的效应强于各药物单用及AST IV+PNS组（P<0.05、0.01）。模型组大鼠缺血皮质区VEGF蛋白表达显著增加（P<0.05），NICD、Notch1蛋白表达无显著变化；冰片+AST IV+PNS组VEGF、NICD、Notch1蛋白表达显著上调（P<0.01），且3种药物配伍的效应强于各药物单用及AST IV+PNS（P<0.05、0.01）。结论 冰片、AST IV、PNS具有抗脑缺血再灌注后神经元和脑微血管损伤的作用，3种药物配伍的作用强于各药物单用及AST IV+PNS，其作用可能与激活Notch信号通路、上调VEGF表达，从而发挥对缺血脑组织的保护作用有关。

Neuroprotective effect of borneol combined with astragaloside IV and Panax notoginseng saponins in cerebral ischemia reperfusion injury rat model through Notch signaling pathway

Objective To observe the neuroprotective effect of borneol combined with astragaloside IV (AST IV) and Panax notoginseng saponins (PNS) on cerebral ischemia reperfusion injury (CIRI) rat model through Notch signaling pathway. Methods SD rats were randomly divided into sham group, model group, borneol (7.5 mg/kg) group, AST IV (25 mg/kg) group, PNS (10 mg/kg) group, AST IV (10 mg/kg) + PNS (25 mg/kg) group, borneol (7.5 mg/kg) + AST IV (25 mg/kg) + PNS (10 mg/kg) low dose group, borneol (15 mg/kg) + AST IV (20 mg/kg) + PNS (50 mg/kg) high dose group and edaravone (4 mg/kg) group. Rats in sham group and model group were ig 0.5% CMC-Na, edaravone group was ip drug, and the other groups were ig corresponding drugs, twice a day with an interval of 12 h. The right middle cerebral artery of rat was blocked by a suture method 2 h after last administration to establish a CIRI model. After 2 h of ischemia and 22 h of reperfusion, the eurological function scores were scored and pathological changes of ischemic cortex in brain tissues of rats were observed by HE staining. The expressions of neuron specific nuclear (NeuN) and endothelial barrier antigen (EBA) in ischemic cortex of brain tissue were detected by immunohistochemistry. The expressions of vascular endothelial growth factor (VEGF), Notch1, and intracellular domain of Notch (NICD) in ischemic cortex of brain tissue were detected by Western blotting. Results The score of neural dysfunction and cell damage rate in model group were significantly increased (P<0.01); The score of nerve function defect and rate of cell damage in each administration group were significantly reduced (P<0.05, 0.01), the effect of borneol + AST IV + PNS group was better than that of single drug and AST IV + PNS group (P<0.05, 0.01). NeuN and EBA protein expressions were significantly decreased in the ischemic cortex of model group (P<0.01), while NeuN and EBA protein expressions were significantly enhanced in each administration group (P<0.05, 0.01), and the effect of borneol + AST IV + PNS group was better than that of single drug and AST IV + PNS group (P<0.05, 0.01). In model group, VEGF protein expression was increased significantly (P<0.05), while NICD and Notch1 protein expression had no significant change. The expression of VEGF, NICD and Notch1 protein were significantly up-regulated in borneol + AST IV + PNS group (P<0.01), and the effect of combination of three drugs was better than that of single drug and AST IV + PNS (P<0.05, 0.01). Conclusion Borneol, AST IV, and PNS have the effects of preventing neuronal and cerebral microvascular damage after CIRI, and the effect of combination of three drugs was better than that of single drug and AST IV + PNS, which may be related to the activation of the Notch signaling pathway and up-regulation of VEGF expression, thereby, exerting protective effects on ischemic brain tissue.