体外培育牛黄配伍冰片对脑缺血再灌注模型大鼠脑血管及神经元的保护作用

Protective Effects of In Vitro Cultured Calculus Bovis Combined with Borneol on Cerebral Vessels and Neurons in Rats with Cerebral Ischemia-reperfusion

目的考察体外培育牛黄、冰片及两药配伍对脑缺血再灌注模型大鼠的脑保护作用,并探讨其机制。方法采用改良线栓法制备短暂大脑中动脉阻塞(MCAO)模型,缺血90min后再灌注,MCAO后3d采用Zea-Longa及mNSS评分法评价大鼠神经功能,TTC染色后计算其脑梗死率;ELISA法检测大鼠血清中VEGF、Ang-1及bFGF水平与缺血侧脑组织中GDNF、NGF及BDNF的水平;免疫荧光双染观察其对神经元及血管新生的影响;透射电镜观察其对神经元及脑微血管超微结构的影响。结果与溶剂模型组相比,体外培育牛黄、冰片及两药配伍可不同程度地降低大鼠神经功能评分、降低脑梗死率;升高血清VEGF、Ang-1水平,增强缺血侧脑组织中GDNF、NGF及BDNF的表达水平;还可增加缺血侧皮层区神经元数量及新生血管密度;改善缺血侧神经元及脑微血管的超微结构。结论体外培育牛黄、冰片及两药配伍通过促进MCAO模型大鼠血管新生和神经元保护,表现出“开窍醒神”功效,并体现“相使”增效的配伍关系,为临床合理用药提供参考。

Objective To observe the brain protective effects of in vitro cultured Calculus Bovis(ICCB)and borneol,alone or in combination,on cerebral ischemia-reperfusion(MCAO)model rats,and to explore its molecular mechanism.Methods The model of transient middle cerebral artery occlusion(MCAO)was established by modified suture.Reperfusion was performed after 90min of ischemic treatment.The neurological function was evaluated 3dafter operation through Zea-Longa and mNSS scale.The cerebral infarction rate was calculated after TTC staining.The levels of VEGF,Ang-1and bFGF in rat serum and GDNF,NGF and BDNF in ischemic brain tissue were detected by ELISA.Immunofluorescence double staining was used to observe effects of drugs on neurons and angiogenesis.The ultrastructural changes of cerebral microvessels were observed by transmission electron microscope.Results Compared with the solvent model group,ICCB,the neurological function score and cerebral infarction rate in Calculus Bovis group,borneol group and combination group were reduced.The levels of VEGF,Ang-1,GDNF,NGF and BDNF were increased.The density of neurons and neovascularization in ischemic cortex was increased.The ultrastructure of cerebral microvessels was improved.Conclusion In vitro cultured Calculus Bovis and borneol,no matter alone or in combination,were proved to have the effect of"orifice opening and mind wakening"by promoting angiogenesis and neuron protection in MCAO model rats.The compatibility relationship of"mutual promotion"and synergistic effect are observed,which provide reference for rational drug use in clinical practice.