【摘要】 目的探讨黄芪皂甙Ⅳ联合骨髓间充质干细胞(BMSCs)移植对缺血再灌注大鼠海马神经细胞凋亡及Bcl-2、Bax、Caspase-3表达的影响。方法实验动物随机分为假手术组、模型组、BMSCs组、BMSCs+黄芪组。采用线栓法建立大鼠大脑中动脉缺血(MCAO)模型,BMSCs组、BMSCs+黄芪组分别于建模成功后3h经尾静脉注射已制备好的PKH26标记BMSCs悬液。另外BMSCs+黄芪组于缺血前5min、缺血后12h,24h腹腔注射黄芪皂甙Ⅳ。缺血再灌注72h后,各组进行神经功能评分,应用荧光显微镜观察海马PKH-26标记的移植BMSCs分布,采用免疫组化法和实时荧光定量PCR方法检测大鼠海马Bcl-2、Bax、caspase-3的表达,运用TUNEL方法检测神经元细胞凋亡指数。结果 PKH-26标记的BMSCs在海马有表达。与模型组比较,BMSCs组与BMSCs+黄芪组均能够使大鼠神经功能损害评分降低(P<0.01),BMSCs+黄芪组神经功能恢复最为明显(P<0.05)。模型组Bcl-2、Caspase-3、Bax蛋白表达及细胞凋亡与假手术组差异显著(P<0.01)。与模型组比较,BMSCs+黄芪组和BMSCs组均可下调Caspase-3、Bax蛋白表达和细胞凋亡指数(P<0.05),上调Bcl-2蛋白表达及Bcl-2/Bax比值(P<0.05),其中以BMSCs+黄芪组作用更为显著(P<0.05)。实时荧光定量PCR显示:与模型组和BMSCs组比较,BMSCs+黄芪组能显著下调Caspase-3、Baxm RNA相对表达(P<0.05),上调Bcl-2mRNA相对表达(P<0.05)。结论黄芪皂甙Ⅳ联合BMSCs移植对脑缺血再灌注神经元细胞凋亡有协同抑制作用,其作用机制可能与黄芪皂甙Ⅳ促进BMSCs抑制Bax、caspase-3表达,增强Bcl-2表达和Bcl-2/Bax比值有关。更多还原

【Abstract】 Objective To investigate the effects of Astragaloside IV combined with bone mesenchymal stem cells(BMSCs)on the apoptosis of neural cells and expressions of Bcl-2,Bax and caspase-3 in hippocampus of cerebral ischemia-reperfusion rats.Methods 42 healthy SD rats were randomly divided into sham operation,model,BMSCs and BMSCs+ Astragaloside Ⅳgroups.The rat model were established by middle cerebral artery occlusion(MCAO),PBS and PKH-26 labeled BMSCs suspension were injected via caudal vein 3 hours after MCAO.Astragaloside IV was administered 5min before cerebral ischemia,and 12h,24h after cerebral ischemia in BMSCs+ Astragaloside Ⅳgroup.72h after reperfusion,neurological function score were observed,the expressions of Bcl-2,Bax and caspase-3 were determined by immunohistochemistry and real-time fluorescent quantitative PCR.The apoptosis of neural cells was detected by TUNEL method.Results PKH-26 labeled BMSCs were observed in hippocampus,the neurological scores in BMSCs+ Astragaloside Ⅳgroup significantly decreased compared with model and BMSCs group,(P<0.05).The number of apoptotic neural cells and the expression of Bcl-2,Bax and caspase-3 proteins were significantly increased in model than in sham(P<0.01).Compared with model group,the number of apoptotic neural cells and the expression of Bax and caspase-3 proteins were significantly lower in BMSCs group and BMSCs+ Astragaloside Ⅳgroup (P<0.05),but the expression of bcl-2 proteins was significantly increased (P<0.05).Moreover,the effect of BMSCs+ Astragaloside Ⅳ is more powerful than BMSCs(P<0.05).The relative expression of Caspase-3,Bax mRNA was significantly lower in BMSCs+ Astragaloside Ⅳgroup than in model and BMSCs groups (P<0.05),but the relative expression of Bcl-2 mRNA significantly increased by real-time fluorescent quantitative PCR (P<0.05).Conclusion BMSCs+ Astragaloside Ⅳ could play coordinate inhibition on neural cell apoptosis in cerebral ischemia-reperfusion rats,which may be related to Bcl-2 and Caspase-3 and Bax expressions.更多还原