黄芪甲苷配伍三七总皂苷对脑缺血大鼠BMSCs移植后神经修复的影响  

Effect of astragaloside IV combined with Panax notoginsengsaponins on nerve repair after BMSCs transplantation in rats with cerebral ischemia

作　　者：李艳玲[1] 丁煌[1] 傅馨莹 唐三 陆展辉 杨芙蓉 刘晓丹[1] 邓常清[1] LIYan-ling;DING Huang;FU Xin-ying;TANG San;LU Zhan-hui;YANG Fu-rong;LIU Xiao-dan;DENG Chang-qing(Hunan Provincial Key Laboratory of Cell Biology and Molecular Technology,Hunan Provincial Key Laboratory of Integrated Traditional Chinese and Western Medicine,Hunan University of Traditional Chinese Medicine,Changsha 410208,China)

机构地区：[1]湖南中医药大学,中西医结合心脑疾病防治湖南省重点实验室,细胞生物学与分子技术湖南省高校重点实验室,湖南长沙410208

出　　处：《中草药》2021年第21期6537-6544,共8页Chinese Traditional and Herbal Drugs

基　　金：国家自然科学基金资助项目(81904181);湖南省自然科学基金资助项目(2018JJ3382);湖南省教育厅优秀青年项目(18B236);湖南省科技厅科技创新平台与人才计划项目(2017SK4005);湖南中医药大学研究生创新课题(2020CX63)。

摘　　要：目的探讨黄芪甲苷IV(astragalosideIV,ASTIV)与三七总皂苷(Panaxnotoginsengsaponins,PNS)配伍联合骨髓间充质干细胞(bone marrowmesenchymal stem cells,BMSCs)移植对脑缺血大鼠神经功能修复的影响。方法大鼠随机分为对照组、模型组、ASTIV(10 mg/kg)与PNS(25 mg/kg)低剂量组、ASTIV(20 mg/kg)与PNS(50 mg/kg)高剂量组、BMSCs输注组、BMSCs输注联合ASTIV(10 mg/kg)与PNS(25 mg/kg)低剂量组、BMSCs输注联合ASTIV(20 mg/kg)与PNS(50mg/kg)高剂量组。全骨髓贴壁法分离、纯化BMSCs,流式细胞术检测BMSCs表面标志物CD29、CD90、CD34、CD45阳性表达率。各给药组给予药物进行干预,采用大脑中动脉阻塞(middle cerebral artery occlusion,MCAO)建立局灶性脑缺血模型,采用Longa法测定各组大鼠神经功能缺损症状;采用苏木素-伊红(HE)染色与尼氏染色测定各组大鼠脑组织病理变化;采用免疫荧光法检测各组大鼠海马区BMSCs和神经元特异性烯醇化酶(neuronspecific enolase,NSE)阳性表达;采用Western blotting法测定各组大鼠脑组织脑源性神经营养因子(brain-derived neurotrophic factor,BDNF)和胶质细胞源性神经营养因子(glial cell line derived neurotrophic factor,GDNF)蛋白表达。结果成功分离培养BMSCs,表面标志物CD29、CD90、CD34、CD45鉴定符合BMSCs特征。大鼠脑缺血后出现神经功能缺损症状及脑组织病理损伤,模型组大鼠神经功能缺损评分和脑组织细胞损伤率显著升高(P<0.01),尼氏体数量显著减少(P<0.01);各给药组均能够不同程度减轻上述病理改变,其中效应最强为BMSCs输注联合ASTⅣ+PNS高剂量组(P<0.01),优于单用药物和单用BMSCs输注。大鼠脑缺血后神经元损伤,输注BMSCs后,细胞可在缺血侧脑组织增殖并分化为神经元,药物联合BMSCs输注能够增强BMSCs在大鼠脑内的增殖和分化。大鼠脑缺血后,BDNF和GDNF蛋白表达增加,各药物组均能够不同程度上调其表达,其中效应最强为BMSCs输注联合ASTⅣ+PObjectiveTo investigate the effect of astragaloside IV(AST IV)and Panax notoginsengsaponins(PNS)combined with bone marrow mesenchymal stem cells(BMSCs)transplantation on neural function repair in rats with cerebral ischemia.Methods Rats were randomly divided into controlgroup,model group,AST IV(10 mg/kg)+PNS(25 mg/kg)lowdose group,AST IV(20 mg/kg)+PNS(50 mg/kg)highdose group,BMSCs infusion group,BMSCs infusion combined with AST IV(10 mg/kg)and PNS(25 mg/kg)lowdose group,BMSCs infusion combined with AST IV(20 mg/kg)and PNS(50 mg/kg)highdose group.BMSCs were isolated and purified by whole bone marrow adherent method.The positive expression rates of CD29,CD90,CD34 and CD45 were detected by flow cytometry.Each drug administration group was given drugs for intervention,model of focal cerebralischemia was established by middle cerebral artery occlusion(MCAO).Longa method was used to determine the symptoms of neurological deficits in each group of rats;Hematoxylin-eosin(HE)staining and Nissl staining were used to determine the pathologicalchanges in brain tissue of each group of rats;Immunofluorescence was used to detect positive expressionsof neuron specific enolase(NSE)and BMSCs in hippocampus of rats in each group;Western blotting was used to determine brain-derived neurotrophic factor(BDNF)and glial cell-derived neurotrophic factor(GDNF)protein expressionsin brain tissue of rats in each group.Results BMSCs were isolated and cultured successfully.The surface markers CD29,CD90,CD34 and CD45 were identified to be in accordance with the characteristics of BMSCs.Neurologic defects and pathological damage of brain tissue occurred after cerebral ischemia.Neurological deficit score and brain tissue cell damage rate in model group were significantly increased(P<0.01),and number of Nissl bodies was significantly reduced(P<0.01).Each administration group alleviatedthe above-mentioned pathological changes to varying degrees.Among them,the strongest effect wasBMSCs infusion combined with ASTⅣ+PNS highdose group(P<0.01),which w

关 键 词：黄芪甲苷 三七总皂苷 骨髓间充质干细胞 脑缺血 增殖 迁移 神经分化 

分 类 号：R285.5[医药卫生—中药学]