扎里奴思方干预骨髓间充质干细胞移植对脑缺血再灌注损伤大鼠黏附分子及NeuN表达的影响

Effect of Zhali Nusi Fang( ZLNF) Intervening Bone Marrow Mesenchymal Stem Cells Transplantation on Adhesion Molecules and Neu N Expression in Rats After Cerebral Ischemia Reperfusion Injury

目的:观察扎里奴思方(扎方)干预骨髓间充质干细胞(BMSCs)移植对大脑中动脉阻塞(MCAO)模型大鼠黏附分子及神经元核抗原(Neu N)表达的影响,并探讨其作用机制。方法:SD大鼠随机分为假手术组、模型组、扎方组、移植组和联合组;线栓法制备大鼠MCAO模型,体外全骨髓贴壁筛选法培养及扩增BMSCs;大鼠ig给药(14.6 g·kg^(-1)·d^(-1)),BMSCs悬浮液经颈内动脉移植入脑;移植后1,3,7,14 d取材,评价神经功能积分(NS),逆转录-聚合酶链式反应(RT-PCR)技术检测血管细胞间黏附分子^(-1)(VCAM^(-1))和整合素α4mRNA表达,免疫组化法测Neu N表达。结果:MCAO模型大鼠NS及Neu N表达较假手术组显著降低(P<0.01),VCAM^(-1)及整合素α4mRNA表达显著增高(P<0.01);与模型组比较,扎方1,3,14 d移植3,7,14 d及联合各组NS增加(P<0.01,P<0.05),扎方、移植及联合各组Neu N增加(P<0.01),VCAM^(-1)及整合素α4mRNA降低(P<0.01);与移植组比较,扎方7 d组NS减小(P<0.05),联合1,3,14 d组NS增加(P<0.01),扎方1,3 d组VCAM^(-1)mRNA减少(P<0.01),扎方1,7,14 d组整合素α4mRNA减少(P<0.01),联合各组VCAM^(-1)及整合素α4mRNA均降低(P<0.01),扎方及联合各组Neu N增高(P<0.01);扎方与联合组比较,联合各组NS及Neu N表达增加(P<0.01),VCAM^(-1)和整合素α4mRNA表达降低(P<0.01);同组间比较,各组NS,Neu N及VCAM^(-1) mRNA均以7 d组变化显著,14 d有明显改善(P<0.01),整合素α4mRNA以3 d组变化显著,14 d可改善(P<0.01)。结论:脑缺血后神经功能及神经元均出现不同程度损伤;扎方和BMSCs移植均可不同程度改善脑缺血后神经功能及脑组织神经元状态,以二者联合作用显著,其机制可能与干预VCAM^(-1)及整合素α4动态表达有关。

Objective： To observe the effect of Hui Medicine Zhali Nusi Fang（ ZLNF） intervening bone marrow mesenchymal stem cells（ BMSCs） transplantation on Adhesion molecules and neuronal nuclei（ Neu N） expression in rats after cerebral ischemia reperfusion injury（ CIRI）. Method： SD rats were divided randomly into Sham-operated,model,ZLNF,BMSCs transplantation and ZLNF combined with BMSCs groups;middle cerebral artery occlusion（ MCAO） model was duplicated with nylon thread,BMSCs were cultured and amplified by the whole bone marrow adherence method; drugs were given to the rats by intragastric administration（ 14. 6 g ·kg-1·d-1）,BMSCs suspension solution were transplanted into brain through carotid artery; rat neurological scores（ NS） were evaluated and brain was taken out at 1,3,7,14 days after transplantation;vascular cell adhesion molecular（ VCAM-1） and integrin α4mRNA were detected by real time-PCR and immunohistochemical method to measure the expression of Neu N. Result： The NS and Neu N in model groups were obviously decreased than the sham-operated groups（ P〈0. 01）,and the expression of VCAM-1 and integrin α4mRNA were significantly increased（ P〈0. 01）; compared with the model groups,the NS at 1,3,14 days in ZLNF and at 3,7,14 days in transplantation groups and in all combination groups were increased（ P〈0. 01,P〈0. 05）,the Neu N in all of the groups were increased（ P〈0. 01）,the VCAM-1 and integrin α4mRNA in all of the groups were decreased（ P〈0. 01）; compared with the transplantation groups,the NS at 7 day in ZLNF group was decreased（ P〈0. 05） and were significantly increased at 1,3,14 days in combination groups（ P〈0. 01）,the VCAM-1 mRNA at 1,3 days in ZLNF groups were decreased（ P〈0. 01）,and the integrin α4mRNA at 1,7,14 days in ZLNF groups were decreased（ P〈0. 01）, the VCAM-1 and integrin α4mRNA in all of the combination groups were decreased（ P〈0. 01）,the Neu N in all of the ZLNF groups and combination groups were increased（ P〈0. 01）; Compared with the ZLNF groups,the NS and Neu N in combination groups were obviously increased and the expression of VCAM-1 and Integrin α4mRNA were significantly decreased（ P〈0. 01）.Compared with the same group,the changes of NS,Neu N and VCAM-1 mRNA at all groups were remarkable in 7day group,and were improved in 14 days group,the changes of integrin α4mRNA were remarkable in 3 day group,and can improved in 14 days group. Conclusion： The neurological function and neurons were damaged to some degree after cerebral ischemia; both ZLNF and BMSCs transplantation can improved the neurological function and the condition of neurons,the effect was superior by the combination of them, and its mechanism may be related to the regulation of the dynamic expression of VCAM-1 and Integrin α4.