猕猴骨髓间质干细胞移植修复脊髓损伤  

Transplantation of rhesus bone marrow mesenchymal stem cells for the repair of spinal cord injury

作　　者：邓宇斌[1] 原清涛[1] 刘晓刚[1] 毕小彬[1] 

机构地区：[1]中山大学中山医学院病理生理教研室,广东省广州市510080

出　　处：《中国组织工程研究与临床康复》2007年第20期3964-3967,共4页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：广东省科技计划资助项目(2005B50301012)~~

摘　　要：目的:观察猕猴骨髓间质干细胞的诱导分化及其对损伤脊髓功能恢复的影响。方法:实验于2004-09/2007-01在中山大学实验动物中心完成,选用健康雄性恒河猴10只。①将体外分离传代获得的猕猴骨髓间质干细胞中加入Hoechst33342标记液,然后应用隐丹参酮进行诱导分化。②按照改良Allen氏法制作猕猴脊髓损伤模型,按随机数字表法分为2组,实验组(n=5)在脊髓损伤处头尾两端注射已初步诱导的猴骨髓间质干细胞,对照组(n=5)注射磷酸盐缓冲液。③细胞移植后1周至3个月,采用Tarlov评分方法(0￣4级)评价猴损伤脊髓的功能恢复情况:0级为肢体完全瘫痪;4级为正常。④采用Trueblue逆行追踪实验评估轴突通路的再建情况。⑤移植术后3个月处死动物,将脊髓组织经苏木精-伊红染色后行组织学观察。结果:实验组5只动物均进入结果分析,对照组2只进入结果分析。①猴骨髓间质干细胞诱导分化半小时后,神经元样细胞的胞体和轴突呈神经元特异性烯醇化酶强阳性表达。细胞移植3个月后,实验组猴子的脊髓损伤段冰冻切片中有大量Hoechst33342标记阳性的细胞。②实验组术后1周起即可见动物双下肢有轻微动作,至2个月左右,猴子的脊髓功能基本得以稳定,此时Tarlov评分介于2级与3级之间。与实验组相比,对照组动物脊髓功能无恢复。③实验组可见脊髓组织形态完整,细胞无变性衰亡。④实验组动物的胸段脊髓和大脑感觉运动区的冰冻切片上均发现了Trueblue标记阳性的细胞。结论:骨髓间质干细胞来源的神经元能够促进脊髓损伤猕猴的运动功能恢复。AIM： To observe the effects of the transplantation of bone marrow mesenchymal stem cells （MSCs）-derived neurons on the functional recovery of injured spinal cord in rhesus. METHODS： The experiment was conducted in the Experimental Animal Center of Sun Yat-sen University between September 2004 and January 2007. （1） The MSCs isolated in vitro from ten healthy male rhesuses were put in Hoechst33342 labeling buffer, and induced to differentiate into neurons by cryptotanshinone. （2）The spinal cord injuries were made by the modified Allen method, then the animals were randomly divided into two groups： the animals in the experimental group （n =5） were injected with the primarily induced MSCs in both of the proximal and distal sides of the epicenter, and the control group （n =5） was injected with phosphate buffer. （3）Tadov classification with 4 grades was used to evaluate the recovery of motor function at 1 week, 1, 2 and 3 months after transplantation： complete paralysis scored grade 0, normal scored grade 4. （4）True blue chloride retrograde tracing test was carried out to evaluate the reestablishment of axons pathway. （5） After the animals were sacrificed three months post-transplantation, the hematoxylin-eosin （HE） staining and immunohistochemistry were performed. RESULTS： All 5 rhesuses of the experimental group entered the result analysis, while only 2 of the control group were involved in the result analysis except 3 died at 6 week after modeling. （1）Immunocytochemistry suggested that after differentiated for half an hour, the neuron specific enolase was strongly positively expressed in the cell body and axon of neuron-like cells; three months after cell transplantation in the experimental group, a plenty of Hoechst33342 labeled positively cells were found in the frozen sections of injured segment, with nucleus sending out blue fluorescence. （2）At one week after operation, slight movement in the limbs of the experimental group was found, and to two month, Tarlov

关 键 词：细胞移植 骨髓细胞 间质细胞 脊髓损伤 猕猴 

分 类 号：R339.4[医药卫生—人体生理学]