过表达NeuroD1对脊髓反应性星形胶质细胞转分化为神经元的影响  

The influence of overexpression of Neuro D1 on transdifferentiation of spinal cord reactive astrocytes into neurons

作　　者：康文博[1] 陈翀[1] 李晓红[1] 王景景[1] 涂悦[1] 张赛[1] 梁海乾[1] 

机构地区：[1]武警后勤学院附属医院脑科医院、脑创伤与神经疾病研究所、天津市神经创伤修复重点实验室,天津300162

出　　处：《中国神经精神疾病杂志》2015年第9期564-568,共5页Chinese Journal of Nervous and Mental Diseases

基　　金：国家自然科学基金项目(编号:81301050,81271392,81401067);天津市应用基础与前沿技术研究计划(编号:14JCQNJC10200,15JCYBJC28100);中国博士后基金项目(编号:2013M542583)

摘　　要：目的探讨过表达Neuro D1对脊髓反应性星形胶质细胞转分化为神经元的影响。方法体外原代培养SD大鼠脊髓星形胶质细胞,以划痕实验制备反应性星形胶质细胞。实验分为空白组(NV组)、对照病毒组(GFP组)和Neuro D1病毒组(Neuro D1组)。各组行划痕处理7 d后,NV组不感染病毒,GFP组感染携带GFP基因的逆转录病毒,Neuro D1组感染同时携带GFP和Neuro D1基因的逆转录病毒,24 h后同时更换神经元条件培养基。各组在1 d、2 d、3 d、5 d、7 d、14 d观察细胞形态,并采用免疫荧光染色方法分别在感染病毒后7 d观察细胞DCX阳性率和14 d观察细胞Neu N阳性率。结果更换神经元条件培养基后,各组细胞形态发生改变,胞核明显饱满,胞浆减少,突起减少并延长。与Neuro D1组相比,NV组和GFP组细胞突起短而分枝多,胞核较小。感染病毒后7 d,NV组和GFP组细胞部分形态逐渐恢复以前形态而Neuro D1组维持变化后形态。与NV组和GFP组相比,Neuro D1组出现DCX(9.84%±2.06%)(F=40.107)和Neu N(8.25%±2.78%)阳性细胞(F=21.73),结果差异都有统计学意义(P<0.05)。结论在体外培养,Neuro D1的过表达可以介导体外培养脊髓反应性星形胶质细胞转分化为神经元。ObjectiveTo investigate the effect of the overexpression of Neuro D1 on mediating transdifferentiation of spinal reactive astrocytes into neurons.MethodsSpinal cord astrocytes were cultured from the SD rat, and reactive astrocytes were prepared by scratches treatment. Cells were divided into blank groups（NV group）, control virus group（GFP group） and Neuro D1 virus group（Neuro D1 group）. At 7 d after scratches treatment, GFP and Neuro D1 groups were infected with retroviruses carrying the GFP gene and and GFP gene plus Neuro D1 gene, respectively,whereas NV group was not infected with the virus. Twenty-four hours late, the culture medium were replaced by neuron conditioned medium. Cell morphology was examined at 1, 2, 3, 5, 7 and 14 d. DCX positive and Neu N positive cells were detected at 7 d and 14 d after infection by using immunofluorescence staining method, respectively.ResultsAfter replacement with the neuron conditioned medium, the nucleus was obviously plump, the cytoplasm was thin and neurites was reduced and extended. Compared with the Neuro D1 group, neurites of NV group and GFP group were shorter with many branches andthe nucleus was smaller. At 7 d after infection, cell morphology of NV group and GFP group gradually recovered, but cell morphology of Neuro D1 group did not. Compared with NV group and GFP group, Neuro D1 group had more DCX（9.84±2.06%）and Neu N（8.25±2.78%）positive cells [F values 40.107 for DCX and 21.73 for Neu N（P〈0.05）].Conclusion The overexpression of Neuro D1 can mediate the transdifferentiation of spinal reactive astrocytes into neurons.

关 键 词：NeuroD1 转分化 反应性星形胶质细胞 神经元 划痕 

分 类 号：R651.2[医药卫生—外科学]