脉冲强磁场诱导体外新生大鼠神经干细胞向神经元方向分化  

High-intensity pulsed electromagnetic field induces the differentiation of neural stem cells from neonatal rats into neurons in vitro

作　　者：蒋婷[1] 许涛[1] 向威[2] 龙兴蓝[3] 李冰冰[1] 彭涛[4] 孙衢骎 

机构地区：[1]华中科技大学同济医学院附属同济医院康复科,武汉430030 [2]华中科技大学同济医学院附属同济医院骨科,武汉430030 [3]湖北省宜昌市第一人民医院 [4]华中科技大学脉冲强磁场中心

出　　处：《中华物理医学与康复杂志》2014年第10期740-744,共5页Chinese Journal of Physical Medicine and Rehabilitation

基　　金：国家自然科学基金（面上项目）资助（81271732）

摘　　要：目的 探讨脉冲强磁场对体外新生大鼠神经干细胞（NSCs）向神经元方向分化的作用.方法 体外分离培养新生3d内的SD大鼠脑室下区的NCSs,无血清培养14d后,随机分为实验组和对照组.实验组置于脉冲强磁场条件下用前期探索的促增殖参数（0.1 Hz、4T、8次）进行干预,每次脉冲放电20 ms.对照组置于相同条件下但不做干预处理.干预后第1天,采用10％胎牛血清诱导贴壁分化,显微镜下观察不同时间段细胞形态学变化;贴壁分化后第7天,通过免疫荧光染色、蛋白免疫印迹法（Western Blot）及实时定量聚合酶链反应技术（RT-PCR）检测NCSs分化后第7天神经元βⅢ微管蛋白（TUJ1）及星形胶质细胞相关特异性标志神经胶质纤维酸性蛋白（GFAP）的表达.结果 干预后,免疫荧光染色示实验组TUJ1阳性细胞数为（33.4±5.1）％,较对照组[（26.5 ±7.0）％]明显增多（P＜0.05）,实验组的GFAP阳性细胞率[（23.9±5.0）％]较对照组[（36.2±2.2）％]明显减少（P＜0.05）.RT-PCR检测显示,实验组TUJ1的mRNA表达量是对照组的（1.682 ±0.086）倍,GFAP的mRNA相对表达量是对照组的（0.590±0.157）倍,且组间差异有统计学意义（P＜0.05）.Western Blot检测结果与RT-PCR检测一致,实验组TUJ1蛋白表达较对照组增多,GFAP蛋白表达较对照组减少.结论 脉冲强磁场具有促进NCSs向神经元分化,并抑制其向星形胶质细胞分化的作用.Objective To Investigate the influence of high-intensity pulsed electromagnetic field （HIPEMF） on neural differentiation of neonatal rats neural stem cells in vitro.Methods Neural stem cells （NSCs） were isolated from the subventricular zone of 3-day-old neonatal rats and cultured with serum-free condition medium for 14 days.All the NSCs were then randomly classified into an experimental group which received the stimulation of HIPEMF （0.1 Hz,4 T,8 pulses） and a control group which received no special intervention.Differentiation of the culture was induced by addition of 10％ fetal bovine serum on the first day after intervention,the morphological changes of cells were observed under the microscope at different time points.The NSCs adhered to the wall and differentiated for seven days,the immunofluorescence was employed to observed and calculate the ratio of differentiated cells with astrocyte marker GFAP or neuronal markers TUJ1.RT-PCR and western blotting were used to measure the expression levels of the differentiated cells based on gene and protein levels,respectively.Results Immunofluorescence staining showed the number of TUJI positive cells in the experimental group（33.4％ ± 5.1％）was significantly more than the control group （26.5％ ± 7.0％）,while the number of GFAP positive cells was decreased（23.9％ ± 5.0％） as compared with the control group（36.2％ ± 2.2％）.RT-PCR showed that the TUJ1 mRNA expression levels in the experimental group was （1.682 ± 0.086） times of the control group.Western blot showed that the expression of TUJ1 （0.729 ±0.061） in the experimental group was higher than in the control group （0.590 ± 0.157）,while the expression of GFAP in the experimental group （0.566 ± 0.056） was less than in the control group（1.034 ± 0.051）.Conclusions HIPEMF facilitates differentiation of neural stem cells to neurons,at the cost of reducing astrocytic differentiation.

关 键 词：神经干细胞 脉冲强磁场 细胞分化 神经元 

分 类 号：R741[医药卫生—神经病学与精神病学]