基质细胞衍生因子-1诱导神经干细胞靶向性迁移的实验研究  

Stromal cell derived factor-1 inducing targeted migration of neural stem cells

作　　者：陈二涛[1] 潘栋超[1] 冯东福[1] 毕永延[1] 汪洋[2] 朱志安[1] 

机构地区：[1]上海交通大学医学院附属第三人民医院神经外科,上海201900 [2]复旦大学上海医学院解剖与组织胚胎学系,上海200032

出　　处：《中华神经医学杂志》2010年第6期561-565,570,共6页Chinese Journal of Neuromedicine

基　　金：上海市教委自然科学基金（04BB23、07ZZ42）;上海市自然科学基金（09ZR1417500）

摘　　要：目的 观察基质细胞衍生因子-1（SDF-1）对神经干细胞（NSCs）迁移的影响.方法 由胚胎大鼠脑组织获取NSCs并进行传代培养和分化鉴定,使用流式细胞术检测NSCs纯度及SDF-1特异性受体CXCR4的表达情况,之后利用Blind-Well小室体外迁移体系观察不同浓度的SDF-1（0 ng/L、1 ng/mL、10 ng/mL、50 ng/mL、100 ng/mL、500 ng/mL和1000 ng/mL）对NSCs迁移数量的影响,并使用μ-载片观察SDF-1对NSCs迁移距离的影响.结果成功分离培养得到了能够在体外不断分裂增殖、具有多向分化潜能的NSCs,连续传3代后绝大部分细胞nestin表达阳性,nestin和CXCR4的共表达率达到80%左右.细胞趋化实验结果表明,SDF-1对NSCs有较强的趋化作用,随着SDF-1浓度的升高,发生迁移的细胞数量也随之增加,并于SDF-1浓度为500ng/mL时达到最高峰[（256.79±38.27）个细胞/每高倍视野].在μ-载片细胞生长通道两侧的SDF-1浓度梯度（500ng/mL→0ng/mL）作用下,由细胞克隆球迁出的细胞呈不对称分布,通常有更多的迁出细胞分布于趋化因子浓度较高的一侧,且该侧细胞的最大迁移距离也比对侧远.结论 SDF-1与其特异性受体CXCR4相巨作用能够诱导NSCs发生靶向性迁移.Objective To investigate the effect ofstromal cell derived factor-1 （SDF-1） on the regulation of neural stem cells （NSCs） migration.Methods NSCs were obtained from the cerebral cortex of embryonic rats and cultured in serum-free medium,and their stem cell properties were assessed by means of induced differentiation in vitro into neurons and astrocytes.After in vitro cell culture,the purity of NSCs and the co-expression rate of CXCR4/nestin were detected by flow cytometry.Blind-well chambers were employed to detect the chemotactic effects of SDF-1 by counting the cells which had crossed a 8 μm pore membrane when confronted with varying concentrations of SDF-1 （0,1,10,50,100,500 and 1000 ng/mL）,and the distribution of cells migrated out of the same neurosphere was overviewed by μ-slides in the persistent concentration gradient of SDF-1.Results Neurospheres were formed by persistent proliferation of NSCs, which were capable of differentiating into neurons （β-tubulin＋） and astrocytes （GFAP＋） in media without mitogens,and flow cytometry analyses showed that most of the cultured cells expressed nestin and the co-expression rate of CXCR4/nestin was nearly 80%.SDF-1 showed great chemotaxis to NSCs,and the amount of cells having migrated through the membrane in 500 ng/ml SDF-1 group was higher than that in other groups （P〈0.05）.When the cells were confronted with a linear concentration gradient （from 500 to 0 ng/mL）,which was generated by diffusion and stable for at least 48 h,the cells migrated out ofa neruosphere could distribute irregularly with more cells locating in the region of higher concentration of SDF-1 and longer migration distance away from the center of the neurosphere than the opposite.Conclusion SDF-1 binding to its specific receptor CXCR4 was capable of inducing NSCs migrating directionally to the source of SDF-1.

关 键 词：神经干细胞 基质细胞衍生因子-1 细胞迁移 Blind-Well小室 

分 类 号：R329.28[医药卫生—人体解剖和组织胚胎学]