大鼠原代背根神经元在聚甲基丙烯酸甲酯电纺纳米纤维支架上培养的微观形态研究  

Growth pattern of rat dorsal root ganglion neurons cultured on random or aligned electrospun polymethylmethacrylate nanofibers plus its microstructure research

作　　者：夏海坚[1] 孙晓川[1] 王伯初[2] 刘丹[3] 王亚洲[2] 郝石磊[2] 钟东[1] 夏永智[1] 

机构地区：[1]重庆医科大学附属第一医院神经外科,重庆400016 [2]重庆大学生物工程学院生物流变科学与技术教育部重点实验室,重庆400044 [3]重庆市中山医院药剂科,重庆400013

出　　处：《重庆医科大学学报》2016年第5期438-442,共5页Journal of Chongqing Medical University

基　　金：国家自然科学基金青年基金资助项目(编号:81100906);重庆市科委前沿与应用基础研究计划资助项目(编号:cstc2014jcyj A 10061);国家临床重点专科建设经费资助项目(编号:财社【2011】170);重庆大学"生物流变科学与技术"教育部重点实验室访问学者基金资助项目(编号:CQKLBST-2014-004)

摘　　要：目的:探讨原代大鼠背根神经元(dorsal root ganglion neurons,DRGn)在随机或轴向有序排列聚甲基丙烯酸甲酯(polymethylmethacrylate,PMMA)电纺纳米纤维材料上的生长特点及其微观形态。方法:利用静电纺丝技术构建具随机或轴向有序拓扑结构的PMMA纳米纤维、及作为对照的PMMA薄膜,研究分为PMMA薄膜组、随机及有序PMMA电纺纤维组共3组;纯化大鼠DRGn,利用所转染的绿色荧光蛋白作为显色手段,分析DRGn在各型PMMA电纺纳米纤维上,其神经突的生长能力及生长形式;并通过扫描电镜探索DRGn在电纺纤维上的微观形态特点;本研究采用不同批次构建的PMMA材料重复5次。结果:随机及有序PMMA电纺纤维的直径分别为(0.49±0.04)μm和(0.47±0.03)μm,两者无统计学差异(t=1.079,P=0.474);DRGn能够在PMMA各型材料上贴壁及生长,培养第2天,DRGn在PMMA薄膜、随机或有序电纺纳米纤维上平均分别生成(3.3±1.6)、(2.9±1.2)、(3.1±1.3)个神经突,神经突平均长度分别为(143.7±45.2)μm、(134.2±38.7)μm、(138.4±40.6)μm,和薄膜环境相比,在检查时间点无平均神经突数量(F=8.230,P=0.138)及神经突长度(F=5.470,P=0.195)的差异,有序纳米纤维上的DRGn其神经突能够和基质纤维的延伸方向保持一致;电镜检查证实电纺纤维对于DRGn神经突的生长具有明显的接触引导作用,而DRGn能够敏感感知培养局部的拓扑环境特点并作出相应反应,其具有较强的把握周围纤维信号的能力。结论:PMMA电纺纳米纤维具有作为神经损伤后植入性支架材料的潜力。Objective：To evaluate the growth pattern of rat dorsal root ganglion neurons（DRGn）cultured on random or aligned polymethylmethacrylate（PMMA）electrospun nanofibers and to exhibit the microstructure between neurites and fibers. Methods：Random and aligned electrospun nanofibers were fabricated to culture DRGn on them. The growth manner of DRGn on different substrates were showed,with transfected GFP as the reporting gene and PMMA film as the control. The microstructure between neurites and fibers were analyzed by scanning electron microscope（SEM）. Results：The diameters of the random and aligned PMMA electrospun nanofibers were（0.49±0.04）μm and（0.47±0.03）μm,without statistical differences（t=1.079,P=0.474）. DRGn could adhere to and grow on the nanofibers;on 2nd day of culture,no difference in average neurite number（F=8.23,P=0.138）and neurite length（F=5.47,P =0.195）was found. On aligned nanofibers DRGn could form aligned neurites along the orientation of substrate nanofi-bers. Through analysis of SEM,it was revealed that PMMA electrospun nanofibers had strong contact guidance on the neurite growth of DRGn,and DRGn could also perceive and response to local topographic surrounding sensitively. Conclusion：PMMA electrospun nanofibers is potential scaffold after neural injury.

关 键 词：静电纺丝 纳米纤维 背根神经元 细胞支架 接触引导 

分 类 号：R318.08[医药卫生—生物医学工程]