中国组织工程研究 ›› 2016, Vol. 20 ›› Issue (29): 4328-4333.doi: 10.3969/j.issn.2095-4344.2016.29.010

• 组织构建细胞学实验 cytology experiments in tissue construction • 上一篇    下一篇

大鼠皮质少突胶质细胞培养条件的优化

杨  凯1,2,3,李一鹏4,刘英富5,程远驰1,2,3,汤锋武3,梁  冰3,徐忠伟4,陈旭义2,3   

  1. 1锦州医科大学武警后勤学院附属医院研究生培养基地,辽宁省锦州市  121000;2武警后勤学院附属医院脑创伤与神经疾病研究所,天津市  300162;3武警后勤学院附属医院,天津市  300162;4天津中医药大学,天津市  300193;5武警后勤学院,天津市  300162
  • 收稿日期:2016-04-23 出版日期:2016-07-08 发布日期:2016-07-08
  • 通讯作者: 陈旭义,博士,硕士生导师,武警后勤学院附属医院中国武警脑科医院,天津市 300162
  • 作者简介:杨凯,男,1987年生,河北省曲阳县人,锦州医科大学在读硕士,主要从事神经细胞生物的研究。
  • 基金资助:

    国家自然科学基金(11102235);天津市科技支撑项目(14ZCZDGX00500);天津市卫生局课题(2013KZ134,2014KZ135);武警后勤学院附属医院种子基金(FYM201417,FYM201432,FYM201542,WHJ2015018);武警后勤学院博士启动基金(WHB201417,WHB201514);武警后勤学院中心实验室开放基金(2015ZXKF01)

Optimization of culture conditions for oligodendrocytes of the rat cerebral cortex

Yang Kai1, 2, 3, Li Yi-peng4, Liu Ying-fu5, Cheng Yuan-chi1, 2, 3, Tang Feng-wu3, Liang Bing3, Xu Zhong-wei4, Chen Xu-yi2, 3   

  1. 1Postgraduate Training Basement, the Affiliated Hospital of Logistics University of Chinese People’s Armed Police Force, Jinzhou Medical University, Jinzhou 121000, Liaoning Province, China; 2Institute of Traumatic Brain Injury and Neurology, the Affiliated Hospital of Logistics University of Chinese People’s Armed Police Force, Tianjin 300162, China; 3the Affiliated Hospital of Logistics University of Chinese People’s Armed Police Force, Tianjin 300162, China; 4Tianjin University of Traditional Chinese Medicine, Tianjin 300193, China; 5Logistics University of Chinese People’s Armed Police Force, Tianjin 300162, China
  • Received:2016-04-23 Online:2016-07-08 Published:2016-07-08
  • Contact: Chen Xu-yi, M.D., Master’s supervisor, Institute of Traumatic Brain Injury and Neurology, the Affiliated Hospital of Logistics University of Chinese People’s Armed Police Force, Tianjin 300162, China; the Affiliated Hospital of Logistics University of Chinese People’s Armed Police Force, Tianjin 300162, China
  • About author:Yang Kai, Studying for master’s degree, Postgraduate Training Basement, the Affiliated Hospital of Logistics University of Chinese People’s Armed Police Force, Jinzhou Medical University, Jinzhou 121000, Liaoning Province, China; Institute of Traumatic Brain Injury and Neurology, the Affiliated Hospital of Logistics University of Chinese People’s Armed Police Force, Tianjin 300162, China; the Affiliated Hospital of Logistics University of Chinese People’s Armed Police Force, Tianjin 300162, China
  • Supported by:

    the National Natural Science Foundation of China, No. 11102235; Tianjin Science and Technology Support Project, No. 14ZCZDGX00500; a grant from Health Bureau of Tianjin, China, No. 2013KZ134, 2014KZ135; the Seed Fund of the Affiliated Hospital of Logistics College of Chinese People’s Armed Police Force, No. FYM201417, FYM201432, FYM201542, WHJ2015018; Doctoral Startup Fund of Logistics University of Chinese People’s Armed Police Force, No. WHB201417, WHB201514; Open Fund of Center Laboratory of Logistics University of Chinese People’s Armed Police Force, No. 2015ZXKF01

摘要:

文章快速阅读:

文题释义:
神经胶质:是神经组织中除神经元外的另一大类细胞,分布在神经元之间,形成网状支架。其数量比神经元多10-50倍。神经胶质细胞也具有多突起,但无树突和轴突之分。胞质内不含尼氏小体和神经原纤维,没有感受刺激和传导冲动的功能。但它们参与神经元的活动,对神经元具有支持、保护、营养、形成髓鞘和修复等多种功能。
轴突:是指神经元传导神经冲动离开细胞体的细而长的突起。
摘要
背景:
由于少突胶质细胞主要来自少突前体细胞,实验通过提取少突胶质前体细胞获取少突胶质细胞的过程中,选择合适的培养基和细胞接种密度对生长及形态学观察具有重要影响。
目的:对比优化少突胶质细胞的培养条件。
方法:取48 h内的SD新生鼠大脑皮质前体少突胶质细胞。分别用DMEM/高糖、DMEM/F12培养基培养少突胶质前体细胞,每组接种密度分别2×104/cm2,4×104/cm2,8×104/cm2,16×104/cm2,32×104/cm2,64×104/cm2;分别于少突胶质前体细胞贴壁72 h后,进行诱导分化,分化7 d后的少突胶质前体细胞在光镜下观察细胞形态变化并通过免疫荧光技术对细胞进行鉴定。
结果与结论:DMEM/高糖、DMEM/F12 培养基以2×104/cm2,4×104/cm2,8×104/cm2接种密度时,可辨识完整少突胶质前体细胞形态。诱导分化7 d后,免疫荧光鉴定提示3组少突胶质前体细胞均可呈现髓鞘碱性蛋白阳性,F12组中2×104/cm2,4×104/cm2,8×104/cm2接种密度条件下细胞均数分别为16.40±3.30,49.95±2.33,76.95±4.86,高于相应条件下的高糖组12.65±2.53,32.10±1.17,54.05± 1.56(P < 0.05)。结果表明,DMEM/F12较DMEM/高糖培养基适于少突胶质细胞培养,在一定范围内,少突胶质前体细胞分化为少突胶质细胞随接种密度成梯度增多,接种密度在4×104/cm2-8×104/cm2范围对观察细胞形态观察较为适合。

中国组织工程研究杂志出版内容重点:组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松组织工程
ORCID: 0000-0002-0298-2947(杨凯)

关键词: 组织构建, 组织工程, 少突胶质前体细胞, 少突胶质细胞, 细胞密度, 培养基, 分化, 国家自然科学基金

Abstract:

BACKGROUND: Oligodendrocytes are mostly differentiated from oligodendrocyte precursor cells. A suitable medium and cell seeding density have a significant impact on the process of the isolation of oligodendrocyte precursor cells to obtain oligodendrocytes.
OBJECTIVE: To explore the optimization of oligodendrocyte culture conditions.
METHODS: Oligodendrocyte precursor cells isolated from the newborn rats 48 hours after birth were cultured in DMEM/high glucose medium or DMEM/F12 medium using seeding densities of 2×104 cells/cm2, 4×104 cells/cm2, 8×104 cells/cm2, 16×104 cells/cm2, 32×104 cells/cm2, and 64×104 cells/cm2, respectively. Oligodendrocyte precursor cells were induced to differentiate into oligodendrocytes at 72 hours after cell adhesion. Morphology of differentiated oligodendrocyte precursor cells were observed under a light microscope, and the differentiation results were identified by immunofluorescence staining after 7-day induced differentiation.
RESULTS AND CONCLUSION: Morphology of oligodendrocyte precursor cells were recognized when cultured in DMEM/high glucose medium or DMEM/F12 medium using seeding densities of 2×104 cells/cm2, 4×104 cells/cm2, and 8×104 cells/cm2, respectively. Immunofluorescence staining showed that myelin basic protein-positive cells were found after 7-day induced differentiation, and the positive cell number were 16.40±3.30, 49.95±2.33, and 76.95±4.86 in DMEM/F12 medium, and 12.65±2.53, 32.10±1.17, and 54.05±1.56 in DMEM/high glucose medium (P < 0.05). These findings indicate that DMEM/F12 medium is more suitable for culturing oligodendrocyte precursor cells compared with DMEM/high glucose medium to some extent. The number of differentiated oligodendrocytes was gradually increased with the enhanced seeding density of oligodendrocyte precursor cells, and the seeding densities from 4×104 to 8×104 cells/cm2 were appropriate for the observation of cell morphology.

中国组织工程研究杂志出版内容重点:组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松组织工程

Key words: Oligodendroglia, Cells, Cultured, Rats

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