中国组织工程研究与临床康复2008，Vol.12Issue(16)：3189-3192,4.
胚胎大鼠原代神经干细胞培养方法的改进
A new method to culture primary neural stem cells of embryonic rats
摘要
Abstract
BACKGROUND:Both in vivo and vitro microenvironment can influence the proliferation and differentiation of neural stem cells (NSCs). In addition, cell culture solution plays variable roles in cell proliferation and differentiation.OBJECTIVE:To develop a convenient and rapid method to promote NSC primary culture by modifying traditional serum-free medium. DESIGN, TIME AND SETTING:Randomized controlled cell trial was performed at Institute of Brain Science, Qingdao University Medical School from November 2005 to September 2006.MATERIALS:Ten Wistar rat of gestation for 12-16 days; cell suspension fron brain tissue of embryonic rat. METHODS:Cell suspension were seeded into four 50-mL culture flasks with cell density of 1×106 mL-1, and divided into 2 groups with 2 flasks in each group. The control cells (flasks A and B) were cultured in serum-free medium containing DMEM/F12, B27 (2%), basic fibroblast growth factor (20 μg/L), and epidermal growth factor (20μg/L), and the experimental cells (flasks C and D) were firstly cultured with DMEM/F12 containing fetal bovine serum (FBS, 5%), following by the same serum-free medium 2 to 3 days later. MAIN OUTCOME MEASURES:The proliferation of NSCs was observed by inverted microscopy and immunocytochemistry.RESULTS:①Most of the cells in two culture conditions expressed nestin, the specific marker of NSCs, and were immunocytochemically positive. ②Neural stem cells cultured with FBS formed neurospheres 3-4 days earlier than those without FBS. ③There were no significant differences in cell number, but the neurospheres under the experimental condition were larger than those in control group. Some of the cells were neurone specific enolase-positive after serum-conditioned induction, and some were glial fibrillary acidic protein-positive, indicating NSCs had differentiated into neuron-like cells and neurogliocytes.CONCLUSION:FBS-conditioned pre-culture can accelerate the proliferation of neural stem cells.关键词
神经干细胞/细胞培养/大鼠分类
医药卫生引用本文复制引用
邢成名,耿秀丽,刘玉霞..胚胎大鼠原代神经干细胞培养方法的改进[J].中国组织工程研究与临床康复,2008,12(16):3189-3192,4.基金项目
青岛市卫生局重点课题"神经干细胞的培养和移植在治疗缺血性脑血管病中的应用"(2004-wsxk)the Key Subject of Qingdao Health Bureau, No.2004-wsxk （2004-wsxk）
