摘要
目的:建立一种简便的胚胎小鼠肠道神经元分离和培养方法。方法:分离12～13 d胎鼠肠道组织,制备单细胞悬液,用神经元专用培养基进行培养,观察神经元的生长情况。培养第3、5、7、9、14 d使用免疫细胞化学染色法,以β微管蛋白(Tu J1)反映神经元突起形态并用Sholl分析进行量化,以神经元核心抗原(Neu N)为标志物反映神经元的纯度。结果:培养24 h神经元贴壁,随培养天数延长神经元突起长度增加,数目增多,形态逐渐复杂,与邻近细胞突起相互连接,形成网络。与培养第3 d相比,第7、9、14 d神经元纯度明显增高(P <0.05)。结论:本方法分离培养的神经元纯度高,在神经元专用培养基中存活状态良好,未见胶质细胞明显增生。
Objective: To establish the method of isolation and culture of embryonic mice intestinal neurons. Methods:Guts of E12. 5 were dissociated and made single cell suspension,which were cultured in neuron specific medium observing the growth of neurons. The neurons were stained with immunocytochemistry on the day 3,5,7,9 and 14 of culturing in vitro. The morphological changes of neurons were displayed by anti-β-III tubulin( Tu J1) and analyzed quantitatively by Sholl analysis. Neuron specific protein( Neu N) was used as a marker to reflect the purity of neurons.Results: The neurons adhered to the wall at 24 hours after culturing in vitro. With the increase of the length and number of neurites,the number of primary neurites increased,and the morphology became more and more complicated. The neurites connected with the adjacent cells to form a network after several days. Compared with the 3 rd day of culture,the purity of neurons on the 7 th,9 th and 14 th day were significantly higher( P < 0. 05). Conclusion: Neurons isolated and cultured using this mothed had a high-purity. They were in good conditions without glial cells in neuron specific medium.
作者
李娜
程波
李岩松
袁伟
苗继文
王强
李爽
Li Na;Cheng Bo;Li Yansong;Yuan Wei;Miao Jiwen;Wang Qiang;Li Shuang(Deptment of Anesthesiology,The First Affiliated Hospital,School of Medicine,Xi’an Jiaotong University,Xi’an,710061 China)
出处
《神经解剖学杂志》
CAS
CSCD
北大核心
2019年第2期182-186,共5页
Chinese Journal of Neuroanatomy
基金
国家自然科学基金(81804190)
关键词
肠道神经元
胚胎
原代培养
小鼠
intestinal neurons
embryo
primary culture
mouse
