解剖学报

解剖学报 ›› 2020, Vol. 51 ›› Issue (6): 832-938.doi: 10.16098/j.issn.0529-1356.2020.06.005

• 神经生物学 • 上一篇    下一篇

骨形态发生蛋白/维甲酸诱导的神经特异性蛋白3在丙戊酸钠诱导神经干细胞向神经元分化过程中的作用

何辉 李雯 刘娟 姚翔 薛成 秦建兵 田美玲 金国华*   

  1. 南通大学医学院人体解剖学系，江苏省神经再生重点实验室，神经再生协同创新中心，江苏 南通 226001
  • 收稿日期:2020-04-19 修回日期:2020-05-17 出版日期:2020-12-06 发布日期:2020-12-06
  • 通讯作者: 金国华 E-mail:jguohua@ntu.edu.cn
  • 基金资助:
    江苏省自然科学基金青年基金项目;南通大学大学生创新创业训练计划项目

Role of bone morphogenetic protein/retinoic acid-inducible neural-specific protein 3 in the differentiation of neural stem cells into neurons induced by valproate

HE Hui LI Wen LIU Juan YAO Xiang XUE Cheng QIN Jian-bing TIAN Mei-ling JIN Guo-hua*   

  1. Department of Human Anatomy, Medical College of Nantong University, Jiangsu Key Laboratory of Neuroregeneration, Collaborative Innovation Center of Neuroregeneration, Jiangsu Nantong 226001, China
  • Received:2020-04-19 Revised:2020-05-17 Online:2020-12-06 Published:2020-12-06
  • Contact: JIN Guo-hua E-mail:jguohua@ntu.edu.cn

PDF

摘要：

目的 探讨骨形态发生蛋白/维甲酸诱导的神经特异性蛋白3（Brinp3）在丙戊酸钠（VPA）诱导神经干细胞向神经元分化过程中的作用。  方法 体外培养大鼠海马神经干细胞，运用Real-time PCR和Western blotting技术在VPA诱导神经干细胞分化后24 h和48 h检测Brinp3的表达；Real-time PCR检测Brinp3在成年大鼠各组织中以及在神经干细胞、星形胶质细胞和神经元中的表达水平；在神经干细胞中转染Brinp3小干扰RNA（siRNA）并诱导分化24 h后，运用Real-time PCR、Western blotting和免疫荧光技术检测Brinp3和神经元标志分子的表达。以上实验均包含5次生物学重复。  结果 与对照组相比，VPA处理组中Brinp3的mRNA和蛋白水平在24 h和48 h均显著上调（P<0.05）；Brinp3在脑组织中呈优势表达；Brinp3在星形胶质细胞中表达较低，而在神经元中表达较高（P<0.001）；在神经干细胞中转染Brinp3 siRNA，诱导分化24 h后，Brinp3的表达被显著抑制（P<0.001），神经元标志分子的表达均显著下调（P<0.01），第4天分化成的神经元比例减少（P<0.001）。  结论 Brinp3表达的上调可能介导了VPA促神经干细胞向神经元分化的功能。

关键词: 骨形态发生蛋白/维甲酸诱导的神经特异性蛋白3, 丙戊酸钠, 神经干细胞, 神经元, 免疫印迹法, 大鼠

Abstract:

Objective  To explore the role of bone morphogenetic protein/retinoic acid-inducible neural-specific protein 3 (Brinp3) in the differentiation of neural stem cells into neurons induced by valproate (VPA).   Methods  The neural stem cells of rat hippocampi were cultured in vitro. The expression of Brinp3 was detected by Real-time PCR and Western blotting at 24 hour and 48 hour after VPA induced neural stem cells differentiation. Real-time PCR was used to detect the expression of Brinp3 in various tissues of adult rats, as well as in neural stem cells, astrocytes and neurons. After neural stem cells were transfected with Brinp3 small interfering RNA (siRNA) and then induced differentiation for 24 hours, the expression of Brinp3 and neuron-specific markers were detected by Real-time PCR, Western blotting and immunofluorescence technique. All the above experiments included 5 biological repeats.   Results Compared with the control group, the mRNA and protein levels of Brinp3 in the VPA group increased significantly at 24 hour and 48 hour (P<0.05). Brinp3 showed dominant expression in brain tissue, and its expression was lower in astrocytes but higher in neurons (P<0.001). The expression of Brinp3 and neuron-specific markers was significantly inhibited after transfected Brinp3 siRNA into neural stem cells and induced differentiation for 24 hours (P<0.001 and P<0.01, respectively). In addition, the proportion of neurons after differentiation decreased at day 4 (P<0.001).   Conclusion  The upregulation of Brinp3 may mediate the role of VPA promoting the differentiation of neural stem cells into neurons.

Key words: Bone morphogenetic protein/retinoic acid-inducible neural-specific protein 3, Valproate, Neural stem cell, Neuron, Western blotting, Rat

中图分类号: