《中国康复理论与实践》

《中国康复理论与实践》

• 基础研究 • 上一篇    下一篇

碱性成纤维细胞生长因子-壳聚糖载体诱导神经干细胞向神经元分化的机制①

段红梅1，王聪2，杨朝阳1,2，郝鹏2，尚俊奎2，李晓光1,2   

  1. 1. 北京航空航天大学生物与医学工程学院，北京市100191；2.首都医科大学神经生物学系，北京市100069。
  • 出版日期:2016-05-25 发布日期:2016-07-04

Mechanism of Basic Fibroblast Growth Factor-chitosan Carrier Inducing Neural Stem Cells to Differentiate into Neurons

DUAN Hong-mei1, WANG Cong2, YANG Zhao-yang1,2, HAO Peng2, SHANG Jun-kui2, LI Xiao-guang1,2   

  1. 1. Department of Biomedical Engineering, School of Biological Science and Medical Engineering, Beihang University, Beijing 100191, China; 2. Department of Neurobiology, Capital Medical University, Beijing 100069, China
  • Published:2016-05-25 Online:2016-07-04

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摘要： 目的探讨碱性成纤维细胞生长因子(bFGF)-壳聚糖载体诱导神经干细胞高比例向神经元分化的潜在机制。方法纯化后神经干细胞分别与单纯壳聚糖、可溶性bFGF和bFGF-壳聚糖载体共培养。在共培养3 h、24 h、3 d 和7 d 后，Nestin、β-Ⅲ tubulin、微管相关蛋白2(MAP2)和成纤维细胞生长因子受体1(FGFR1)的免疫荧光双标记观察受体的表达情况；实时荧光定量PCR(qRT-PCR)及Western blotting 分别检测不同时间点神经干细胞诱导后相关基因的RNA水平变化。结果诱导后3 h，FGFR1 的表达量在三组中无明显差异；诱导后24 h，bFGF-壳聚糖载体组FGFR1 的表达量显著高于单纯壳聚糖组和可溶性bFGF组(P<0.001)；诱导后3 d 和7 d，FGFR1 的表达量在单纯壳聚糖组和可溶性bFGF组显著减少(P<0.001)，而在bFGF-壳聚糖载体组仍维持较高水平；qPT-PCR及Western blotting 结果显示，bFGF-壳聚糖载体组中与Erk/MAPK信号通路相关基因的表达水平显著高于单纯壳聚糖组和可溶性bFGF 组(P<0.001)。结论bFGF-壳聚糖载体通过缓释bFGF，可能上调FGFR1 的表达，进而激活Erk/MAPK 信号通路，促进神经干细胞向神经元的分化。

关键词: 碱性成纤维细胞生长因子, 壳聚糖, 神经元, 神经干细胞, 分化, 大鼠

Abstract: Objective To investigate the potential mechanism of basic fibroblast growth factor (bFGF)-chitosan carrier to induce neural stem cells to differentiate into neurons. Methods After purification, the neural stem cells were cocultured with chitosan, soluble bFGF and bFGF-chitosan carrier. Three hours, twenty-four hours, three days and seven days after induction, immunofluorescence staining of Nestin, beta tubulin III, microtubule-associated protein-2 (MAP2), and fibroblast growth factor receptor 1 (FGFR1) were used to observe the expression of FGFR1; real-time fluorescent quantitative polymerase chain reaction (qRT-PCR) and Western blotting were used to detect RNA and protein level changes after induction. Results Three hours after induction, there was no significant difference in the expression of FGFR1 among three groups. Twenty-four hours after induction, the expression level of FGFR1 was significantly higher in the bFGF-chitosan carrier group than in the chitosan group and the soluble bFGF group (P<0.001); three days and seven days after induction, the expression of FGFR1 decreased significantly in the chitosan group and soluble bFGF group (P<0.001), however, it was still higher in the bFGF-chitosan carrier group; moreover, the expression of genes associated with the pathway of extracellular regulated protein kinases/mitogen activated protein kinase (Erk/MAPK) was significantly higher in the bFGF- chitosan carrier group than in the chitosan group and soluble bFGF group (P< 0.001). Conclusion bFGF-chitosan carrier might upregulate the expression of FGFR1, then activate Erk/MAPK signal pathways, and finally promote the differentiation of neural stem cells into neurons.

Key words: basic fibroblast growth factor, chitosan, neuron, neural stem cell, differentiation, rats