中国组织工程研究

中国组织工程研究 ›› 2016, Vol. 20 ›› Issue (41): 6092-6097.doi: 10.3969/j.issn.2095-4344.2016.41.002

• 骨髓干细胞 bone marrow stem cells • 上一篇    下一篇

细胞传代对骨髓间充质干细胞向神经干细胞分化的影响

梁  维1，刘  洲1，许志恩1，林力峰1，方洪明2   

  1. 1广东医科大学附属医院神经内科，广东省湛江市  524001
    2普宁市人民医院神经内科，广东省普宁市  515300
  • 修回日期:2016-08-02 出版日期:2016-10-07 发布日期:2016-10-07
  • 通讯作者: 刘洲，博士，副主任医师，硕士生导师，广东医科大学附属医院神经内科，广东省湛江市 524001
  • 作者简介:梁维，女，1985年生，广东省湛江市人，汉族，2012年广东医学院毕业，硕士，医师，主要从事神经干细胞和脑血管病方面的研究。

Effect of cell passage on differentiation of bone marrow mesenchymal stem cells into neural stem cells

Liang Wei1, Liu Zhou1, Xu Zhi-en1, Lin Li-feng1, Fang Hong-ming2   

  1. 1Department of Neurology, Affiliated Hospital of Guangdong Medical University, Zhanjiang 524001, Guangdong Province, China
    2Department of Neurology, Puning People’s Hospital, Puning 515300, Guangdong Province, China
  • Revised:2016-08-02 Online:2016-10-07 Published:2016-10-07
  • Contact: Liu Zhou, M.D., Associate chief physician, Master’s supervisor, Department of Neurology, Affiliated Hospital of Guangdong Medical University, Zhanjiang 524001, Guangdong Province, China
  • About author:Liang Wei, Master, Physician, Department of Neurology, Affiliated Hospital of Guangdong Medical University, Zhanjiang 524001, Guangdong Province, China

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摘要：

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文题释义：
传代培养：是指需要将培养物分割成小的部分，重新接种到另外的培养器皿(瓶)内，再进行培养的过程。对单层培养而言，80%汇合或刚汇合的细胞是较理想的传代阶段。
神经干细胞：是一种来源于神经组织及神经组织发源地，具有干细胞特征，终生保持自我更新能力并能分化为神经元、星形胶质细胞和少突胶质细胞的一类细胞。神经干细胞可直接来源于胚胎、成年哺乳动物的脑组织，也可来源于非神经组织如：骨髓、脂肪、皮肤等组织的干细胞。

 

摘要
背景：体外持续细胞传代是否影响骨髓间充质干细胞分化为神经干细胞的机制尚未明确。
目的：观察细胞传代对骨髓间充质干细胞向神经干细胞分化的影响。
方法：采用全骨髓贴壁法培养大鼠骨髓间充质干细胞，取传代至第3，6，9，12代的骨髓间充质干细胞，在无血清神经干细胞培养基中诱导培养7，14 d，观察细胞生物学特征，通过流式细胞仪检测Nestin的表达情况观察骨髓间充质干细胞向神经干细胞分化能力，再进一步诱导分化，通过免疫组织化学荧光技术检测神经烯醇化酶及胶质酸性蛋白表达，以鉴定神经干细胞的多向分化能力。
结果与结论：各组骨髓间充质干细胞加入诱导剂后均分化成神经干细胞，Nestin表达阳性，各组细胞分化的比例差异有显著性意义(P < 0.05)，其中第6代骨髓间充质干细胞分化能力最强，诱导7 d及14 d Nestin阳性细胞的比例分别为(93.7±2.3)%，(96.2±1.8)%，第6，9代明显较第3，12代骨髓间充质干细胞向神经干细胞分化的比例高，且所得神经干细胞球经贴壁分化后可表达神经烯醇化酶、胶质酸性蛋白。结果表明，骨髓间充质干细胞向神经干细胞分化与细胞传代有关，过早或过晚均不利于分化。

 

 

关键词: 干细胞, 骨髓干细胞, 细胞传代, 骨髓间充质干细胞, 神经干细胞, 分化

Abstract:

BACKGROUND: It is unclear whether serial cell passage in vitro influences the differentiation of bone marrow mesenchymal stem cells into neural stem cells.
OBJECTIVE: To investigate the effect of cell passage on the differentiation of bone marrow mesenchymal stem cells into neural stem cells.
METHODS: Rat bone marrow mesenchymal stem cells were isolated and cultured by the whole bone marrow adherence method. Bone marrow mesenchymal stem cells at passages 3, 6, 9, 12 were incubated in serum-free medium. After culture for 7 and 14 days, cell biological characterization was observed and differenitaiton ability into neural stem cells was observed by detecting Nestin expression in cells using flow cytometry. Then, the cells were further induced to differentiate and cell multipotential differentiation capacity was detected by measurement of nerve enolase and glial acidic protein expression.
RESULTS AND CONCLUSION: Under induction, bone marrow mesenchymal stem cells at different passages were all differentiated into Nestin-positive neural stem cells. However, there was a significant difference in differentiation proportion of cells at different passages (P < 0.05). Strongest differentiation ability was found in the passage 6 cells, with the Nestin expression up to (93.7±2.3)% at 7 days of induction and (96.2±1.8)% at 14 days of induction. The proportion of differentiated cells at passages 6 and 9 was signfiicantly higher than that at passages 3 and 12. Moreover, adherent cells were positive for nerve enolase and glial acidic protein. All these findings indicate that the differentiation of bone marrow mesenchymal stem cells into neural stem cells is correlated with cell passage. Cells at lower or higher passages are both detrimental to cell differentiation.

 

 

Key words: Bone Marrow, Mesenchymal Stem Cells, Serial Passage, Cell Differentiation, Neural Stem Cells, Tissue Engineering

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