中国组织工程研究

中国组织工程研究 ›› 2010, Vol. 14 ›› Issue (6): 985-991.doi: 10.3969/j.issn.1673-8225.2010.06.007

• 干细胞培养与分化 • 上一篇    下一篇

移植神经干细胞的存活与迁移：电刺激小脑顶核可提高移植效率吗？

黄作义1，吴成吉1，朱晓峰2，董淑欣1，魏春杰1   

  1. 1佳木斯大学附属第一医院神经三科，黑龙江省佳木斯市  154002； 
    2佳木斯大学神经科学研究所，黑龙江省佳木斯市  154002
  • 出版日期:2010-02-05 发布日期:2010-02-05
  • 通讯作者: 朱晓峰，教授，佳木斯大学神经科学研究所，黑龙江省佳木斯市 154002
  • 作者简介:黄作义，男，1963年生，黑龙江省密山市人，汉族，1987年佳木斯医学院毕业，硕士，教授，主要从事神经干细胞基础与应用的研究。 wuchengji12@163.com
  • 基金资助:

    2006国家自然科学基金：神经干细胞共移植体的构建及其在局灶性脑缺血治疗中的应用(30540062)；
    2006黑龙江省自然基金：神经干细胞移植治疗缺血性脑血管病及移植环境优化的实验研究(D2006-17)。

Survival and migration of transplanted neural stem cells: Can it elevate the efficiency of transplantation by cerebellar fastigial nucleus stimulation?

Huang Zuo-yi1, Wu Cheng-ji1, Zhu Xiao-feng2, Dong Shu-xin1, Wei Chun-jie1   

  1. 1Third Department of Neurology, First Affiliated Hospital of Jiamusi University, Jiamusi  154002, Heilongjiang Province, China;
    2Institute of Neuroscience, Jiamusi University, Jiamusi  154002, Heilongjiang Province, China
  • Online:2010-02-05 Published:2010-02-05
  • Contact: Zhu Xiao-feng, Professor, Institute of Neuroscience, Jiamusi University, Jiamusi 154002, Heilongjiang Province, China
  • About author:Huang Zuo-yi, Master, Professor, Third Department of Neurology, First Affiliated Hospital of Jiamusi University, Jiamusi 154002, Heilongjiang Province, China wuchengji12@163.com
  • Supported by:

    the National Natural Science Foundation of China, No. 30540062*;
    Natural Science Foundation of Heilongjiang Province, No. D2006-17*

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摘要：

背景：缺血早期半暗带中神经元和内皮细胞的坏死和凋亡无法得到缓解，加之移植后的存活率低，向功能细胞的分化率低，是单纯神经干细胞移植的缺陷。课题组提出整体干预理念，期望给移植细胞提供优化的整体环境。

目的：观察神经干细胞移植与电刺激小脑顶核相结合整体干预对移植神经干细胞存活与迁移的影响。

方法：体外分离、培养新生Wistar大鼠海马表皮生长因子反应性神经干细胞，Brdu标记，并用胎牛血清诱导，观察其多向分化潜能；80只Wistar大鼠分为4组：顶核刺激+神经干细胞移植组(n=32)：左侧小脑顶核刺激24 h后行右侧大脑中动脉梗死，再24 h后将神经干细胞立体定向注入右侧侧脑室内；顶核刺激组(n=8)：PBS代替神经干细胞，其余同顶核刺激+神经干细胞移植组；单纯神经干细胞移植组(n=32)：同心圆电极插入小脑顶核，不通电其余同顶核刺激+神经干细胞移植组；对照组(n=8)：同心圆电极插入小脑顶核后不通电，其余同顶核刺激组。记录梗死后6，24 h，3，7，14，28 d大鼠的神经功能；分别在移植后3，7，14，28 d处死大鼠，以梗死灶为中心切片，Brdu免疫组织化学染色，观察移植神经干细胞的存活与迁移。

结果与结论：分离培养的表皮生长因子反应性细胞表达nestin抗原，并有自我更新及多向分化潜能，在胎牛血清诱导下可分化为神经胶质细胞和神经元。经Brdu标记后，85%以上神经干细胞表达Brdu抗原。移植28 d内顶核刺激+神经干细胞移植组功能评分明显优于与其他3组(P < 0.05~0.01)。顶核刺激+神经干细胞移植组在移植14，28 d存活细胞数明显多于单纯神经干细胞移植组；提示电刺激小脑顶核可显著提高移植神经干细胞的存活率及大脑中动脉梗死大鼠的神经功能评分；优化的整体环境可提高移植神经干细胞对局灶性脑缺血损伤细胞的替代作用。

关键词: 电刺激, 小脑顶核, 神经干细胞, 移植, 存活, 迁移

Abstract:

BACKGROUND: The discrepancy of neural stem cells (NSCs) transplantation is low cell survival rate and poor differentiation, which can not relieve the large apoptosis of neuron and endothelial cells in ischemic penumbra. This paper proposes a principle of overall intervention to provide an optimal environment for transplanted cells. 

OBJECTIVE: To observe the effect of cerebellar fastigial nucleus electrical stimulation (FNS) on survival and migration of NSCs.

METHODS: The EGF-responsive hippocampal NSCs of neonate Wistar rats were isolated and cultured in vitro. They were labeled by Brdu and induced by embryo cattles blood serum. The multi-differentiation potential was studied. Totally 80 rats were assigned into 4 groups. FNS+NSCs transplantation (n=32): at 24 hours FNS, right middle cerebral artery occlusion (MCAO) was prepared and received NSCs transplantation. FNS group (n =8): received the same procedure as FNS+NSCs transplantation except substitute PBS for NSCs. NSCs transplantation group (n=32): concentric electrode was inserted without electrify, the remained procedure was the same to FNS+NSCs transplantation group. Control group (n=8): concentric electrode was inserted without electrify, the remained procedure was the same to FNS group. The neural functions were recorded at hours 6, 24 and days 3, 7, 14, 28 after infarction. Rats were sacrificed at days 3, 7, 14 and 28 after transplantation, and the survival and migration of NSCs were investigated by Brdu immunocytochemical staining.

RESULTS AND CONCLUSION: Isolated and purified epidermal growth factor-responsive hippocampal NSCs were Nestin-positive and had ability of self-renewing and multi-directional differentiation. More than 85% NSCs expressed the antigen of Brdu after Brdu labeled. The functional scores of the FNS+NSCs transplantation group were significant better than those of the other 3 groups at 28 days after transplantation (P < 0.05-0.01). The number of survival cells in the FNS+NSCs transplantation group was significantly greater than that of NSCs transplantation group at days 14 and 28 after transplantation. FNS can improve the survival rate of transplanted NSCs and the functional scores following MCAO. The grading-up global environment can improve the substitution role of transplanted NSCs at local cerebral infarction damage.

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