中国组织工程研究

中国组织工程研究 ›› 2010, Vol. 14 ›› Issue (14): 2539-2544.doi: 10.3969/j.issn.1673-8225.2010.14.014

• 干细胞移植 stem cell transplantation • 上一篇    下一篇

神经干细胞NgR基因沉默立体定向移植治疗大鼠脑损伤

王  东，张建军，马景鑑   

  1. 天津医科大学总医院，天津市  300052
  • 出版日期:2010-04-02 发布日期:2010-04-02
  • 通讯作者: 马景鑑，博士生导师，天津医科大学总医院，天津市 300052
  • 作者简介:王 东★，男，1971年生，河北省乐亭县人，汉族，天津医科大学在读硕士，副主任医师，主要从事脑外伤，脊髓损伤的研究。 5609@hotmail.com

Stereotaxic intracerebral transplantation of neural stem cells with Nogo-66 receptor gene silencing for treating traumatic brain injury in rats 

Wang Dong, Zhang Jian-jun, Ma Jing-jian   

  1. General Hospital, Tianjin Medical University, Tianjin   300052, China
  • Online:2010-04-02 Published:2010-04-02
  • Contact: Ma Jing-jian, Doctoral supervisor, General Hospital, Tianjin Medical University, Tianjin 300052, China
  • About author:Wang Dong★, Studying for master’s degree, Associate chief physician, General Hospital, Tianjin Medical University, Tianjin 300052, China 5609@hotmail.com

PDF

441

被引次数

23

摘要：

背景：神经干细胞具有自我增殖能力和多向分化潜能，一定条件下可以分化成神经系统的各种细胞，因此在神经损伤修复方面有着良好的应用前景。而RNA干扰避免了永久基因沉默的弊病，最有希望与神经干细胞移植相结合治疗颅脑损伤。
目的：检测是否可以通过沉默NgR基因的方法提高神经干细胞立体定向移植对重型颅脑损伤大鼠的治疗效果。
方法：60只雄性Wistar大鼠制成重型液压颅脑损伤模型后随机区组法分为3组，每组20只。实验组：造模24 h后向损伤的大鼠脑组织内注射NgR基因沉默的神经干细胞悬液6 μL；对照组：同法注射等量的神经干细胞悬液；空白组：同法注射等量的不含干细胞的培养液。伤后24 h，3 d，1，2周行动物神经学缺损评分。2周后处死行免疫组织化学和苏木精-伊红染色。
结果与结论：转染小分子干扰RNA后，与对照组相比，实验组NgR基因蛋白表达量明显降低，移植后1周和2周，接受神经干细胞移植的大鼠神经学缺损评分明显低于对照组(P < 0.05)；且其脑组织切片中的神经元数量较对照组明显增多(P < 0.01)。伤后2周苏木精-伊红染色空白组可见损伤处脑组织断裂，为瘢痕连接，有明显空洞形成；对照组在移植部位出现典型的神经细胞样形态学改变；实验组出现典型的神经细胞样形态学改变且空洞消失。免疫组织化学染色观察空白组BrdU标记的阳性细胞为(37.92±16.02)个/高倍视野，对照组为(89.68±15.34)个/高倍视野，实验组为(102.67±13.52)个/高倍视野，各组间两两比较，差异均有显著性意义(P < 0.01)。提示神经干细胞NgR基因沉默后立体定向移植治疗大鼠脑损伤可明显改善重型颅脑损伤后大鼠的神经学功能。

关键词: 神经干细胞, NgR, RNA干扰, 脑内移植, 颅脑损伤

Abstract:

BACKGROUND: Neural stem cells (NSCs) have the potential of self-proliferation and multiple directional differentiation, and can differentiate into various cells in the neural system under a certain condition. Therefore, NSCs have good prospect in repair of nerve injury. However, RNA interference avoids the abuse of permanent gene silencing, and is hopeful to combine with NSC transplantation for treating craniocerebral injury.
OBJECTIVE: To determine whether the Nogo-66 receptor (NgR) gene silencing in NSCs can enhance curative effects of stereotaxic intracerebral transplantation of NSCs on traumatic brain injury (TBI) in rats.
METHODS: A total of 60 male Wistar rats following TBI establishment were randomly assigned to 3 groups (n = 20). In the experimental group, NgR gene silencing NSC suspension (6 μL) was injected into rat brain tissue following 24 hours of model induction. In the control group, an equal volume of NSC suspension was infused by the same method. In the blank group, an equal volume of medium without stem cells was infused by the same method. At 24 hours, 3 days, 1 and 2 weeks following injury, neurological deficits were scored. Two weeks later, animals were sacrificed and subjected to immunohistochemistry and hematoxylin-eosin staining.
RESULTS AND CONCLUSION: Following transfection of small interfering RNA, compared with control group, NgR gene protein expression was significantly reduced in the experimental group. At 1 and 2 weeks following transplantation, neurological deficit score was significantly less in animals undergoing NSC transplantation in the experimental group compared with the control group (P < 0.05). Moreover, neuron number in the brain tissue sections of experimental group was significantly more than in the control group (P < 0.01). At 2 weeks following injury, hematoxylin-eosin staining showed that brain tissue breakage at damaged site as scar connection, remarkable porosis in the blank group; typical morphological changes as neural cells at the transplanted site in the control group; typical morphological changes as neural cells without cavity in the experimental group. Immunohistochemistry showed (37.92±16.02) BrdU-labeled positive cells/high-power field in the blank group, (89.68±15.34) cells/high-power field in the control group, and (102.67±13.52) cells/high-power field in the experimental group. Significant differences were detected between groups (P < 0.01). The above-mentioned results indicated that the NSCs of NgR gene silencing transplanted into the injured cerebral tissues can significantly improve the neurological function in rats with TBI.

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