天津医药

天津医药 ›› 2017, Vol. 45 ›› Issue (1): 21-24.doi: 10.11958/1089

• 实验研究 • 上一篇    下一篇

人脐带间充质干细胞源的多巴胺分泌细胞治疗 帕金森病模型大鼠的研究

谢姜 1， 2， 王斌 1， 董方 1， 张宏炜 1， 张力伟 1， 白钢 2△， 刘拥军 1△   

  1. 1 天津市干细胞再生医学转化企业重点实验室， 天津和泽干细胞科技有限公司博士后科研工作站（邮编 300304）； 2 南开大学 药学院， 南开大学博士后流动站
  • 收稿日期:2016-10-09 修回日期:2016-11-21 出版日期:2017-01-15 发布日期:2017-01-15
  • 通讯作者: 白钢，刘拥军 E-mail:bioprs_ts@163.com
  • 基金资助:
    干细胞药研制技术平台建设;脐带间充质干细胞抗肝纤维化制剂

Study of dopamine-producing cells derived from human umbilical cord mesenchymal stem cells in treatment in rats with Parkinson’ s disease

XIE Jiang1,2, WANG Bin1, DONG Fang1, ZHANG Hong-wei1, ZHANG Li-wei1, BAI Gang2△, LIU Yong-jun1△   

  1. 1 Alliancells Key Institute of Stem Cells and Translational Regenerative Medicine, Tianjin Allian Stem Cell Biotech Co., Ltd., Post-Doctoral Research Station, Tianjin 300304, China; 2 College of Pharmacy, Nankai University, Nankai University Post-Doctoral Programme
  • Received:2016-10-09 Revised:2016-11-21 Published:2017-01-15 Online:2017-01-15

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摘要： 摘要： 目的 探讨人脐带间充质干细胞（hUC-MSCs）源的多巴胺分泌细胞（hUC-MSCs-DPCs）对帕金森病（PD） 模型大鼠的疗效及机制。方法 体外培养并诱导 hUC-MSCs 分化为 DPCs， 采用 ELISA 法通过检测多巴胺 （DA）、 胶 质细胞源性神经营养因子 （GDNF） 表达水平对诱导后的 DPCs 进行鉴定。SD 大鼠大脑右侧黑质区注射 6-羟基多巴 胺（6-OHDA）制备 PD 模型。60 只 PD 模型鼠随机分为 hUC-MSCs 组（移植 5×105个 hUC-MSCs 至纹状体， 20 只）、 hUC-MSCs-DPCs 组（移植 5×105个 hUC-MSCs-DPCs， 20 只）、 对照组［注射等体积磷酸盐缓冲液（PBS）， 20 只）］， 采 用 CM-Dil 标记移植细胞。移植后 4、 8 和 12 周， 皮下注射阿扑吗啡， 观察其诱导的旋转行为； ELISA 法检测 3 组 GDNF 表达情况。移植后 12 周处死大鼠， 制备脑冰冻切片， 免疫荧光染色观察酪氨酸羟化酶（TH）表达情况， FLUORO-JADE® C 染色观察神经元生长情况。结果 hUC-MSCs-DPCs 诱导成功并且高表达 DA 和 GDNF。移植 后 4、 8、 12 周， 相比对照组和 hUC-MSCs 组， hUC-MSCs-DPCs 组大鼠旋转行为明显改善， GDNF 表达量明显升高 （P＜0.05）。移植后 12 周， hUC-MSCs-DPCs 组大鼠毁损侧纹状体以及双侧黑质区可见 TH+细胞， 变性神经元数目较 对照组明显减少 （P＜0.05）。结论 hUC-MSCs-DPCs 可改善阿扑吗啡诱发的 PD 模型大鼠的旋转行为， 这可能与提 高毁损侧纹状体内 DA、 GDNF 表达水平， 保护神经元有关。

关键词: 间质干细胞, 脐血干细胞移植, 帕金森病, 多巴胺, 纹状体黑质变性, 大鼠, Sprague-Dawley, 胶质细胞源 性神经营养因子

Abstract: Abstract: Objective To investigate the efficacy of dopamine producing cells (DPCs) derived from human umbilical cord- derived mesenchymal stem cells (hUC- MSCs) in treatment in model rats with Parkinson’ s disease (PD). Methods The cultured hUC-MSCs were induced into DPCs in vitro. The dopamine (DA) and glial cell line-derived neurotrophic factor (GDNF) expression levels were detected by ELISA to identify the DPCs. The PD rat model was established by injecting 6- OHDA into the right substantia nigra (SN). A total of 60 successfulled PD model rats were randomized into hUC-MSCsDPCs group (5×105 hUC-MSCs-DPCs were transplanted into right striatum, n=20), hUC-MSCs group (5×105 hUC-MSCs were transplanted, n=20) and control group (same volume PBS, n=20). All the transplanted cells were labeled with CM-Dil. The apomorphine induced rotation behavior was assessed at 4, 8 and 12 weeks after cell transplantation. The rats were executed after 12 weeks. The immunofluorescence staining was used to detect the tyrosine hydroxylase (TH) expression level, and FLUORO-JADE® C staining was used to test the apoptotic neurons in brain of rats. Results The hUC-MSCs-DPCs were induced successfully in vitro, which showed a high expression level of DA and GDNF. Furthermore, at 4, 8 and 12 weeks after cell transplantation, the rotation behavior was improved, and expression levels of GDNF were significantly higher in hUC-MSCs-DPCs group than those of hUC-MSCs group and control group (P＜0.05). In addition, we found that most of the transplanted TH + hUC-MSCs-DPCs at the right striatum and a few cells around both the left and the right substantia nigra at 12 weeks after transplantation. The apoptotic neurons were decreased after cell transplantation in hUC-MSCs-DPCs group than that of control group (P＜0.05). Conclusion The hUC- MSCs- DPCs can improve the rotational behavior induced by apomorphine in PD model rats, which may be involved in improving levels of DA and GDNF in damaged striatum and protecting neurons.

Key words: mesenchymal stem cells, cord blood stem cell transplantation, Parkinson’s disease, dopamine, striatonigral degeneration, rats, Sprague-Dawley, Glial cell line-derived neurotrophic factor