中国组织工程研究

中国组织工程研究 ›› 2012, Vol. 16 ›› Issue (10): 1803-1807.doi: 10.3969/j.issn.1673-8225.2012.10.021

• 干细胞培养与分化 stem cell culture and differentiation • 上一篇    下一篇

小分子干扰RNA介导的RhoA沉默优化胎肝干细胞培养*★

赵  戈，刘卫辉，王  涛，王  兴，夏  宁，杨  平，寇明文，张  宁，陶开山   

  1. 解放军第四军医大学西京医院肝胆外科，陕西省西安市  710032
  • 收稿日期:2011-12-15 修回日期:2012-01-31 出版日期:2012-03-04 发布日期:2012-03-04
  • 通讯作者: 陶开山，解放军第四军医大学西京医院肝胆外科，陕西省西安市 710032 taokaishan@yahoo.com.cn
  • 作者简介:赵戈★，男，1982年生，吉林省永吉县人，汉族，2007年河北医科大学毕业，解放军第四军医大学在读硕士，医师，主要从事胎肝干细胞增殖分化研究。zhaoge1982119@163.com
  • 基金资助:

    国家自然科学基金面上项目，项目号：81170419。

Small interfering RNA-mediated RhoA silencing optimizes the culture of fetal liver stem cells 

Zhao Ge, Liu Wei-hui, Wang Tao, Wang Xing, Xia Ning, Yang Ping, Kou Ming-wen, Zhang Ning, Tao Kai-shan   

  1. Department of Hepatobiliary Surgery, Xijing Hospital, Fourth Military Medical University, Xi’an   710032, Shaanxi Province, China
  • Received:2011-12-15 Revised:2012-01-31 Online:2012-03-04 Published:2012-03-04
  • Contact: author: Tao Kai-shan, Department of Hepatobiliary Surgery, Xijing Hospital, Fourth Military Medical University, Xi’an 710032, Shaanxi Province, China taokaishan@yahoo.com.cn
  • About author:Zhao Ge★, Studying for master’s degree, Physician, Department of Hepatobiliary Surgery, Xijing Hospital, Fourth Military Medical University, Xi’an 710032, Shaanxi Province, China zhaoge1982119@ 163.com
  • Supported by:

    the National Natural Science Foundation of China, No. 81170419*

PDF

300

摘要：

背景：细胞移植治疗是目前的研究热点之一，寻找良好的细胞来源并有效扩增，是大量临床应用的基础。
目的：通过沉默RhoA基因优化胎肝干细胞培养方法。
方法：体外培养胎肝干细胞，经小分子干扰RNA转染以沉默RhoA基因表达，分为3组：A组为干细胞组，B组为经随机打乱顺序的小分子干扰RNA转染干细胞组，C组为经小分子干扰RNA转染的干细胞组。应用反转录-聚合酶链反应、Western blot法检测胎肝干细胞在转染前后RhoA基因和蛋白的表达；四甲基偶氮唑蓝比色法观察细胞生长的优化作用；流式细胞术测定细胞周期分布的变化。
结果与结论：转染小分子干扰RNA后C组与A、B组相比，RhoA基因和蛋白表达量明显降低(P < 0.05)，细胞的生长速度明显增快，细胞周期G0/G1期减少，S期细胞数增多，各组间差异有显著性意义(P < 0.05)。提示通过沉默RhoA基因的方法可以促进胎肝干细胞增殖，对其培养有优化作用。
关键词：胎肝干细胞；细胞培养；RhoA；RNA干扰；细胞增殖
doi:10.3969/j.issn.1673-8225.2012.10.021

关键词: 胎肝干细胞, 细胞培养, RhoA, RNA干扰, 细胞增殖

Abstract:

BACKGROUND: Cell transplantation for disease treatment is one of interesting topics. To find a better source of seed cells and a more efficient amplification method is the basis for clinical application.
OBJECTIVE: To determine whether the RhoA gene silencing in fetal liver stem cells can optimize liver stem cell cultures.
METHODS: Fetal liver stem cells were in vitro cultured and were transfected by small interfering RNA (siRNA) to knock down the expression of RhoA. Then the cells were randomly divided into three groups. In the group A, fetal liver stem cells were non-treated. In the group B, fetal liver stem cells were transfected with randomly sequenced siRNA. In the group C, fetal liver stem cells were transfected with siRNA. RhoA gene and protein expression prior to and after transfection was detected by RT-PCR and western blot methods, respectively. Cellular proliferation was determined by MTT assay. The cell cycle was analyzed by flow cytometry.
RESULTS AND CONCLUSION: Compared with groups A, B, the expression of RhoA gene and protein in the group C was significantly decreased (P < 0.05), the cell growth speed was significantly increased, cells in the G0/G1 phase of the cell cycle were decreased, and cells in the S phase were increased (P < 0.05). These findings suggest that the RhoA gene silencing can promote the proliferation of fetal liver stem cells and optimize the culture of fetal liver stem cells. 

中图分类号: