中国组织工程研究

中国组织工程研究 ›› 2012, Vol. 16 ›› Issue (19): 3551-3555.doi: 10.3969/j.issn.1673-8225.2012.19.027

• 干细胞转基因表达 transgenic expression in stem cells • 上一篇    下一篇

RNA干扰抑制骨髓间充质干细胞中caspase-3基因的表达**★

刘立宝1，华  平1，杨淞然2，刘家良1，江慧琦1，王  萌1，曾  宽1，敖  翔1，杨艳旗1   

  1. 1中山大学孙逸仙纪念医院心胸外科，广东省广州市 510120；2广州市第一人民医院脑内科，广东省广州市  510180
  • 收稿日期:2011-10-30 修回日期:2012-01-28 出版日期:2012-05-06 发布日期:2012-05-06
  • 通讯作者: 华平，博士，副教授，副主任医师，硕士生导师，中山大学孙逸仙纪念医院心胸外科，广东省广州市 510120 huaping88@sina.com
  • 作者简介:刘立宝★，男，1983年生，安徽省凤阳县人，汉族，中山大学孙逸仙纪念医院心胸外科在读硕士，主要从事干细胞治疗心肌缺血方面的研究。hefeiliulibao@sina.com
  • 基金资助:

    广东省自然科学基金资助项目(7001619)；广东省科技计划项目(2007B050200011)。

RNA interference inhibits caspase-3 gene expression in bone marrow mesenchymal stem cells   

Liu Li-bao1, Hua Ping1, Yang Song-ran2, Liu Jia-liang1, Jiang Hui-qi1, Wang Meng1, Zeng Kuan1, Ao Xiang1, Yang Yan-qi1   

  1. 1Department of Cardiothoracic Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, Guangdong Province, China; 2Department of Neurology, Guangzhou First Municipal People’s Hospital, Guangzhou 510180, Guangdong Province, China
  • Received:2011-10-30 Revised:2012-01-28 Online:2012-05-06 Published:2012-05-06
  • Contact: Hua Ping, M.D., Associate professor, Associate chief physician, Master’s supervisor, Department of Cardiothoracic Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, Guangdong Province, China huaping88@sina.com
  • About author:Liu Li-bao★, Studying for master’s degree, Department of Cardiothoracic Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, Guangdong Province, China
  • Supported by:

     the Natural Science Foundtion of Guangdong Province, No. 7001619*; the Social Development Science and Technology Program of Guangdong Province, No. 2007B050200011*

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摘要：

背景：研究发现，骨髓间充质干细胞移植入心肌梗死区域后易发生凋亡导致移植细胞存活率较低。
目的：构建caspase-3基因的小分子干扰RNA片段(small interfering RNAs，siRNAs)表达质粒，探讨其在体外对骨髓间充质干细胞中caspase-3基因表达的抑制作用。
方法：根据siRNA设计原则，针对caspase-3基因设计并合成siRNAs，构建caspase-3siRNA表达载体并利用脂质体介导方法瞬时转染骨髓间充质干细胞。实验分为正常对照组，阴性序列对照组(si-ncg)和3个干扰组(si-caspase-3-1、si-caspase-3-2和si-caspase-3-3)。
结果与结论：成功构建了caspase-3siRNA表达载体并转染骨髓间充质干细胞。Western blot和RT-PCR检测转染后的骨髓间充质干细胞中caspase-3蛋白和mRNA的表达下降，以转染si-caspase-3-3的骨髓间充质干细胞下降最明显。结果提示构建的caspase-3siRNA表达质粒在体外能抑制骨髓间充质干细胞中caspase-3基因的表达。
 

关键词: 骨髓间充质干细胞, 凋亡, caspase-3, RNA干扰, 抑制效应

Abstract:

BACKGROUND: Poor viability of bone mesenchymal stem cells after transplantation into the infarcted region has limited the reparative capacity of these cells because of the apoptosis.
OBJECTIVE: To construct eukaryotic vector expressing small interfering RNAs against caspase-3 and investigate the inhibitory effect on the expression of caspase-3 in BMSCs in vitro.
METHODS: According to the principle of siRNA design, siRNAs against caspase-3 gene were designed and synthesized. Eukaryotic vector expressing caspase-3siRNA was constructed and transferred into BMSCs with the liposome conduct. There were 5 groups, including normal cell group, negative sequence group and 3 interfering groups (si-caspase-3-1, si-caspase-3-2 and si-caspase-3-3).
RESULTS AND CONCLUSION: Eukaryotic vector expressing caspase-3siRNA was constructed and transferred into BMSCs successfully. Western blot analysis and reverse-transcription polymerase chain reaction demonstrated that levels of of caspase-3 protein and mRNA in transfected BMSCs decreased, with the most significant decline in the group with si-caspase-3-3. Eukaryotic vector expressing caspase-3 siRNA could inhibit the expression of caspase-3 gene in BMSCs in vitro.

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