c-Met基因过表达脐带间充质干细胞株的构建*

doi:10.3969/j.issn.1672-5069.2018.03.020

实用肝脏病杂志 ›› 2018, Vol. 21 ›› Issue (3): 397-400.doi: 10.3969/j.issn.1672-5069.2018.03.020

c-Met基因过表达脐带间充质干细胞株的构建*

张永婷, 朱传龙, 李军, 章莉莉

210029 南京市南京医科大学第一附属医院感染病科

收稿日期:2017-10-09 出版日期:2018-05-10 发布日期:2018-05-25
通讯作者: 朱传龙,E-mail:zhuchuanlong@jsph.org.cn ;章莉莉,E-mail: dr.zhanglili@aliyun.com
作者简介:张永婷,女,26岁,硕士研究生。主要从事病毒性肝炎防治研究
基金资助:
*国家自然科学基金资助项目（编号：81770591）; 江苏省高层次卫生人才“六个一工程”资助项目（编号：LGY2016005）

Construction of overexpression of c-met in human umbilical cord mesenchymal stem cells with lenti-vector transfection in vitro

Zhang Yongting, Zhu Chuanlong, Li Jun, et al

Department of Infectious Disease,First Affiliated Hospital,Nanjing Medical University,Nanjing 210029,Jiangsu Province,China

Received:2017-10-09 Online:2018-05-10 Published:2018-05-25

摘要/Abstract

摘要： 目的建立c-Met慢病毒载体转染人脐带间充质干细胞（hUMSC）,为治疗肝衰竭作为种子细胞。方法培养hUMSC,经流式细胞技术检测细胞表面表型,转染c-Met慢病毒载体,在荧光显微镜下观察转染效率,确定最佳多重感染复数（MOI）。采用嘌呤霉素抗性筛选稳定表达c-Met的hUMSC细胞系,采用Western-blot法检测细胞c-met蛋白表达量。结果 P5代细胞高表达CD44、CD90和CD105抗原,不表达CD31、CD45和CD34相关造血细胞抗原,符合人脐带间充质干细胞的特性;构建成功的c-Met慢病毒载体转染人脐带间充质干细胞最佳MOI=80,经嘌呤霉素筛药,在荧光显微镜下观察发现荧光阳性率为100%,且经Western-blot法检测证实该细胞过表达c-Met蛋白。结论成功构建过表达c-Met基因的脐带间充质干细胞,为进一步实验打下了基础。

关键词: 人脐带间充质干细胞, c-Met基因, 转染, 体外

Abstract: Objective To construct the overexpression of c-met in human umbilical cord mesenchymal stem cells（hUMSCs） with lenti-vector transfection in vitro. Methods The phenotype of hUMSCs was determined by Flow cytometry analysis, and the hUMSCs were transfected by different multiple of infection（MOI with c-met lenti-vector. The optimal MOI was determined under fluorescence microscopy. The c-Met protein was detected by Western blot. Results P5 gegeration of hUMSCs were CD44/CD90/CD105-positive,and CD31/CD45/CD34-negative, indicating the hUMSCs were obtained with specific surface markers. The optimal MOI was 80, and after puromycin screening,the transfected efficiency was up to 100%. The c-Met protein was overexpressed in the transfected hUMSCs by Western blot detection. Conclusion c-Met overexpressing hUMSCs were successfully constructed, which might be beneficial for further study.

Key words: Human umbilical cord mesenchymal stem cells, c-Met, Transduction, In vitro

张永婷, 朱传龙, 李军, 章莉莉. c-Met基因过表达脐带间充质干细胞株的构建*[J]. 实用肝脏病杂志, 2018, 21(3): 397-400.

Zhang Yongting, Zhu Chuanlong, Li Jun, et al. Construction of overexpression of c-met in human umbilical cord mesenchymal stem cells with lenti-vector transfection in vitro[J]. JOURNAL OF PRACTICAL HEPATOLOGY, 2018, 21(3): 397-400.

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c-Met基因过表达脐带间充质干细胞株的构建*

Construction of overexpression of c-met in human umbilical cord mesenchymal stem cells with lenti-vector transfection in vitro

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