An improved method for generating integration-free human induced pluripotent stem cells.
10.7534/j.issn.1009-2137.2014.03.002
- Author:
Shu-Ping LIU
1
;
Yan-Xin LI
2
;
Jing XU
1
;
Hai-Hui GU
1
;
Hong-Yan ZHANG
1
;
Hao-Yue LIANG
1
;
Han-Zhi LIU
1
;
Xiao-Bing ZHANG
1
;
Tao CHENG
1
;
Wei-Ping YUAN
1
Author Information
1. State Key Laboratory of Experimental Hematology, Institute of Hematology and Blood Diseases Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin 300020, China.
2. State Key Laboratory of Experimental Hematology, Institute of Hematology and Blood Diseases Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin 300020, China. E-mail:liyanxincau@gmail.com.
- Publication Type:Journal Article
- MeSH:
Cell Culture Techniques;
methods;
Cellular Reprogramming;
Genetic Vectors;
Humans;
Induced Pluripotent Stem Cells;
cytology;
Plasmids;
Transfection
- From:
Journal of Experimental Hematology
2014;22(3):580-587
- CountryChina
- Language:Chinese
-
Abstract:
The genome instability and tumorigenicity of induced pluripotent stem cells (iPSC) hinder their great potentials for clinical application. Using episomal vectors to generate iPSC is the best way to solve safety issues at present. This method is simple and the exogenous gene was not integrated into the host genome. However, the reprogramming efficiency for this method is very low and thus limits its usage. This study was purposed to improve episomal method for generating induced pluripotent stem cells from cord blood mononuclear cells (CB MNC), to establish integration-free iPSC technology system, and to lay the foundation for individualized iPSC for future clinical uses. To improve the reprogramming efficiency for iPSC, episomal method was used at various combinations of episomal vectors, pre-stimulating culture mediums and oxygen condition were tested to optimize the method. The results showed that using erythroid culture medium for culturing 8 days, transfecting with episomal vectors with SFFV (spleen focus forming virus) promoter under the hypoxic condition (3%), CB MNC could be mostly efficiently reprogrammed with the efficiency 0.12%. Furthermore, the results showed that erythroblasts (CD36(+)CD71(+)CD235a(low)) were the cells that are reprogrammed with high efficiency after culture for 8 days. It is concluded that a highly efficient and safe method for generation of integration-free iPSC is successfully established, which is useable in clinical study.
