Efficient human iPS cell derivation by a non-integrating plasmid from blood cells with unique epigenetic and gene expression signatures

被引:336
作者
Chou, Bin-Kuan [1 ,2 ]
Mali, Prashant [1 ,3 ]
Huang, Xiaosong [1 ,4 ]
Ye, Zhaohui [1 ,4 ]
Dowey, Sarah N. [1 ,4 ]
Resar, Linda M. S. [4 ]
Zou, Chunlin [1 ,5 ,6 ]
Zhang, Y. Alex [5 ,6 ]
Tong, Jay [7 ]
Cheng, Linzhao [1 ,2 ,4 ]
机构
[1] Johns Hopkins Univ, Sch Med, Stem Cell Program, Inst Cell Engn, Baltimore, MD 21205 USA
[2] Johns Hopkins Univ, Sch Med, Grad Program Cellular & Mol Med, Baltimore, MD 21205 USA
[3] Johns Hopkins Univ, Sch Med, Grad Program Biomed Engn, Baltimore, MD 21205 USA
[4] Johns Hopkins Univ, Sch Med, Dept Med, Div Hematol, Baltimore, MD 21205 USA
[5] Capital Med Univ, Beijing, Peoples R China
[6] Xuanwu Hosp, Cell Therapy Ctr, Beijing, Peoples R China
[7] All Cells LLC, Emeryville, CA USA
关键词
iPS Cells; reprogramming; cord blood; episomal vectors; epigenetics; DNA methylation; sickle cell disease; PLURIPOTENT STEM-CELLS; GENERATION;
D O I
10.1038/cr.2011.12
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
To identify accessible and permissive human cell types for efficient derivation of induced pluripotent stem cells (iPSCs), we investigated epigenetic and gene expression signatures of multiple postnatal cell types such as fibroblasts and blood cells. Our analysis suggested that newborn cord blood (CB) and adult peripheral blood (PB) mononuclear cells (MNCs) display unique signatures that are closer to iPSCs and human embryonic stem cells (ESCs) than agematched fibroblasts to iPSCs/ESCs, thus making blood MNCs an attractive cell choice for the generation of integration-free iPSCs. Using an improved EBNA1/OriP plasmid expressing 5 reprogramming factors, we demonstrated highly efficient reprogramming of briefly cultured blood MNCs. Within 14 days of one-time transfection by one plasmid, up to 1000 iPSC-like colonies per 2 million transfected CB MNCs were generated. The efficiency of deriving iPSCs from adult PB MNCs was approximately 50-fold lower, but could be enhanced by inclusion of a second EBNA1/OriP plasmid for transient expression of additional genes such as SV40 T antigen. The duration of obtaining bona fide iPSC colonies from adult PB MNCs was reduced to half (similar to 14 days) as compared to adult fibroblastic cells (2830 days). More than 9 human iPSC lines derived from PB or CB blood cells are extensively characterized, including those from PB MNCs of an adult patient with sickle cell disease. They lack V(D)J DNA rearrangements and vector DNA after expansion for 10-12 passages. This facile method of generating integration-free human iPSCs from blood MNCs will accelerate their use in both research and future clinical applications.
引用
收藏
页码:518 / 529
页数:12
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