Generation of non-integrating induced pluripotent stem cells with episomal plasmids and differentiation into neural stem cells
LIN Cheng-kai, RONG Li-ming, LIU Bin
Chinese Journal of Clinical Anatomy ›› 2016, Vol. 34 ›› Issue (6) : 647-654.
Generation of non-integrating induced pluripotent stem cells with episomal plasmids and differentiation into neural stem cells
Objective To study the induction of non-intergrated induced pluripotent stem cells (iPSCs) with episomal plasmid vectors, and differentiating into neural stem cells in vitro. Methods Non-integrating mouse iPSCs were induced from mouse embryonic fibroblasts (MEFs) with plasmid vectors pCEP4-EO2S-ET2K. For the differentiation of neural stem cells (NSCs), iPSCs were cultured in suspension and adherent cultural plate for 2 times with different mediums. The characterization of iPSCs and NSCs were then investigated. Results Our results demonstrated that mouse iPSC could be effectively generated and the pluripotency of iPSC was similar to that of mESCs. In addition, no vector integration was found in iPSCs of 10th passages. Quantitative real-time PCR showed that the neural stem cell marker gene expression was significantly improved in iPSC-derived NSCs, which was similar to that of wild-type NSCs. Immunofluorescence demonstrated that iPSC-derived NSCs expressed NESTIN and PAX6 and could differentiated into neurons, oligodendrocytes and astrocytes in vitro. Conclusions Non-integrated iPSCs could be generated with episomal plasmid vectors and differentiated into NSCs which could be the ideal seeding cells for the treatment of spinal cord injury.
Episomal plasmid vectors / Non-integrating induced pluripotent stem cells / Neural stem cells
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