Using low-risk factors to generate non-integrated human induced pluripotent stem cells from urine-derived cells

Qiang Li,Libing Song,Bin Wang,Linli Wang,Wei Wu,Zhiju Zhao,Yuehua Chen,Jianguo Yang,Xiaohui Yang,Jian Mo,Jun Li,Chunyan Guan

doi:10.1186/s13287-017-0698-8

Abstract

BackgroundBecause the lack of an induced pluripotent stem cell (iPSC) induction system with optimal safety and efficiency limits the application of these cells, development of such a system is important.MethodsTo create such an induction system, we screened a variety of reprogrammed plasmid combinations and multiple compounds and then verified the system’s feasibility using urine cells from different individuals. We also compared large-scale iPSC chromosomal variations and expression of genes associated with genomic stability between this system and the traditional episomal system using karyotype and quantitative reverse transcription polymerase chain reaction analyses.ResultsWe developed a high-efficiency episomal system, the 6F/BM1-4C system, lacking tumorigenic factors for human urine-derived cell (hUC) reprogramming. This system includes six low-risk factors (6F), Oct4, Glis1, Klf4, Sox2, L-Myc, and the miR-302 cluster. Transfected hUCs were treated with four compounds (4C), inhibitor of lysine-demethylase1, methyl ethyl ketone, glycogen synthase kinase 3 beta, and histone deacetylase, within a short time period. Comparative analysis revealed significantly decreased chromosomal variation in iPSCs and significantly increased Sirt1 expression compared with iPSCs induced using the traditional episomal system.ConclusionThe 6F/BM1-4C system effectively induces reprogramming of urine cells in samples obtained from different individuals. iPSCs induced using the 6F/BM1-4C system are more stable at the cytogenetic level and have potential value for clinical application.

Highlights

Because the lack of an induced pluripotent stem cell induction system with optimal safety and efficiency limits the application of these cells, development of such a system is important
When we used the xeno-free Human embryonic stem cell (hESC) E8 medium [38] to induce human urine-derived cell (hUC) reprogramming based on episomal vectors, we found it to be unsuitable after the addition of certain compounds, with deformed cells all dying
We developed a safe method based on an episomal vector for inducing induced pluripotent stem cell (iPSC) from hUCs

Summary

Introduction

Because the lack of an induced pluripotent stem cell (iPSC) induction system with optimal safety and efficiency limits the application of these cells, development of such a system is important. Human iPSCs (hiPSCs) are primarily induced using retroviral or lentiviral vectors carrying reprogramming factors [4, 5], and exogenous DNA fragments can randomly insert into genomic DNA and induce cell transformation, thereby preventing the clinical application of iPSCs. To date, many non-integrating methods have been generated, including mRNA and protein transfection [6, 7], Sendai virus (SeV) [8], piggyback (PB) transposons [9], and episomal vectors [10]. As induction efficiency varies when the same method is used to reprogram different types of somatic cells or when different methods are applied to reprogram the same type of somatic

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Discussion

Conclusion