A microRNA-Based System for Selecting and Maintaining the Pluripotent State in Human Induced Pluripotent Stem Cells

Abstract

Induced pluripotent stem cell (iPSC) technology has provided researchers with a unique tool to derive disease-specific stem cells for the study and possible treatment of degenerative disorders with autologous cells. The low efficiency and heterogeneous nature of reprogramming is a major impediment to the generation of personalized iPSC lines. Here, we report the generation of a lentiviral system based on a microRNA-regulated transgene that enables for the efficient selection of mouse and human pluripotent cells. This system relies on the differential expression pattern of the mature form of microRNA let7a in pluripotent versus committed or differentiated cells. We generated microRNA responsive green fluorescent protein and Neo reporters for specific labeling and active selection of the pluripotent cells in any culture condition. We used this system to establish Rett syndrome and Parkinson's disease human iPSCs. The presented selection procedure represents a straightforward and powerful tool for facilitating the derivation of patient-specific iPSCs.

Synergetic Cooperation of microRNAs with Transcription Factors in iPS Cell Generation

“…These miRNA methods were more rapid and efficient than the use of OSKM [12], [28]. A miRNA-based system for selecting and maintaining the pluripotent state in human iPS cells has also been reported recently and represents a straightforward and powerful tool to facilitate the derivation of patient-specific iPS cells [29]. Further study of the specific roles and mechanisms of these signaling pathways in iPS generation and the regulation by candidate miRNAs of developmental signaling pathways will provide further experimental evidence regarding the specific role of miRNAs in iPS cell generation.…”

Glioblastomas and the Special Role of Adhesion Molecules in Their Invasion

“…Also, some miRNAs may cause chemosensitivity and radioresistance in GBM (Shi et al 2010;Chan et al 2012;Ujifuku et al 2010;Zhang et al 2012). A cancer stem cell (CSC)-specific miRNA expression profile has been identified in cancer stem/progenitor cells (Chu et al 2013;Di Stefano et al 2011;Gonzalez-Gomez et al 2011;Sharma and Wu 2013;Wilson et al 2009). Based on the abundant evidence for the role of miRNAs in biological mechanisms and tumorigenesis, currently there is a consensus that miRNAs are promising molecular markers and therapeutic agents that could be used in parallel to current GBM treatment (for details see Chap.…”

Functional Studies of microRNAs in Neural Stem Cells: Problems and Perspectives

“…Injection of a miR-124 reporter vector into the brain will separate reporter gene expression between different cell types such as neurons and astrocytes (Colin et al, 2009). The system has been used to segregate differentiated neural cells in pluripotent cell cultures, based on the expression of miR-292 that is expressed in embryonic stem cells but not in NSCs (Sachdeva et al, 2010), as well as the opposite where a miRNA let-7a reporter was used to select undifferentiated cells from more differentiated cells (Di Stefano et al, 2011). In NSCs in vivo , sensor vectors have been used to track the expression of miR-132 in the DG (Luikart et al, 2011).…”