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Non-genetic Purification of Ventricular Cardiomyocytes from Differentiating Embryonic Stem Cells through Molecular Beacons Targeting IRX-4

Authors
 Kiwon Ban  ;  Brian Wile  ;  Kyu-Won Cho  ;  Sangsung Kim  ;  Ming-Ke Song  ;  Sang Yoon Kim  ;  Jason Singer  ;  Anum Syed  ;  Shan Ping Yu  ;  Mary Wagner  ;  Gang Bao  ;  Young-sup Yoon 
Citation
 STEM CELL REPORTS, Vol.5(6) : 1239-1249, 2015 
Journal Title
 STEM CELL REPORTS 
Issue Date
2015
MeSH
Action Potentials ; Animals ; Base Sequence ; Cell Differentiation ; Cell Separation/methods* ; Cells, Cultured ; Embryonic Stem Cells/cytology* ; Flow Cytometry ; Heart Ventricles/cytology* ; Homeodomain Proteins/genetics* ; Mice ; Myocytes, Cardiac/cytology* ; Oligonucleotide Probes/genetics ; RNA, Messenger/genetics
Keywords
IRX4 ; cell selection ; molecular beacons ; pluripotent stem cell ; ventricular cardiomyocyte
Abstract
Isolation of ventricular cardiomyocytes (vCMs) has been challenging due to the lack of specific surface markers. Here we show that vCMs can be purified from differentiating mouse embryonic stem cells (mESCs) using molecular beacons (MBs) targeting specific intracellular mRNAs. We designed MBs (IRX4 MBs) to target mRNA encoding Iroquois homeobox protein 4 (Irx4), a transcription factor specific for vCMs. To purify mESC vCMs, IRX4 MBs were delivered into cardiomyogenically differentiating mESCs, and IRX4 MBs-positive cells were FACS-sorted. We found that, of the cells isolated, ~98% displayed vCM-like action potentials by electrophysiological analyses. These MB-purified vCMs continuously maintained their CM characteristics as verified by spontaneous beating, Ca(2+) transient, and expression of vCM-specific proteins. Our study shows the feasibility of isolating pure vCMs via cell sorting without modifying host genes. The homogeneous and functional ventricular CMs generated via the MB-based method can be useful for disease investigation, drug discovery, and cell-based therapies.
DOI
10.1016/j.stemcr.2015.10.021
Appears in Collections:
1. College of Medicine (의과대학) > BioMedical Science Institute (의생명과학부) > 1. Journal Papers
Yonsei Authors
Yoon, Young Sup(윤영섭)
URI
https://ir.ymlib.yonsei.ac.kr/handle/22282913/156951
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