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


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.

SUBMITTER: Ban K 

PROVIDER: S-EPMC4682289 | biostudies-literature | 2015 Dec

REPOSITORIES: biostudies-literature

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

Ban Kiwon K   Wile Brian B   Cho Kyu-Won KW   Kim Sangsung S   Song Ming-Ke MK   Kim Sang Yoon SY   Singer Jason J   Syed Anum A   Yu Shan Ping SP   Wagner Mary M   Bao Gang G   Yoon Young-Sup YS  

Stem cell reports 20151201 6


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, a  ...[more]

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