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A systemic evaluation of cardiac differentiation from mRNA reprogrammed human induced pluripotent stem cells.
- Source :
-
PloS one [PLoS One] 2014 Jul 28; Vol. 9 (7), pp. e103485. Date of Electronic Publication: 2014 Jul 28 (Print Publication: 2014). - Publication Year :
- 2014
-
Abstract
- Genetically unmodified cardiomyocytes mandated for cardiac regenerative therapy is conceivable by "foot-print free" reprogramming of somatic cells to induced pluripotent stem cells (iPSC). In this study, we report generation of foot-print free hiPSC through messenger RNA (mRNA) based reprograming. Subsequently, we characterize cardiomyocytes derived from these hiPSC using molecular and electrophysiological methods to characterize their applicability for regenerative medicine. Our results demonstrate that mRNA-iPSCs differentiate ontogenetically into cardiomyocytes with increased expression of early commitment markers of mesoderm, cardiac mesoderm, followed by cardiac specific transcriptional and sarcomeric structural and ion channel genes. Furthermore, these cardiomyocytes stained positively for sarcomeric and ion channel proteins. Based on multi-electrode array (MEA) recordings, these mRNA-hiPSC derived cardiomyocytes responded predictably to various pharmacologically active drugs that target adrenergic, sodium, calcium and potassium channels. The cardiomyocytes responded chronotropically to isoproterenol in a dose dependent manner, inotropic activity of nifidipine decreased spontaneous contractions. Moreover, Sotalol and E-4031 prolonged QT intervals, while TTX reduced sodium influx. Our results for the first time show a systemic evaluation based on molecular, structural and functional properties of cardiomyocytes differentiated from mRNA-iPSC. These results, coupled with feasibility of generating patient-specific iPSCs hold great promise for the development of large-scale generation of clinical grade cardiomyocytes for cardiac regenerative medicine.
- Subjects :
- Carbachol pharmacology
Cardiotonic Agents pharmacology
Cells, Cultured
Cellular Reprogramming drug effects
Cellular Reprogramming genetics
Cholinergic Agonists pharmacology
Fibroblasts cytology
Fibroblasts drug effects
Fibroblasts metabolism
Gene Expression
Humans
Induced Pluripotent Stem Cells cytology
Induced Pluripotent Stem Cells physiology
Ion Channels drug effects
Ion Channels physiology
Isoproterenol pharmacology
Male
Membrane Potentials drug effects
Microscopy, Confocal
Middle Aged
Myocytes, Cardiac cytology
Myocytes, Cardiac physiology
Octamer Transcription Factor-3 genetics
Octamer Transcription Factor-3 metabolism
Pluripotent Stem Cells cytology
Pluripotent Stem Cells metabolism
Pluripotent Stem Cells physiology
Reverse Transcriptase Polymerase Chain Reaction
Stage-Specific Embryonic Antigens genetics
Stage-Specific Embryonic Antigens metabolism
Cell Differentiation genetics
Cellular Reprogramming Techniques methods
Induced Pluripotent Stem Cells metabolism
Myocytes, Cardiac metabolism
RNA, Messenger genetics
Subjects
Details
- Language :
- English
- ISSN :
- 1932-6203
- Volume :
- 9
- Issue :
- 7
- Database :
- MEDLINE
- Journal :
- PloS one
- Publication Type :
- Academic Journal
- Accession number :
- 25068310
- Full Text :
- https://doi.org/10.1371/journal.pone.0103485