Abstract
BackgroundHuman embryonic stem cells (hESCs) have the potential to provide an unlimited source of cardiomyocytes, which are invaluable resources for drug or toxicology screening, medical research, and cell therapy. Currently a number of obstacles exist such as the insufficient efficiency of differentiation protocols, which should be overcome before hESC-derived cardiomyocytes can be used for clinical applications. Although the differentiation efficiency can be improved by the genetic manipulation of hESCs to over-express cardiac-specific transcription factors, these differentiated cells are not safe enough to be applied in cell therapy. Protein transduction has been demonstrated as an alternative approach for increasing the efficiency of hESCs differentiation toward cardiomyocytes.MethodsWe present an efficient protocol for the differentiation of hESCs in suspension by direct introduction of a LIM homeodomain transcription factor, Islet1 (ISL1) recombinant protein into the cells.ResultsWe found that the highest beating clusters were derived by continuous treatment of hESCs with 40 µg/ml recombinant ISL1 protein during days 1–8 after the initiation of differentiation. The treatment resulted in up to a 3-fold increase in the number of beating areas. In addition, the number of cells that expressed cardiac specific markers (cTnT, CONNEXIN 43, ACTININ, and GATA4) doubled. This protocol was also reproducible for another hESC line.ConclusionsThis study has presented a new, efficient, and reproducible procedure for cardiomyocytes differentiation. Our results will pave the way for scaled up and controlled differentiation of hESCs to be used for biomedical applications in a bioreactor culture system.
Highlights
Cardiomyocytes derived from human embryonic stem cells potentially offer large numbers of cells for biomedical and industrial applications
Direct protein transduction into the cells both in vitro and in vivo is an efficient alternative to genetic manipulation, which leads to the production of safe cells required for cell therapy
We have demonstrated that the transduction of rISL1 protein enhanced Human embryonic stem cells (hESCs) differentiation into the beating cardiomyocytes phenotype
Summary
Cardiomyocytes derived from human embryonic stem cells (hESCs) potentially offer large numbers of cells for biomedical and industrial applications. Current protocols for differentiation of cardiomyocytes from hESCs are time consuming, have low yield, and lack reproducibility (for review see, ref [1]). For the applicability of these cells in biomedicine it is necessary to produce sufficient numbers of functional cardiomyocytes or their progenitors. This requires the development of large-scale expansion of hESCs and their controlled differentiation protocols. Human embryonic stem cells (hESCs) have the potential to provide an unlimited source of cardiomyocytes, which are invaluable resources for drug or toxicology screening, medical research, and cell therapy. Protein transduction has been demonstrated as an alternative approach for increasing the efficiency of hESCs differentiation toward cardiomyocytes
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