Generation of electrophysiologically functional cardiomyocytes from mouse induced pluripotent stem cells

•In this study we show that iPS derived cardiomyocytes (iPS-CMs) express cardiac specific markers, e.g. MHC and cTnT, and importantly express membrane connexin-43, a key gap junction protein that is responsible for synchronous communication between cardiac cells.•Further investigations demonstrated that these iPS-CMs display cardiac electrophysiological activity verified by patch-clamp in single contracting cells, and optical mapping with RH237, a voltage-sensitive dye, in iPS-CM clusters.•These results indicate that iPS cells might be a promising cell source for the generation of patient-specific cardiomyocytes even for patients with severe heart disease, because the cells can be made available in unlimited amounts and are capable of cardiomyocyte generation.

Induced pluripotent stem (iPS) cells can efficiently differentiate into the three germ layers similar to those formed by differentiated embryonic stem (ES) cells. This provides a new source of cells in which to establish preclinical allogeneic transplantation models. Our iPS cells were generated from mouse embryonic fibroblasts (MEFs) transfected with the Yamanaka factors, the four transcription factors (Oct4, Sox2, Klf4 and c-Myc), without antibiotic selection or MEF feeders. After the formation of embryoid bodies (EBs), iPS cells spontaneously differentiated into Flk1-positive cardiac progenitors and cardiomyocytes expressing cardiac-specific markers such as alpha sarcomeric actinin (α-actinin), cardiac alpha myosin heavy chain (α-MHC), cardiac troponin T (cTnT), and connexin 43 (CX43), as well as cardiac transcription factors Nk2 homebox 5 (Nkx2.5) and gata binding protein 4 (gata4). The electrophysiological activity of iPS cell-derived cardiomyocytes (iPS-CMs) was detected in beating cell clusters with optical mapping and RH237 a voltage-sensitive dye, and in single contracting cells with patch-clamp technology. Incompletely differentiated iPS cells formed teratomas when transplanted into a severe combined immunodeficiency (SCID) mouse model of myocardial infarction. Our results show that somatic cells can be reprogrammed into pluripotent stem cells, which in turn spontaneously differentiate into electrophysiologically functional mature cardiomyocytes expressing cardiac-specific makers, and that these cells can potentially be used to repair myocardial infarction (MI) in the future.