Induction and Enhancement of Cardiac Cell Differentiation from Mouse and Human Induced Pluripotent Stem Cells with Cyclosporin-A

Abstract

Induced pluripotent stem cells (iPSCs) are novel stem cells derived from adult mouse and human tissues by reprogramming. Elucidation of mechanisms and exploration of efficient methods for their differentiation to functional cardiomyocytes are essential for developing cardiac cell models and future regenerative therapies. We previously established a novel mouse embryonic stem cell (ESC) and iPSC differentiation system in which cardiovascular cells can be systematically induced from Flk1+ common progenitor cells, and identified highly cardiogenic progenitors as Flk1+/CXCR4+/VE-cadherin− (FCV) cells. We have also reported that cyclosporin-A (CSA) drastically increases FCV progenitor and cardiomyocyte induction from mouse ESCs. Here, we combined these technologies and extended them to mouse and human iPSCs. Co-culture of purified mouse iPSC-derived Flk1+ cells with OP9 stroma cells induced cardiomyocyte differentiation whilst addition of CSA to Flk1+ cells dramatically increased both cardiomyocyte and FCV progenitor cell differentiation. Spontaneously beating colonies were obtained from human iPSCs by co-culture with END-2 visceral endoderm-like cells. Appearance of beating colonies from human iPSCs was increased approximately 4.3 times by addition of CSA at mesoderm stage. CSA-expanded human iPSC-derived cardiomyocytes showed various cardiac marker expressions, synchronized calcium transients, cardiomyocyte-like action potentials, pharmacological reactions, and ultra-structural features as cardiomyocytes. These results provide a technological basis to obtain functional cardiomyocytes from iPSCs.

Pluripotent Stem Cell-Engineered Cell Sheets Reassembled with Defined Cardiovascular Populations Ameliorate Reduction in Infarct Heart Function Through Cardiomyocyte-Mediated Neovascularization

“…S8). We recently reported functional CM induction from human iPSCs [60]. We further established a more efficient CM induction as well as purification methods in human iPSCs [61].…”

Potential for pharmacological manipulation of human embryonic stem cells

“…Cyclosporin A (CSA) treatment of hiPSCs at the mesoderm differentiation stage in visceral endodermal stromal cell co-culture-mediated cardiomyocyte differentiation (Mummery et al, 2003) led to an increased number of beating colonies, although direct treatment of the undifferentiated hiPSCs themselves yielded no effect (Fujiwara et al, 2011). CSA is an immunosuppressant and a calcineurin inhibitor that is thought to function through the inhibition of nuclear factor of activated T cells (NFAT) signalling in T cells (Crabtree and Olson, 2002).…”

“…It has also been shown to have some effects on cardiac myocytes through decreased hypertrophy (Lim et al, 2000). CSA-treated human iPSC-derived cardiomyocytes have the same various cardiac marker expressions, synchronized Ca 2+ transients, cardiomyocyte-like action potentials, pharmacological reactions and ultrastructural features as usual cardiomyocytes (Fujiwara et al, 2011). Treatment of hESCs with cobalt chloride boosts the differentiation of cardiomyocytes to functionally mature cardiomyocytes by inducing the stabilization of HIF-1a (Ng et al, 2011), thereby chemically mimicking a reduction in oxygen concentration.…”

Human induced pluripotent stem cells derived endothelial cells mimicking vascular inflammatory response under flow

“…Recently, hiPSCs have been differentiated into cardiomyocytes, pancreatic b-like cells, endothelial cells, and other cell types. [11][12][13] ECs derived from hiPSCs have great potential as a source for vascular regeneration and tissue engineering.…”