Induced pluripotent stem cells differentiation into cardiomyocytes by rotating suspension culture and in specific plates with microwells

Abstract

In order to enhance the differentiation of induced pluripotent cells into cardiomyocytes, we compared two methods of embryoid bodies formation: differentiation in rotating suspension culture and formation of embryoid bodies from a predetermined numbers of pluripotent stem cells in microwells of AggreWell plates. We used transgenic murine induced pluripotent stem cell line AT25. Cell line expressed IRES-flanked enhanced green fluorescent protein (eGFP) under the control of cardiac alpha myosin heavy chain promoter (αMHC). We applied flow cytometry and fluorescence microscopyin order to test the efficiency of differentiation processes. Thus, differentiation of pluripotent stem cells in AggreWell plates without adding differentiation factors was more effective than differentiation in rotating suspension culture. However, we obtained the most amounts of cardiomyocytes on the 11-th day in rotating suspension culture with ascorbic acid,after we applied dorsomorfin, DMSO, ascorbic acid, G-CSF with the above-mentioned methods. The amount of GFP + cells was 2,71 ± 0,07%.

Keywords

• pluripotent stem cells, induced pluripotent stem cells, embryoid bodies, cardiomyocytes, differentiation