Biological pacemakers: characterization in an in vitro coculture model

Background

Biological pacemakers could be an alternative or complement to electronic pacemakers. Embryonic stem cells (ESCs) can be differentiated in vitro to spontaneously active cells. Although numerous studies show that ESC-derived cardiomyocytes (ESC-CMs) and other cell types are capable to exert pacemaker function in vivo, detailed analyses of pattern and safety of conduction on a tissue level are rare.

Methods

Murine ESCs (mESCs) expressing enhanced green fluorescent protein and puromycin resistance under control of the promoter of α-myosin (heavy chain) were differentiated to cardiomyocytes (mESC-CMs) and purified by negative antibiotic selection. Ventricles of mouse embryonic hearts (embryonic day 16.5) were embedded in agarose and sliced along the short axis. Clusters of mESC-CMs and the murine, vital heart slices were cocultured on multielectrode arrays for 4 days. Field potentials and videos were recorded daily to investigate beating behavior and excitation spreading within the slice.

Results

On the first day of coculture, the mean beating rate of the tissue slices cocultured with mESC-CMs (n = 19) did not differ significantly from the beating rate of control slices (n = 19) (37 ± 10 versus 19 ± 7 bpm, P = .133). After 4 days of coculture, beating rates were significantly higher in cocultures than in control slices (154 ± 22 versus 49 ± 8 bpm, P < .001). On day 4, 1:1 coupling could be found in 1 of 19 preparations; 2:1, 3:1, or 4:1 coupling in another 4 of 19 preparations; 14 of 19 propagation patterns were irregular.

Conclusion

In this in vitro model, the increase of the beating rate suggests that purified mESC-CMs can pace native heart tissue, albeit with low efficiency.