Abstract 16119: Enhanced Embryonic Stem Cell Differentiation to Cardiac Pacemaker Cells by Transduction with a Single Transcription Factor
Embryonic stem cells (ESCs) can randomly differentiate into heterogeneous aggregates of cardiomyocytes with atrial, ventricular and pacemaker properties. In an effort to create efficient biological pacemakers, we sought to direct ESCs to differentiate into pacemaker cardiomyocytes by overexpressing SHOX2, a transcription factor critical for the patterning of the sinoatrial node (SAN) during development. To this end, mouse ESCs were transduced with an adenoviral vector expressing SHOX2. Transient overexpression of SHOX2 significantly increased the quantity of spontaneously-beating EBs by 8-fold (80±27%) compared to control (9±6%). Within each beating EB, twice as many beating foci (2.1±0.8 per EB) were found in SHOX2-EBs compared to control (1.0±0.6 per EB). Na+/Ca2+ exchanger (NCX1) and HCN4 play a prominent role in cardiac pacemaker electrophysiology. NCX1 protein level increased >6-fold in SHOX2-EBs compared to control at the early and late stages of differentiation. HCN4 protein level was significantly up-regulated at all differentiation stages, with heightened cell surface expression in SHOX2-EBs. Action potentials are mainly propagated by connexin (Cx) 45 but not by Cx43 in the SAN, which is mirrored in the atrial/ventricular myocardium. SHOX2-overexpression led to an increase in the transcript (∼1.5-fold) and protein (∼3-fold) levels of Cx45. In contrast, Cx43 transcript and protein levels were reduced in SHOX2-EBs by 33±5% and 30±10%, respectively, compared to control. In order to examine functional efficacy of the enhanced pacemaker properties of SHOX2 overexpression, we injected 10 randomly-selected SHOX2-EBs (or GFP-EBs) into the apex of a rat heart. Two days after the in vivo cell transplantation, Langendorff-perfused, SHOX2-EB-injected hearts demonstrated ectopic ventricular beats upon complete heart block with higher rate than the slow junctional rhythm exhibited in control hearts (injected with GFP-EBs) under the same condition. The data provide a novel and efficient platform to develop biological pacemakers from pluripotent cells with enriched pacemaker myocyte population.
- © 2012 by American Heart Association, Inc.