Abstract 13066: Targeting of Heterocellular Coupling in Fibrotic Neonatal Rat Myocardial Cultures by Lentiviral Connexin43 Knockdown is Anti-Arrhythmic
Objective: Cardiac fibrosis is associated with an increased risk of cardiac arrhythmias, while treatment of these arrhythmias is suboptimal and underlying pro-arrhythmic mechanisms are still incompletely understood. This study investigated the pro-arrhythmic effect of myofibroblast (MFB) cardiomyocyte (CMC) coupling and the anti-arrhythmic potential of Cx43 downregulation in MFBs, which inhibits such coupling.
Methods: Co-cultures of CMCs and MFBs in a 1:1 ratio as a model of cardiac fibrosis, and purified cultures of CMCs were studied for arrhythmogeneity by optical mapping. Dyetransfer, immunocytological staining and patchclamp were also performed. Heterocellular coupling in MFB-CMC co-cultures was reduced by transducing MFB's with a lentiviral vector encoding shRNA for Cx43 prior to co-culturing. In control groups, MFBs were either transduced with a lentiviral vector encoding shRNA for pLuc or no shRNA was used.
Results: Conduction velocity (CV) was significantly lower in fibrotic cultures compared to controls (10.9±2.4 cm/s vs 21.6±3.6 cm/s p<0.0001), while ectopic activity was increased in fibrotic cultures. Of these cultures (n=30) 30.0% showed spontaneous reentrant tachyarrhythmias compared to 3.2% in control cultures (n=63 in total). CMC membrane potential was significantly depolarized in fibrotic cultures, and was associated with diminished upstroke velocity (42.3%, p<0.0001). Knockdown of Cx43 expression in MFBs decreased transfer of dye and thereby functional coupling. Also, heterocellular MFB-CMC Cx43 expression was decreased by 69%, while CMC-CMC Cx43 expression remained unaffected by Cx43 knockdown in MFBs. Importantly, Cx43 knockdown in MFBs increased CV by 37.5% (p<0.001), reduced depolarization of adjacent CMCs and ectopic activity, thereby reducing the incidence of reentry by 41.1%.
Conclusions: Electrotonic coupling of MFBs to CMCs in myocardial cultures gives rise to spontaneous reentrant tachyarrhythmias. Targeting of this heterocellular coupling, by reducing the expression of Cx43 in MFBs, limits conduction slowing and ectopic activity and thereby prevents such arrhythmias. This study could provide a rationale for the development of more substrate-oriented anti-arrhythmic therapies.
- © 2011 by American Heart Association, Inc.