Abstract
In an earlier study exploring the potential of gene transfer to repair myocardial conduction defects, we observed that myotubes, generated by forced expression of MyoD, exhibit reduced excitability when also modified to express connexin43 (Cx43). We hypothesized that this effect was caused by gap junction-mediated coupling between myotubes and the underlying fibroblast feeder layer. This intriguing possibility has important implications for ongoing efforts to develop strategies for repairing myocardial conduction defects by gene transfer, and also provides novel insights into the electrophysiological function of naturally occurring heterologous cell coupling within the heart. Although a conductive function for fibroblasts through heterologous coupling has previously been reported, the current study provides novel evidence that fibroblasts can modulate cardiomyocyte excitability in a Cx43-dependent manner. In a co-culture study system, neonatal rat cardiomyocytes were grown on monolayers of mouse fibroblasts with genetically altered Cx43 expression and the effect on intrinsic beat frequency examined. Cardiomyocytes grown on wild-type (WT) fibroblasts expressing native levels of Cx43 beat significantly slower than cells grown on fibroblasts devoid of this molecule (germline knockout) or with dominant-negative functional suppression. Expression of Cx43 in fibroblasts from Cx43 knockout mice restored cardiomyocyte beat frequency, to rates comparable with those observed in co-culture with WT fibroblasts.
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Acknowledgements
We thank Professor Inder Verma (Salk Institute) for providing lentiviral reagents and Dr Alan Lau (University of Hawaii) for supplying fibroblasts from WT and CxKO mice. We also thank Professor Dale Laird (University of Western Ontario) and Dr Vladimir Krutovskikh (International Agency for Research on Cancer) for providing the cDNA for the rat Cx43-GFP fusion gene and a 21-bp Cx43 deletion mutant, respectively. SLG is supported by the Noel Dowling Research Fellowship. EK was the recipient of a Postgraduate Scholarship from the National Heart Foundation of Australia.
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Kizana, E., Ginn, S., Smyth, C. et al. Fibroblasts modulate cardiomyocyte excitability: implications for cardiac gene therapy. Gene Ther 13, 1611–1615 (2006). https://doi.org/10.1038/sj.gt.3302813
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DOI: https://doi.org/10.1038/sj.gt.3302813
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