Inhibition of Wnt/β-catenin signalling upregulates Nav1.5 channels in Brugada syndrome iPSC-derived cardiomyocytes

Abstract

The voltage-gated Nav1.5 channels mediate the fast Na+ current (INa) in cardiomyocytes initiating action potentials and cardiac contraction. Downregulation of INa, as occurs in Brugada Syndrome (BrS), causes ventricular arrhythmias. The present study aims to investigate whether the Wnt/-catenin signalling (Wnt signalling) regulates Nav1.5 expression in human cardiomyocytes and whether inhibition of Wnt signalling upregulates Nav1.5 in BrS cardiomyocytes. In human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) that were derived from two healthy volunteers (one male and one female), pharmacological activation of Wnt/-catenin signalling via CHIR-99021 treatment reduced (p<0.01) both Nav1.5 protein and SCN5A mRNA. In iPSC-CMs derived from a BrS patient, both Nav1.5 protein and peak INa density were reduced compared to those in healthy iPSC-CMs. Treatment of BrS iPSC-CMs with Wnt-C59, a small-molecule inhibitor of Wnt signalling, led to a 2.1-fold increase in Nav1.5 protein (p=0.0005) but surprisingly did not affect SCN5A mRNA (p=0.146). Markers of myocyte maturation, α-sarcomeric actinin protein, Kir2.1 protein and KCNJ2 mRNA were not affected by Wnt-C59. Similarly, inhibition of Wnt signalling using shRNA-mediated -catenin knockdown in BrS iPSC-CMs led to a 4.4-fold increase in Nav1.5, which was associated with a 4.9-fold increase in peak INa density but only a 2.1-fold increase in SCN5A mRNA. The upregulation of Nav1.5 by -catenin knockdown was verified in iPSC-CMs derived from a second BrS patient. This study demonstrated that Wnt/-catenin signalling inhibits Nav1.5 expression in both male and female human iPSC-CMs, and inhibition of Wnt signalling upregulates Nav1.5 in BrS iPSC-CMs through both transcriptional and post-transcriptional mechanisms.

Keywords: Nav1.5, Wnt/-catenin signalling, Cardiomyocytes, Induced pluripotent stem cells