Striking In Vivo Phenotype of a Disease-Associated Human SCN5A Mutation Producing Minimal Changes in Vitro

Abstract

Background The D1275N SCN5A mutation has been associated with a range of unusual phenotypes including conduction disease and dilated cardiomyopathy (DCM) as well as atrial and ventricular tachyarrhythmias. However, when D1275N is studied in heterologous expression systems, most studies show near-normal sodium channel function. Thus, the relationship of the variant to the clinical phenotypes remains uncertain. Methods and results We identified D1275N in a patient with atrial flutter, atrial standstill, conduction disease, and sinus node dysfunction. There was no major difference in biophysical properties between wild-type and D1275N channels expressed in CHO or tsA201 cells in the absence or presence of β1 subunits. To determine D1275N function in vivo, the Scn5a locus was modified to knock out the mouse gene, and the full-length wild-type (H) or D1275N (DN) human SCN5A cDNAs were then inserted at the modified locus using recombinase mediated cassette exchange. Mice carrying the DN allele displayed slow conduction, heart block, atrial fibrillation, ventricular tachycardia, and a DCM phenotype, with no significant fibrosis or myocyte disarray on histological examination. The DN allele conferred gene-dose dependent increases in SCN5A mRNA abundance, but reduced sodium channel protein abundance and peak sodium current amplitudes (H/H, −41.0±2.9 pA/pF at −30mV; DN/H, 19.2±3.1 pA/pF, P<0.001 versus H/H; DN/DN, −9.3±1.1 pA/pF, P<0.001 versus H/H). Conclusions Although D1275N produces near normal currents in multiple heterologous expression experiments, our data establish this variant as a pathological mutation that generates conduction slowing, arrhythmias, and a DCM phenotype by reducing cardiac sodium current.

Gain-of-Function Mutation of the SCN5A Gene Causes Exercise-Induced Polymorphic Ventricular Arrhythmias

“…As an example, although many studies have reported several varied phenotypes associating with a SCN5A p.D1275N missense mutation, for example, atrial standstill, dilated cardiomyopathy with conduction disease, atrial flutter/fibrillation, sick sinus syndrome, and ventricular dilation and dysfunction, the mutation does not induce marked Na v 1.5 dysfunction in vitro. However, when studied in genetically engineered mice, in vivo, the mutation generates extensive aberration of channel function, 21 underscoring that functional phenotypes may be influenced substantially by the experimental systems used.…”

Development of a Patient-Derived Induced Pluripotent Stem Cell Model for the Investigation of <i>SCN5A</i>-D1275N-Related Cardiac Sodium Channelopathy

“…11- 13 The reported electrophysiological properties of SCN5A-D1275N channels vary with experimental system; studies using heterologous expression systems showed no major differences between the mutant and wild-type (WT) channels, 14,15 whereas peak sodium current densities were reduced in SCN5A-D1275N knock-in mice than in WT ones. 16 The functional properties of SCN5A-D1275N channels in human cardiomyocytes (CMs) are currently unclear. Thus, the present study aimed to investigate the biophysical properties of SCN5A-D1275N channels using human-induced pluripotent stem cell-derived CMs (hiPSCCMs) generated from a patient with familial CCD who carried the SCN5A-D1275N mutation.…”

“…15 Recently, a study using a human SCN5A-D1275N knock-in mouse model reported that sodium channel protein levels and peak sodium current densities were lower in D1275N knock-in mice than in Controls. 16 Additionally, a study using a transgenic zebrafish model of SCN5A-D1275N demonstrated reduced heart rate, sinus pause, and AV block. 26 In order to assess the biophysical properties of SCN5A-D1275N channels in a human CM model, we generated hiPSCs from a patient carrying SCN5A-D1275N who had presented with SND and an AV block.…”

Cardiac Sodium Channel Disease Modeling Using Patient-Derived Induced Pluripotent Stem Cells

“…In this issue of the Journal, Hayano et al 8 report on the generation of iPSCs from the cells of a patient with SCN5A missense mutation D1275N, which did not result in major differences in the biophysical properties of the channel, compared with the wild-type channels in vitro. 9 The clinical manifestations of this mutation were sinus node dysfunction and atrioventricular block. The authors report that SCN5A-D1275N hiPSC-derived CMs showed reduced sodium channel protein expression and maximum sodium conductance.…”