Development of a patient-derived induced pluripotent stem cell model for the investigation of SCN5A-D1275N-related cardiac sodium channelopathy

Abstract

Background: The SCN5A gene encodes the α subunit of the cardiac voltage-gated sodium channel, NaV1.5. The missense mutation, D1275N, has been associated with a range of unusual phenotypes associated with reduced NaV1.5 function, including cardiac conduction disease and dilated cardiomyopathy. Curiously, the reported biophysical properties of SCN5A-D1275N channels vary with experimental system. Methods and Results: First, using a human embryonic kidney (HEK) 293 cell-based heterologous expression system, the SCN5A-D1275N channels showed similar maximum sodium conductance but a significantly depolarizing shift of activation gate (+10 mV) compared to wild type. Second, we generated human-induced pluripotent stem cells (hiPSCs) from a 24-year-old female who carried heterozygous SCN5A-D1275N and analyzed the differentiated cardiomyocytes (CMs). Although SCN5A transcript levels were equivalent between D1275N and control hiPSC-CMs, both the total amount of NaV1.5 and the membrane fractions were reduced approximately half in the D1275N cells, which were rescued by the proteasome inhibitor MG132 treatment. Electrophysiological assays revealed that maximum sodium conductance was reduced to approximately half of that in control hiPSC-CMs in the D1275N cells, and maximum upstroke velocity of action potential was lower in D1275N, which was consistent with the reduced protein level of NaV1.5. Conclusions: This study successfully demonstrated diminished sodium currents resulting from lower NaV1.5 protein levels, which is dependent on proteasomal degradation, using a hiPSC-based model for SCN5A-D1275N-related sodium channelopathy.