Novel insights in the pathomechanism of Brugada syndrome and fever-related type 1 ECG changes in a preclinical study using human-induced pluripotent stem cell-derived cardiomyocytes.

BACKGROUND: Brugada syndrome (BrS) is causing sudden cardiac death (SCD) mainly at young age. Studying the underlying mechanisms associated with BrS type I electrocardiogram (ECG) changes in the presence of fever and roles of autophagy for BrS remains lacking. OBJECTIVES: We sought to study the pathogenic role of an SCN5A gene variant for BrS with fever-induced type 1 ECG phenotype. In addition, we studied the role of inflammation and autophagy in the pathomechanism of BrS. METHODS: Human-induced pluripotent stem cell (hiPSC) lines from a BrS patient harboring a pathogenic variant (c.3148G>A/p. Ala1050Thr) in SCN5A and two healthy donors (non-BrS) and a CRISPR/Cas9 site-corrected cell line (BrS-corr) were differentiated into cardiomyocytes (hiPSC-CMs) for the study. RESULTS: Reductions of Nav 1.5 expression, peak sodium channel current (INa ) and upstroke velocity (Vmax ) of action potentials with an increase in arrhythmic events were detected in BrS compared to non-BrS and BrS-corr cells. Increasing the cell culture temperature from 37 to 40°C (fever-like state) exacerbated the phenotypic changes in BrS cells. The fever-effects were enhanced by protein kinase A (PKA) inhibitor but reversed by PKA activator. Lipopolysaccharides (LPS) but not increased temperature up to 40°C enhanced the autophagy level in BrS-hiPSC-CMs by increasing reactive oxidative species and inhibiting PI3K/AKT signalling, and hence exacerbated the phenotypic changes. LPS enhanced high temperature-related effect on peak INa shown in BrS hiPSC-CMs. Effects of LPS and high temperature were not detected in non-BrS cells. CONCLUSIONS: The study demonstrated that the SCN5A variant (c.3148G>A/p.Ala1050Thr) caused loss-of-function of sodium channels and increased the channel sensitivity to high temperature and LPS challenge in hiPSC-CMs from a BrS cell line with this variant but not in two non-BrS hiPSC-CM lines. The results suggest that LPS may exacerbate BrS phenotype via enhancing autophagy, whereas fever may exacerbate BrS phenotype via inhibiting PKA-signalling in BrS cardiomyocytes with but probably not limited to this variant.

A Preclinical Study on Brugada Syndrome with a CACNB2 Variant Using Human Cardiomyocytes from Induced Pluripotent Stem Cells.

Aims: Some gene variants in the sodium channels, as well as calcium channels, have been associated with Brugada syndrome (BrS). However, the investigation of the human cellular phenotype and the use of drugs for BrS in presence of variant in the calcium channel subunit is still lacking. Objectives: The objective of this study was to establish a cellular model of BrS in the presence of a CACNB2 variant of uncertain significance (c.425C > T/p.S142F) using human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and test drug effects using this model. Methods and results: This study recruited cells from a patient with Brugada syndrome (BrS) and recurrent ventricular fibrillation carrying a missense variant in CACNB2 as well as from three healthy independent persons. These cells (hiPSC-CMs) generated from skin biopsies of healthy persons and the BrS patient (BrS-hiPSC-CMs) as well as CRISPR/Cas9 corrected cells (isogenic control, site-variant corrected) were used for this study. The hiPSC-CMs from the BrS patient showed a significantly reduced L-type calcium channel current (ICa-L) compared with the healthy control hiPSC-CMs. The inactivation curve was shifted to a more positive potential and the recovery from inactivation was accelerated. The protein expression of CACNB2 of the hiPSC-CMs from the BrS-patient was significantly decreased compared with healthy hiPSC-CMs. Moreover, the correction of the CACNB2 site-variant rescued the changes seen in the hiPSC-CMs of the BrS patient to the normal state. These data indicate that the CACNB2 gene variant led to loss-of-function of L-type calcium channels in hiPSC-CMs from the BrS patient. Strikingly, arrhythmia events were more frequently detected in BrS-hiPSC-CMs. Bisoprolol (beta-blockers) at low concentration and quinidine decreased arrhythmic events. Conclusions: The CACNB2 variant (c.425C > T/p.S142F) causes a loss-of-function of L-type calcium channels and is pathogenic for this type of BrS. Bisoprolol and quinidine may be effective for treating BrS with this variant.