Influence of estrogen on sodium channel function in human induced stem cell derived cardiomyocytes from a patient with Brugada Syndrome

Background Brugada Syndrome is an autosomal dominant inherited cardiac arrhythmia disorder. The incidence of the syndrome is significantly higher in males. The potential influence of estrogen on the phenotype of Brugada Syndrome and its underlying mechanisms remains to be elucidated. This study aims to explore the influence of estrogen on SCN5A gene variants in Brugada Syndrome and elucidate potential mechanisms.

 

Methods Human-induced pluripotent stem cell (hiPSC) lines were generated from fibroblasts obtained from a Brugada Syndrome (BrS) patient with SCN5A variants (c.3148G>A/p.Ala1050Thr), a healthy donor (WT), and a site-corrected hiPSC line using CRISPR/Cas9 technology (Isogenic). These hiPSC lines were differentiated into cardiomyocytes (hiPSC-CMs), followed by a 24-hour treatment with 17β-Estradiol (E2, 1/100nM). Subsequent Western blot and patch clamp analyses were conducted for characterization.

 

Results Brs-hiPSC-CMs exhibited lower SCN5A expression level and smaller peak sodium channel current (INa). Treatment with 100nM of E2 on BrS hiPSC-CMs resulted in a significant upregulation in the expression of Nav1.5 and an increase in INa, and the maximal depolarization velocity (Vmax) of action potentials (APs). The effects of E2 were mimicked by the Estrogen Receptor (ER)β agonist diarylpropionitrile (DPN), but not the ERα agonist propyl pyrazole triol (PPT). Furthermore, the G protein-coupled estrogen receptor (GPER) agonist G1 did not mimic the effects of E2, and the GPER antagonist G15 could not abolish the effects of E2. Remarkably, E2 did not elicit any significant impact on Nav1.5 expression and INa in both the WT and isogenic hiPSC-CMs. Additionally, E2 led to a reduction of autophagy levels. Activation of autophagy through rapamycin led to a decrease in the expression of Nav1.5 and INa within BrS hiPSC-CMs. Conversely, the inhibition of autophagy resulted in opposite effects, highlighting the pivotal role of autophagy modulation. Furthermore, E2 treatment decreased phosphorylation levels of AMP-activated protein kinase (AMPK) and increased the phosphorylation levels of mammalian target of rapamycin (mTOR) in BrS hiPSC-CMs. 

 

Conclusion Estrogen rescues the BrS phenotype in hiPSC-CMs with SCN5A variants through autophagy regulation. The m-TOR/AMPK signaling pathway plays a pivotal role in mediating these effects.