Résumé
Human induced pluripotent stem cells derived-cardiomyocytes (hiPSC-CMs) are widely recognized as a model to study several cardiac disorders. In this perspective, differentiation of hiPSCs into sinoatrial node (SAN)-like cells may represent a significant advance in providing an in vitro platform for exploring SAN dysfunction (SND) and evaluate new therapeutic approaches. Recently, hiPSC-CMs derived from patient carrying mutation in the cardiac type 2 ryanodine receptor (RyR2-H29D) have been linked to catecholaminergic polymorphic ventricular tachycardia (CPVT), in association with SND-dependent severe bradycardia.
We aimed to obtain hiPSCs-derived cardiac pacemaker cells (SAN-hiPSCs) from RyR2-H29D mutated patient and characterize these cells at electrophysiological level.
hiPSCs derived from patients with CPVT and healthy subjects were differentiated into SAN-hiPSCs using a 2D matrix sandwich method, modulating Wnt signaling. Moreover, cells were treated with triiodothyronine, dexamethasone and intracellular cyclic AMP (DTA) “cocktail” to improve cell maturity, including intracellular Ca2+ handling. At day 40 post-differentiation, electrical activity was assessed by patch-clamp.
Significant decrease in action potential (AP) frequency was observed in SAN-hiPSCs derived from CPVT patients compared to control (Ctrl) ones (Ctrl: 157.5±9.9 bpm; CPVT: 38.4±13.6 bpm). RyR2-H29D mutation did not affect pacemaker “funny” current (If) (@–135mV Ctrl: –34.9±16.7 pA/pF; CPVT: –21.8±2.8 pA/pF).
SAN-hiPSCs derived from CPVT patient present a bradycardic phenotype compared to SAN-hiPSCs derived from healthy patient and consistent with patient clinical feature. Bradycardia is not related to alteration of If. Nevertheless, G protein-gated inwardly rectifying potassium current (IKACh), L- and T-type Ca2+ currents as well as intracellular Ca2+ handling, involved in cardiac automaticity, remain to be evaluated. Finally, recent studies have demonstrated that the genetic ablation or pharmacological inhibition of IKACh prevents SND-dependent bradycardia. In the future, this therapeutic strategy could be tested to rescue bradycardia in our model of SAN-hiPSCs derived from CPVT patient.