hiPSC-derived cardiac models for Arrhythmogenic Cardiomyopathy: characterization and preliminary insights into ncRNA modulation

Arrhythmogenic cardiomyopathy (ACM) is a genetic heart disease, characterized by the degeneration of ventricular cardiomyocytes with fibrofatty replacement of myocardial tissue. Clinical manifestations include syncope and ventricular arrhythmias that can lead to sudden death. ACM is a genetically heterogenous disorder, mainly autosomal with incomplete penetrance and variable expressivity. Most mutations involve genes encoding major components of cardiac intercalated discs, which are implicated in cell-cell adhesion junctions. Despite advances in understanding its genetic basis, the underlining mechanisms are not clarified yet. Among the putative regulatory factors, long non-coding RNAs (lnc-RNAs) have recently emerged as modulators of cardiac pathophysiology and were found deregulated in ACM models. In this thesis we generated 2D and 3D in vitro models to better characterize the disease and to investigate the feasibility of lnc-RNA-targeted therapeutic strategies. We successfully differentiated hiPSC-CMs, hiPSC-FBs, hiPSC-ECs and assembled 3D MTs comprising these cell types. Differentiation was validated by immunofluorescence of cell-specific markers. From their analysis, significant differences were observed in contractile behaviour in mutated hiPSC-CMs ad in desmosomes length in mutated MTs. To explore the therapeutic potential of antisense oligonucleotides (GapmeRs) for ACM, as a proof of concept, we performed a preliminary study to target NEAT1, a lnc-RNA previously found dysregulated in ACM models, in patient-derived hiPSC-CM. These experiments provided data on NEAT1 expression in ACM hiPSC-CMs and MTs. However, further optimization of both dose and delivery conditions is still necessary to fully evaluate their potential.