Abstract
Background: Arrhythmogenic cardiomyopathy (ACM) is a disease characterized by ventricular arrhythmias, progressive fibrofatty substitution of the myocardium and sudden cardiac death. Many of the genes identified encode proteins of intercalated disks. Patient-specific induced pluripotent stem cell derived cardiomyocytes (iPSC-CMs) were previously used to model ACM. Material and methods: iPSCs were generated from 3 ACM patients carrying a deletion of whole exon4 in PKP2 gene, 2 asymptomatic PKP2 mutation carriers and 1 control (CTR), belonging to the same family. After cardiac differentiation the purified iPSC-CMs were cultured in basal (BM) and adipogenic medium (AM), as previously reported in literature. Results: Only the wild-type PKP2 allele was identified in ACM and asymptomatic iPSC-CMs indicating a condition of haploinsufficiency. ACM iPSC-CMs, cultured in AM, showed higher lipid accumulation compared to iPSC-CMs from CTR (1580±219 (n=16) vs 615±199 (n=4); p=0.0351) and asymptomatic cases (1580±219(n=16) vs 689±146(n=8); p=0.0059). In AM, ACM iPSC-CMs presented also a higher sarcomeric disorganization compared to CTR (0.0191±0.0015(n=11) vs 0.0280±0.0031(n=4); p=0.039) and asymptomatic (0.0191±0.0015(n=11) vs 0.0273±0.0018 (n=6); p=0.010) iPSC-CMs. Interestingly, a lower expression of the non-phosphorylated (active) form of β-catenin in ACM and asymptomatic iPSC-CMs compared to CTR iPSC-CMs has been shown not only in AM (0.136±0.027(n=9) vs 0.355±0.070(n=6); p=0.023), (0.127±0.108(n=8) vs 0.355±0.070(n=6); p=0.014) but also in BM (0.932±0.178(n=9) vs 1.307±0.096(n=8); p=0.057), (0.758±0.105(n=9) vs 1.307±0.096(n=8); p=0.0057). Transcriptome analysis revealed that pathways like oxidoreduction coenzyme metabolic process, receptor metabolic process, inorganic anion transport, response to reactive oxygen species and gap junction assembly are differentially regulated only in asymptomatic iPSC-CMs when exposed to AM. Conclusions: These data for the first time indicate that asymptomatic iPSC-CMs, although showing a condition of haploinsufficiency with a downregulation of active β-catenin, do not recapitulate the disease phenotype as ACM iPSC-CMs do. Specific pathways altered only in asymptomatic iPSC-CMs could explain reduced penetrance in healthy PKP2 mutation carriers.