Patient-specific induced pluripotent stem cell properties implicate Ca2+-homeostasis in clinical arrhythmia associated with combined heterozygous RYR2 and SCN10A variants

We illustrate the use of induced pluripotent stem cells (iPSCs) as platforms for investigating cardiomyocyte phenotypes in a human family pedigree exemplified by novel heterozygous RYR2-A1855D and SCN10A-Q1362H variants occurring alone and in combination. The proband, a four-month-old boy, presented with polymorphic ventricular tachycardia (PVT). Genetic tests revealed double novel heterozygous RYR2-A1855D and SCN10A-Q1362H variants inherited from his father (F) and mother (M) respectively. His father showed ventricular premature beats (VPB); his mother was asymptomatic. Molecular biological characterisations demonstrated greater TNNT2 mRNA expression in the iPSCs-induced cardiomyocytes (iPS-CMs) than in the iPSCs. cTNTs became progressively organised, but cytoplasmic RYR2 and SCN10A aggregations occurred in the iPS-CMs. Proband-specific iPS-CMs showed decreased RYR2 and SCN10A mRNA expression. The RYR2-A1855D variant resulted in premature spontaneous sarcoplasmic reticular (SR) Ca2+ tr..., 1. Family pedigree
The investigation was conducted following principles defined by the Helsinki Declaration and approved by the Ethics Committee of Xi’an Children’s Hospital (No. 2019-599), Affiliated Children’s Hospital of Xi’an Jiaotong University. The family pedigree (Fig. 1A) of a four-month-old boy with CPVT was obtained from the Department of Cardiology, Xi’an Children’s Hospital in 2019. Clinical phenotypes of the pedigree were deduced from the clinical history and physical, electrocardiographic (ECG) and ultrasoundcardiographic (UCG) examination. Written informed consent was obtained from the parents. We term the proband father as proband-F and the proband mother as proband-M
2. Genetic analysis
Genomic DNA was extracted and clinical whole exons were tested using next-generation sequencing (NGS) target capture (SinoPath Company, China). The pathological characteristics of suspicious variants were predicted using multiple bioinformatics software, including Polyphen-2, Provean, ..., Prism, clampit, Origin8