RAD-Deficient Human Cardiomyocytes Develop Hypertrophic Cardiomyopathy Phenotypes Due to Calcium Dysregulation

Abstract

Ras associated with diabetes (RAD) is a membrane protein that acts as a calcium channel regulator by interacting with cardiac L-type Ca 2+ channels (LTCC). RAD defects can disrupt intracellular calcium dynamics and lead to cardiac hypertrophy. However, due to the lack of reliable human disease models, the pathological mechanism of RAD deficiency leading to cardiac hypertrophy is not well understood. In this study, we created a RRAD −/− H9 cell line using CRISPR/Cas9 technology. RAD disruption did not affect the ability and efficiency of cardiomyocytes differentiation. However, RAD deficient hESC-CMs recapitulate hypertrophic phenotype in vitro. Further studies have shown that elevated intracellular calcium level and abnormal calcium regulation are the core mechanisms by which RAD deficiency leads to cardiac hypertrophy. More importantly, management of calcium dysregulation has been found to be an effective way to prevent the development of cardiac hypertrophy in vitro.

Comprehensive transcriptome-wide analysis of spliceopathy correction of myotonic dystrophy using CRISPR-Cas9 in iPSCs-derived cardiomyocytes

“…Notably, 1 of the 26 different retroviral ISs (Ch16; −66927012) was located upstream of the RRAD (Ras-related glycolysis inhibitor and calcium channel regulator) gene that plays a role in electrical and cytoskeletal function and Ca 2+ handling in cell cardiomyocytes. 30 , 31 However, it is reassuring that the retroviral integration was not in the RRAD open reading frame itself and consequently the exonic regions of the RRAD gene remained unaffected.…”

Ion Channel Impairment and Myofilament Ca2+ Sensitization: Two Parallel Mechanisms Underlying Arrhythmogenesis in Hypertrophic Cardiomyopathy

“…In particular, the onset of impaired Ca 2+ homeostasis preceded the hypertrophic phenotype, suggesting that a Ca 2+ handling impairment plays a central role in the pathogenesis of HCM. This is further highlighted by a chronic verapamil treatment (10 days, 100 nM), showing that the blocking of L-type Ca 2+ channel restored the HCM phenotype; while Ca 2+ handling homeostasis was reverted by normalizing SR Ca 2+ storage, cell size and p-CaMKII significantly decreased [105].…”

Natural genetic variation quantitatively regulates heart rate and dimension

“…Intriguingly, using heart rate metrics for phenotype-genotype mapping, we identified genes with dual roles affecting heart muscle mass and rhythm; e.g., rrad combining regulative functions on cardiac muscle strength and electrophysiology. Rrad is known to inhibit cardiac hypertrophy through the CaMKII pathway with implications for heart failure (32), and is associated with hypertrophic cardiomyopathy (HCM) phenotype in RRAD-deficient cell line (33). Rrad mutations detected in a specific form of familiar arrhythmia (Brugada syndrome) trigger cytoskeleton and electrophysiological abnormalities in iPSC-CMs (30), which we validated in vivo , demonstrating a specific atrioventricular block in medaka rrad crispants, base-edited embryos and mutants.…”