Human Induced Pluripotent Stem Cells-Derived Cardiomyocytes Carrying CALM1-F142l Mutation Recapitulate LQTS Phenotype in Vitro

Abstract

Background: Calmodulin (CaM), a small essential Ca2+ binding protein, is encoded by 3 genes, namely CALM1, CALM2, CALM3, and exerts its modulatory role depending on intracellular Ca2+ concentration changes. In the heart, CaM mutations in these genes led to recurrent cardiac arrest in infants associated with severe QT prolongation (LQTS-CaM).Aims: To explore how the heterozygous CALM1-F142L mutation can support the reported extremely severe cardiac phenotype. To maintain the correct allele balance we studied the functional consequences of this mutation by using patient-specific human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CMs).Methods: Skin fibroblast of a patient carrying CALM1-F142L mutation were reprogrammed to iPSC and differentiated into CMs; control individuals were 2 unrelated healthy donors without history of cardiac disease. We focus our attention on ICaL biophysics, electrical activity and intracellular Ca2+ dynamics. To make the iPSC-CMs a more reliable model to investigate mechanisms underlying cardiac arrhythmia, in-silico IK1 was injected with a Dynamic-Clamp approach.Results: CALM1-F142L mutation induced impaired Ca2+-dependent inactivation (CDI) and incomplete inactivation of ICaL, thus causing increased Ca2+ influx and consequent action potential prolongation. Therapeutic concentration of verapamil markedly shortened QT in mutated cells. F142L iPSC-CMs failed to adapt to high pacing rates, pointing this as a likely arrhythmogenic mechanism for the mutation. Intracellular Ca2+ handling was instead marginally affected by the mutation. Indeed, sarcoplasmic reticulum (SR) Ca2+content and Ca2+ induced Ca2+ release (CICR)-gain were not affected by the mutation.Conclusions: LQTS-F142L CaM mutation is highly arrhythmogenic mainly because of impaired ICaL CDI; accordingly, its pathogenicity might be limited by the therapeutic use of Ca2+ channel blockers like verapamil.