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Calsequestrin mutation and catecholaminergic polymorphic ventricular tachycardia: a simulation study of cellular mechanism.

ABSTRACT: Patients with a missense mutation of the calsequestrin 2 gene (CASQ2) are at risk for catecholaminergic polymorphic ventricular tachycardia. This mutation (CASQ2(D307H)) results in decreased ability of CASQ2 to bind Ca2+ in the sarcoplasmic reticulum (SR). In this theoretical study, we investigate a potential mechanism by which CASQ2(D307H) manifests its pro-arrhythmic consequences in patients.Using simulations in a model of the guinea pig ventricular myocyte, we investigate the mutation's effect on SR Ca2+ storage, the Ca2+ transient (CaT), and its indirect effect on ionic currents and membrane potential. We model the effects of isoproterenol (ISO) on Ca(V)1.2 (the L-type Ca2+ current, I(Ca(L))) and other targets of beta-adrenergic stimulation.ISO increases I(Ca(L)), prolonging action potential (AP) duration (Control: 172 ms, +ISO: 207 ms, at cycle length of 1500 ms) and increasing CaT (Control: 0.79 microM, +ISO: 1.61 microM). ISO increases I(Ca(L)) by reducing the fraction of channels which undergo voltage-dependent inactivation and increasing transitions from a non-conducting to conducting mode of channel gating. CASQ2(D307H) reduces SR storage capacity, thereby reducing the magnitude of CaT (Control: 0.79 microM, CASQ2(D307H): 0.52 microM, at cycle length of 1500 ms). The combined effect of CASQ2(D307H) and ISO elevates SR free Ca2+ at a rapid rate, leading to store-overload-induced Ca2+ release and delayed afterdepolarization (DAD). If resting membrane potential is sufficiently elevated, the Na+-Ca2+ exchange-driven DAD can trigger I(Na) and I(Ca(L)) activation, generating a triggered arrhythmogenic AP.The CASQ2(D307H) mutation manifests its pro-arrhythmic consequences due to store-overload-induced Ca2+ release and DAD formation due to excess free SR Ca2+ following rapid pacing and beta-adrenergic stimulation.

SUBMITTER: Faber GM

PROVIDER: S-EPMC2030636 | biostudies-literature | 2007 Jul

REPOSITORIES: biostudies-literature

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Calsequestrin mutation and catecholaminergic polymorphic ventricular tachycardia: a simulation study of cellular mechanism.

Faber Gregory M GM Rudy Yoram Y

Cardiovascular research 20070421 1

<h4>Objectives</h4>Patients with a missense mutation of the calsequestrin 2 gene (CASQ2) are at risk for catecholaminergic polymorphic ventricular tachycardia. This mutation (CASQ2(D307H)) results in decreased ability of CASQ2 to bind Ca2+ in the sarcoplasmic reticulum (SR). In this theoretical study, we investigate a potential mechanism by which CASQ2(D307H) manifests its pro-arrhythmic consequences in patients.<h4>Methods</h4>Using simulations in a model of the guinea pig ventricular myocyte, ...[more]

PMID: 17531962

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Project description:RationaleAutosomal-dominant mutations in ryanodine receptor type 2 ( RYR2) are responsible for ≈60% of all catecholaminergic polymorphic ventricular tachycardia. Dysfunctional RyR2 subunits trigger inappropriate calcium leak from the tetrameric channel resulting in potentially lethal ventricular tachycardia. In vivo CRISPR/Cas9-mediated gene editing is a promising strategy that could be used to eliminate the disease-causing Ryr2 allele and hence rescue catecholaminergic polymorphic ventricular tachycardia.ObjectiveTo determine if somatic in vivo genome editing using the CRISPR/Cas9 system delivered by adeno-associated viral (AAV) vectors could correct catecholaminergic polymorphic ventricular tachycardia arrhythmias in mice heterozygous for RyR2 mutation R176Q (R176Q/+).Methods and resultsGuide RNAs were designed to specifically disrupt the R176Q allele in the R176Q/+ mice using the SaCas9 ( Staphylococcus aureus Cas9) genome editing system. AAV serotype 9 was used to deliver Cas9 and guide RNA to neonatal mice by single subcutaneous injection at postnatal day 10. Strikingly, none of the R176Q/+ mice treated with AAV-CRISPR developed arrhythmias, compared with 71% of R176Q/+ mice receiving control AAV serotype 9. Total Ryr2 mRNA and protein levels were significantly reduced in R176Q/+ mice, but not in wild-type littermates. Targeted deep sequencing confirmed successful and highly specific editing of the disease-causing R176Q allele. No detectable off-target mutagenesis was observed in the wild-type Ryr2 allele or the predicted putative off-target site, confirming high specificity for SaCas9 in vivo. In addition, confocal imaging revealed that gene editing normalized the enhanced Ca2+ spark frequency observed in untreated R176Q/+ mice without affecting systolic Ca2+ transients.ConclusionsAAV serotype 9-based delivery of the SaCas9 system can efficiently disrupt a disease-causing allele in cardiomyocytes in vivo. This work highlights the potential of somatic genome editing approaches for the treatment of lethal autosomal-dominant inherited cardiac disorders, such as catecholaminergic polymorphic ventricular tachycardia.

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Calsequestrin mutation and catecholaminergic polymorphic ventricular tachycardia: a simulation study of cellular mechanism.

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Calsequestrin mutation and catecholaminergic polymorphic ventricular tachycardia: a simulation study of cellular mechanism.

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