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Dantrolene rescues arrhythmogenic RYR2 defect in a patient-specific stem cell model of catecholaminergic polymorphic ventricular tachycardia.

Jung CB, Moretti A, Mederos y Schnitzler M, Iop L, Storch U, Bellin M, Dorn T, Ruppenthal S, Pfeiffer S, Goedel A, Dirschinger RJ, Seyfarth M, Lam JT, Sinnecker D, Gudermann T, Lipp P, Laugwitz KL - EMBO Mol Med (2012)

Bottom Line: In patient iPSC-derived cardiomyocytes, catecholaminergic stress led to elevated diastolic Ca(2+) concentrations, a reduced SR Ca(2+) content and an increased susceptibility to DADs and arrhythmia as compared to control myocytes.Dantrolene, a drug effective on malignant hyperthermia, restored normal Ca(2+) spark properties and rescued the arrhythmogenic phenotype.This suggests defective inter-domain interactions within the RYR2 channel as the pathomechanism of the S406L mutation.

View Article: PubMed Central - PubMed

Affiliation: Klinikum rechts der Isar, Technische Universität München, I. Medizinische Klinik, Kardiologie, München, Germany.

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Myocytic subtypes of iPSC-derived myocytes and their Ca2+ spark propertiesPercentage of ventricular-, atrial- and nodal-like myocytes after 3–4 month cardiac iPSC differentiation based on single cell electrophysiological measurements of action potentials (n = 47–50 cells) and expression of specific myocytic lineage markers (MLC2v, for ventricular cells, and MLC2a, for atrial cells) by immunohistochemistry (n = 100 cells). Scale bars, 10 µm. Dotted lines in the action potential traces indicate 0 mV.Summary of Ca2+ spark characteristics from control (black) and CPVT (red) cells under basal conditions when all myocytes or specifically ventricular (MLC2v+) and non-ventricular (MLC2v−) subtypes are analysed. Fold changes are relative to all myocytes control. Staining for MLC2v was performed after Ca2+ spark imaging. Between 21 and 113 cells from three iPSC lines were analysed per group. Data are means ± SEM from three independent experiments; p-values from two-tailed t-test.
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fig05: Myocytic subtypes of iPSC-derived myocytes and their Ca2+ spark propertiesPercentage of ventricular-, atrial- and nodal-like myocytes after 3–4 month cardiac iPSC differentiation based on single cell electrophysiological measurements of action potentials (n = 47–50 cells) and expression of specific myocytic lineage markers (MLC2v, for ventricular cells, and MLC2a, for atrial cells) by immunohistochemistry (n = 100 cells). Scale bars, 10 µm. Dotted lines in the action potential traces indicate 0 mV.Summary of Ca2+ spark characteristics from control (black) and CPVT (red) cells under basal conditions when all myocytes or specifically ventricular (MLC2v+) and non-ventricular (MLC2v−) subtypes are analysed. Fold changes are relative to all myocytes control. Staining for MLC2v was performed after Ca2+ spark imaging. Between 21 and 113 cells from three iPSC lines were analysed per group. Data are means ± SEM from three independent experiments; p-values from two-tailed t-test.

Mentions: Since in CPVT patients tachycardia is restricted to the ventricles under stress condition, it might be expected that CPVT is a disease of ventricular cardiomyocytes. Our cardiac differentiation protocol leads to the generation of all three subtypes of cardiomyocytes, namely ventricular-like, atrial-like and nodal-like cells, which can be distinguished by the expression of specific myocytic lineage markers and by the shape of the action potential (Moretti et al, 2010b). Immunohistochemical analysis for ventricular and atrial myosin light chain 2 (MLC2v and MLC2a) proteins and electrophysiological measurements of action potentials in single iPSC-derived myocytes demonstrated that the ventricular subtype is largely predominant, accounting for 70–80% of all myocytes similarly in both control and CPVT groups (Fig 5A). To assess whether Ca2+ spark properties were specifically altered only in ventricular cells, we stained the cells with an antibody against MLC2v retrospectively and analysed spark data from ventricular (MLC2v+ cells) and non-ventricular (MLC2v− cells) myocytes separately. The observed differences in Ca2+ spark properties between control and CPVT cells persisted in the ventricular and non-ventricular subpopulations (Fig 5B), indicating that the mutated S406L-RYR2 channels are dysfunctional in all myocytes.


Dantrolene rescues arrhythmogenic RYR2 defect in a patient-specific stem cell model of catecholaminergic polymorphic ventricular tachycardia.

Jung CB, Moretti A, Mederos y Schnitzler M, Iop L, Storch U, Bellin M, Dorn T, Ruppenthal S, Pfeiffer S, Goedel A, Dirschinger RJ, Seyfarth M, Lam JT, Sinnecker D, Gudermann T, Lipp P, Laugwitz KL - EMBO Mol Med (2012)

Myocytic subtypes of iPSC-derived myocytes and their Ca2+ spark propertiesPercentage of ventricular-, atrial- and nodal-like myocytes after 3–4 month cardiac iPSC differentiation based on single cell electrophysiological measurements of action potentials (n = 47–50 cells) and expression of specific myocytic lineage markers (MLC2v, for ventricular cells, and MLC2a, for atrial cells) by immunohistochemistry (n = 100 cells). Scale bars, 10 µm. Dotted lines in the action potential traces indicate 0 mV.Summary of Ca2+ spark characteristics from control (black) and CPVT (red) cells under basal conditions when all myocytes or specifically ventricular (MLC2v+) and non-ventricular (MLC2v−) subtypes are analysed. Fold changes are relative to all myocytes control. Staining for MLC2v was performed after Ca2+ spark imaging. Between 21 and 113 cells from three iPSC lines were analysed per group. Data are means ± SEM from three independent experiments; p-values from two-tailed t-test.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3376852&req=5

fig05: Myocytic subtypes of iPSC-derived myocytes and their Ca2+ spark propertiesPercentage of ventricular-, atrial- and nodal-like myocytes after 3–4 month cardiac iPSC differentiation based on single cell electrophysiological measurements of action potentials (n = 47–50 cells) and expression of specific myocytic lineage markers (MLC2v, for ventricular cells, and MLC2a, for atrial cells) by immunohistochemistry (n = 100 cells). Scale bars, 10 µm. Dotted lines in the action potential traces indicate 0 mV.Summary of Ca2+ spark characteristics from control (black) and CPVT (red) cells under basal conditions when all myocytes or specifically ventricular (MLC2v+) and non-ventricular (MLC2v−) subtypes are analysed. Fold changes are relative to all myocytes control. Staining for MLC2v was performed after Ca2+ spark imaging. Between 21 and 113 cells from three iPSC lines were analysed per group. Data are means ± SEM from three independent experiments; p-values from two-tailed t-test.
Mentions: Since in CPVT patients tachycardia is restricted to the ventricles under stress condition, it might be expected that CPVT is a disease of ventricular cardiomyocytes. Our cardiac differentiation protocol leads to the generation of all three subtypes of cardiomyocytes, namely ventricular-like, atrial-like and nodal-like cells, which can be distinguished by the expression of specific myocytic lineage markers and by the shape of the action potential (Moretti et al, 2010b). Immunohistochemical analysis for ventricular and atrial myosin light chain 2 (MLC2v and MLC2a) proteins and electrophysiological measurements of action potentials in single iPSC-derived myocytes demonstrated that the ventricular subtype is largely predominant, accounting for 70–80% of all myocytes similarly in both control and CPVT groups (Fig 5A). To assess whether Ca2+ spark properties were specifically altered only in ventricular cells, we stained the cells with an antibody against MLC2v retrospectively and analysed spark data from ventricular (MLC2v+ cells) and non-ventricular (MLC2v− cells) myocytes separately. The observed differences in Ca2+ spark properties between control and CPVT cells persisted in the ventricular and non-ventricular subpopulations (Fig 5B), indicating that the mutated S406L-RYR2 channels are dysfunctional in all myocytes.

Bottom Line: In patient iPSC-derived cardiomyocytes, catecholaminergic stress led to elevated diastolic Ca(2+) concentrations, a reduced SR Ca(2+) content and an increased susceptibility to DADs and arrhythmia as compared to control myocytes.Dantrolene, a drug effective on malignant hyperthermia, restored normal Ca(2+) spark properties and rescued the arrhythmogenic phenotype.This suggests defective inter-domain interactions within the RYR2 channel as the pathomechanism of the S406L mutation.

View Article: PubMed Central - PubMed

Affiliation: Klinikum rechts der Isar, Technische Universität München, I. Medizinische Klinik, Kardiologie, München, Germany.

Show MeSH
Related in: MedlinePlus