Analysis of calcium transients in cardiomyocytes derived from hiPSCs: the variant p. Y4734C in RYR2 vs. unrelated healthy controls

ŠVECOVÁ, Olga, Martin KRÁL, Štefan ZELENÁK, Jiří PACHERNÍK, Tomáš BÁRTA, Samuel LIETAVA, Iva SYNKOVÁ, Jana ZÍDKOVÁ, Tomáš NOVOTNÝ and Markéta BÉBAROVÁ. Analysis of calcium transients in cardiomyocytes derived from hiPSCs: the variant p. Y4734C in RYR2 vs. unrelated healthy controls. In 15th New Frontiers in Basic Cardiovascular Research: A France-New EU Members Symposium. 2024.

Basic information

• Original name: Analysis of calcium transients in cardiomyocytes derived from hiPSCs: the variant p. Y4734C in RYR2 vs. unrelated healthy controls
• Authors: ŠVECOVÁ, Olga, Martin KRÁL, Štefan ZELENÁK, Jiří PACHERNÍK, Tomáš BÁRTA, Samuel LIETAVA, Iva SYNKOVÁ, Jana ZÍDKOVÁ, Tomáš NOVOTNÝ and Markéta BÉBAROVÁ.
• Edition: 15th New Frontiers in Basic Cardiovascular Research: A France-New EU Members Symposium, 2024.

Other information

• Original language: English
• Type of outcome: Conference abstract
• Country of publisher: France
• Confidentiality degree: is not subject to a state or trade secret
• Organization unit: Faculty of Medicine
• Keywords (in Czech): calcium transient, hiPSC-derived cardiomyocytes

Abstract

Introduction: the regulation of intracellular calcium levels is crucial for excitation-contraction coupling. The release of calcium into the intracellular space is controlled by the ryanodine receptor type 2 (RYR2) located on the sarcoplasmic reticulum. Dysfunction of RYR2 is involved in the pathogenesis of inherited and non-inherited diseases such as cardiac arrhythmias, ventricular fibrillation, ventricular tachycardia, etc. The variant p. Y4734C in RYR2 was found in a patient with idiopathic ventricular fibrillation without structural changes in the heart and signs of arrhythmia on clinical examination. Preliminary data show calcium transient of patient-specific cardiomyocytes derived from human-induced pluripotent stem cells (hiPSC-CM). Methods: calcium transients of hiPSC-CM (Y4734C) and hiPSC-CM unrelated healthy controls (WT) were measured using the Myocyte Calcium and Contractility System (IonOptix LLC). Cell clusters were loaded with Fura-2 (Molecular Probes, Invitrogen) at a final concentration of 1 mM. Cells were incubated for 15 min in Tyrode solution with 1 μmol/L Fura-2-am at 37 °C and then washed repeatedly with Tyrode solution followed by incubation for 10 min in Tyrode solution at 37 °C. Measurements were performed in Tyrode solution at 37 ± 0.5 °C. Cells were not stimulated. Analysis of calcium transients was performed using CytoSolver software (IonOptix LLC). Results: calcium transient parameters and frequency of Y4734C and WT were evaluated. Time to peak and Time constant were significantly longer in Y4734C (0,24±xx and 0,20±xx s, respectively; n=8; P˂0.004 and P˂0.048) than in WT (0,09±xx and 0,11±xx s, respectively; n=4). A nonsignificant change was observed in the amplitude of the calcium transient, Y4734C (0,22±xx; n=8) and WT (0,15±xx; n=4). Cell clusters with the variant Y4734C have higher frequency then WT (40±xx and 28±xx b/min, respectively). Conclusions: the preliminary data showed delayed release of calcium from the sarcoplasmic reticulum in cells with the variant Y4734C and prolonged reabsorption of calcium back into the sarcoplasmic reticulum.

Links

• MUNI/A/1547/2023, interní kód MU: Name: Analýza (dys)funkce: od molekul k živému organismu

Investor: Masaryk University, Analysis of (dys)function: from molecules to the living organism

• NU22-02-00348, research and development project: Name: Funkční hodnocení genetických variant u případů klinicky „skutečné“ idiopatické fibrilace komor: in vitro a in silico modelování s cílem odhalit arytmogenní mechanismus

Investor: Ministry of Health of the CR, Subprogram 1 - standard