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

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00216224%3A14110%2F24%3A00136408" target="_blank" >RIV/00216224:14110/24:00136408 - isvavai.cz</a>

Výsledek na webu

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DOI - Digital Object Identifier

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Jazyk výsledku

angličtina

Název v původním jazyce

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

Popis výsledku v původním jazyce

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.

Název v anglickém jazyce

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

Popis výsledku anglicky

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.

Druh

O - Ostatní výsledky

CEP obor

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OECD FORD obor

30201 - Cardiac and Cardiovascular systems

Projekt

<a href="/cs/project/NU22-02-00348" target="_blank" >NU22-02-00348: Funkční hodnocení genetických variant u případů klinicky „skutečné“ idiopatické fibrilace komor: in vitro a in silico modelování s cílem odhalit arytmogenní mechanismus</a><br>

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)<br>S - Specificky vyzkum na vysokych skolach

Rok uplatnění

2024

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů