Long-term effects of empagliflozin on excitation-contraction-coupling in human induced pluripotent stem cell cardiomyocy
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ORIGINAL ARTICLE
Long-term effects of empagliflozin on excitation-contraction-coupling in human induced pluripotent stem cell cardiomyocytes Steffen Pabel 1 & Florian Reetz 1 & Nataliya Dybkova 2 & Orr Shomroni 3 & Gabriela Salinas 3 & Julian Mustroph 1 & Karin P. Hammer 1 & Gerd Hasenfuss 2 & Nazha Hamdani 4 & Lars S. Maier 1 & Katrin Streckfuss-Bömeke 2 & Samuel Sossalla 1,2 Received: 13 May 2020 / Revised: 11 September 2020 / Accepted: 1 October 2020 # The Author(s) 2020
Abstract The SGLT2 inhibitor empagliflozin improved cardiovascular outcomes in patients with diabetes. As the cardiac mechanisms remain elusive, we investigated the long-term effects (up to 2 months) of empagliflozin on excitation-contraction (EC)-coupling in human cardiomyocytes derived from induced pluripotent stem cells (iPSC-CM) in a blinded manner. IPSC from 3 donors, differentiated into pure iPSC-CM (4 differentiations), were treated with a clinically relevant concentration of empagliflozin (0.5 μmol/l) or vehicle control. Treatment, data acquisition, and analysis were conducted externally blinded. Epifluorescence microscopy measurements in iPSC-CM showed that empagliflozin has neutral effects on Ca2+ transient amplitude, diastolic Ca2+ levels, Ca2+ transient kinetics, or sarcoplasmic Ca2+ load after 2 weeks or 8 weeks of treatment. Confocal microscopy determining possible effects on proarrhythmogenic diastolic Ca2+ release events showed that in iPSC-CM, Ca2+ spark frequency and leak was not altered after chronic treatment with empagliflozin. Finally, in patch-clamp experiments, empagliflozin did not change action potential duration, amplitude, or resting membrane potential compared with vehicle control after long-term treatment. Next-generation RNA sequencing (NGS) and mapped transcriptome profiles of iPSC-CMs untreated and treated with empagliflozin for 8 weeks showed no differentially expressed EC-coupling genes. In line with NGS data, Western blots indicate that empagliflozin has negligible effects on key EC-coupling proteins. In this blinded study, direct treatment of iPSC-CM with empagliflozin for a clinically relevant duration of 2 months did not influence cardiomyocyte EC-coupling and electrophysiology. Therefore, it is likely that other mechanisms independent of cardiomyocyte EC-coupling are responsible for the beneficial treatment effect of empagliflozin. Key messages & This blinded study investigated the clinically relevant long-term effects (up to 2 months) of empagliflozin on cardiomyocyte excitation-contraction (EC)-coupling. & Human cardiomyocytes derived from induced pluripotent stem cells (iPSC-CM) were used to study a human model including a high repetition number of experiments. & Empagliflozin has neutral effects on cardiomyocyte Ca2+ transients, sarcoplasmic Ca2+ load, and diastolic sarcoplasmic Ca2+ leak. & In patch-clamp experiments, empagliflozin did not change the action potential. & Next-generation RNA sequencing, mapped transcriptome profiles, and Western blots of iPSC-CM untreated and treated with empagliflozi
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