Molecular characterization and functional properties of induced pluripotent stem cells-derived cardiomyocytes from healthy and diseased individuals. Models for investigating inherited cardiac diseases
Joint Event on 3rd World Congress on Cardiology & 16th International Conference on Nutrition and Fitness
October 29-30, 2018 | London, UK
Ofer Binah
Technion-Israel Institute of Technology, Israel
Keynote : J Nutr Hum Health
Abstract:
Introduction: Duchenne Muscular Dystrophy (DMD) caused by
mutations in the DMD gene encoding the dystrophin protein, is
an X-linked disease affecting boys and teenagers and rarely adult
heterozygous females. DMD is characterized by progressive
muscle degeneration and weakness, loss of ambulation and
death by the late 20’s or early 30’s. Dilated cardiomyopathy
(DCM) is a major cause of morbidity and mortality in DMD
patients.
Hypothesis: Induced pluripotent stem cell-derived cardiomyocytes
(iPSC-CMs) generated from the DMD patients exhibit intracellular
[Ca2+]i handling and mechanical abnormalities. Our goal was to
decipher the mechanical and molecular mechanisms underlying
the abnormal [Ca2+]i handling and contraction in DMD patients.
Methods: Dystrophinâmutated iPSC-CMs were generated from
male and female DMD patients. To test the hypothesis, [Ca2+]i
transients and contractions were recorded from stimulated iPSCCMs
clusters using fura-2 fluorescence and video edge detector,
in the absence and presence of the β-adrenergic agonist
isoproterenol, which increases SR Ca2+ release through PKAregulated
Ryanodine (RyR2) channels. Specifically, we measured
the inotropic response to 10-9-10-6 M of isoproterenol using
the IonOptix calcium and contractility system. In addition,
metabolic indices were evaluated using liquid chromatography
followed by mass spectrometry and Seahorse XF analyser.
Results: Our experiments showed a concentration-dependent
positive inotropic and lusitropic effects in healthy iPSC-CMs, on
both [Ca2+]i transient and contraction parameters. In contrast,
compared to healthy iPSC-CMs, the female and male DMD
iPSC-CMs displayed a markedly depressed inotropic response
to isoproterenol. To decipher the underlying mechanism, we
determined SR Ca2+ release and capacity in DMD iPSC-CMs by
means of a brief application of caffeine (10 mM) which serves
as an opener of the RyR2 channel. In control iPSC-CMs, caffeine
caused an abrupt increase in [Ca2+]i, followed by a gradual
decline in [Ca2+]i level. In marked contrast to control iPSC-CMs,
the male DMD iPSC-CMs exhibited a much shorter response to
caffeine, while only 50% of the female DMD iPSC-CMs displayed
abnormal [Ca2+]i handling in response to caffeine. The caffeineinduced
Ca2+ signal area of DMD iPSC-CMs (male and 50%
of female) was smaller than control. In addition, the caffeineinduced
Ca2+ signal amplitude of DMD iPSC-CM (female) was
significantly smaller than control. In addition, Seahorse XF
analyser demonstrated decreased oxidative phosphorylation
accompanied by a correlated increase in glycolysis in DMD
iPSC-CMs. Accordingly, mass spectrometry analysis showed a
dramatic fall in phosphocreatine levels in DMD iPSC-CMs.
Conclusion: DMD iPSC-CMs exhibit an attenuated β-adrenergic
inotropic response, metabolic deficits and reduced energy stores.
Biography:
Ofer Binah is Chair of Physiology, Biophysics and Systems Biology, at the Ruth & Bruce Rappaport Faculty of Medicine, Technion, Israel. He is a cardiac physiologist working for the past 33 years on research topics related to cellular electrophysiology, mechanics, signalling pathways and arrhythmias. In addition, he investigated the cellular mechanisms whereby cytotoxic T lymphocytes destroy cardiomyocytes in the course of heart transplant rejection and inflammatory heart diseases. Since 2001 he is investigating the functional properties of human embryonic stem cell-derived cardiomyocytes, and have published several papers in this area. Over 10 years ago he has begun investigating iPSC-derived cardiomyocytes generated from both dermal fibroblasts and keratinocytes, from healthy volunteers and from patients with inherited cardiac pathologies, including inherited arrhythmias and cardiomyopathies such as Duchenne Muscular Dystrophy.
E-mail: binah@technion.ac.il
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