Comparison of contractile behavior of native murine ventricular tissue and cardiomyocytes derived from embryonic or induced pluripotent stem cells
ABSTRACT Cardiomyocytes generated from embryonic stem cells (ESCs) and induced pluripotent stem (iPS) cells are suggested for repopulation of destroyed myocardium. Because contractile properties are crucial for functional regeneration, we compared cardiomyocytes differentiated from ES cells (ESC-CMs) and iPS cells (iPS-CMs). Native myocardium served as control. Murine ESCs or iPS cells were differentiated 11 d in vitro and cocultured 5-7 d with irreversibly injured myocardial tissue slices. Vital embryonic ventricular tissue slices of similar age served for comparison. Force-frequency relationship (FFR), effects of Ca(2+), Ni(2+), nifedipine, ryanodine, beta-adrenergic, and muscarinic modulation were studied during loaded contractions. FFR was negative for ESC-CMs and iPS-CMs. FFR was positive for embryonic tissue and turned negative after treatment with ryanodine. In all groups, force of contraction and relaxation time increased with the concentration of Ca(2+) and decreased with nifedipine. Force was reduced by Ni(2+). Isoproterenol (1 microM) increased the force most pronounced in embryonic tissue (207+/-31%, n=7; ESC-CMs: 123+/-5%, n=4; iPS-CMs: 120+/-4%, n=8). EC(50) values were similar. Contractile properties of iPS-CMs and ESC-CMs were similar, but they were significantly different from ventricular tissue of comparable age. The results indicate immaturity of the sarcoplasmic reticulum and the beta-adrenergic response of iPS-CMs and ESC-CMs.
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ABSTRACT: Long-QT syndrome mutations can cause syncope and sudden death by prolonging the cardiac action potential (AP). Ion channels affected by mutations are various, and the influences of cellular calcium cycling on LQTS cardiac events are unknown. To better understand LQTS arrhythmias, we performed current-clamp and intracellular calcium ([Ca2+]i) measurements on cardiomyocytes differentiated from patient-derived induced pluripotent stem cells (iPS-CM). In myocytes carrying an LQT2 mutation (HERG-A422T), APs and [Ca2+]i transients were prolonged in parallel. APs were abbreviated by nifedipine exposure and further lengthened upon releasing intracellularly stored Ca2+. Validating this model, control iPS-CM treated with HERG-blocking drugs recapitulated the LQT2 phenotype. In LQT3 iPS-CM, expressing NaV1.5-N406K, APs and [Ca2+]i transients were markedly prolonged. AP prolongation was sensitive to tetrodotoxin and to inhibiting Na+-Ca2+ exchange. These results suggest that LQTS mutations act partly on cytosolic Ca2+ cycling, potentially providing a basis for functionally targeted interventions regardless of the specific mutation site.Stem Cell Reports 08/2014; 3(2). DOI:10.1016/j.stemcr.2014.06.003
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ABSTRACT: Human stem cell-derived cardiomyocytes hold promise for heart repair, disease modeling, drug screens and questions of fundamental biology. All of these applications can be improved by assessing the contractility of cardiomyocytes at the single-cell level. We have developed an in vitro platform for assessing the contractile performance of stem cell-derived cardiomyocytes that is compatible with other common endpoints such as microscopy and molecular biology. Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were seeded onto elastomeric micropost arrays in order to characterize the contractile force, velocity, and power produced by these cells. We assessed contractile function by tracking the deflection of all of the microposts underneath an individual hiPSC-CM with optical microscopy. Immunofluorescent staining of these cells was used with the microposts to assess their spread area, nucleation, and sarcomeric structure. Following seeding of hiPSC-CMs onto microposts coated with fibronectin, laminin, and collagen IV, we found that hiPSC-CMs on laminin coatings demonstrated higher attachment, spread area, and contractile force than those seeded on fibronectin or collagen IV coatings. Under optimized conditions, hiPSC-CMs spread to an area of approximately 420 square microns, generated total systolic force of 15 nanonewtons per cell, showed contraction and relaxation rates of 6.5 microns per second and 5.25 microns per second, respectively, and had a peak upstroke power of 29 femtowatts. Thus, elastomeric micropost arrays can be used to study the contractile strength and kinetics of hiPSC-CMs. This system should facilitate studies of hiPSC-CM maturation, disease modeling and drug screens as well as fundamental studies of human cardiac contraction.Journal of Biomechanical Engineering 03/2014; 136(5). DOI:10.1115/1.4027145 · 1.75 Impact Factor
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ABSTRACT: An in vitro heart disease model is a promising model used for identifying the genes responsible for the disease, evaluating the effects of drugs, and regenerative medicine. We were interested in disease models using a patient-induced pluripotent stem (iPS) cell-derived cardiomyocytes because of their similarity to a patient's tissues. However, as these studies have just begun, we would like to review the literature in this and other related fields and discuss the path for future models of molecular biology that can help to diagnose and cure diseases, and its involvement in regenerative medicine. The heterogeneity of iPS cells and/or differentiated cardiomyocytes has been recognized as a problem. An in vitro heart disease model should be evaluated using molecular biological analyses, such as mRNA and micro-RNA expression profiles and proteomic analysis.03/2012; 2012:439219. DOI:10.1155/2012/439219