Comparison of contractile behavior of native murine ventricular tissue and cardiomyocytes derived from embryonic or induced pluripotent stem cells

Institute for Neurophysiology, University of Cologne, Cologne, Germany.
The FASEB Journal (Impact Factor: 5.04). 04/2010; 24(8):2739-51. DOI: 10.1096/fj.09-145177
Source: PubMed


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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    • "In LQT3 iPS-CM, we observed similar results to controls, where nifedipine caused AP shortening and arrested beating (Figure S4C). The more sustained automaticity in nifedipine-treated LQT2 iPS-CM may reflect a specific genetic process in that individual, or perhaps compensatory automaticity unmasked by the I Kr insufficiency (Itzhaki et al., 2011; Xi et al., 2010). When a cardiac AP is triggered, I Ca evokes a [Ca 2+ ] i "
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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 · 5.37 Impact Factor
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    • "Traditional video-based CM analysis methods [10,11] may not, however, be optimal for the study of single iPS cell-derived CMs. The sarcomere structure of iPS cell-derived CMs is not fully organized [12] and, therefore, their beating is less uniform with no main contraction direction. Thus, better methods are required that are robust in the detection of movement signals from these types of cells. "
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    ABSTRACT: The functionality of a cardiomyocyte is primarily measured by analyzing the electrophysiological properties of the cell. The analysis of the beating behavior of single cardiomyocytes, especially ones derived from stem cells, is challenging but well warranted. In this study, a video-based method that is non-invasive and label-free is introduced and applied for the study of single human cardiomyocytes derived from induced pluripotent stem cells. The beating of dissociated stem cell-derived cardiomyocytes was visualized with a microscope and the motion was video-recorded. Minimum quadratic difference, a digital image correlation method, was used for beating analysis with geometrical sectorial cell division and radial/tangential directions. The time series of the temporal displacement vector fields of a single cardiomyocyte was computed from video data. The vector field data was processed to obtain cell-specific, contraction-relaxation dynamics signals. Simulated cardiomyocyte beating was used as a reference and the current clamp of real cardiomyocytes was used to analyze the electrical functionality of the beating cardiomyocytes. Our results demonstrate that our sectorized image correlation method is capable of extracting single cell beating characteristics from the video data of induced pluripotent stem cell-derived cardiomyocytes that have no clear movement axis, and that the method can accurately identify beating phases and time parameters. Our video analysis of the beating motion of single human cardiomyocytes provides a robust, non-invasive and label-free method to analyze the mechanobiological functionality of cardiomyocytes derived from induced pluripotent stem cells. Thus, our method has potential for the high-throughput analysis of cardiomyocyte functions.
    BioMedical Engineering OnLine 04/2014; 13(1):39. DOI:10.1186/1475-925X-13-39 · 1.43 Impact Factor
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    • "Others are computationally complex or require mathematical estimations in order to determine twitch forces [17] [18] [19] [20]. The contractile properties of a population of cardiomyocytes have also been approximated by seeding these cells within a three-dimensional construct [33] [34] [35] [36] [37] [38] [39]. However, in this configuration , it is difficult to determine the actual contractile force produced by individual cells, since a portion of this contraction is lost to the rigidity of the bulk tissue and/or through its cell-cell contacts. "
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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 · 1.78 Impact Factor
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