Informace o publikaci
ATOMIC FORCE MICROSCOPY AND HUMAN PLURIPOTENT STEM CELL-DERIVED CARDIOMYOCYTES FORMS A BIOSENSOR OF PHYSIOLOGICAL CHANGES AND ADRENERGIC MODUALTION THROUGH DYNAMIC FORCE-CONTRACTION ANALYSIS
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| Rok publikování | 2015 |
| Druh | Konferenční abstrakty |
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| Popis | A differentiation of human pluripotent stem cells (PSCs) into the functional cardiomyocytes presents a powerful tool for cardiac diseases modeling when coupled with sensitive and robust detection of physiological response of cardiac cells. Although the contractions of the cell clusters resemble heart functionality and the phenotype description of stem cell-derived cardiomyocytes (SC-CMs) is crucial parameter for understanding of physiology and further modeling of the disease, the analysis of drug effects and the environmental conditions on the maturity and functional characteristics of SC-CMs usually depends on the technologically challenging techniques, such as patch clamp or Ca2+ imaging disqualifying the method from the routine use. We thus developed a novel biosensor combining the atomic force microscope (AFM) with SC-CMs differentiated in vitro from human embryonic SCs and human induced PSCs. Methods: Defined number of undifferentiated single cells in AggreWell plates formed homogeneous spheroids. The differentiation process was achieved using defined growth factors (ActivinA, BMP4, IWR2, VEGF, FGF2) at different stages to enhance mesodermal differentiation and production of cardiac progenitors. Molecular and functional characterization confirmed the cardiac identity of the resulting cells. AFM experiments were performed in Tyrode’s solution; different ion concentrations and temperature effects were tested, as well as drugs modulating the beta-adrenergic receptors were applied. Dynamics of uniform-sized and homogenous spheroid embryonic and induced PSCs-CMs were well measurable using the AFM-based technique. The beat rates were comparable among measured clusters and did not show high variability when thermal stability was assured. Spheroids responded comparably upon stimulation or inhibition of the beta-adrenergic pathway. Thermal and ionic dependency curves were obtained. Mechano-biological properties of homogenous beating spheroids, containing SC-CMs can be investigated by atomic force microscopy. The method allows testing of various physiological conditions as well as novel drugs on 3D homogeneous spheroid containing beating human cardiomyocytes. |