Application of Patient-Specific iPSCs for Modelling and Treatment of X-Linked Cardiomyopathies

Figures

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  Figure 1. Generation of isogenic induced pluripotent stem cells (iPSCs) with distinctive X-chromosome inactivation status from the heterozygous female with X-linked mutation.

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  Figure 2. Differentiation of iPSCs into cardiomyocytes using commercially available kits from Thermo Fisher scientific: (A) the iPSCs were cultured as a monolayer in differentiation medium A (day 0-day 2), medium B (day 2-day 4), and cardiomyocyte maintenance medium (from day 4). Beating clusters usually can be observed by day 8, scale bar: 200 µm; (B) flow cytometry analysis indicates that the cardiac troponin-T (cTnT)-SERCA2-double positive cells constitute approximately 84%; (C) immunostaining of the cardiomyocytes dissociated from the beating clusters, scale bar: 50 µm.

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  Figure 3. MEA and optical mapping for studying the electrophysiology of the iPSC-derived cardiomyocytes: (A) microelectrode array (MEA) analysis for evaluating the field potential changes across the monolayer surface of iPSC-derived cardiomyocytes; (B) optical mapping-based methods for evaluating the changes in membrane potentials of a single iPSC-derived cardiomyocyte. EAD: early after depolarisation.

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  Figure 4. IonOptix recording of the change in cell length during contraction.

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  Figure 5. Generation of iPSC-derived cardiomyocytes from Fabry disease patient: (A) pedigree analysis of a family carrying the GLA-IVS4–919 G > A allele; (B) the iPSC line established from non-disease individual and FD-patient 1 were differentiated into cardiomyocytes and subjected to Western blot analysis; (C) a-GAL enzyme activity of the corresponding iPSC-derived cardiomyocytes. *: p < 0.05 between the indicated groups, N = 4.

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  Figure 6. Application of the iPSCs with a silenced wildtype LAMP2 allele for drug screening: (A) the iPSCs with silenced LAMP2 allele were cultured on 96-well plates and treated with DiscoveryProbe™ Epigenetics Compound Library (APExBio). The reactivation of X-chromosomes will be detected using Alexa488-conjugated antibodies specific to human LAMP2; (B) primary screening results.