Biomechanical stress unmasks a fibroblast-dependent hypercontractile-disarray phenotype in MYBPC3 truncation HCM [dataset]
Description
This dataset contains experimental data supporting the study “Biomechanical Stress Unmasks a Fibroblast-Dependent Hypercontractile-Disarray Phenotype in MYBPC3 Truncation HCM.” Data were generated using engineered human micro-heart tissues (microHTs) composed of human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) and primary human cardiac fibroblasts (cFBs). Cardiomyocytes were derived from isogenic control and MYBPC3 frameshift mutant (MYBPC3fs) iPSC lines. The dataset includes quantitative measurements of tissue contractile function, mechanical properties, calcium transients, tissue morphology, and cellular organization. Key measurements include active force, resting tension, contraction kinetics, estimated contractile work, strain, stress, tissue dimensions, GCaMP6f-based calcium transient parameters, contraction directional uniformity, sarcomeric alignment, α-actinin-positive area, and α-smooth muscle actin (α-SMA) expression. Data were collected across experimental conditions varying cardiomyocyte genotype, added fibroblast fraction (0%, 5%, and 25%), and mechanical afterload (soft versus stiff posts). Additional experiments evaluated tissue responses to transforming growth factor-β1 (TGF-β1) stimulation. The deposited materials comprise 51 data files organized into 10 folder, containing CSV. The data originate from in vitro experiments using human iPSC-derived cardiomyocytes and primary human cardiac fibroblasts, rather than animal models or direct patient measurements. These datasets support the main and supplementary figures and enable evaluation of genotype-, mechanical load-, and fibroblast-dependent changes in engineered cardiac tissue function and structure.
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Steps to reproduce
Human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) were generated from isogenic control and MYBPC3 frameshift mutant (MYBPC3fs; G1206fs) cell lines carrying a GCaMP6f calcium reporter. iPSCs were differentiated through temporal modulation of Wnt signaling, followed by lactate-based cardiomyocyte purification. Purified iPSC-CMs were combined with primary human ventricular cardiac fibroblasts (cFBs) at targeted added-cFB fractions of 0%, 5%, or 25%, while maintaining constant total cell input. Cell mixtures were embedded in a collagen-based extracellular matrix and seeded into microfabricated wells surrounding paired flexible poly(dimethylsiloxane) (PDMS) posts. Mechanical afterload was controlled by adjusting PDMS post-bending stiffness to establish soft- and stiff-post conditions. In separate profibrotic experiments, tissues were treated with recombinant TGF-β1 (10 ng/mL) from culture day 2 through day 10. Functional measurements were performed on day 10 using a Nikon Eclipse Ts2R epifluorescence microscope equipped with a Hamamatsu ORCA-Flash4.0 V2 camera. Brightfield and GCaMP6f fluorescence recordings were acquired at 100 frames per second, with electrical field pacing at 1 Hz for paced measurements. Custom Python scripts tracked post displacement to calculate active force, resting tension, contraction kinetics, strain, and estimated contractile work using calibrated post stiffness. Tissue dimensions were measured from calibrated images to estimate cross-sectional area and contractile stress. Stiff-post stress calculations incorporated side-view thickness measurements, while soft-post calculations used a width-based circular approximation. GCaMP6f fluorescence recordings were analyzed using custom MATLAB scripts to quantify normalized calcium transient amplitude (ΔF/F0), upstroke time, and decay kinetics. For structural assessment, tissues were fixed and immunostained for sarcomeric α-actinin, vimentin, α-smooth muscle actin (α-SMA), and nuclei, followed by confocal microscopy. ImageJ was used to quantify marker-positive areas and fluorescence intensity. Sarcomeric organization was assessed using the orientational order parameter derived from α-actinin staining. Statistical analyses and graphical representations were performed using GraphPad Prism 9.3.1. Experiments included at least three independent batches. Detailed experimental procedures are described in the open access article Jiang et al., APL Bioengineering (2026), DOI: 10.1063/5.0336866.
Institutions
- Washington University in St. LouisMissouri, St Louis
Categories
Funders
- National Heart, Lung, and Blood InstituteNational Institutes of HealthMaryland, BethesdaGrant ID: R01HL159094
- National Heart, Lung, and Blood InstituteNational Institutes of HealthMaryland, BethesdaGrant ID: R01HL170079
Additional Metadata for Digital Commons Data@Becker
| Keywords | Hypertrophic Cardiomyopathy, MYBPC3, Cardiac Fibroblasts, Induced Pluripotent Stem Cells, Biomechanics, Myocardial Contraction, Engineered Cardiac Tissues, Calcium Signaling, Transforming Growth Factor beta1 |