Dataset Information

Investigation of the Role of Myocyte Orientations in Cardiac Arrhythmia Using Image-Based Models.

ABSTRACT: Cardiac electrical excitation-propagation is influenced by myocyte orientations (cellular organization). Quantitatively understanding this relationship presents a significant research challenge, especially during arrhythmias in which excitation patterns become complex. Tissue-scale simulations of cardiac electrophysiology, incorporating both dynamic action potential behavior and image-based myocardial architecture, provide an approach to investigate three-dimensional (3D) propagation of excitation waves in the heart. In this study, we aimed to assess the importance of natural variation in myocyte orientations on cardiac arrhythmogenesis using 3D tissue electrophysiology simulations. Three anatomical models (i.e., describing myocyte orientations) of healthy rat ventricles-obtained using diffusion tensor imaging at 100 ?m resolution-were registered to a single biventricular geometry (i.e., a single cardiac shape), in which the myocyte orientations could be represented by each of the diffusion tensor imaging data sets or by an idealized rule-based description. The Fenton-Karma cellular excitation model was modified to reproduce rat ventricular action potential duration restitution to create reaction-diffusion cardiac electrophysiology models. Over 250 3D simulations were performed to investigate the effects of myocyte orientations on the following: 1) ventricular activation, 2) location-dependent arrhythmia induction via rapid pacing, and 3) dynamics of re-entry averaged over multiple episodes. It was shown that 1) myocyte orientation differences manifested themselves in local activation times, but the influence on total activation time was small; 2) differences in myocyte orientations could critically affect the inducibility and persistence of arrhythmias for specific stimulus-location/cycle-length combinations; and 3) myocyte orientations alone could be an important determinant of scroll wave break, although no significant differences were observed in averaged arrhythmia dynamics between the four myocyte orientation scenarios considered. Our results show that myocyte orientations are an important determinant of arrhythmia inducibility, persistence, and scroll wave break. These findings suggest that where specificity is desired (for example, when predicting location-dependent, patient-specific arrhythmia inducibility), subject-specific myocyte orientations may be important.

SUBMITTER: Whittaker DG

PROVIDER: S-EPMC6990390 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature

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Investigation of the Role of Myocyte Orientations in Cardiac Arrhythmia Using Image-Based Models.

Whittaker Dominic G DG Benson Alan P AP Teh Irvin I Schneider Jürgen E JE Colman Michael A MA

Biophysical journal 20191008 12

Cardiac electrical excitation-propagation is influenced by myocyte orientations (cellular organization). Quantitatively understanding this relationship presents a significant research challenge, especially during arrhythmias in which excitation patterns become complex. Tissue-scale simulations of cardiac electrophysiology, incorporating both dynamic action potential behavior and image-based myocardial architecture, provide an approach to investigate three-dimensional (3D) propagation of excitati ...[more]

PMID: 31679763

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Project description:RationaleLoss-of-function studies in cardiac myocytes (CMs) are currently limited by the need for appropriate conditional knockout alleles. The factors that regulate CM maturation are poorly understood. Previous studies on CM maturation have been confounded by heart dysfunction caused by whole organ gene inactivation.ObjectiveTo develop a new technical platform to rapidly characterize cell-autonomous gene function in postnatal murine CMs and apply it to identify genes that regulate transverse tubules (T-tubules), a hallmark of mature CMs.Methods and resultsWe developed CRISPR/Cas9/AAV9-based somatic mutagenesis, a platform in which AAV9 delivers tandem guide RNAs targeting a gene of interest and cardiac troponin-T promoter-driven Cre to RosaCas9GFP/Cas9GFP neonatal mice. When directed against junctophilin-2 (Jph2), a gene previously implicated in T-tubule maturation, we achieved efficient, rapid, and CM-specific JPH2 depletion. High-dose AAV9 ablated JPH2 in 64% CMs and caused lethal heart failure, whereas low-dose AAV9 ablated JPH2 in 22% CMs and preserved normal heart function. In the context of preserved heart function, CMs lacking JPH2 developed T-tubules that were nearly morphologically normal, indicating that JPH2 does not have a major, cell-autonomous role in T-tubule maturation. However, in hearts with severe dysfunction, both adeno-associated virus-transduced and nontransduced CMs exhibited T-tubule disruption, which was more severe in the transduced subset. These data indicate that cardiac dysfunction disrupts T-tubule structure and that JPH2 protects T-tubules in this context. We then used CRISPR/Cas9/AAV9-based somatic mutagenesis to screen 8 additional genes for required, cell-autonomous roles in T-tubule formation. We identified RYR2 (Ryanodine Receptor-2) as a novel, cell-autonomously required T-tubule maturation factor.ConclusionsCRISPR/Cas9/AAV9-based somatic mutagenesis is a powerful tool to study cell-autonomous gene functions. Genetic mosaics are invaluable to accurately define cell-autonomous gene function. JPH2 has a minor role in normal T-tubule maturation but is required to stabilize T-tubules in the failing heart. RYR2 is a novel T-tubule maturation factor.

Project description:Recent advances have made it possible to readily derive cardiac myocytes from human induced pluripotent stem cells (hiPSC-CMs). HiPSC-CMs represent a valuable new experimental model for studying human cardiac muscle physiology and disease. Many laboratories have devoted substantial effort to examining the functional properties of isolated hiPSC-CMs, but to date, force production has not been adequately characterized. Here, we utilized traction force microscopy (TFM) with micro-patterning cell printing to investigate the maximum force production of isolated single hiPSC-CMs under varied culture and assay conditions. We examined the role of length of differentiation in culture and the effects of varied extracellular calcium concentration in the culture media on the maturation of hiPSC-CMs. Results show that hiPSC-CMs developing in culture for two weeks produced significantly less force than cells cultured from one to three months, with hiPSC-CMs cultured for three months resembling the cell morphology and function of neonatal rat ventricular myocytes in terms of size, dimensions, and force production. Furthermore, hiPSC-CMs cultured long term in conditions of physiologic calcium concentrations were larger and produced more force than hiPSC-CMs cultured in standard media with sub-physiological calcium. We also examined relationships between cell morphology, substrate stiffness and force production. Results showed a significant relationship between cell area and force. Implementing directed modifications of substrate stiffness, by varying stiffness from embryonic-like to adult myocardium-like, hiPSC-CMs produced maximal forces on substrates with a lower modulus and significantly less force when assayed on increasingly stiff adult myocardium-like substrates. Calculated strain energy measurements paralleled these findings. Collectively, these findings further establish single cell TFM as a valuable approach to illuminate the quantitative physiological maturation of force in hiPSC-CMs.

Dataset Information

Investigation of the Role of Myocyte Orientations in Cardiac Arrhythmia Using Image-Based Models.

Publications

Investigation of the Role of Myocyte Orientations in Cardiac Arrhythmia Using Image-Based Models.

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