Chronic Myotrope Treatment Induces Contractile and Myofibrillar Remodeling in Cardiomyocytes

Introduction: : Hypertrophic and dilated cardiomyopathies (HCM and DCM) are hereditary diseases that impair cardiac function, characterized by hallmark cellular features including altered force generation and disrupted myofibril structure. These conditions are increasingly treated with cardiac myotropes such as Mavacamten and Omecamtiv Mecarbil (OM). Mavacamten reduces contractility and is approved by the FDA for patients with HCM. In contrast, OM enhances contractility and is a promising candidate for DCM, although it has not yet received regulatory approval. In our previous work, a 1-hour treatment with 50 nM Mavacamten reduced contractile force in human induced pluripotent stem cell–derived cardiomyocytes (hiPSC-CMs), as measured by traction force microscopy (TFM) (Pardon, Nature Communications, 2024). Similarly, Ribeiro (Ribeiro, Circulation Research, 2017) showed that 100 nM OM rapidly disrupted myofibril structure in hiPSC-CMs. Other studies have applied myotropes treatments in vitro for up to 12 hours (Solis, FEBS Journal, 2023), but most reports use shorter, 1-hour exposures (Toepfer, Circulation Research, 2019). Despite widespread use of short-term treatments, the chronic effects of myotropes on contractile force modulation and cytoskeletal organization in cardiomyocytes remain largely unexplored in vitro.

Materials and

Methods: : To investigate chronic effects—here defined as a 5-day continuous exposure—we used hiPSC-CMs endogenously labeled with α-actinin 2, a myofibril protein present in the Z-line. Cells were cultured on single-cell micropatterned polyacrylamide gels (1500 μm² area, 7:1 aspect ratio) with a physiological stiffness of 10 kPa and treated for 5 days with Mavacamten (10 nM or 50 nM), OM (100 nM [OM-100] or 1 µM [OM-1000]), or DMSO (control). After treatment, contractile force was measured via TFM in Mavacamten and OM-treated and control groups. Cells were then fixed, and myofibril morphology was analyzed by segmenting Z-line width and number using Labkit, a Fiji plugin.

Results, Conclusions, and Discussions:: Preliminary results showed that OM-1000 increased contractile force by approximately 600 nN compared to DMSO in the mean value, and led to a 34% increase in Z-line width. OM-100 did not significantly affect either contractile force or Z-line morphology. Additionally, the number of Z-lines per cell did not differ significantly among treatment groups after the 5-day exposure, as illustrated in Figure 1. Ongoing work includes additional OM treatment batches and continued analysis of chronic Mavacamten data.

Acknowledgements and/or References (Optional):: Acknowledgments: This research project was supported by the National Science Foundation(NSF) Graduate Research Fellowship, NSF Research Traineeship (DGE-2125644), Fulbright Spain Scholarship, NSF CMMI-2227509, and the National Institute of Health under grants NIH 5UG3TR002588-02 and 1RM1GM131981-01.