Key result
Fluorescent cell cycle reporters in 3D cardiac models enable real-time tracking of cardiomyocyte proliferation.
Why the study?
Quantification of cardiomyocyte proliferation in high-throughput screens has remained problematic because conventional antibody-based staining is costly, technically challenging, and cannot discriminate division from failure in karyokinesis or cytokinesis.
Integrating advanced live cell cycle fluorescent reporters with 3D human iPSC-derived cardiac organoids and myocardial slices enables accurate, high-throughput screening for cardiac regenerative therapeutics by distinguishing true cardiomyocyte cytokinesis from multinucleation and polyploidization.
Cardiomyocytes in the adult human heart are quiescent and those lost following heart injury are not replaced by proliferating survivors. Considerable effort has been made to understand the mechanisms underlying cardiomyocyte cell cycle exit and re-entry, with view to discovering therapeutics that could stimulate cardiomyocyte proliferation and heart regeneration. The advent of large compound libraries and robotic liquid handling platforms has enabled the screening of thousands of conditions in a single experiment but success of these screens depends on the appropriateness and quality of the model used. Quantification of (human) cardiomyocyte proliferation in high throughput has remained problematic because conventional antibody-based staining is costly, technically challenging and does not discriminate between cardiomyocyte division and failure in karyokinesis or cytokinesis. Live cell imaging has provided alternatives that facilitate high-throughput screening but these have other limitations. Here, we (i) review the cell cycle features of cardiomyocytes, (ii) discuss various cell cycle fluorescent reporter systems, and (iii) speculate on what could improve their predictive value in the context of cardiomyocyte proliferation. Finally, we consider how these new methods can be used in combination with state-of-the-art three-dimensional human cardiac organoid platforms to identify pro-proliferative signalling pathways that could stimulate regeneration of the human heart.
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Salmenov et al. (2024) conducted a review in Cardiomyocyte proliferation and heart regeneration. Live cell cycle fluorescent reporter systems (e.g., FUCCI variants, localization-based markers, genetic tracing systems) vs. Conventional antibody-based staining (e.g., BrdU, EdU, Ki-67, PCNA, AurkB, pHH3) was evaluated on Real-time visualization and discrimination of cardiomyocyte cell cycle progression, karyokinesis, and cytokinesis. Live cell cycle fluorescent reporter systems integrated with 3D human cardiac models overcome traditional antibody staining limitations to track real-time cardiomyocyte proliferation.
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