A modular organoid platform decodes human stage-specific cardiogenesis
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo Sapiens
SUBMITTER: 
Deling Kong
PROVIDER: PXD065083 | iProX | Mon Jun 16 00:00:00 BST 2025
REPOSITORIES: iProX
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Project description:Cardiac maturation is an important developmental phase where there are profound biological and functional changes after birth in mammals. Herein, we use our profiling of human heart maturation in vivo to identify key drivers of maturation in our human cardiac organoid (hCO) model. After screening of various metabolism modulating factors, we established a directed maturation (DM) protocol to induce mature cardiac expression and compared the proteomic changes to our original serum free (SF) protocol. In this dataset, we compared 4 replicates of DM to 4 replicates of SF derived cardiac organoids using global DIA-MS/MS.
2025-04-17 | PXD059940 | Pride
Project description:Human induced pluripotent stem cell (hiPSC)-derived cardiovascular cells are promising cell source for cell therapy to repair the heart. Cardiac microtissue consisted of cardiomyocytes and fibroblast cells exhibited much better physiological functions. How different cardiovascular cell types interact and evolve in 3D microenvironment is unknown. In this study, we performed single-cell transcriptome profiling of hiPSC-derived mini-cardiac organoid consisted of cardiomyocytes, endothelial cells and smooth muscle cells. Our analysis showed that cardiac fibroblasts emerged spontaneously in 3D microenvironment which in turn facilitated the maturation of cardiomyocytes. HiPSC-derived cardiomyocytes, endothelial cells and smooth muscle cells assembled into mini-cardiac organoid in collagen-matrigel after 2 weeks. Single-cell study uncovered significant cell fate shift and improvement in cardiomyocyte maturation status upon-multilineage co-culture. Ligand-receptor analysis identified DLK1-Notch signaling to be one of the most upregulated pathways in the fibroblast population. Modulate the activity of DLK1-Notch signaling affected the assembly of the mini-cardiac organoid and the expression of immune regulatory genes. Interestingly, transplantation of trilineage mini-cardiac organoid into a rat model of myocardial infarction leads to significant improvement of cardiac function. Collectively, our single-cell analysis of mini-cardiac organoid provided rich information about cell fate dynamics and multilineage cross-talks occurred in the 3D microenvironment, which bring new insight on the molecular mechanism that promotes cardiomyocyte maturation and heart repair.
2023-02-09 | GSE166462 | GEO
Project description:Rationale: Human pluripotent stem cells-derived cardiomyocytes (hPSC-CMs) exhibit the properties of fetal CMs, which limit their applications. Various methods have been used to promote maturation of hPSC-CMs; however, there is a lack of an unbiased and comprehensive method for accurate benchmarking of hPSC-CM maturation.
Objective: We aim to develop an unbiased proteomics method integrating high-throughput top-down targeted proteomics and bottom-up global proteomics for accurate and comprehensive assessment of hPSC-CM maturation.
Methods and Results: Utilizing hPSC-CMs from early- and late-stage two-dimensional monolayer culture and three-dimensional engineered cardiac tissue, we demonstrated high reproducibility and reliability of the top-down proteomics method, which enabled simultaneous quantification of contractile protein isoform expressions and their PTMs. This method allowed for the detection of known maturation-associated contractile protein alterations, and for the first time, identified contractile protein PTMs as promising new markers of maturation. By employing a global proteomics strategy, we identified candidate maturation markers important for sarcomere organization, cardiac excitability, and Ca2+ homeostasis; and validated these markers in the developing mouse cardiac ventricles.
Conclusions: We established an unbiased proteomics method that can provide accurate and specific benchmarking of hPSC-CM maturation, and identified new markers of maturation. Furthermore, this integrated proteomics strategy laid a strong foundation for uncovering molecular basis underlying cardiac development and disease using hPSC-CMs.
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Project description:Cardiac maturation is an important developmental phase where there are profound biological and functional changes after birth in mammals. Herein, we use our profiling of human heart maturation in vivo to identify key drivers of maturation in our human cardiac organoid (hCO) model. After screening of various metabolism modulating factors, we established a directed maturation protocol to induce mature cardiac expression. We next compared directed maturation treatment to electrical pacing using phosphoproteomics in order to assess the similarities in the induction of maturation. The electrical pacing protocol utilized a custom platform, where we added Heart-Dyno inserts into C-pace system in 24-well plates enabling 120 bpm pacing for 5 minutes without causing toxicity. In this dataset, we compared 3 replicates of CTRL (our original serum free organoid protocol), electrical pacing (the standard protocol for maturation of cardiac stem cells), and directed maturation protocol (DM, new protocol) through phosphoproteomics.
2025-04-17 | PXD054810 | Pride
Project description:Crosstalk between cardiac cells is critical during heart development but its role in organ maturation is still largely uncharacterised. Here, we show that endothelial cells increase the force of contraction and enhance the expression of mature sarcomeric proteins and extracellular matrix (ECM) components in human pluripotent stem cell derived cardiac organoids (hCO). Endothelial cells regulate cardiac maturation and function both directly through secretion of ECM molecules and indirectly via paracrine signaling. Laminin α5, an endothelial enriched ECM protein, was identified as a key regulator of cardiac maturation and contractility in vitro. In vivo loss-of-function studies in mice confirmed that Lama5 was required for myocardial expansion during heart development in vivo. In addition, paracrine PDGF signaling was identified as a mediator of increased ECM deposition and cardiac contractility in hCO. This study uncovers matrix regulatory functions of endothelial cells governing cardiac maturation and highlights the importance of multicellularity for organoid models.
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Project description:Reactivating the human epicardium post-cardiac injury holds promise for cardiac tissue regeneration. Despite successful differentiation protocols yielding pure, self-renewing epicardial cells from induced pluripotent stem cells (iPSCs), these cells maintain an embryonic, proliferative state, impeding adult epicardial reactivation investigation. We introduce an optimized method that employs mammalian target of rapamycin (mTOR) signaling inhibition in embryonic epicardium, inducing a quiescent state that enhances multi-step epicardial maturation. This yields functionally mature epicardium, valuable for modeling adult epicardial reactivation. Furthermore, we assess cardiac organoids with cardiomyocytes and mature epicardium, probing molecular mechanisms governing epicardial quiescence during cardiac maturation. Our results highlight iPSC-derived mature epicardium's potential in investigating adult epicardial reactivation, pivotal for effective cardiac regeneration. Additionally, the cardiac organoid model offers insight into intricate cardiomyocyte-epicardium interactions in cardiac development and regeneration.
2025-06-30 | GSE252590 | GEO
Project description:PLK inhibitors identified by high content phenotypic screening promote maturation of human iPSC-derived cardiomyocytes
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Project description:Single-cell analysis of human iPSC-derived multi-lineage mini-cardiac organoid reveals molecular pathways to promote cardiomyocyte maturation and heart repair
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Project description:Cardiac maturation is an important developmental phase where there are profound biological and functional changes after birth in mammals. Herein, we use our profiling of human heart maturation in vivo to identify key drivers of maturation in our human cardiac organoid (hCO) model. In this dataset, we exemplified the applicability of our mature organoids in modelling cardiac contraction. Three calsequesterin-2 (CASQ2) knock out (-/-) hCO lines were generated to demonstrate the effect of sarcoplasmic reticulum Ca2+ leakiness on contraction. In this dataset, we evaluate the proteomic remodelling induced by CASQ2 (-/-, n=3) versus CTRL mature hCOs (n=1).
2025-04-17 | PXD055266 | Pride
Project description:The mammalian heart undergoes major transitions during postnatal life to acquire the physiological properties of an adult organ. Postnatal life imposes numerous adaptations including electrophysiological, structural and metabolic maturation of cardiomyocytes1, which occur coincident with loss of proliferative capacity and regenerative potential2,3. The discovery of key upstream drivers of cardiomyocyte maturation and cell cycle arrest remains one of the most important unanswered questions in cardiac biology. Discovery of these drivers would facilitate current attempts to promote cardiomyocyte maturation in vitro for drug discovery and to de-differentiate adult cardiomyocytes in vivo for regenerative medicine. A recent study has suggested that the shift from a low oxygen environment in utero towards a high oxygen environment after birth acts as a key trigger for cardiomyocyte cell cycle exit4. Moreover, it was recently demonstrated that proliferative adult cardiomyocytes reside in a hypoxic niche5 and that exposure of adult mice to gradual hypoxemia is sufficient to drive cell cycle re-entry and regeneration following infarction6. However, it is currently unclear whether postnatal changes in oxygen tension or the associated shifts in cardiomyocyte metabolism are sufficient to promote maturation and cell cycle arrest as human pluripotent stem cell (hPSC)-derived cardiomyocytes fail to mature when cultured at 21% oxygen7,8. There are considerable changes in metabolic substrate provision during early postnatal life. The mammalian heart relies on high concentrations of carbohydrates and the presence of insulin in utero but later switches to fatty acid dominated substrates present in milk and low insulin levels post-birth9. In order to adapt to these changes in substrates, cardiomyocytes upregulate the genes required for fatty acid oxidation after birth10. The importance of these metabolic adaptations for cardiomyocyte maturation has been difficult to study because genetic disruption of fatty acid oxidation components in vivo can have a broad range of negative health impacts11. Therefore, there is a need to develop alternative approaches for studying the impact of cardiomyocyte metabolism on the maturation process. hPSCs are now widely used for the generation of defined human somatic cell types, including cardiomyocytes. These cardiomyocytes have now been used extensively for developmental studies, drug screening, disease modeling, and heart repair. However, lack of maturity and inappropriate responses to pharmacological agents have been identified as limitations in 2D or embryoid body based differentiation strategies12. To improve maturity of hPSC-derived cardiomyocytes, long-term culture can be used13, although long-term cultures may not be amenable to high-throughput screening applications and adult-like maturity is still not achieved14. In an effort to better simulate heart muscle structure and function, cardiac tissue engineering to form 3D engineered heart tissue has been used15-19. However, despite these recent advances in human cardiac tissue engineering, cardiac tissues derived from hPSC still lack many features of fully mature adult heart tissue20. Moreover, engineered heart tissue fabrication, culture, mechanical loading and pacing protocols, and analysis methods using organ baths are costly, labor intensive, and the multiple handling steps induce variability. In order to facilitate higher-throughput experiments, platforms for engineered heart tissue production have been miniaturized, however, screening experiments using semi-automated force of contraction analyses have only been published in 24-well plate formats21. Therefore, we developed a novel 96-well device, the heart dynamometer (Heart-Dyno), for high-throughput functional screening of human cardiac organoids (hCOs) to facilitate screening on a larger scale. The Heart-Dyno is designed to facilitate automated formation of dense muscle bundles from minimal cells and reagents while also facilitating culture and automated force of contraction analysis without any tissue handling. Using the Heart-Dyno, we define serum-free 3D culture conditions that promote structural, electrophysiological, metabolic and proliferative maturation of hPSC-derived cardiac organoids. Furthermore, we uncover a metabolic mechanism governing cardiomyocyte cell cycle arrest through repression of a β-catenin and YAP1 dependent signalling.
2017-11-21 | PXD005736 | Pride