The structural and functional maturation of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) is essential for pharmaceutical testing, disease modeling, and ultimately therapeutic use. Multicellular 3D-tissue platforms have improved the functional maturation of hiPSC-CMs, but probing cardiac contractile properties in a 3D environment remains challenging, especially at depth and in live tissues. Using small-angle X-ray scattering (SAXS) imaging, we show that hiPSC-CMs matured and examined in a 3D environment exhibit a periodic spatial arrangement of the myofilament lattice, which has not been previously detected in hiPSC-CMs. The contractile force is found to correlate with both the scattering intensity (
Clinical translation of stem cell therapies for heart disease requires electrical integration of transplanted cardiomyocytes. Generation of electrically matured human induced pluripotent stem cell–derived cardiomyocytes (hiPSC-CMs) is critical for electrical integration. Here, we found that hiPSC-derived endothelial cells (hiPSC-ECs) promoted the expression of selected maturation markers in hiPSC-CMs. Using tissue-embedded stretchable mesh nanoelectronics, we achieved a long-term stable map of human three-dimensional (3D) cardiac microtissue electrical activity. The results revealed that hiPSC-ECs accelerated the electrical maturation of hiPSC-CMs in 3D cardiac microtissues. Machine learning–based pseudotime trajectory inference of cardiomyocyte electrical signals further revealed the electrical phenotypic transition path during development. Guided by the electrical recording data, single-cell RNA sequencing identified that hiPSC-ECs promoted cardiomyocyte subpopulations with a more mature phenotype, and multiple ligand-receptor interactions were up-regulated between hiPSC-ECs and hiPSC-CMs, revealing a coordinated multifactorial mechanism of hiPSC-CM electrical maturation. Collectively, these findings show that hiPSC-ECs drive hiPSC-CM electrical maturation via multiple intercellular pathways.
more » « less- Award ID(s):
- 2038603
- NSF-PAR ID:
- 10480053
- Publisher / Repository:
- AAAS
- Date Published:
- Journal Name:
- Science Advances
- Volume:
- 9
- Issue:
- 10
- ISSN:
- 2375-2548
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Abstract R 2 = 0.44) and lattice spacing (R 2 = 0.46). The scattering intensity also correlates with lattice spacing (R 2 = 0.81), suggestive of lower noise in our structural measurement than in the functional measurement. Notably, we observed decreased myofilament ordering in tissues with a myofilament mutation known to lead to hypertrophic cardiomyopathy (HCM). Our results highlight the progress of human cardiac tissue engineering and enable unprecedented study of structural maturation in hiPSC-CMs. -
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Cardiomyocytes (CMs) generated from human pluripotent stem cells (hPSCs) are immature in their structure and function, limiting their potential in disease modeling, drug screening, and cardiac cellular therapies. Prior studies have demonstrated that coculture of hPSC‐derived CMs with other cardiac cell types, including endothelial cells (ECs), can accelerate CM maturation. To address whether the CM differentiation stage at which ECs are introduced affects CM maturation, the authors coculture hPSC‐derived ECs with hPSC‐derived cardiac progenitor cells (CPCs) and CMs and analyze the molecular and functional attributes of maturation. ECs have a more significant effect on acceleration of maturation when cocultured with CPCs than with CMs. EC coculture with CPCs increases CM size, expression of sarcomere, and ion channel genes and proteins, the presence of intracellular membranous extensions, and chronotropic response compared to monoculture. Maturation is accelerated with an increasing EC:CPC ratio. This study demonstrates that EC incorporation at the CPC stage of CM differentiation expedites CM maturation, leading to cells that may be better suited for in vitro and in vivo applications of hPSC‐derived CMs.
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Abstract Background Cardiac pathological outcome of metabolic remodeling is difficult to model using cardiomyocytes derived from human-induced pluripotent stem cells (hiPSC-CMs) due to low metabolic maturation.
Methods hiPSC-CM spheres were treated with AMP-activated protein kinase (AMPK) activators and examined for hiPSC-CM maturation features, molecular changes and the response to pathological stimuli.
Results Treatment of hiPSC-CMs with AMPK activators increased ATP content, mitochondrial membrane potential and content, mitochondrial DNA, mitochondrial function and fatty acid uptake, indicating increased metabolic maturation. Conversely, the knockdown of AMPK inhibited mitochondrial maturation of hiPSC-CMs. In addition, AMPK activator-treated hiPSC-CMs had improved structural development and functional features—including enhanced Ca2+transient kinetics and increased contraction. Transcriptomic, proteomic and metabolomic profiling identified differential levels of expression of genes, proteins and metabolites associated with a molecular signature of mature cardiomyocytes in AMPK activator-treated hiPSC-CMs. In response to pathological stimuli, AMPK activator-treated hiPSC-CMs had increased glycolysis, and other pathological outcomes compared to untreated cells.
Conclusion AMPK activator-treated cardiac spheres could serve as a valuable model to gain novel insights into cardiac diseases.
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Costameres, as striated muscle-specific cell adhesions, anchor both M-lines and Z-lines of the sarcomeres to the extracellular matrix. Previous studies have demonstrated that costameres intimately participate in the initial assembly of myofibrils. However, how costamere maturation cooperates with myofibril growth is still underexplored. In this work, we analyzed zyxin (costameres), α-actinin (Z-lines) and myomesin (M-lines) to track the behaviors of costameres and myofibrils within the cardiomyocytes derived from human induced pluripotent stem cells (hiPSC-CMs). We quantified the assembly and maturation of costameres associated with the process of myofibril growth within the hiPSC-CMs in a time-dependent manner. We found that asynchrony existed not only between the maturation of myofibrils and costameres, but also between the formation of Z-costameres and M-costameres that associated with different structural components of the sarcomeres. This study helps us gain more understanding of how costameres assemble and incorporate into the cardiomyocyte sarcomeres, which sheds a light on cardiomyocyte mechanobiology.more » « less
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Cardiomyocytes (CMs) are heart cells responsible for heart contraction and relaxation. CMs can be derived from human induced pluripotent stem cells (hiPSCs) with high yield and purity. Mature CMs can potentially replace dead and dysfunctional cardiac tissue and be used for screening cardiac drugs and toxins. However, hiPSCs-derived CMs (hiPSC-CMs) are immature, which limits their utilization. Therefore, it is crucial to understand how experimental variables, especially tunable ones, of hiPSC expansion and differentiation phases affect the hiPSC-CM maturity stage. This study applied clustering algorithms to day 30 cardiac differentiation data to investigate if any maturity-related cell features could be related to the experimental variables. The best models were obtained using k-means and Gaussian mixture model clustering algorithms based on the evaluation metrics. They grouped the cells based on eccentricity and elongation. The cosine similarity between the clustering results and the experimental parameters revealed that the Gaussian mixture model results have strong similarities of 0.88, 0.94, and 0.93 with axial ratio, diameter, and cell concentration.more » « less
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1126/sciadv.ade8513
