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Bioacoustic-enabled patterning of human iPSC-derived cardiomyocytes into 3D cardiac tissue.

ABSTRACT: The creation of physiologically-relevant human cardiac tissue with defined cell structure and function is essential for a wide variety of therapeutic, diagnostic, and drug screening applications. Here we report a new scalable method using Faraday waves to enable rapid aggregation of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) into predefined 3D constructs. At packing densities that approximate native myocardium (108-109 cells/ml), these hiPSC-CM-derived 3D tissues demonstrate significantly improved cell viability, metabolic activity, and intercellular connection when compared to constructs with random cell distribution. Moreover, the patterned hiPSC-CMs within the constructs exhibit significantly greater levels of contractile stress, beat frequency, and contraction-relaxation rates, suggesting their improved maturation. Our results demonstrate a novel application of Faraday waves to create stem cell-derived 3D cardiac tissue that resembles the cellular architecture of a native heart tissue for diverse basic research and clinical applications.

SUBMITTER: Serpooshan V

PROVIDER: S-EPMC5446052 | biostudies-other | 2017 Jul

REPOSITORIES: biostudies-other

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Bioacoustic-enabled patterning of human iPSC-derived cardiomyocytes into 3D cardiac tissue.

Serpooshan Vahid V Chen Pu P Wu Haodi H Lee Soah S Sharma Arun A Hu Daniel A DA Venkatraman Sneha S Ganesan Adarsh Venkataraman AV Usta Osman Berk OB Yarmush Martin M Yang Fan F Wu Joseph C JC Demirci Utkan U Wu Sean M SM

Biomaterials 20170328

The creation of physiologically-relevant human cardiac tissue with defined cell structure and function is essential for a wide variety of therapeutic, diagnostic, and drug screening applications. Here we report a new scalable method using Faraday waves to enable rapid aggregation of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) into predefined 3D constructs. At packing densities that approximate native myocardium (10<sup>8</sup>-10<sup>9</sup> cells/ml), these hiPSC-CM-d ...[more]

PMID: 28376365

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Project description:Natural polymer hydrogels are used ubiquitously as scaffold materials for cardiac tissue engineering as well as for soft tissue engineering more broadly because of FDA approval, minimal immunogenicity, and well-defined physiological clearance pathways. However, the relationships between natural polymer hydrogels and resident cell populations in directing the development of engineered tissues are poorly defined. This interaction is of particular concern for tissues prepared with iPSC-derived cell populations, in which population purity and batch-to-batch variability become additional critical factors to consider. Herein, the design space for a blended fibrin and collagen scaffold is characterized for applications in creating engineered myocardium with human iPSC-derived cardiomyocytes. Stiffness values of the acellular hydrogel formulations approach those of native myocardium in compression, but deviate significantly in tension when compared to rat myocardium in both transverse and longitudinal fiber orientations. A response surface methodology approach to understanding the relationship between collagen concentration, fibrin concentration, seeding density, and cardiac purity found a statistically significant predictive model across three repeated studies that confirms that all of these factors contribute to tissue compaction. In these constructs, increased fibrin concentration and seeding density were each associated with increased compaction, while increased collagen concentration was associated with decreased compaction. Both the lowest (24.4% cTnT+) and highest (60.2% cTnT+) cardiomyocyte purities evaluated were associated with decreased compaction, whereas the greatest compaction was predicted to occur in constructs prepared with a 40-50% cTnT+ population. Constructs prepared with purified cardiomyocytes (≥75.5% cTnT+) compacted and formed syncytia well, although increased fibrin concentration in these groups was associated with decreased compaction, a reversal of the trend observed in unpurified cardiomyocytes. This study demonstrates an analytical approach to understanding cell-scaffold interactions in engineered tissues and provides a foundation for the development of more sophisticated and customized scaffold platforms for human cardiac tissue engineering.

Project description:Purpose: Both cardiomyocytes and cardiac fibroblasts (CF) play essential roles in cardiac development, function, and remodeling. Properties of 3D co-cultures are incompletely understood. Hence, 3D co-culture of cardiomyocytes and CF was characterized, and selected features compared with single-type and 2D culture conditions. Methods: Human cardiomyocytes derived from induced-pluripotent stem cells (hiPSC-CMs) were obtained from Cellular Dynamics or Ncardia, and primary human cardiac fibroblasts from ScienCell. Cardiac spheroids were investigated using cryosections and whole-mount confocal microscopy, video motion analysis, scanning-, and transmission-electron microscopy (SEM, TEM), action potential recording, and quantitative PCR (qPCR). Results: Spheroids formed in hanging drops or in non-adhesive wells showed spontaneous contractions for at least 1 month with frequent media changes. SEM of mechanically opened spheroids revealed a dense inner structure and no signs of blebbing. TEM of co-culture spheroids at 1 month showed myofibrils, intercalated disc-like structures and mitochondria. Ultrastructural features were comparable to fetal human myocardium. We then assessed immunostained 2D cultures, cryosections of spheroids, and whole-mount preparations by confocal microscopy. CF in co-culture spheroids assumed a small size and shape similar to the situation in ventricular tissue. Spheroids made only of CF and cultured for 3 weeks showed no stress fibers and strongly reduced amounts of alpha smooth muscle actin compared to early spheroids and 2D cultures as shown by confocal microscopy, western blotting, and qPCR. The addition of CF to cardiac spheroids did not lead to arrhythmogenic effects as measured by sharp-electrode electrophysiology. Video motion analysis showed a faster spontaneous contraction rate in co-culture spheroids compared to pure hiPSC-CMs, but similar contraction amplitudes and kinetics. Spontaneous contraction rates were not dependent on spheroid size. Applying increasing pacing frequencies resulted in decreasing contraction amplitudes without positive staircase effect. Gene expression analysis of selected cytoskeleton and myofibrillar proteins showed more tissue-like expression patterns in co-culture spheroids than with cardiomyocytes alone or in 2D culture. Conclusion: We demonstrate that the use of 3D co-culture of hiPSC-CMs and CF is superior over 2D culture conditions for co-culture models and more closely mimicking the native state of the myocardium with relevance to drug development as well as for personalized medicine.

Dataset Information

Bioacoustic-enabled patterning of human iPSC-derived cardiomyocytes into 3D cardiac tissue.

Publications

Bioacoustic-enabled patterning of human iPSC-derived cardiomyocytes into 3D cardiac tissue.

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