3D bioprinting of high cell-density heterogeneous tissue models through spheroid fusion within self-healing hydrogels

Abstract

Cellular models are needed to study human development and disease in vitro, including the screening of drugs for toxicity and efficacy. However, current approaches are limited in the engineering of functional tissue models with requisite cell densities and heterogeneity to appropriately model cell and tissue behaviors. Here, we develop a new bioprinting approach to transfer spheroids into self-healing support hydrogels at high resolution, which enables their patterning and fusion into high-cell density microtissues of prescribed spatial organization. As an example application, we bioprint induced pluripotent stem cell-derived cardiac microtissue models with spatially controlled cardiomyocyte and fibroblast cell ratios to replicate the structural and functional features of scarred cardiac tissue that arise following myocardial infarction, including reduced contractility and irregular electrical activity. The bioprinted in vitro model is combined with functional readouts to probe how various pro-regenerative microRNA treatment regimes influence tissue regeneration and recovery of function as a result of cardiomyocyte proliferation. This method is useful for a range of biomedical applications, including the development of precision models to mimic diseases and for the screening of drugs, particularly where high cell densities and heterogeneity are important.

Emergence of FRESH 3D printing as a platform for advanced tissue biofabrication

“… 38–40 Daly and co-workers have also explored 3D printing in yield-stress support baths using a range of chemistries and hydrogel microparticles. 41 The impact of FRESH printing is also apparent by the large number of research labs that have adopted the technique. A few examples include the FRESH printing of nanocellulose, 42 conductive hydrogels, 43 scaffolds for stem cell growth, 44 and ventricle-like heart chambers composed of beating cardiomyocytes.…”

Advances in Engineering Human Tissue Models

“…Moreover, AD approaches, and specifically 3DP techniques, based on coupling multimaterial printing with high performance bioinks (i.e., hydrogel solutions that act both as cell carriers, and structural components to control and direct cell activity and fate; Chimene et al, 2020 ) and biomolecules have been developed to obtain highly customisable, biofunctional, and mechanically compliant scaffolds (Chimene et al, 2016 ). The potential of bioprinting for building such biorelevant models is highlighted in various recent studies, seeking to develop highly biomimetic and functional tissues for disease modeling and drug testing (Kolesky et al, 2014 ; Horvath et al, 2015 ; Lee A. et al, 2019 ; Lee H. et al, 2019 ; Theodoridis et al, 2019 ; Daly et al, 2020 ). However, there are still challenges and limitations to be addressed before this novel approach is fully adopted by researchers.…”

Complex 3D bioprinting methods

“…A similar approach is reported to bioprint spheroids within hydrogels with shear-thinning and rapid self-healing property. 195 Cardiac microtissue models for disease modeling applications were fabricated using induced pluripotent stem cell-derived cardiac spheroids with spatially controlled cardiomyocyte and fibroblast cell ratios to replicate healthy and scarred cardiac tissue. Overall, these aspiration-assisted freeform bioprinting is an emerging technology that enables bioprinting of high-density tissues with precise control over microstructure and cellular heterogeneity.…”