Hydrogels are attractive materials for tissue engineering, but efforts to date have shown limited ability to produce the microstructural features necessary to promote cellular self-organization into hierarchical three-dimensional (3D) organ models. Here we develop a hydrogel ink containing prefabricated gelatin fibres to print 3D organ-level scaffolds that recapitulate the intra- and intercellular organization of the heart. The addition of prefabricated gelatin fibres to hydrogels enables the tailoring of the ink rheology, allowing for a controlled sol–gel transition to achieve precise printing of free-standing 3D structures without additional supporting materials. Shear-induced alignment of fibres during ink extrusion provides microscale geometric cues that promote the self-organization of cultured human cardiomyocytes into anisotropic muscular tissues in vitro. The resulting 3D-printed ventricle in vitro model exhibited biomimetic anisotropic electrophysiological and contractile properties.
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Data generated or analysed during this study are included in the Article and its Supplementary Information files and publicly available at figshare at https://doi.org/10.6084/m9.figshare.22787714. Additional data may be obtained from the corresponding author upon reasonable request. Source data are provided with this paper.
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We thank M. Rosnach for photography and illustrations and A. G. Kleber for discussions regarding cardiac physiology. This work was sponsored by the John A. Paulson School of Engineering and Applied Sciences at Harvard University, the National Science Foundation through the Harvard University Materials Research Science and Engineering Center (DMR-1420570, DMR-2011754 to K.K.P.) and the National Institutes of Health and National Center for Advancing Translational Sciences (UH3HL141798 to W.T.P. and K.K.P.; UG3TR003279 to W.T.P. and K.K.P.) and the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIT) (RS-2023-00248477, 2022R1A2C2012738 to K.Y.L.). This work was also performed in part at the Harvard University Center for Nanoscale Systems, a member of the National Nanotechnology Coordinated Infrastructure Network, which is supported by the National Science Foundation under award no. ECCS-2025158. Microcomputed tomography imaging reported in this publication was supported by Harvard University, Center for Nanoscale Systems under National Institutes of Health award no. S10OD023519. We gratefully acknowledge financial support by the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (grant agreement no. 810104-PoInt to A.R.B.) and the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation, Project ID No. 201269156—SFB 1032 to A.R.B.). H.A.M.A. thanks the American Chemical Society for support through the Irving S. Sigal Postdoctoral Fellowship. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Harvard College and Sogang University are patent applicants for one pending utility patent, application number 16/756,214, with L.A.M. and K.K.P. included among the inventors. This patent application describes three-dimensional biological tissue formation using macromolecular-fibre-infused additive manufacturing inks. The remaining authors declare no competing interests.
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Supplementary Figs. 1–12, note and references.
Direct 3D printing of ventricle-shaped scaffolds without using supporting materials.
Beating hiPSC-CMs after the differentiation process is finished.
Optical mapping images for the Ca2+ transient propagation of NRVM tissues cultured on the printed Gel–Alg hydrogel (left) and FIG scaffolds (right). A 1 Hz point electrical stimulation was applied at the left top corner of each tissue.
Optical mapping images for the Ca2+ transient propagation of hiPSC-CM tissues cultured on the printed FIG scaffolds under 1 Hz (left) and 2 Hz (right) point electrical stimulation at the left top corner.
Optical mapping images for the Ca2+ wave propagation occurred through the printed FIG patterns, showing sequential muscle activation.
Contraction of NRVM tissue on the 3D-printed rectangular scaffolds printed in the parallel, angled and perpendicular directions to the long side of the scaffold geometry, showing different contractile motions—rolling, twisting and bending, respectively—under 0.5 Hz field electrical stimulation.
Spontaneous contraction of 3D in vitro hiPSC-CM ventricle model in the RPMI/B27(–) media.
Spontaneous contraction of 3D in vitro NRVM ventricle model in Tyrode’s solution.
Ca2+ transient propagation from a 3D in vitro NRVM ventricle model under 1 Hz point electrical stimulation. The point stimulation was applied where the propagation started.
Spontaneous Ca2+ transient propagation from a 3D in vitro hiPSC-CM ventricle model observed at the top, middle and bottom planes.
Deformation mapping analysis from a 3D in vitro hiPSC-CM ventricle model. Movement of fluorescent beads adhered on the ventricle (left panel) was analysed during the systole and diastole cycle of the 3D in vitro ventricle tissue model (right panel).
Contractility assessment of a 3D in vitro hiPSC-CM ventricle tissue model using PIV analysis. Bright-field micrograph of a 3D in vitro hiPSC-CM ventricle model (left) with a green region of interest box (left), fluorescent bead movement from the corresponding region of interest (top, right) and phase-averaged velocity map from the region of interest (bottom, right).
G-code files used for the 3D-printed structures in Fig. 1.
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Choi, S., Lee, K.Y., Kim, S.L. et al. Fibre-infused gel scaffolds guide cardiomyocyte alignment in 3D-printed ventricles. Nat. Mater. 22, 1039–1046 (2023). https://doi.org/10.1038/s41563-023-01611-3