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3D extrusion bioprinting

Nature Reviews Methods Primers volume 1, Article number: 75 (2021) Cite this article

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Abstract

Three-dimensional (3D) bioprinting strategies use computer-aided processes to enable automated simultaneous spatial patterning of cells and/or biomaterials. These technologies are suitable for a broad range of biomedical applications owing to their capability to produce structurally sophisticated and functionally relevant tissue constructs. Extrusion-based 3D bioprinting strategies were among the first modalities developed and are now arguably the most widely used for producing 3D tissue constructs. These technologies have rapidly evolved over the past two decades, providing a powerful tool set for the biofabrication of tissues that can facilitate translational efforts in the field. In this Primer, we describe the methodology of 3D extrusion bioprinting, focusing on the selection of hardware, software and bioinks. We expand upon recent advances in 3D extrusion bioprinting by illustrating the key variations that promote its biofabrication abilities. Finally, we provide an outlook on possible future refinements of the technology.

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Fig. 1: Extrusion bioprinting methods.
Fig. 2: Bioink designs.
Fig. 3: Variations on extrusion bioprinting strategies.
Fig. 4: Exemplary printability assessments.
Fig. 5: Shear stress assessment.
Fig. 6: In situ extrusion bioprinting strategies.
Fig. 7: Advances in extrusion (bio)printing methods.
Fig. 8: Artificial intelligence integration with extrusion (bio)printing methods.

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References

  1. Guillemot, F., Mironov, V. & Nakamura, M. Bioprinting is coming of age: report from the international conference on bioprinting and biofabrication in bordeaux (3b′09). Biofabrication 2, 010201 (2010).

    ADS  Google Scholar 

  2. Groll, J. et al. Biofabrication: reappraising the definition of an evolving field. Biofabrication 8, 013001 (2016).

    ADS  Google Scholar 

  3. Landers, R., Hübner, U., Schmelzeisen, R. & Mülhaupt, R. Rapid prototyping of scaffolds derived from thermoreversible hydrogels and tailored for applications in tissue engineering. Biomaterials 23, 4437–4447 (2002).

    Google Scholar 

  4. Jiang, T., Munguia-Lopez, J. G., Flores-Torres, S., Kort-Mascort, J. & Kinsella, J. M. Extrusion bioprinting of soft materials: an emerging technique for biological model fabrication. Appl. Phys. Rev. 6, 011310 (2019).

    ADS  Google Scholar 

  5. Ozbolat, I. T. & Hospodiuk, M. Current advances and future perspectives in extrusion-based bioprinting. Biomaterials 76, 321–343 (2016).

    Google Scholar 

  6. Ning, L. & Chen, X. A brief review of extrusion-based tissue scaffold bio-printing. Biotechnol. J. 12, 1600671 (2017).

    Google Scholar 

  7. Gao, G., Kim, B. S., Jang, J. & Cho, D.-W. Recent strategies in extrusion-based three-dimensional cell printing toward organ biofabrication. ACS Biomater. Sci. Eng. 5, 1150–1169 (2019).

    Google Scholar 

  8. Ouyang, L., Highley, C. B., Rodell, C. B., Sun, W. & Burdick, J. A. 3D printing of shear-thinning hyaluronic acid hydrogels with secondary cross-linking. ACS Biomater. Sci. Eng. 2, 1743–1751 (2016).

    Google Scholar 

  9. Wilson, S. A., Cross, L. M., Peak, C. W. & Gaharwar, A. K. Shear-thinning and thermo-reversible nanoengineered inks for 3D bioprinting. ACS Appl. Mater. Interfaces 9, 43449–43458 (2017).

    Google Scholar 

  10. Murphy, S. V. & Atala, A. 3D bioprinting of tissues and organs. Nat. Biotechnol. 32, 773–785 (2014).

    Google Scholar 

  11. Moroni, L. et al. Biofabrication strategies for 3D in vitro models and regenerative medicine. Nat. Rev. Mater. 3, 21–37 (2018).

    ADS  Google Scholar 

  12. Murphy, S. V., De Coppi, P. & Atala, A. Opportunities and challenges of translational 3D bioprinting. Nat. Biomed. Eng. 4, 370–380 (2019).

    Google Scholar 

  13. Heinrich, M. A. et al. 3D bioprinting: from benches to translational applications. Small 15, 1805510 (2019).

    Google Scholar 

  14. Levato, R. et al. From shape to function: the next step in bioprinting. Adv. Mater. 32, 1906423 (2020).

    Google Scholar 

  15. Peng, W., Unutmaz, D. & Ozbolat, I. T. Bioprinting towards physiologically relevant tissue models for pharmaceutics. Trends Biotechnol. 34, 722–732 (2016).

    Google Scholar 

  16. Moroni, L. et al. Biofabrication: a guide to technology and terminology. Trends Biotechnol. 36, 384–402 (2018).

    Google Scholar 

  17. Cui, X. et al. Advances in extrusion 3D bioprinting: a focus on multicomponent hydrogel-based bioinks. Adv. Healthc. Mater. 9, 1901648 (2020).

    Google Scholar 

  18. Schaffner, M., Rühs, P. A., Coulter, F., Kilcher, S. & Studart, A. R. 3D printing of bacteria into functional complex materials. Sci. Adv. 3, eaao6804 (2017).

    Google Scholar 

  19. Garciamendez-Mijares, C. E., Agrawal, P., García Martínez, G., Cervantes Juarez, E. & Zhang, Y. S. State-of-art affordable bioprinters: a guide for the DIY community. Apppl. Phys. Rev. 8, 031312 (2021).

    Google Scholar 

  20. Ravanbakhsh, H. et al. Emerging technologies in multi-material bioprinting. Adv. Mater. https://doi.org/10.1002/adma.202104730 (2021).

    Article  Google Scholar 

  21. Wu, Y., Ravnic, D. J. & Ozbolat, I. T. Intraoperative bioprinting: repairing tissues and organs in a surgical setting. Trends Biotechnol. 38, 594–605 (2020).

    Google Scholar 

  22. Li, H., Cheng, F., Orgill, D. P., Yao, J. & Zhang, Y. S. Handheld bioprinting strategies for in situ wound-dressing. Essays Biochem. 65, 533–543 (2021).

    Google Scholar 

  23. Kosik-Kozioł, A. et al. 3D bioprinted hydrogel model incorporating β-tricalcium phosphate for calcified cartilage tissue engineering. Biofabrication 11, 035016 (2019).

    ADS  Google Scholar 

  24. Sithole, M. N. et al. A 3D bioprinted in situ conjugated-co-fabricated scaffold for potential bone tissue engineering applications. J. Biomed. Mater. Res. A 106, 1311–1321 (2018).

    Google Scholar 

  25. Qiu, K. et al. 3D printed organ models with physical properties of tissue and integrated sensors. Adv. Mater. Technol. 3, 1700235 (2018).

    Google Scholar 

  26. Haghiashtiani, G. et al. 3D printed patient-specific aortic root models with internal sensors for minimally invasive applications. Sci. Adv. 6, eabb4641 (2020).

    ADS  Google Scholar 

  27. Lee, A. et al. 3D bioprinting of collagen to rebuild components of the human heart. Science 365, 482 (2019).

    ADS  Google Scholar 

  28. Kamio, T., Suzuki, M., Asaumi, R. & Kawai, T. Dicom segmentation and stl creation for 3D printing: a process and software package comparison for osseous anatomy. 3D Print. Med. 6, 1–12 (2020).

    Google Scholar 

  29. Mitsouras, D. et al. Medical 3D printing for the radiologist. Radiographics 35, 1965–1988 (2015).

    Google Scholar 

  30. Rocca, M., Fragasso, A., Liu, W., Heinrich, M. A. & Zhang, Y. S. Embedded multi-material extrusion bioprinting. SLAS Technol. 23, 154–163 (2017).

    Google Scholar 

  31. Lee, J. M. & Yeong, W. Y. Design and printing strategies in 3D bioprinting of cell-hydrogels: a review. Adv. Healthc. Mater. 5, 2856–2865 (2016).

    Google Scholar 

  32. Bader, C. et al. Making data matter: voxel printing for the digital fabrication of data across scales and domains. Sci. Adv. 4, eaas8652 (2018).

    ADS  Google Scholar 

  33. Skylar-Scott, M. A., Mueller, J., Visser, C. W. & Lewis, J. A. Voxelated soft matter via multimaterial multinozzle 3D printing. Nature 575, 330–335 (2019).

    ADS  Google Scholar 

  34. Wang, H. et al. Valve-based consecutive bioprinting method for multimaterial tissue-like constructs with controllable interfaces. Biofabrication 13, 035001 (2021).

    Google Scholar 

  35. Ma, K. et al. Application of robotic-assisted in situ 3D printing in cartilage regeneration with HAMA hydrogel: an in vivo study. J. Adv. Res. 23, 123–132 (2020).

    Google Scholar 

  36. Zhu, Z., Park, H. S. & Mcalpine, M. C. 3D printed deformable sensors. Sci. Adv. 6, eaba5575 (2020).

    ADS  Google Scholar 

  37. Zhu, Z. et al. 3D printed functional and biological materials on moving freeform surfaces. Adv. Mater. 30, 1707495 (2018).

    Google Scholar 

  38. Adib, A. A. et al. Direct-write 3D printing and characterization of a GelMA-based biomaterial for intracorporeal tissue engineering. Biofabrication 12, 045006 (2020).

    Google Scholar 

  39. Moncal, K. K. et al. Intra-operative bioprinting of hard, soft, and hard/soft composite tissues for craniomaxillofacial reconstruction. Adv. Funct. Mater. 31, 2010858 (2021).

    Google Scholar 

  40. Theus, A. S. et al. Bioprintability: physiomechanical and biological requirements of materials for 3D bioprinting processes. Polymers 12, 2262 (2020).

    Google Scholar 

  41. Chopin-Doroteo, M., Mandujano-Tinoco, E. A. & Krötzsch, E. Tailoring of the rheological properties of bioinks to improve bioprinting and bioassembly for tissue replacement. Biochim. Biophys. Acta Gen. Subj. 1865, 129782 (2020).

    Google Scholar 

  42. Kyle, S., Jessop, Z. M., Al-Sabah, A. & Whitaker, I. S. ‘Printability’ of candidate biomaterials for extrusion based 3D printing: state-of-the-art. Adv. Healthc. Mater. 6, 1700264 (2017).

    Google Scholar 

  43. Pakhomova, C., Popov, D., Maltsev, E., Akhatov, I. & Pasko, A. Software for bioprinting. Int. J. Bioprint. 6, 279 (2020).

    Google Scholar 

  44. Leppiniemi, J. et al. 3D-printable bioactivated nanocellulose–alginate hydrogels. ACS Appl. Mater. Interfaces 9, 21959–21970 (2017).

    Google Scholar 

  45. Göhl, J. et al. Simulations of 3D bioprinting: predicting bioprintability of nanofibrillar inks. Biofabrication 10, 034105 (2018).

    ADS  Google Scholar 

  46. Lemarié, L., Anandan, A., Petiot, E., Marquette, C. & Courtial, E.-J. Rheology, simulation and data analysis toward bioprinting cell viability awareness. Bioprinting 21, e00119 (2021).

    Google Scholar 

  47. Robu, A., Robu, N. & Neagu, A. in 2018 IEEE 12th International Symposium on Applied Computational Intelligence and Informatics (SACI) 1–5 (IEEE, 2018).

  48. Robu, A. & Stoicu-Tivadar, L. Simmmc — an informatic application for modeling and simulating the evolution of multicellular systems in the vicinity of biomaterials. Romanian J. Biophys. 26, 145–162 (2016).

    Google Scholar 

  49. Groll, J. et al. A definition of bioinks and their distinction from biomaterial inks. Biofabrication 11, 013001 (2018).

    ADS  Google Scholar 

  50. Gungor-Ozkerim, P. S., Inci, I., Zhang, Y. S., Khademhosseini, A. & Dokmeci, M. R. Bioinks for 3D bioprinting: an overview. Biomater. Sci. 6, 915–946 (2018).

    Google Scholar 

  51. Skardal, A. & Atala, A. Biomaterials for integration with 3-D bioprinting. Ann. Biomed. Eng. 43, 730–746 (2015).

    Google Scholar 

  52. Hospodiuk, M., Dey, M., Sosnoski, D. & Ozbolat, I. T. The bioink: a comprehensive review on bioprintable materials. Biotechnol. Adv. 35, 217–239 (2017).

    Google Scholar 

  53. Nulty, J., Schipani, R., Burdis, R. & Kelly, D. J. in Polymer-Based Additive Manufacturing (ed. Devine, D. M.) 187–218 (Springer, 2019).

  54. Kim, B. S., Das, S., Jang, J. & Cho, D.-W. Decellularized extracellular matrix-based bioinks for engineering tissue-and organ-specific microenvironments. Chem. Rev. 120, 10608–10661 (2020).

    Google Scholar 

  55. Morgan, F. L. C., Moroni, L. & Baker, M. B. Dynamic bioinks to advance bioprinting. Adv. Healthc. Mater. 9, 1901798 (2020).

    Google Scholar 

  56. Ravanbakhsh, H., Bao, G., Luo, Z., Mongeau, L. G. & Zhang, Y. S. Composite inks for extrusion printing of biological and biomedical constructs. ACS Biomater. Sci. Eng. 7, 4009–4026 (2021).

    Google Scholar 

  57. Loebel, C., Rodell, C. B., Chen, M. H. & Burdick, J. A. Shear-thinning and self-healing hydrogels as injectable therapeutics and for 3D-printing. Nat. Protoc. 12, 1521 (2017).

    Google Scholar 

  58. Müller, S. J. et al. Flow and hydrodynamic shear stress inside a printing needle during biofabrication. PLoS ONE 15, e0236371 (2020).

    Google Scholar 

  59. Chen, N. et al. Hydrogel bioink with multilayered interfaces improves dispersibility of encapsulated cells in extrusion bioprinting. ACS Appl. Mater. Interfaces 11, 30585–30595 (2019).

    Google Scholar 

  60. Ghavaminejad, A., Ashammakhi, N., Wu, X. Y. & Khademhosseini, A. Crosslinking strategies for 3D bioprinting of polymeric hydrogels. Small 16, 2002931 (2020).

    Google Scholar 

  61. Ying, G., Jiang, N., Yu, C. & Zhang, Y. S. Three-dimensional bioprinting of gelatin methacryloyl (GelMA). Biodesign Manuf. 1, 215–224 (2018).

    Google Scholar 

  62. Skardal, A. et al. A hydrogel bioink toolkit for mimicking native tissue biochemical and mechanical properties in bioprinted tissue constructs. Acta Biomater. 25, 24–34 (2015).

    Google Scholar 

  63. Bertlein, S. et al. Thiol–ene clickable gelatin: a platform bioink for multiple 3D biofabrication technologies. Adv. Mater. 29, 1703404 (2017).

    Google Scholar 

  64. Tytgat, L. et al. Additive manufacturing of photo-crosslinked gelatin scaffolds for adipose tissue engineering. Acta Biomater. 94, 340–350 (2019).

    Google Scholar 

  65. Fisch, P., Broguiere, N., Finkielsztein, S., Linder, T. & Zenobi-Wong, M. Bioprinting of cartilaginous auricular constructs utilizing an enzymatically crosslinkable bioink. Adv. Funct. Mater. 31, 2008261 (2021).

    Google Scholar 

  66. Hong, S., Kim, J. S., Jung, B., Won, C. & Hwang, C. Coaxial bioprinting of cell-laden vascular constructs using a gelatin–tyramine bioink. Biomater. Sci. 7, 4578–4587 (2019).

    Google Scholar 

  67. Ouyang, L., Highley, C. B., Sun, W. & Burdick, J. A. A generalizable strategy for the 3D bioprinting of hydrogels from nonviscous photo-crosslinkable inks. Adv. Mater. 29, 1604983 (2017).

    Google Scholar 

  68. Berg, J. et al. Optimization of cell-laden bioinks for 3D bioprinting and efficient infection with influenza A virus. Sci. Rep. 8, 13877 (2018).

    ADS  Google Scholar 

  69. Gao, T. et al. Optimization of gelatin–alginate composite bioink printability using rheological parameters: a systematic approach. Biofabrication 10, 034106 (2018).

    ADS  Google Scholar 

  70. Yang, X. et al. Collagen-alginate as bioink for three-dimensional (3D) cell printing based cartilage tissue engineering. Mater. Sci. Eng. C. 83, 195–201 (2018).

    Google Scholar 

  71. Li, C. et al. Rapid formation of a supramolecular polypeptide–DNA hydrogel for in situ three-dimensional multilayer bioprinting. Angew. Chem. Int. Ed. 54, 3957–3961 (2015).

    Google Scholar 

  72. Susapto, H. H. et al. Ultrashort peptide bioinks support automated printing of large-scale constructs assuring long-term survival of printed tissue constructs. Nano Lett. 21, 2719–2729 (2021).

    ADS  Google Scholar 

  73. Markstedt, K. et al. 3D bioprinting human chondrocytes with nanocellulose–alginate bioink for cartilage tissue engineering applications. Biomacromolecules 16, 1489–1496 (2015).

    Google Scholar 

  74. Distler, T. et al. 3D printed oxidized alginate-gelatin bioink provides guidance for C2C12 muscle precursor cell orientation and differentiation via shear stress during bioprinting. Biofabrication 12, 045005 (2020).

    Google Scholar 

  75. Chimene, D. et al. Nanoengineered ionic–covalent entanglement (nice) bioinks for 3D bioprinting. ACS Appl. Mater. Interfaces 10, 9957–9968 (2018).

    Google Scholar 

  76. Ouyang, L. et al. Expanding and optimizing 3D bioprinting capabilities using complementary network bioinks. Sci. Adv. 6, eabc5529 (2020).

    ADS  Google Scholar 

  77. Zhu, K. et al. A general strategy for extrusion bioprinting of bio-macromolecular bioinks through alginate-templated dual-stage crosslinking. Macromol. Biosci. 18, 1800127 (2018).

    Google Scholar 

  78. Ying, G.-L. et al. Aqueous two-phase emulsion bioink-enabled 3D bioprinting of porous hydrogels. Adv. Mater. 30, 1805460 (2018).

    Google Scholar 

  79. Bao, G. et al. Triggered micropore-forming bioprinting of porous viscoelastic hydrogels. Mater. Horiz. 7, 2336–2347 (2020).

    Google Scholar 

  80. Ying, G. et al. Bioprinted injectable hierarchically porous gelatin methacryloyl hydrogel constructs with shape-memory properties. Adv. Funct. Mater. 30, 2003740 (2020).

    Google Scholar 

  81. Ying, G. et al. An open-source handheld extruder loaded with pore-forming bioink for in situ wound dressing. Mater. Today Bio 8, 100074 (2020).

    Google Scholar 

  82. Highley, C. B., Song, K. H., Daly, A. C. & Burdick, J. A. Jammed microgel inks for 3D printing applications. Adv. Sci. 6, 1801076 (2019).

    Google Scholar 

  83. Kessel, B. et al. 3D bioprinting of macroporous materials based on entangled hydrogel microstrands. Adv. Sci. 7, 2001419 (2020).

    Google Scholar 

  84. Jang, J. et al. Tailoring mechanical properties of decellularized extracellular matrix bioink by vitamin B2-induced photo-crosslinking. Acta Biomater. 33, 88–95 (2016).

    Google Scholar 

  85. Kim, H. et al. Light-activated decellularized extracellular matrix-based bioinks for volumetric tissue analogs at the centimeter scale. Adv. Funct. Mater. 31, 2011252 (2021).

    Google Scholar 

  86. Wu, W., Deconinck, A. & Lewis, J. A. Omnidirectional printing of 3D microvascular networks. Adv. Mater. 23, H178–H183 (2011).

    Google Scholar 

  87. Kolesky, D. B. et al. 3D bioprinting of vascularized, heterogeneous cell-laden tissue constructs. Adv. Mater. 26, 3124–3130 (2014).

    Google Scholar 

  88. Homan, K. A. et al. Bioprinting of 3D convoluted renal proximal tubules on perfusable chips. Sci. Rep. 6, 34845 (2016).

    ADS  Google Scholar 

  89. Kolesky, D. B., Homan, K. A., Skylar-Scott, M. A. & Lewis, J. A. Three-dimensional bioprinting of thick vascularized tissues. Proct. Natl Acad. Sci. USA 113, 3179–3184 (2016).

    ADS  Google Scholar 

  90. Lin, N. Y. C. et al. Renal reabsorption in 3D vascularized proximal tubule models. Proct. Natl Acad. Sci. USA 116, 5399–5404 (2019).

    ADS  Google Scholar 

  91. Miller, J. S. et al. Rapid casting of patterned vascular networks for perfusable engineered three-dimensional tissues. Nat. Mater. 11, 768–774 (2012).

    ADS  Google Scholar 

  92. Bertassoni, L. E. et al. Hydrogel bioprinted microchannel networks for vascularization of tissue engineering constructs. Lab. Chip 14, 2202–2211 (2014).

    Google Scholar 

  93. Massa, S. et al. Bioprinted 3D vascularized tissue model for drug toxicity analysis. Biomicrofluidics 11, 044109 (2017).

    Google Scholar 

  94. Duchamp, M. et al. Sacrificial bioprinting of a mammary ductal carcinoma model. Biotechnol. J. 14, 1700703 (2019).

    Google Scholar 

  95. Liu, T. et al. Investigating lymphangiogenesis in a sacrificially bioprinted volumetric model of breast tumor tissue. Methods 190, 72–79 (2021).

    Google Scholar 

  96. Lee, V. K. et al. Creating perfused functional vascular channels using 3D bio-printing technology. Biomaterials 35, 8092–8102 (2014).

    Google Scholar 

  97. Ozturk, M. S. et al. High-resolution tomographic analysis of in vitro 3D glioblastoma tumor model under long-term drug treatment. Sci. Adv. 6, eaay7513 (2020).

    ADS  Google Scholar 

  98. Hinton, T. J. et al. Three-dimensional printing of complex biological structures by freeform reversible embedding of suspended hydrogels. Sci. Adv. 1, e1500758 (2015).

    ADS  Google Scholar 

  99. Bhattacharjee, T. et al. Writing in the granular gel medium. Sci. Adv. 1, e1500655 (2015).

    ADS  Google Scholar 

  100. Highley, C. B., Rodell, C. B. & Burdick, J. A. Direct 3D printing of shear-thinning hydrogels into self-healing hydrogels. Adv. Mater. 27, 5075–5079 (2015).

    Google Scholar 

  101. Jeon, O., Lee, Y. B., Hinton, T. J., Feinberg, A. W. & Alsberg, E. Cryopreserved cell-laden alginate microgel bioink for 3D bioprinting of living tissues. Mater. Today Chem. 12, 61–70 (2019).

    Google Scholar 

  102. Ning, L. et al. Embedded 3D bioprinting of gelatin methacryloyl-based constructs with highly tunable structural fidelity. ACS Appl. Mater. Interfaces 12, 44563–44577 (2020).

    Google Scholar 

  103. Hull, S. M. et al. 3D bioprinting using universal orthogonal network (union) bioinks. Adv. Funct. Mater. 31, 2007983 (2021).

    Google Scholar 

  104. Skylar-Scott, M. A. et al. Biomanufacturing of organ-specific tissues with high cellular density and embedded vascular channels. Sci. Adv. 5, eaaw2459 (2019).

    ADS  Google Scholar 

  105. Compaan, A. M., Song, K., Chai, W. & Huang, Y. Cross-linkable microgel composite matrix bath for embedded bioprinting of perfusable tissue constructs and sculpting of solid objects. ACS Appl. Mater. Interfaces 12, 7855–7868 (2020).

    Google Scholar 

  106. Luo, G. et al. Freeform, reconfigurable embedded printing of all-aqueous 3D architectures. Adv. Mater. 31, 1904631 (2019).

    Google Scholar 

  107. Chao, Y. & Shum, H. C. Emerging aqueous two-phase systems: from fundamentals of interfaces to biomedical applications. Chem. Soc. Rev. 49, 114–142 (2020).

    Google Scholar 

  108. Ma, Q. et al. Cell-inspired all-aqueous microfluidics: from intracellular liquid–liquid phase separation toward advanced biomaterials. Adv. Sci. 7, 1903359 (2020).

    Google Scholar 

  109. Duarte Campos, D. F. et al. Three-dimensional printing of stem cell-laden hydrogels submerged in a hydrophobic high-density fluid. Biofabrication 5, 015003 (2012).

    ADS  Google Scholar 

  110. Shin, S. et al. Solid matrix-assisted printing for three-dimensional structuring of a viscoelastic medium surface. Nat. Commun. 10, 4650 (2019).

    ADS  Google Scholar 

  111. Mccormack, A., Highley, C. B., Leslie, N. R. & Melchels, F. P. W. 3D printing in suspension baths: keeping the promises of bioprinting afloat. Trends Biotechnol. 38, 584–593 (2020).

    Google Scholar 

  112. Shiwarski, D. J., Hudson, A. R., Tashman, J. W. & Feinberg, A. W. Emergence of fresh 3D printing as a platform for advanced tissue biofabrication. APL. Bioeng. 5, 010904 (2021).

    Google Scholar 

  113. Onoe, H. et al. Metre-long cell-laden microfibres exhibit tissue morphologies and functions. Nat. Mater. 12, 584–590 (2013).

    ADS  Google Scholar 

  114. Kang, E. et al. Digitally tunable physicochemical coding of material composition and topography in continuous microfibres. Nat. Mater. 10, 877–883 (2011).

    ADS  Google Scholar 

  115. Costantini, M., Colosi, C., S´wie˛szkowski, W. & Barbetta, A. Co-axial wet-spinning in 3D bioprinting: state of the art and future perspective of microfluidic integration. Biofabrication 11, 012001 (2018).

    ADS  Google Scholar 

  116. Zhang, Y., Yu, Y. & Ozbolat, I. T. Direct bioprinting of vessel-like tubular microfluidic channels. J. Nanotechnol. Eng. Med. 4, 020902 (2013).

    Google Scholar 

  117. Zhang, Y. et al. In vitro study of directly bioprinted perfusable vasculature conduits. Biomater. Sci. 3, 134–143 (2015).

    Google Scholar 

  118. Jia, W. et al. Direct 3D bioprinting of perfusable vascular constructs using a blend bioink. Biomaterials 106, 58–68 (2016).

    Google Scholar 

  119. Colosi, C. et al. Microfluidic bioprinting of heterogeneous 3D tissue constructs using low-viscosity bioink. Adv. Mater. 28, 677–684 (2015).

    Google Scholar 

  120. Costantini, M. et al. Microfluidic-enhanced 3D bioprinting of aligned myoblast-laden hydrogels leads to functionally organized myofibers in vitro and in vivo. Biomaterials 131, 98–110 (2017).

    Google Scholar 

  121. Gao, G. et al. Tissue-engineering of vascular grafts containing endothelium and smooth-muscle using triple-coaxial cell printing. Appl. Phys. Rev. 6, 041402 (2019).

    ADS  Google Scholar 

  122. Colosi, C. et al. Rapid prototyping of chitosan-coated alginate scaffolds through the use of a 3D fiber deposition technique. J. Mater. Chem. B 2, 6779–6791 (2014).

    Google Scholar 

  123. Pi, Q. et al. Digitally tunable microfluidic bioprinting of multilayered cannular tissues. Adv. Mater. 30, 1706913 (2018).

    Google Scholar 

  124. Gao, G. et al. Construction of a novel in vitro atherosclerotic model from geometry-tunable artery equivalents engineered via in-bath coaxial cell printing. Adv. Funct. Mater. 31, 2008878 (2021).

    Google Scholar 

  125. Liu, W. et al. Coaxial extrusion bioprinting of 3D microfibrous constructs with cell-favorable gelatin methacryloyl microenvironments. Biofabrication 10, 024102 (2018).

    ADS  Google Scholar 

  126. Wu, D. et al. A 3D-bioprinted multiple myeloma model. Adv. Healthc. Mater. https://doi.org/10.1002/adhm.202100884 (2021).

  127. Wang, Y. et al. Coaxial extrusion of tubular tissue constructs using a gelatin/gelma blend bioink. ACS Biomater. Sci. Eng. 5, 5514–5524 (2019).

    Google Scholar 

  128. Tavafoghi, M. et al. Multimaterial bioprinting and combination of processing techniques towards the fabrication of biomimetic tissues and organs. Biofabrication 13, 042002 (2021).

    Google Scholar 

  129. Freeman, F. E. et al. 3D bioprinting spatiotemporally defined patterns of growth factors to tightly control tissue regeneration. Sci. Adv. 6, eabb5093 (2020).

    ADS  Google Scholar 

  130. Liu, W. et al. Rapid continuous multimaterial extrusion bioprinting. Adv. Mater. 29, 1604630 (2017).

    Google Scholar 

  131. Hardin, J. O., Ober, T. J., Valentine, A. D. & Lewis, J. A. Microfluidic printheads for multimaterial 3D printing of viscoelastic inks. Adv. Mater. 27, 3279–3284 (2015).

    Google Scholar 

  132. Lee, C.-Y., Lin, C.-H. & Fu, L.-M. in Encyclopedia of Microfluidics and Nanofluidics (ed. Dongqing, L.) 1602–1610 (Springer, 2008).

  133. Chang, C.-C., Fu, L.-M. & Yang, R.-J. in Encyclopedia of Microfluidics and Nanofluidics (ed. Dongqing, L.) 33–38 (Springer, 2008).

  134. Ober, T. J., Foresti, D. & Lewis, J. A. Active mixing of complex fluids at the microscale. Proct. Natl Acad. Sci. USA 112, 12293–12298 (2015).

    ADS  Google Scholar 

  135. Idaszek, J. et al. 3d bioprinting of hydrogel constructs with cell and material gradients for the regeneration of full-thickness chondral defect using a microfluidic printing head. Biofabrication 11, 044101 (2019).

    ADS  Google Scholar 

  136. Chávez-Madero, C. et al. Using chaotic advection for facile high-throughput fabrication of ordered multilayer micro- and nanostructures: continuous chaotic printing. Biofabrication 12, 035023 (2020).

    ADS  Google Scholar 

  137. Ceballos-González, C. F. et al. High-throughput and continuous chaotic bioprinting of spatially controlled bacterial microcosms. ACS Biomater. Sci. Eng. 7, 2408–2419 (2021).

    Google Scholar 

  138. Bolívar-Monsalve, E. J. et al. Continuous chaotic bioprinting of skeletal muscle-like constructs. Bioprinting 21, e00125 (2021).

    Google Scholar 

  139. Samandari, M. et al. Controlling cellular organization in bioprinting through designed 3D microcompartmentalization. Appl. Phys. Rev. 8, 021404 (2021).

    ADS  Google Scholar 

  140. Datta, P., Vyas, V., Dhara, S., Chowdhury, A. R. & Barui, A. Anisotropy properties of tissues: a basis for fabrication of biomimetic anisotropic scaffolds for tissue engineering. J. Bionic Eng. 16, 842–868 (2019).

    Google Scholar 

  141. Malda, J. et al. 25th anniversary article: engineering hydrogels for biofabrication. Adv. Mater. 25, 5011–5028 (2013).

    Google Scholar 

  142. Fu, Z. et al. Printability in extrusion bioprinting. Biofabrication https://doi.org/10.1088/1758-5090/abe7ab (2021).

    Article  Google Scholar 

  143. Gillispie, G. et al. Assessment methodologies for extrusion-based bioink printability. Biofabrication 12, 022003 (2020).

    ADS  Google Scholar 

  144. Kesti, M. et al. A versatile bioink for three-dimensional printing of cellular scaffolds based on thermally and photo-triggered tandem gelation. Acta Biomater. 11, 162–172 (2015).

    Google Scholar 

  145. Ouyang, L., Yao, R., Zhao, Y. & Sun, W. Effect of bioink properties on printability and cell viability for 3D bioplotting of embryonic stem cells. Biofabrication 8, 035020 (2016).

    ADS  Google Scholar 

  146. Lee, J. M., Ng, W. L. & Yeong, W. Y. Resolution and shape in bioprinting: strategizing towards complex tissue and organ printing. Appl. Phys. Rev. 6, 011307 (2019).

    ADS  Google Scholar 

  147. Ribeiro, A. et al. Assessing bioink shape fidelity to aid material development in 3D bioprinting. Biofabrication 10, 014102 (2017).

    ADS  Google Scholar 

  148. Lee, J. M. & Yeong, W. Y. Engineering macroscale cell alignment through coordinated toolpath design using support-assisted 3D bioprinting. J. R. Soc. Interface 17, 20200294 (2020).

    Google Scholar 

  149. Tan, E. Y. S. & Yeong, W. Y. Concentric bioprinting of alginate-based tubular constructs using multi-nozzle extrusion-based technique. Int. J. Bioprinting 1, 49–56 (2015).

    Google Scholar 

  150. Soltan, N., Ning, L., Mohabatpour, F., Papagerakis, P. & Chen, X. Printability and cell viability in bioprinting alginate dialdehyde-gelatin scaffolds. ACS Biomater. Sci. Eng. 5, 2976–2987 (2019).

    Google Scholar 

  151. Wang, L., Xu, M., Zhang, L., Zhou, Q. & Luo, L. Automated quantitative assessment of three-dimensional bioprinted hydrogel scaffolds using optical coherence tomography. Biomed. Opt. Express 7, 894–910 (2016).

    Google Scholar 

  152. Petta, D., Grijpma, D. W., Alini, M., Eglin, D. & D’este, M. Three-dimensional printing of a tyramine hyaluronan derivative with double gelation mechanism for independent tuning of shear thinning and postprinting curing. ACS Biomater. Sci. Eng. 4, 3088–3098 (2018).

    Google Scholar 

  153. Mahmodi, H., Piloni, A., Utama, R. & Kabakova, I. Mechanical mapping of bioprinted hydrogel models by brillouin microscopy. Preprint at bioRxiv https://doi.org/10.1101/2021.02.18.431535 (2021).

    Article  Google Scholar 

  154. Boularaoui, S., Al Hussein, G., Khan, K. A., Christoforou, N. & Stefanini, C. An overview of extrusion-based bioprinting with a focus on induced shear stress and its effect on cell viability. Bioprinting 20, e00093 (2020).

    Google Scholar 

  155. Blaeser, A. et al. Controlling shear stress in 3D bioprinting is a key factor to balance printing resolution and stem cell integrity. Adv. Healthc. Mater. 5, 326–333 (2016).

    Google Scholar 

  156. Liu, W. et al. Extrusion bioprinting of shear-thinning gelatin methacryloyl bioinks. Adv. Healthc. Mater. 6, 1601451 (2017).

    Google Scholar 

  157. Guvendiren, M., Lu, H. D. & Burdick, J. A. Shear-thinning hydrogels for biomedical applications. Soft Matter 8, 260–272 (2012).

    ADS  Google Scholar 

  158. Du Chatinier, D., Figler, K. P., Agrawal, P. & Zhang, Y. S. The potential of microfluidics-enhanced extrusion bioprinting. Biomicrofluidics 15, 041304 (2021).

    Google Scholar 

  159. Hölzl, K. et al. Bioink properties before, during and after 3D bioprinting. Biofabrication 8, 032002 (2016).

    ADS  Google Scholar 

  160. Zhang, Y. S. et al. Bioprinting 3D microfibrous scaffolds for engineering endothelialized myocardium and heart-on-a-chip. Biomaterials 110, 45–59 (2016).

    Google Scholar 

  161. Kupfer, M. E. et al. In situ expansion, differentiation, and electromechanical coupling of human cardiac muscle in a 3D bioprinted, chambered organoid. Circul. Res. 127, 207–224 (2020).

    Google Scholar 

  162. Song, Y. et al. Engineering of brain-like tissue constructs via 3D cell-printing technology. Biofabrication 12, 035016 (2020).

    ADS  Google Scholar 

  163. Bhise, N. S. et al. A liver-on-a-chip platform with bioprinted hepatic spheroids. Biofabrication 8, 014101 (2016).

    ADS  Google Scholar 

  164. Kang, D. et al. Bioprinting of multiscaled hepatic lobules within a highly vascularized construct. Small 16, 1905505 (2020).

    Google Scholar 

  165. Yang, H. et al. Three-dimensional bioprinted hepatorganoids prolong survival of mice with liver failure. Gut 70, 567–574 (2021).

    Google Scholar 

  166. Derr, K. et al. Fully three-dimensional bioprinted skin equivalent constructs with validated morphology and barrier function. Tissue Eng. C 25, 334–343 (2019).

    Google Scholar 

  167. Madden, L. R. et al. Bioprinted 3D primary human intestinal tissues model aspects of native physiology and adme/tox functions. iScience 2, 156–167 (2018).

    ADS  Google Scholar 

  168. Cao, X. et al. A tumor-on-a-chip system with bioprinted blood and lymphatic vessel pair. Adv. Funct. Mater. 29, 1807173 (2019).

    Google Scholar 

  169. Kang, H.-W. et al. A 3D bioprinting system to produce human-scale tissue constructs with structural integrity. Nat. Biotechnol. 34, 312–319 (2016).

    Google Scholar 

  170. Kesti, M. et al. Bioprinting complex cartilaginous structures with clinically compliant biomaterials. Adv. Funct. Mater. 25, 7406–7417 (2015).

    Google Scholar 

  171. Duan, B., Hockaday, L. A., Kang, K. H. & Butcher, J. T. 3D bioprinting of heterogeneous aortic valve conduits with alginate/gelatin hydrogels. J. Biomed. Mater. Res. A. 101, 1255–1264 (2013).

    Google Scholar 

  172. De Ruijter, M., Ribeiro, A., Dokter, I., Castilho, M. & Malda, J. Simultaneous micropatterning of fibrous meshes and bioinks for the fabrication of living tissue constructs. Adv. Healthc. Mater. 8, 1800418 (2019).

    Google Scholar 

  173. Kundu, J., Shim, J. H., Jang, J., Kim, S. W. & Cho, D. W. An additive manufacturing-based PCL–alginate–chondrocyte bioprinted scaffold for cartilage tissue engineering. J. Tissue Eng. Regen. Med. 9, 1286–1297 (2015).

    Google Scholar 

  174. Schuurman, W. et al. Bioprinting of hybrid tissue constructs with tailorable mechanical properties. Biofabrication 3, 021001 (2011).

    ADS  Google Scholar 

  175. Shim, J.-H., Lee, J.-S., Kim, J. Y. & Cho, D.-W. Bioprinting of a mechanically enhanced three-dimensional dual cell-laden construct for osteochondral tissue engineering using a multi-head tissue/organ building system. J. Micromech. Microeng. 22, 085014 (2012).

    ADS  Google Scholar 

  176. Rhee, S., Puetzer, J. L., Mason, B. N., Reinhart-King, C. A. & Bonassar, L. J. 3D bioprinting of spatially heterogeneous collagen constructs for cartilage tissue engineering. ACS Biomater. Sci. Eng. 2, 1800–1805 (2016).

    Google Scholar 

  177. Kilian, D. et al. 3D bioprinting of osteochondral tissue substitutes — in vitro-chondrogenesis in multi-layered mineralized constructs. Sci. Rep. 10, 1–17 (2020).

    Google Scholar 

  178. Critchley, S. et al. 3D printing of fibre-reinforced cartilaginous templates for the regeneration of osteochondral defects. Acta Biomater. 113, 130–143 (2020).

    Google Scholar 

  179. Peiffer, Q. C. et al. Melt electrowriting onto anatomically relevant biodegradable substrates: resurfacing a diarthrodial joint. Mater. Des. 195, 109025 (2020).

    Google Scholar 

  180. Diloksumpan, P. et al. Combining multi-scale 3D printing technologies to engineer reinforced hydrogel-ceramic interfaces. Biofabrication 12, 025014 (2020).

    ADS  Google Scholar 

  181. Daly, A. C., Pitacco, P., Nulty, J., Cunniffe, G. M. & Kelly, D. J. 3D printed microchannel networks to direct vascularisation during endochondral bone repair. Biomaterials 162, 34–46 (2018).

    Google Scholar 

  182. Shao, L. et al. Synchronous 3D bioprinting of large-scale cell-laden constructs with nutrient networks. Adv. Healthcare Mater. 9, 1901142 (2019).

    Google Scholar 

  183. Gao, Q., He, Y., Fu, J.-Z., Liu, A. & Ma, L. Coaxial nozzle-assisted 3D bioprinting with built-in microchannels for nutrients delivery. Biomaterials 61, 203–215 (2015).

    Google Scholar 

  184. Mirabella, T. et al. 3D-printed vascular networks direct therapeutic angiogenesis in ischaemia. Nat. Biomed. Eng. 1, 0083 (2017).

    Google Scholar 

  185. Nulty, J. et al. 3D bioprinting of prevascularised implants for the repair of critically-sized bone defects. Acta Biomater. 126, 154–169 (2021).

    Google Scholar 

  186. Willemen, N. G. A. et al. Oxygen-releasing biomaterials: current challenges and future applications. Trends Biotechnol. https://doi.org/10.1016/j.tibtech.2021.01.007 (2021).

    Article  Google Scholar 

  187. Correia, C. R. & Mano, J. F. 3D-bioprinted constructs that breathe. Matter 4, 15–17 (2021).

    Google Scholar 

  188. Erdem, A. et al. 3D bioprinting of oxygenated cell-laden gelatin methacryloyl constructs. Adv. Healthc. Mater. 9, 1901794 (2020).

    Google Scholar 

  189. Farzin, A. et al. Self-oxygenation of tissues orchestrates full-thickness vascularization of living implants. Adv. Funct. Mater. https://doi.org/10.1002/adfm.202100850 (2021).

    Article  Google Scholar 

  190. Lode, A. et al. Green bioprinting: fabrication of photosynthetic algae-laden hydrogel scaffolds for biotechnological and medical applications. Report No. 1618-0240 (Wiley Online Library, 2015).

  191. Maharjan, S. et al. Symbiotic photosynthetic oxygenation within 3D-bioprinted vascularized tissues. Matter 4, 217–240 (2021).

    Google Scholar 

  192. Di Bella, C. et al. In situ handheld three-dimensional bioprinting for cartilage regeneration. J. Tissue Eng. Regen. Med. 12, 611–621 (2018).

    Google Scholar 

  193. D O’connell, C. et al. Development of the biopen: a handheld device for surgical printing of adipose stem cells at a chondral wound site. Biofabrication 8, 015019 (2016).

    ADS  Google Scholar 

  194. Duchi, S. et al. Handheld co-axial bioprinting: application to in situ surgical cartilage repair. Sci. Rep. 7, 1–12 (2017).

    Google Scholar 

  195. Onofrillo, C. et al. Biofabrication of human articular cartilage: a path towards the development of a clinical treatment. Biofabrication 10, 045006 (2018).

    ADS  Google Scholar 

  196. Hakimi, N. et al. Handheld skin printer: in situ formation of planar biomaterials and tissues. Lab. Chip 18, 1440–1451 (2018).

    Google Scholar 

  197. Cheng, R. Y. et al. Handheld instrument for wound-conformal delivery of skin precursor sheets improves healing in full-thickness burns. Biofabrication 12, 025002 (2020).

    ADS  Google Scholar 

  198. Russell, C. S. et al. In situ printing of adhesive hydrogel scaffolds for the treatment of skeletal muscle injuries. ACS Appl. Bio Mater. 3, 1568–1579 (2020).

    Google Scholar 

  199. Alarcin, E. et al. Injectable shear-thinning hydrogels for delivering osteogenic and angiogenic cells and growth factors. Biomater. Sci. 6, 1604–1615 (2018).

    Google Scholar 

  200. Zhang, Y. S. & Khademhosseini, A. Systems and methods for in vivo multi-material bioprinting. Patent No. WO2017184839A1 (2017).

  201. Zhao, W. & Xu, T. Preliminary engineering for in situ in vivo bioprinting: a novel micro bioprinting platform for in situ in vivo bioprinting at a gastric wound site. Biofabrication 12, 045020 (2020).

    Google Scholar 

  202. Zhou, C. et al. Ferromagnetic soft catheter robots for minimally invasive bioprinting. Nat. Commun. 12, 5072 (2021).

    ADS  Google Scholar 

  203. Fischbach, C. et al. Engineering tumors with 3D scaffolds. Nat. Methods 4, 855–860 (2007).

    Google Scholar 

  204. Zhang, Y. et al. Chronic label-free volumetric photoacoustic microscopy of melanoma cells in three-dimensional porous scaffolds. Biomaterials 31, 8651–8658 (2010).

    Google Scholar 

  205. Prendergast, M. E. et al. Microphysiological systems: automated fabrication via extrusion bioprinting. Microphysiol. Syst. 2, 3 (2018).

    Google Scholar 

  206. Maharjan, S., Bonilla, D. & Zhang, Y. S. Three-dimensional bioprinting for tissue and disease modeling. Mater. Matters https://www.sigmaaldrich.com/GB/en/technical-documents/technical-article/materials-science-and-engineering/3d-bioprinting/3d-bioprinting-tissue0 (2019).

  207. Miri, A. K. et al. Effective bioprinting resolution in tissue model fabrication. Lab. Chip 19, 2019–2037 (2019).

    Google Scholar 

  208. Li, J., Parra-Cantu, C., Wang, Z. & Zhang, Y. S. Improving bioprinted volumetric tumor microenvironments in vitro. Trends Cancer 6, 745–756 (2020).

    Google Scholar 

  209. Parra-Cantu, C., Li, W., Quiñones-Hinojosa, A. & Zhang, Y. S. 3D bioprinting of glioblastoma models. J. 3D Print. Med. 4, 113–125 (2020).

    Google Scholar 

  210. Nichol, J. W. et al. Cell-laden microengineered gelatin methacrylate hydrogels. Biomaterials 31, 5536–5544 (2010).

    Google Scholar 

  211. Ramon-Azcon, J. et al. Gelatin methacrylate as a promising hydrogel for 3D microscale organization and proliferation of dielectrophoretically patterned cells. Lab. Chip 12, 2959–2969 (2012).

    Google Scholar 

  212. Yoon, H. J. et al. Cold water fish gelatin methacryloyl hydrogel for tissue engineering application. PLoS ONE 11, e0163902 (2016).

    Google Scholar 

  213. Yue, K. et al. Structural analysis of photocrosslinkable methacryloyl-modified protein derivatives. Biomaterials 139, 163–171 (2017).

    Google Scholar 

  214. Shin, S. R. et al. A bioactive carbon nanotube-based ink for printing 2D and 3D flexible electronics. Adv. Mater. 28, 3280–3289 (2016).

    Google Scholar 

  215. Yin, J., Yan, M., Wang, Y., Fu, J. & Suo, H. 3D bioprinting of low-concentration cell-laden gelatin methacrylate (GelMA) bioinks with a two-step cross-linking strategy. ACS Appl. Mater. Interfaces 10, 6849–6857 (2018).

    Google Scholar 

  216. Luo, C. et al. Low-temperature three-dimensional printing of tissue cartilage engineered with gelatin methacrylamide. Tissue Eng. C. 26, 306–316 (2020).

    Google Scholar 

  217. Su, R. et al. 3D printed self-supporting elastomeric structures for multifunctional microfluidics. Sci. Adv. 6, eabc9846 (2020).

    ADS  Google Scholar 

  218. Gough, A., Vernetti, L., Bergenthal, L., Shun, T. Y. & Taylor, D. L. The microphysiology systems database for analyzing and modeling compound interactions with human and animal organ models. Appl. Vitro Toxicol. 2, 103–117 (2016).

    Google Scholar 

  219. Mironov, V. et al. Organ printing: tissue spheroids as building blocks. Biomaterials 30, 2164–2174 (2009).

    Google Scholar 

  220. Norotte, C., Marga, F. S., Niklason, L. E. & Forgacs, G. Scaffold-free vascular tissue engineering using bioprinting. Biomaterials 30, 5910–5917 (2009).

    Google Scholar 

  221. Yu, Y. et al. Three-dimensional bioprinting using self-assembling scalable scaffold-free “tissue strands” as a new bioink. Sci. Rep. 6, 1–11 (2016).

    Google Scholar 

  222. Jeon, O. et al. Individual cell-only bioink and photocurable supporting medium for 3D printing and generation of engineered tissues with complex geometries. Mater. Horiz. 6, 1625–1631 (2019).

    Google Scholar 

  223. Brassard, J. A. & Lutolf, M. P. Engineering stem cell self-organization to build better organoids. Cell Stem Cell 24, 860–876 (2019).

    Google Scholar 

  224. Moldovan, N. I., Hibino, N. & Nakayama, K. Principles of the Kenzan method for robotic cell spheroid-based three-dimensional bioprinting. Tissue Eng. B 23, 237–244 (2017).

    Google Scholar 

  225. Ayan, B. et al. Aspiration-assisted bioprinting for precise positioning of biologics. Sci. Adv. 6, eaaw5111 (2020).

    ADS  Google Scholar 

  226. Daly, A. C., Davidson, M. D. & Burdick, J. A. 3D bioprinting of high cell-density heterogeneous tissue models through spheroid fusion within self-healing hydrogels. Nat. Commun. 12, 1–13 (2021).

    Google Scholar 

  227. Lawlor, K. T. et al. Cellular extrusion bioprinting improves kidney organoid reproducibility and conformation. Nat. Mater. 20, 260–271 (2021).

    ADS  Google Scholar 

  228. Brassard, J. A., Nikolaev, M., Hübscher, T., Hofer, M. & Lutolf, M. P. Recapitulating macro-scale tissue self-organization through organoid bioprinting. Nat. Mater. 20, 22–29 (2021).

    ADS  Google Scholar 

  229. Viola, J. M. et al. Guiding cell network assembly using shape-morphing hydrogels. Adv. Mater. 32, 2002195 (2020).

    Google Scholar 

  230. Behl, M. & Lendlein, A. Shape-memory polymers. Mater. Today 10, 20–28 (2007).

    Google Scholar 

  231. Xia, Y., He, Y., Zhang, F., Liu, Y. & Leng, J. A review of shape memory polymers and composites: mechanisms, materials, and applications. Adv. Mater. 33, 2000713 (2021).

    Google Scholar 

  232. Wang, M., Li, W., Garciamendez-Mijares, C. E. & Zhang, Y. S. Engineering (bio)materials through shrinkage and expansion. Adv. Healthc. Mater. 21, 2100380 (2021).

    Google Scholar 

  233. Gao, B. et al. 4D bioprinting for biomedical applications. Trends Biotechnol. 34, 746–756 (2016).

    Google Scholar 

  234. An, J., Chua, C. K. & Mironov, V. A perspective on 4D bioprinting. Int. J. Bioprint. 2, 3–5 (2016).

    Google Scholar 

  235. Li, Y.-C., Zhang, Y. S., Akpek, A., Shin, S. R. & Khademhosseini, A. 4D bioprinting: the next-generation technology for biofabrication. Biofabrication 9, 012001 (2017).

    ADS  Google Scholar 

  236. Ge, Q., Qi, H. J. & Dunn, M. L. Active materials by four-dimension printing. Appl. Phys. Lett. 103, 131901 (2013).

    ADS  Google Scholar 

  237. Tibbits, S. 4D printing: multi-material shape change. Archit. Des. 84, 116–121 (2014).

    Google Scholar 

  238. Sydney Gladman, A., Matsumoto, E. A., Nuzzo, R. G., Mahadevan, L. & Lewis, J. A. Biomimetic 4D printing. Nat. Mater. 15, 413–418 (2016).

    ADS  Google Scholar 

  239. Gong, J. et al. Complexation-induced resolution enhancement of 3D-printed hydrogel constructs. Nat. Commun. 11, 1267 (2020).

    ADS  Google Scholar 

  240. Kuribayashi-Shigetomi, K., Onoe, H. & Takeuchi, S. Cell origami: self-folding of three-dimensional cell-laden microstructures driven by cell traction force. PLoS ONE 7, e51085 (2012).

    ADS  Google Scholar 

  241. Davidson, M. D. et al. Programmable and contractile materials through cell encapsulation in fibrous hydrogel assemblies. Preprint at bioRxiv https://doi.org/10.1101/2021.04.19.440470 (2021).

    Article  Google Scholar 

  242. Nguyen, P. Q., Courchesne, N.-M. D., Duraj-Thatte, A., Praveschotinunt, P. & Joshi, N. S. Engineered living materials: prospects and challenges for using biological systems to direct the assembly of smart materials. Adv. Mater. 30, 1704847 (2018).

    Google Scholar 

  243. Huang, J. et al. Programmable and printable bacillus subtilis biofilms as engineered living materials. Nat. Chem. Biol. 15, 34–41 (2019).

    ADS  Google Scholar 

  244. Duraj-Thatte, A. M. et al. Programmable microbial ink for 3D printing of living materials produced from genetically engineered protein nanofibers. Preprint at bioRxiv https://doi.org/10.1101/2021.04.19.440538 (2021).

    Article  Google Scholar 

  245. Emmermacher, J. et al. Engineering considerations on extrusion-based bioprinting: interactions of material behavior, mechanical forces and cells in the printing needle. Biofabrication 12, 025022 (2020).

    ADS  Google Scholar 

  246. Zhang, P. & Abate, A. R. High-definition single-cell printing: cell-by-cell fabrication of biological structures. Adv. Mater. 32, 2005346 (2020).

    Google Scholar 

  247. Tang, G. et al. Faithful fabrication of biocompatible multicompartmental memomicrospheres for digitally color-tunable barcoding. Small 16, 1907586 (2020).

    Google Scholar 

  248. Tang, G. et al. Designable dual-power micromotors fabricated from a biocompatible gas-shearing strategy. Chem. Eng. J. 407, 127187 (2021).

    Google Scholar 

  249. Hansen, C. J. et al. High-throughput printing via microvascular multinozzle arrays. Adv. Mater. 25, 96–102 (2013).

    Google Scholar 

  250. Lee, J., Oh, S. J., An, S. H., Kim, W.-D. & Kim, S.-H. Machine learning-based design strategy for 3D printable bioink: elastic modulus and yield stress determine printability. Biofabrication 12, 035018 (2020).

    ADS  Google Scholar 

  251. Mao, Y., He, Q. & Zhao, X. Designing complex architectured materials with generative adversarial networks. Sci. Adv. 6, eaaz4169 (2020).

    ADS  Google Scholar 

  252. Yu, C. & Jiang, J. A perspective on using machine learning in 3D bioprinting. Int. J. Bioprint. 6, 253 (2020).

    Google Scholar 

  253. Conev, A. et al. Machine learning-guided three-dimensional printing of tissue engineering scaffolds. Tissue Engineering Part A 26, 1359–1368 (2020).

    Google Scholar 

  254. Zhu, Z., Ng, D. W. H., Park, H. S. & Mcalpine, M. C. 3D-printed multifunctional materials enabled by artificial-intelligence-assisted fabrication technologies. Nat. Rev. Mater. 6, 27–47 (2020).

    ADS  Google Scholar 

  255. Jin, Z., Zhang, Z. & Gu, G. X. Autonomous in-situ correction of fused deposition modeling printers using computer vision and deep learning. Manuf. Lett. 22, 11–15 (2019).

    Google Scholar 

  256. Sitthi-Amorn, P. et al. Multifab: a machine vision assisted platform for multi-material 3d printing. Acm Trans. Graph. 34, 1–11 (2015).

    Google Scholar 

  257. Jin, Z., Zhang, Z. & Gu, G. X. Automated real-time detection and prediction of interlayer imperfections in additive manufacturing processes using artificial intelligence. Adv. Intell. Syst. 2, 1900130 (2020).

    Google Scholar 

  258. Jin, Z., Zhang, Z., Shao, X. & Gu, G. X. Monitoring anomalies in 3D bioprinting with deep neural networks. ACS Biomater. Sci. Eng. https://doi.org/10.1021/acsbiomaterials.0c01761 (2021).

    Article  Google Scholar 

  259. Eastridge, B. J., Holcomb, J. B. & Shackelford, S. Outcomes of traumatic hemorrhagic shock and the epidemiology of preventable death from injury. Transfusion 59, 1423–1428 (2019).

    Google Scholar 

  260. Schneeberger, K. et al. Converging biofabrication and organoid technologies: the next frontier in hepatic and intestinal tissue engineering? Biofabrication 9, 013001 (2017).

    ADS  Google Scholar 

  261. Parrish, J., Lim, K., Zhang, B., Radisic, M. & Woodfield, T. B. F. New frontiers for biofabrication and bioreactor design in microphysiological system development. Trends Biotechnol. 37, 1327–1343 (2019).

    Google Scholar 

  262. Castilho, M. et al. Multitechnology biofabrication: a new approach for the manufacturing of functional tissue structures? Trends Biotechnol. 38, 1316–1328 (2020).

    Google Scholar 

  263. Di Marzio, N., Eglin, D., Serra, T. & Moroni, L. Bio-fabrication: convergence of 3D bioprinting and nano-biomaterials in tissue engineering and regenerative medicine. Front. Bioeng. Biotechnol. 8, 326 (2020).

    Google Scholar 

  264. Zhang, Y. S. & Khademhosseini, A. Advances in engineering hydrogels. Science 356, eaaf3627 (2017).

    Google Scholar 

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Acknowledgements

Y.S.Z. gratefully acknowledges funding from the NIH (R00CA201603, R21EB025270, R21EB026175, R21EB030257, R01EB028143 and R01HL153857), the National Science Foundation (CBET-EBMS-1936105) and the Brigham Research Institute. J.M. acknowledges support of the Gravitation Program ‘Materials Driven Regeneration’, funded by the Netherlands Organization for Scientific Research (024.003.013; J.M.).

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Authors and Affiliations

  1. Division of Engineering in Medicine, Brigham and Women’s Hospital, Department of Medicine, Harvard Medical School, Cambridge, MA, USA

    Yu Shrike Zhang

  2. Department of Mechanical Engineering, University of Minnesota, Minneapolis, MN, USA

    Ghazaleh Haghiashtiani & Michael C. McAlpine

  3. Laboratory of Stem Cell Bioengineering, Institute of Bioengineering, School of Life Sciences and School of Engineering, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland

    Tania Hübscher & Matthias Lutolf

  4. Trinity Centre for Biomedical Engineering, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland

    Daniel J. Kelly

  5. Department of Mechanical, Manufacturing and Biomedical Engineering, School of Engineering, Trinity College Dublin, Dublin, Ireland

    Daniel J. Kelly

  6. Advanced Materials and Bioengineering Research Centre (AMBER), Royal College of Surgeons in Ireland and Trinity College Dublin, Dublin, Ireland

    Daniel J. Kelly

  7. Department of Anatomy and Regenerative Medicine, Royal College of Surgeons in Ireland, Dublin, Ireland

    Daniel J. Kelly

  8. School of Mechanical and Aerospace Engineering, Nanyang Technological University, Singapore, Singapore

    Jia Min Lee & Wai Yee Yeong

  9. Institute of Chemical Sciences and Engineering, School of Basic Science, EPFL, Lausanne, Switzerland

    Matthias Lutolf

  10. Singapore Centre for 3D Printing (SC3DP), School of Mechanical and Aerospace Engineering, Nanyang Technological University, Singapore, Singapore

    Wai Yee Yeong

  11. HP-NTU Digital Manufacturing Corporate Lab, Nanyang Technological University, Singapore, Singapore

    Wai Yee Yeong

  12. Department of Health Sciences and Technology, ETH Zürich, Zürich, Switzerland

    Marcy Zenobi-Wong

  13. Department of Orthopedics, University Medical Center Utrecht, Utrecht, Netherlands

    Jos Malda

  14. Department of Clinical Sciences, Faculty of Veterinary Medicine, Utrecht University, Utrecht, Netherlands

    Jos Malda

Authors
  1. Yu Shrike Zhang
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  2. Ghazaleh Haghiashtiani
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  3. Tania Hübscher
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  4. Daniel J. Kelly
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  5. Jia Min Lee
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  6. Matthias Lutolf
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  7. Michael C. McAlpine
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  8. Wai Yee Yeong
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  9. Marcy Zenobi-Wong
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  10. Jos Malda
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Contributions

Introduction (Y.S.Z. and J.M.), Experimentation (Y.S.Z., G.H., J.M.L, M.C.M., W.Y.Y., M.Z-W. and J.M.), Results (Y.S.Z., J.M.L., W.Y.Y. and J.M.), Applications (Y.S.Z., D.J.K. and J.M.), Reproducibility and data deposition (Y.S.Z. and J.M.), Limitations and optimizations (Y.S.Z., T.H., M.L. and J.M.), Outlook (Y.S.Z., G.H., M.C.M. and J.M.). Overview of the Primer (Y.S.Z. and J.M.).

Corresponding author

Correspondence to Yu Shrike Zhang.

Ethics declarations

Competing interests

Y.S.Z. sits on the Scientific Advisory Board of Allevi, Inc., which did not participate in this work or bias it in any form. This interest has been reviewed and managed by the Brigham and Women’s Hospital in accordance with its Conflict of Interest policies. M.C.M. serves on the Scientific Advisory Board and holds equity in GRIP Molecular Technologies. M.C.M. is Co-Founder and CSO of Flui3D, Inc. These interests have been reviewed and managed by the University of Minnesota in accordance with its Conflict of Interest policies. The other authors declare no competing interests.

Additional information

Peer review information

Nature Reviews Methods Primers thanks Nureddin Ashammakhi, Bige Deniz Unluturk, Wenguo Cui and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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Simplify3D: https://www.simplify3d.com/

Slic3r: https://slic3r.org/

SolidWorks: https://www.solidworks.com/

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Supplementary information

Glossary

Shape fidelity

The degree to which the bioprinted structure conforms to the digital model.

Toolpath

The trajectory that the nozzle of the 3D bioprinter follows during bioink extrusion.

Infill density

The amount of material to be extruded for constructing the internal volume of a contruct, with 0% yielding a hollow shell structure (empty internally) and 100% yielding a full, solid object.

Shear rate

The rate of change in velocity at which one layer of fluid passes over an adjacent layer.

Amphiphiles

Molecules, either synthetic or natural, that contain both hydrophilic and hydrophobic domains.

Fugitive ink

An ink that serves as a temporary template after being printed, which can be subsequently selectively removed on demand to create a hollow space.

Computer vision

Machine learning or other artificial intelligence algorithms for processing and interpreting the visual data received from cameras or other vision systems, analogous to the human vision system and the brain.

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Zhang, Y.S., Haghiashtiani, G., Hübscher, T. et al. 3D extrusion bioprinting. Nat Rev Methods Primers 1, 75 (2021). https://doi.org/10.1038/s43586-021-00073-8

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