3D Printing in Suspension Baths: Keeping the Promises of Bioprinting Afloat

Abstract

3D printing in suspension media unlocks the full potential of extrusion-based 3D printers by providing a strategy for fabricating non–self-supporting structures from water-rich, low-viscosity bioinks.Biomimetic structures representative of native vascular channels have been printed in suspension media, demonstrating that both omnidirectional printing and printing in discrete arbitrary locations are possible with this printing strategy.Retention of a suspension medium following printing of embedded constructs is achievable through crosslinking. Suspension media are therefore able to double as a 3D cell culture substrate in which printed features such as vessels or cell populations can help with maturing of the engineered tissue. Extrusion-based 3D printers have been adopted in pursuit of engineering functional tissues through 3D bioprinting. However, we are still a long way from the promise of fabricating constructs approaching the complexity and function of native tissues. A major challenge is presented by the competing requirements of biomimicry and manufacturability. This opinion article discusses 3D printing in suspension baths as a novel strategy capable of disrupting the current bioprinting landscape. Suspension baths provide a semisolid medium to print into, voiding many of the inherent flaws of printing onto a flat surface in air. We review the state-of-the-art of this approach and extrapolate toward future possibilities that this technology might bring, including the fabrication of vascularized tissue constructs. Extrusion-based 3D printers have been adopted in pursuit of engineering functional tissues through 3D bioprinting. However, we are still a long way from the promise of fabricating constructs approaching the complexity and function of native tissues. A major challenge is presented by the competing requirements of biomimicry and manufacturability. This opinion article discusses 3D printing in suspension baths as a novel strategy capable of disrupting the current bioprinting landscape. Suspension baths provide a semisolid medium to print into, voiding many of the inherent flaws of printing onto a flat surface in air. We review the state-of-the-art of this approach and extrapolate toward future possibilities that this technology might bring, including the fabrication of vascularized tissue constructs. Will we ever be able to engineer functional tissues and organs suitable for in vivo transplantation? Will 3D printing have a role in helping to achieve this? These questions have come to the forefront of research in the tissue engineering (TE) field over the past few decades, fueled by the demonstration that conventional 3D-printing technologies can be adapted to control the deposition of high-density cellular populations in 3D space. Of the different technologies, extrusion-based 3D printing has been identified as the most likely technique to realize the TE vision. Specifically, the mild processing conditions, which have a limited impact on cell viability, in conjunction with their flexibility in processing materials with a broad range of properties make this technology an attractive candidate. Although extrusion printers have been used extensively in the 3D-bioprinting (see Glossary) field, we are a long way from developing whole functional organs. Therefore, it could be hypothesized that a step change is required to harness the full potential of extrusion-based 3D printing in TE. Looking at the plethora of 3D-bioprinting–related publications, there are currently two predominant, distinct perspectives. The first prioritizes ease of fabrication, leveraging materials that can be extruded into filaments with high shape fidelity to create self-supporting structures. This approach has been extrapolated from 3D printing with stiff plastics for medical devices [1.Rahim T.N.A.T. et al.Recent developments in fused deposition modeling-based 3D printing of polymers and their composites.Polym. Rev. 2019; 59: 589-624Crossref Scopus (64) Google Scholar], where extruded filaments will immediately hold their shape. Regarding TE, this perspective is related to printing polymer-rich constructs that either are initially acellular or restrict encapsulated cells in their ability to develop tissue. The second perspective is tailored toward biomimicry, where replication of cellular and extracellular structures is favored over suitability for fabrication. This second approach has evolved primarily to be the patterning of bioinks. Such structures are more permissive to tissue maturation than is the printing of the previously mentioned polymer-rich constructs. However, bioinks are less suited for use as fabrication materials, due to their innate weak mechanical properties, and thus they are generally avoided for printing structures that are greater than several millimeters in size or require a high structural fidelity. Over the past few years, a new approach has evolved that has demonstrated the potential to marry the two perspectives described above. 3D printing in suspension media provides a platform for the patterning of mechanically weak bioinks into complex, well-defined structures [2.Bhattacharjee T. et al.Writing in the granular gel medium.Sci. Adv. 2015; 1e1500655Crossref PubMed Scopus (255) Google Scholar,3.Hinton T.J. et al.Three-dimensional printing of complex biological structures by freeform reversible embedding of suspended hydrogels.Sci. Adv. 2015; 1e1500758Crossref PubMed Scopus (700) Google Scholar]. This technology uses an extrusion 3D printer that deposits material not on a flat surface in air, but into a bath that suspends the printed material, preventing settling and collapse (Figure 1). Thus, it offers a paradigm shift in bioprinting by diminishing the need to compromise between material biomimicry and manufacturability. Suspension media hold unique traits that are responsible for their ability to suspend and completely encapsulate printed material. First, suspension media exhibit solid-like characteristics in the absence of an applied stress or at very low stresses such as those induced by gravity [2.Bhattacharjee T. et al.Writing in the granular gel medium.Sci. Adv. 2015; 1e1500655Crossref PubMed Scopus (255) Google Scholar,4.Jiang T. et al.Extrusion bioprinting of soft materials: an emerging technique for biological model fabrication.Appl. Phys. Rev. 2019; 6: 011310Crossref Scopus (53) Google Scholar,5.Highley C.B. et al.Direct 3D printing of shear-thinning hydrogels into self-healing hydrogels.Adv. Mater. 2015; 27: 5075-5079Crossref PubMed Scopus (514) Google Scholar]. Application of a stress that overcomes a critical stress, the yield stress, will initiate flow and render the media liquid-like. Second, after the disturbance of a suspension medium’s microstructure by a passing nozzle and its displacement by any deposited material, the microstructure spontaneously recovers. This self-healing capacity permits the medium to transition from a fluidized state back to a solid-like state, thereby encapsulating the deposited material [6.Moxon S.R. et al.Suspended manufacture of biological structures.Adv. Mater. 2017; 29: 1605594Crossref Scopus (39) Google Scholar]. 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