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19 Mar 2025 Given the drawbacks associated with clinically available grafts, ‘bottom-up’ bone tissue engineering emerges as a viable alternative for guided tissue regeneration for osseous defects where physiological bone remodeling capacity is not sufficient to restore the defect, resulting in delayed or non-union of the bone tissue. Osteogenic constructs can be developed as replacements for injured or diseased osseous defects using extrusion-based and light-based additive manufacturing (AM) techniques, including digital light processing (DLP), stereolithography (SLA) and 2-photon polymerization (2PP). However, the potential usage of those AM techniques in clinical practice is limited due to the layer-by-layer printing approach. Since the printing time therefore increases with increasing geometry, the throughput, scalability and cell viability/functionality could potentially be hampered. Additionally, extrusion- and light-based AM techniques are associated with challenges directly related to their respective printing processes. In case of extrusion-based AM, a force is exerted to protrude the ink through the nozzle onto the print bed. The nozzle has the potential to clog depending on the material viscosity, printing speed, and nozzle size. Moreover, the shear stresses exerted during the printing process onto the encapsulated cells potentially lead to cell damage, altered proliferation or cell death. Finally, support structures are needed when overhangs or through holes are printed, requiring dedicated post-processing strategies. Light-based AM techniques create a construct by exposing focused light onto the photo-responsive resin. The techniques are characterized by a superior resolution (<100 µm) compared to their extrusion-based techniques but, nevertheless the printing process remains slow (1–106 mm3 h−1). Moreover, the creation of multi-material constructs is more challenging as compared to extrusion-based AM, for which multiple printheads can be exploited.
Tomographic volumetric printing (VP) is the most recent type of light-based AM. During VP, the rotating resin is irradiated by light patterns, which are constructed through back projections of the object’s radon transforms. Since the construct is printed at once, instead of layer-by-layer, the conventional techniques’ challenges are overcome. Firstly, the printing time does not increase with increasing geometry in case of VP, allowing the production of clinically relevant, cubic centimeter-scale constructs within tens of seconds. Secondly, VP is characterized by the elimination of the need for support structures. However, the key challenge of developing a wider variety of available osteogenic bioinks that allow osteogenic maturation of the encapsulated cells within the construct remains. In this research, a bioink was developed exploiting a step-growth mechanism (norbornene–norbornene functionalized gelatin in combination with thiolated gelatin—GelNBNBSH) which outperformed the bioink exploiting a chain-growth mechanism (gelatin methacryloyl—GelMA), as the necessary photo-initiator concentration was three times lower combined with a more than 50% reduction in required light exposure dose resulting in an improved positive and negative resolution.
Proof of concept perfusion of a 5 w/v% GelNBNBSH (DS 176/72) (0.025 w/v% Li-TPO-L) construct (scale bar: 1 mm).
