Open Access

Ana Mora-Boza, Małgorzata K. Włodarczyk-Biegun, Aránzazu del Campo Bécares, Blanca Vazquez-Lasa, Julio San Roman

• Fabrication of intricate and long-term stable 3D polymeric scaffolds by 3D printing technique is still a challenge. The currently used polymeric materials need long post-printing processes and washing steps. In addition, highly concentrated solutions are necessary for maintaining shape fidelity after 3D deposition. This paper reports the fabrication of dual crosslinked 3D scaffolds using a low concentrated (<10 wt-%) ink of Gelatin Methacryloyl (GelMA)/Chitosan and a novel crosslinking agent, a glycerylphytate (G1Phy) to overcome the current limitations in the 3D printing field using hydrogels. The applied methodology consisted of a first ultraviolet light (UV) photopolymerization followed by a post-printing ionic crosslinking treatment with G1Phy. This crosslinker provides a robust framework and avoids the necessity of neutralization with strong bases. The blend ink showed shear-thinning behavior and excellent printability in the form of straight and homogeneous filament. UV curing was undertaken simultaneously to 3D deposition, which enhanced precision, shape fidelity (resolution ≈ 150 µm), and prevented from collapse of the subsequent printed layers (up to 28 layers). In the second step, the novel G1Phy ionic crosslinker agent provided swelling and long term stability properties to the 3D scaffolds. The multi-layered printed scaffolds were mechanically stable at physiological conditions for at least one month. Preliminary in vitro assays using L929 Fibroblasts showed very promising results in terms of adhesion, spreading, and proliferation in comparison to other phosphate-based traditional crosslinkers (i.e. TPP). We envision that the proposed combination of the blend ink and 3D printing approach can have widespread applications in the regeneration of soft tissues.