Organic acid cross-linked 3D printed cellulose nanocomposite bioscaffolds with controlled porosity, mechanical strength, and biocompatibility

Biocompatible polysaccharide scaffolds with controllable pore size, good mechanical properties and without hazardous chemical crosslinkers, are desirable for long-term tissue engineering applications but their production is highly challenging. Herein, we fabricated three-dimensional (3D) scaffolds using a polysaccharide composite ink composed of nanofibrillated cellulose, carboxymethyl cellulose and citric acid, featuring strong shear thinning behavior and adequate printability. Highly porous and mechanically stable scaffolds were produced by combining direct ink writing 3D printing, freeze-drying and dehydrothermal heat-assisted crosslinking techniques. The last heat-assisted step induces reaction of citric acid, which was chosen as it is a non-hazardous and green crosslinker. Degree of crosslinking was controlled by varying the concentration of citric acid (2.5 – 10 wt.%) to tune the chemical, surface, swelling and degradation properties of the scaffolds in the dry and hydrated states. The scaffolds with the highest porosity (86%) and interconnected pores (100-450 µm) were acquired using the lowest citric acid concentration; and the porosity was significantly reduced at higher citric acid concentration as well as at longer hydration time. The compressive strength, elastic modulus and the shape recovery behavior of the crosslinked scaffolds were increased significantly with increasing crosslinker concentration. The prepared crosslinked scaffolds promoted clustered cell adhesion and showed no cytotoxic effects, as determined by a cell viability assay and live/dead staining with human bone tissue derived osteoblast cells. The water-based and non-hazardous crosslinking method reported here can be extended to all polysaccharide-based materials to develop cell-friendly scaffolds with tailormade properties suitable for various tissue engineering applications in general.