Three-Dimensional Bioprinting of New Hydrogel-based Bioinks into Cell-scaffold Constructs for Articular Cartilage Repair

Articular cartilage exhibits excellent load-bearing capacity and minimal friction that enables essential articulation in human diarthrodial joints. Following disease or injury, its self-repair ability for defects over a critical size diminishes. Tissue engineering strategies involving seeding chondrocytes onto artificial porous scaffolds have shown their efficacy for repairing cartilage defects but have their own limitations including cell penetration and distribution in scaffolds. Three-dimensional bioprinting uses bioinks comprising live cells, extracellular matrix (ECM)-analogous biomaterials and possibly bioactive biomolecules to fabricate living structure with potential high cell viability and with precisely controlled cell distributions for effective body tissue regeneration, and hydrogels based on natural polymers, which may replicate physicochemical and mechanical characteristics of native ECM, are often utilized to create bioinks. In this study, new hydrogel-based printing inks consisting of methacrylate-modified chitosan (CSMA), methacrylate-modified gelatin (GelMA) and hyaluronic acid (HA), with an added photoinitiator, were formulated and with the incorporation of ADTC5 murine chondrogenic cells or rat bone marrow-derived mesenchymal stem cells (rBMSCs), new bioinks were created for bioprinting for cartilage regeneration. Extrusion-based 3D printing or bioprinting was conducted on a bioprinter. Optimal printing allowed inks/bioinks to be extruded smoothly as continuous fine filaments. After printing, printed structures underwent crosslinking by UV light. CSMA, GelMA and HA concentrations were optimized for printing inks via rheological analysis. Results showed GelMA and HA improved printability of CSMA-based hydrogels. Printing parameters such as printing speed, layer height, and nozzle diameter were investigated and optimized, achieving multilayered hydrogel scaffolds with high resolution and fidelity. The morphology, mechanical properties, biodegradation and biocompatibility of 3D printed scaffolds were also assessed. In 3D bioprinted structures, ADTC5 or rBMSC displayed high cell viability after 14-day culture. Overall, 3D bioprinted cell-scaffold constructs showed robust structural integrity, adequate mechanical properties and favorable cellular responses, indicating their potential for cartilage tissue engineering.