Model of a synovial membrane developed by extrusion bioprinting technique

The aim of the present study was to develop a bioartificial synovium model by three dimensional (3D) bioprinting technology. In particular, volumetric extrusion has been adopted to fabricate cellularized scaffolds, usable as in vitro models to study the pathophysiology of some rheumatic diseases such as osteoarthritis (OA) and rheumatoid arthritis (RA) and evaluate new therapeutic approaches, while reducing and replacing animal trials. Synovium is a complex and specialized tissue that surrounds the tendons. It is composed of heterogeneous cell populations (synoviocytes and monocytes), acellular stroma, nerves, lymphatic system and blood vessels. Synovial inflammation plays a critical role in the symptoms and structural progression of joint disease. To realize the synovium model, we utilized a methacrylate gelatin (GelMA) which is a biomaterial widely used in regenerative medicine and tissue engineering due to its tunable mechanical properties, which have been deeply investigated in this study with different characterization methods. GelMA shows a high biocompatibility and a very low cytotoxicity in cell culture. In addition, this biomaterial is very versatile: it is possible to modify its chemical-physical and mechanical characteristics to fabricate scaffolds that mimic various soft tissues (e.g., synovium, cartilage, liver tissue, lung tissue, heart tissue). In particular, GelMA properties depend on temperature and concentration in solution and the Degree of Functionalization (DoF), that can be analyzed by fluorescence assay as was done for this study. GelMA can be easily printed using Additive manufacturing (AM) technologies like the volumetric extrusion. This requires materials with a suitable viscosity to obtain a good print fidelity. The viscosity of GelMA can be easily modified by changing the temperature and by photocross-linking due to an UV source. In this way it is possible to obtain the proper viscosity that allows the GelMA printing and the layer-by-layer process typical of AM technologies. In the present study three different formulations of GelMA with the same DoF were tested (GelMA 7.5% w/v, GelMA 10% w/v and GelMA 12.5% w/v) to find the optimal one that allows the fabrication of a scaffold having the desired geometry and thickness after the 3D printing process. According to the results of the bioprinting optimization, GelMA 10% w/v was chosen as biomaterial useful for the synovial model because this formulation allows to obtain a scaffold with optimal printing and shape fidelity. GelMA 10% w/v has been then combined with an immortalized cell line of fibroblast-like synoviocytes (K4IM) to make the bioink subsequently processed by volumetric extrusion. Square-Grid scaffolds were fabricated by the bioprinting process, placed in culture for 14 days and then analyzed by two different assays showing high cell viability and negligible cytotoxicity. In conclusion, with this work it was possible to demonstrate that 3D bioprinting represents a promising technology for replicating synovial tissue due to its inherent capability to deposit heterogeneous bioink in a defined spatiotemporal manner relevant to biological architecture. However, further research will be needed in order to recapitulate the joint microenvironment to reproduce the interactions between its different tissues.