Three-dimensional printing of cell-laden hydrogels has evolved as a promising approach on the route to patient-specific or complex tissue-engineered constructs. However, it is still challenging to print structures with both, high shape fidelity and cell vitality. Herein, we used a synthetic nanosilicate clay, called Laponite, to build up scaffolds utilising the extrusion-based method 3D plotting. By blending with alginate and methylcellulose, a bioink was developed which allowed easy extrusion, achieving scaffolds with high printing fidelity. Following extrusion, approximately 70%-75% of printed immortalised human mesenchymal stem cells survived and cell viability was maintained over 21 days within the plotted constructs. Mechanical properties of scaffolds comprised of the composite bioink decreased over time when stored under cell culture conditions. Nevertheless, shape of the plotted constructs was preserved even over longer cultivation periods. Laponite is known for its favourable drug delivery properties. Two model proteins, bovine serum albumin and vascular endothelial growth factor were loaded into the bioink. We demonstrate that the release of both growth factors significantly changed to a more sustained profile by inclusion of Laponite in comparison to an alginate-methylcellulose blend in the absence of Laponite. In summary, addition of a synthetic clay, Laponite, improved printability, increased shape fidelity and was beneficial for controlled release of biologically active agents such as growth factors.

For the generation of multi-layered full thickness osteochondral tissue substitutes with an individual geometry based on clinical imaging data, combined extrusion-based 3D printing (3D plotting) of a bioink laden with primary chondrocytes and a mineralized biomaterial phase was introduced. A pasty calcium phosphate cement (CPC) and a bioink based on alginate-methylcellulose (algMC) – both are biocompatible and allow 3D plotting with high shape fidelity – were applied in monophasic and combinatory design to recreate osteochondral tissue layers. The capability of cells reacting to chondrogenic biochemical stimuli inside the algMC-based 3D hydrogel matrix was assessed. Towards combined osteochondral constructs, the chondrogenic fate in the presence of CPC in co-fabricated and biphasic mineralized pattern was evaluated. Majority of expanded and algMC-encapsulated cells survived the plotting process and the cultivation period, and were able to undergo redifferentiation in the provided environment to produce their respective extracellular matrix (ECM) components (i.e. sulphated glycosaminoglycans, collagen type II), examined after 3 weeks. The presence of a mineralized zone as located in the physiological calcified cartilage region suspected to interfere with chondrogenesis, was found to support chondrogenic ECM production by altering the ionic concentrations of calcium and phosphorus in in vitro culture conditions.

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