Supercritical carbon dioxide dried double layer laponite XLS and alginate/polyacrylamide construct and immune...

Fabrication of immunocompatible tissue constructs for bone-cartilage defect regeneration is of prime importance. In this study, a double layer hydrogel was successfully synthesized, where alginate/polyacrylamide were formulated to represent cartilage layer (5-10 % (w/w) total polymer ratio) and laponite XLS (2-5-8% (w/w))/alginate/polyacrylamide formed bone layer. Hydrogels were dried by supercritical CO₂ at 100 and 200 bar, 45 °C, 5 g/min CO₂ flow rate for 2 h. Constructs were treated with collagen, then cellularized and embedded in cell-laden GelMA to mimic the cellular microenvironment. The optimum weight ratio of alginate/polyacrylamide:laponite XLS was 10:5 based on mechanical strength test results. The constructs yielded high porosity (91.50 m²/g) and mesoporous structure, owing to the diffusivity of CO₂ at 200 bar (0.49 × 10^-7  m²/s). Constructs were then treated with collagen to increase cell adhesion and ATDC5 cells were seeded in the cartilage layer, whereas hFOB cells to the bone layer. About 10-15 % higher cell viability was attained. The porous structure of the construct allowed infiltration of macrophages, promoted polarization and positively affected the behavior of macrophages, yielding a decrease in M1 markers, whereas an increase in M2 on day 4. The formulated tissue constructs would be of value in tissue engineering applications.