Macroporous alginate scaffolds with calcium phosphate filler as a 3D in vitro microenvironment supporting bone cancer cells

Macroporous Ca-alginate scaffolds with hydroxyapatite/β-tricalcium phosphate filler were investigated regarding possibilities to provide an adequate environment for 3D bone cancer cell cultures. Two scaffold groups were obtained from starting solutions: 1 wt.% Na-alginate, 1 wt% filler and 0.03 wt.% CaCl₂ (G1) and 2 wt.% Na-alginate, 2 wt.% filler and 0.045 wt.% CaCl₂ (G2). The scaffolds were produced by controlled alginate gelation followed by lyophilization, and further characterized regarding stability in culture medium, mechanical properties and biocompatibility in cultures of murine osteosarcoma cells K7M2-wt. While the scaffold porosity was the same for both groups (∼60%), G2 scaffolds exhibited higher stiffness and higher stability in the culture medium. Next, the cell seeding procedure was optimized followed by cultivation of the seeded scaffolds (1.5 × 10⁶–4 × 10⁶ cells/scaffold) for 7 days under continuous perfusion. The seeding strategy and scaffold architecture facilitated cell infiltration throughout the scaffold. The cells remained viable, while perfusion apparently induced higher cell metabolic activity as compared to static cultures, especially at the higher medium superficial velocity (40 μm/s vs. 15 μm/s). While few literature approaches use bioreactors, our study introduces a robust platform with scaffolds and perfusion to better mimic physiological conditions in 3D models in bone cancer research.