Volumetric Bioprinting of Bone-like Mineralizing Hydrogel Constructs in the Presence of High Cell Densities and Mineral Precursors

A major challenge in bone tissue engineering is the embedding of osteocyte-like cells at high density within a mineralized matrix at the micro-scale and a trabecular-like architecture at the macro-scale. Volumetric bioprinting (VBP) enables rapid creation of complex cell-laden constructs through tomographic light projections. However, integrating both high cell densities and inorganic mineral precursors into VBP processes poses challenges due to light scattering, which can compromise print fidelity. In this study, we aim to combine bioinspired polymer-induced liquid-phase precursor (PILP) mineralization with VBP to fabricate cell-laden gelatin methacryloyl hydrogel constructs with amorphous mineral precursors. By stabilizing amorphous mineral precursors with poly-aspartic acid, light scattering is sufficiently reduced to enable printing. Tuning the refractive index of this mineralizing bioresin allows fast VBP of mineralized bone-like constructs with cell densities of up to 3 million cells ml-1. The constructs display high cell viability (>90%) and enhanced mineralization when cultured in osteogenic conditions with {beta}-glycerophosphate. Encapsulated human mesenchymal stromal cells exhibit an early osteocytic phenotype after 28 days of differentiation. Collectively, this PILP-assisted VBP platform holds promise for the development of advanced in vitro bone models with more physiologically relevant architecture and cellular composition.