A self-organized tri-lineage bone model in liquefied microcapsules captures matrix dynamics in early remodeling

Bone tissue is inherently complex, featuring specialized cells, a dynamic extracellular matrix (ECM), vascular networks, and continuous remodeling. These processes are tightly regulated by cell-cell and cell-ECM interactions, making them difficult to replicate in vitro. Here, we developed a self-regulated, bone-like three-dimensional microenvironment using liquefied microcapsules composed of hydroxypyridinone- and dopamine-modified gelatins, supporting scaffold structure and bioactivity. Bone marrow-derived human mesenchymal stromal cells (BM-hMSCs) and a human monocyte cell line (THP-1) were directly co-cultured in a 3:1 ratio for 35 days within the microcapsules. This heterotypic system, without exogenous differentiating inducers, supported differentiation into osteoblasts, osteocytes, and osteoclasts, alongside ECM deposition, mineralization, and resorption. Proteomic signatures further indicated features associated with bone development and early remodeling. This self-organized tri-lineage bone model provides a simplified tool for studying cellular crosstalk and ECM dynamics, with potential to reduce reliance on animal models.