Engineering macromolecular crowding-driven amyloid-mineral hybrid scaffolds for enhanced bone regeneration

The extracellular matrix (ECM) of native bone features a densely crowded, hierarchically organized architecture composed of collagen fibrils and hydroxyapatite (HAp) nanocrystals, which together confer mechanical strength and biological functionality. However, faithfully replicating this complex organic-inorganic interface in synthetic scaffolds remains a significant challenge. Here, we report a macromolecular crowding (MMC)-driven strategy to construct ECM-mimetic scaffolds using phase-transited lysozyme (PTL) as an amyloid-based protein matrix. By employing a reverse dialysis process to mimic the crowded microenvironment, amyloid proteins undergo aggregation, conformational rearrangement, and a liquid-crystalline-like phase transition, accompanied by reconstruction of the organic-inorganic interface and energetic reorganization, thereby promoting biomineralization. The resulting amyloid-mineral hybrid scaffold exhibits excellent structural stability, mechanical robustness, and bioactivity, supporting bone regeneration comparable to mineralized collagen <i>in vitro</i> and <i>in vivo</i>. Collectively, this study demonstrates that, unlike conventional water-rich and dilute scaffolds, the MMC-driven strategy provides a more biomimetic and functionally versatile platform, highlighting the feasibility of using structurally stable amyloid proteins as substitutes for collagen and offering a powerful design paradigm for next-generation bone tissue engineering scaffolds.