Piezo1 mediated scaffold-free rapid generation of self-mineralized bone organoids via activating Wnt signaling.

Bone organoids (BOs) have emerged as promising models for a wide range of biomedical applications. However, their clinical translation remains constrained by suboptimal mineralization efficiency and protracted fabrication cycles. To address these challenges, we developed an innovative mechanobiology-driven reprogramming strategy for the rapid generation of self-mineralizing BOs directly from Piezo1-activated stem cells from human exfoliated deciduous teeth (SHED), without relying on exogenous scaffolds. This approach synchronously facilitates accelerated self-organization and robust osteogenic differentiation. Notably, under static culture conditions, Piezo1 activation effectively compensates for the lack of persistent mechanical stimulation, thereby enhancing essential cellular functions in the absence of any external mechanical loading devices or dynamically simulated environments Activation of Piezo1 in SHED upregulated cadherin expression, enabling efficient self-assembly into spheroids within 24 h and augmenting osteogenic capacity through the Piezo1-canonical Wnt signaling axis. This method shortens the total manufacturing timeline from single cells to functional organic-inorganic composite BOs to less than one week. Implantation of these pre-mineralized BOs into calvarial defect models significantly enhanced bone regeneration. This study establishes a scaffold-free paradigm for rapid bone organoid biofabrication, offering a promising platform for advanced bone disease modeling and transformative regenerative therapies.