Multiscale technologies and strategies for osteogenic hydrogel microsphere systems: From microscale signal regulation, mesoscale spontaneous assembly, and macroscale technology integration

Background Conventional bone tissue engineering predominantly relies on static preformed scaffolds that lack dynamic microenvironmental adaptability, failing to recapitulate the progressive, endogenous signal-driven tissue evolution characteristic of natural bone repair. To address critical-sized bone defects and repair failures in complex clinical scenarios, the regenerative paradigm is transitioning from static structural filling toward process-driven organoid self-assembly. Hydrogel microspheres have emerged as versatile modular units for building bone-like organoids; however, a comprehensive engineering framework detailing how these microsphere systems exert continuous regulation across micro-, meso-, and macro-scales to yield stable, functional bone tissue has yet to be established. Technology Originating from bio-interface engineering and granular self-assembly principles, this cross-scale technology platform operates across three interconnected levels: (1) Microscale signal regulation, leveraging cell-material interfacial interactions to program cell fate through spatiotemporal factor/ion delivery, localized metabolic/oxygen support, and mechanosensitive signal initiation. This includes the coordinated activation of mechanical transduction (YAP/Piezo1/RhoA), immunoregulatory (NF-κB/mTOR/PI3K-Akt), and master osteogenic (BMP/Wnt/MAPK) signaling cascades; (2) Mesoscale spontaneous assembly, constructing interconnected granular networks via cellular traction forces and dynamic interfacial crosslinking, characterized by spontaneous functional compartmentalization (dense mineralizing cores and loose vascularized channels) and synchronized degradation-remodeling kinetics; and (3) Macroscale technology integration, combining interfacial adaptive crosslinking, non-invasive physical-field actuation (acoustic, optical, electrical, magnetic, and thermal), fluoroscopic guidance, and AI-driven 4D biofabrication. This enables biomechanical stabilization, wireless deep-tissue energetic stimulation, and patient-specific anatomical conformity with host bone. Results By coupling microscale signal regulation, mesoscale spontaneous assembly, and macroscale technology integration, microsphere systems effectively coordinate early immune resolution, microvascular infiltration, and matrix mineralization to achieve robust bone regeneration. Furthermore, this review critically stratifies existing evidence across in vitro platforms, small-animal non-load-bearing models, and large-animal weight-bearing trials, addressing key translational and regulatory barriers to establish a clear engineering roadmap that systematically links localized interfacial signaling, granular organoid assembly, and macroscopic clinical interventions.