Fabrication and Properties of Multi-Functional of Tannic Acid-Modified Sodium Alginate/Chitosan Microspheres for Bone Defect Repair.

Abstract

Conventional bone defect treatments face substantial clinical limitations, underscoring the urgent need for innovative bone regeneration materials. While tannic acid (TA) has shown potential in modifying hydrogel properties, its therapeutic implications in sodium alginate (SA)/chitosan (CS) composite hydrogels, particularly regarding bidirectional regulation of bone remodeling processes remain poorly characterized. This investigation tests the hypothesis that TA-modified SA/CS hydrogel microspheres can concurrently enhance antibacterial efficacy, promote osteogenic differentiation of bone mesenchymal stem cells (BMSCs), and suppress osteoclast differentiation of bone marrow macrophages (BMMs), ultimately accelerating bone repair. We engineered SA/CS/TA hydrogel microspheres through ionic crosslinking, followed by comprehensive material characterization using SEM (morphology), UV-Vis (TA release kinetics), and FT-IR (chemical interactions). Antimicrobial efficacy was quantified against common pathogens using plate coating method. For osteogenic assessment, BMSCs were cultured with microspheres and evaluated through qPCR/Western blot for osteogenic markers, ALP activity quantification, and matrix mineralization via ARS. Osteoclastogenesis was analyzed in BMM cultures using TRAP staining, F-actin staining, and osteoclast-specific gene/protein expression. A rat tibial critical-sized defect model with Micro-CT and histomorphometry served for in vivo validation. The microspheres exhibited sustained TA release and significant antimicrobial activity compared to control groups. Osteogenic assays demonstrated enhanced ALP activity and calcium deposition, accompanied by upregulation of key osteogenic markers. Conversely, the TA-modified microspheres demonstrated the inhibition of osteoclastogenesis throughout the differentiation process: reduction in TRAP activity reflecting impaired osteoclast maturation, and downregulation of osteoclast-specific markers via qPCR and Western blot analysis. In vivo evaluations revealed substantially improved bone regeneration in microsphere-treated defects compared to untreated controls. Our findings confirm the dual functionality of TA-modified SA/CS hydrogel microspheres in bone regeneration, demonstrating simultaneous enhancement of BMSC osteogenesis and suppression of osteoclast differentiation. The coordinated regulation of bone formation and resorption processes, combined with inherent antibacterial properties, establishes SA/CS/TA microspheres as a promising therapeutic platform for critical-sized bone defect reconstruction.