Engineering a Thiolated Polymeric Nanocomposite Scaffold With Chitosan Nanoparticles for Enhanced Osteogenesis

ABSTRACT The development of biomimetic scaffolds capable of providing structural support and promoting osteogenic differentiation remains a major challenge in bone tissue engineering. In the present study, a chitosan nanoparticle‐loaded thiolated Eudragit/alginate/polyethylene glycol nanocomposite scaffold was developed as a potential platform for bone regeneration. Thiolated Eudragit was synthesized through EDC/NHS‐mediated conjugation with l ‐cysteine, while chitosan nanoparticles were prepared by ionic gelation and incorporated into the scaffold matrix via freeze‐drying. Successful thiolation of Eudragit was confirmed by FTIR and Ellman's assay, whereas DLS and AFM verified the formation of stable chitosan nanoparticles. XRD, FESEM, and optical profilometry demonstrated successful nanoparticle incorporation within a highly porous interconnected scaffold architecture. Furthermore, the nanocomposite scaffold exhibited controlled swelling, enhanced gel fraction, sustained degradation, and suitable porosity, indicating favorable physicochemical properties. MTT analysis using MG‐63 osteoblast‐like cells demonstrated excellent cytocompatibility, with cell viability remaining above 85% across all tested concentrations. The scaffold also demonstrated excellent hemocompatibility, exhibiting a hemolysis ratio below the acceptable limit for nonhemolytic biomaterials, while effectively inhibiting the growth of both Staphylococcus aureus and Escherichia coli . Moreover, ALP and ARS staining demonstrated enhanced osteogenic differentiation and extracellular matrix mineralization, while quantitative ARS analysis and hydroxyapatite binding affinity further confirmed increased calcium deposition and improved scaffold–mineral interactions following nanoparticle incorporation. Thus, the developed chitosan nanoparticle‐loaded thiolated Eudragit nanocomposite scaffold represents a promising biomimetic platform for enhancing osteogenesis and facilitating bone tissue regeneration.