In Silico and In Vitro Evaluation of Allicin-Loaded PLGA CompositeScaffolds for Enhanced Osteogenic Potential in Osteoporotic BoneDefects

Introduction: Osteoporosis is characterised by impaired bone remodelling due to increased osteoclast activity and reduced osteoblast function. This study aimed to elucidate the molecular mechanisms of allicin in osteoporosis and to develop an allicinincorporated PLGA/Chitosan (CN)/β-TCP scaffold to enhance osteogenic differentiation for osteoporotic bone regeneration. Methods: Network pharmacology was used to identify common targets between allicin and osteoporosis, followed by hub gene and protein-protein interaction analysis. Molecular docking using estimated active sites was used to evaluate binding affinities. Developmental toxicity was assessed using Danio rerio embryos according to OECD guidelines. Allicin-loaded scaffolds (AL25 and AL75) were fabricated via lyophilisation and characterised for their physicochemical, mechanical, and degradation properties. In vitro cytocompatibility and osteogenic differentiation were assessed in MC3T3-E1 cells using MTT, alkaline phosphatase, and Alizarin Red S assays. Results: Seventy-five overlapping targets were identified, including ten hub genes associated with bone remodelling and inflammatory signalling. Allicin demonstrated strong binding to PIK3CB and BTK (−4.3 kcal/mol). Zebrafish studies confirmed dosedependent biocompatibility. The AL25 scaffold exhibited superior compressive strength and balanced porosity, whereas the AL75 group showed increased degradation. In vitro assays revealed enhanced cell viability, significantly elevated ALP activity, and increased extracellular matrix mineralisation in allicin-incorporated groups. The AL75 group promoted accelerated early osteogenic responses, whereas the AL25 group supported sustained differentiation and mineral deposition at later stages. Discussion: These findings suggest that allicin modulates key molecular pathways and enhances scaffold-mediated osteogenesis in a concentration-dependent manner. However, higher loading may compromise mechanical stability, highlighting the need for dosage optimisation and long-term in vivo validation. Conclusion: Allicin-loaded PLGA/Chitosan/β-TCP scaffolds demonstrate promising osteogenic potential, with the AL25 group providing the most balanced mechanical integrity and sustained osteogenic activity for osteoporotic bone regeneration.