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Introduction and objectives <p>Advanced bone tissue engineering strategies require multifunctional scaffolds that simultaneously support cell viability, exhibit antibacterial properties, and actively induce osteogenic differentiation. The objective of this study was to develop and evaluate an electrospun chitosan-based nanofibrous scaffold reinforced with magnesium-doped hydroxyapatite (Mg-HAp) and bioactive agents, including icariin, lithium chloride, and naringin, to enhance osteogenesis and investigate its association with the upregulation of the Wnt/β-catenin signaling pathway. Specifically, this research aimed to address whether the localized, synergistic co-delivery of these three structurally distinct osteogenic agents within a single biomimetic matrix could effectively overcome the limited regenerative capacity of conventional bone grafts in defect models.</p> Methods <p>The nanocomposite scaffold was fabricated using electrospinning and characterized by Fourier transform infrared spectroscopy and scanning electron microscopy. Physicochemical properties such as porosity, swelling behavior, degradation rate, and mechanical performance were assessed. Biocompatibility and cytotoxicity were evaluated using the MTT assay. Antibacterial activity against <i>Staphylococcus aureus</i> and <i>Escherichia coli</i> was investigated using agar diffusion tests. Osteogenic activity was analyzed by real-time PCR evaluation of Wnt and β‑catenin gene expression. In vivo bone regeneration was assessed using a rat calvarial bone defect model.</p> Results <p>The fabricated scaffold exhibited a highly porous and interconnected nanofibrous structure (~ 87% porosity) with suitable mechanical stability. The scaffold showed excellent biocompatibility, with significantly increased cell viability over time. Effective concentration-dependent antibacterial activity was observed, particularly against S. aureus. Gene expression analysis revealed significant upregulation of Wnt and β‑catenin. In vivo studies demonstrated substantial new bone formation, leading to near-complete defect closure after 12 weeks.</p> Conclusion <p>The bioactive chitosan/Mg‑HAp nanocomposite scaffold effectively promotes bone regeneration by enhancing osteogenic signaling pathways and exhibits strong potential for bone tissue engineering applications.</p>

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Bioactive electrospun chitosan/magnesium-doped hydroxyapatite nanocomposite scaffold co-loaded with icariin, lithium chloride and naringin for enhanced osteogenesis, antibacterial activity and bone regeneration in vitro and in vivo

  • 期刊论文

  • 发表日期：

    2026年7月16日

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  • Sahar Hefzollesan,
  • Hanife Musayeva,
  • Hamed Aghazadeh,
  • Aslan Mammadov,
  • Soheila Ajdary,
  • Rauf Baylarov,
  • Sahra Hefzollesan,
  • Rizvan Mammadov

摘要

Introduction and objectives

Advanced bone tissue engineering strategies require multifunctional scaffolds that simultaneously support cell viability, exhibit antibacterial properties, and actively induce osteogenic differentiation. The objective of this study was to develop and evaluate an electrospun chitosan-based nanofibrous scaffold reinforced with magnesium-doped hydroxyapatite (Mg-HAp) and bioactive agents, including icariin, lithium chloride, and naringin, to enhance osteogenesis and investigate its association with the upregulation of the Wnt/β-catenin signaling pathway. Specifically, this research aimed to address whether the localized, synergistic co-delivery of these three structurally distinct osteogenic agents within a single biomimetic matrix could effectively overcome the limited regenerative capacity of conventional bone grafts in defect models.

Methods

The nanocomposite scaffold was fabricated using electrospinning and characterized by Fourier transform infrared spectroscopy and scanning electron microscopy. Physicochemical properties such as porosity, swelling behavior, degradation rate, and mechanical performance were assessed. Biocompatibility and cytotoxicity were evaluated using the MTT assay. Antibacterial activity against Staphylococcus aureus and Escherichia coli was investigated using agar diffusion tests. Osteogenic activity was analyzed by real-time PCR evaluation of Wnt and β‑catenin gene expression. In vivo bone regeneration was assessed using a rat calvarial bone defect model.

Results

The fabricated scaffold exhibited a highly porous and interconnected nanofibrous structure (~ 87% porosity) with suitable mechanical stability. The scaffold showed excellent biocompatibility, with significantly increased cell viability over time. Effective concentration-dependent antibacterial activity was observed, particularly against S. aureus. Gene expression analysis revealed significant upregulation of Wnt and β‑catenin. In vivo studies demonstrated substantial new bone formation, leading to near-complete defect closure after 12 weeks.

Conclusion

The bioactive chitosan/Mg‑HAp nanocomposite scaffold effectively promotes bone regeneration by enhancing osteogenic signaling pathways and exhibits strong potential for bone tissue engineering applications.