Abstract: Treating critical-size bone defects remains a significant clinical challenge, due to the complexity of achieving adequate immunomodulation, angiogenesis, osteogenic differentiation and matrix mineralization. Successful bone repair requires an orchestrated response in these areas to promote tissue integration and regeneration effectively. In this study, we designed and fabricated a customized, bioactive porous GDM/CeHA@CA scaffold through 3D printing and subsequent UV crosslinking techniques. The scaffold integrating Mn²⁺-chelated deferoxamine (DFO)-grafted gelatin methacryloyl (GDM) with citric acid-modified cerium-doped hydroxyapatite nanowires (CeHA@CA). The controlled Mn²⁺ release from the scaffold strongly modulated macrophages polarization toward the anti-inflammatory M2 phenotype by down-regulating the MAPK signaling pathway and up-regulating the MnSOD signaling pathway. Macrophages maintain the stability of the bone microenvironment and prevent excessive inflammatory responses through immunomodulatory responses, and immunomodulated M2 macrophages promote angiogenesis and osteoblast differentiation by secreting growth factors VEGF and TGF-β. Scaffold degradation also led to the sustained release of covalently bound DFO, along with increased endogenous VEGF levels, promoted robust vascular remodeling. Additionally, the release of Ce^3+/4+, as well as Ca²⁺ and PO₄^3- from CeHA@CA nanowires, in combination with elevated endogenous TGF-β, further boosted osteogenesis. Therefore, GDM/CeHA@CA scaffolds are able to promote angiogenesis and osteogenic differentiation not only through direct degradation, but also indirectly through immunomodulation. Through these synergistic mechanisms of immunomodulation, angiogenesis, osteogenic differentiation and matrix mineralization, the GDM/CeHA@CA scaffold successfully accelerated the repair of the critical-size tibial bone defect in rabbits within 12 weeks. In conclusion, the 3D printed GDM/CeHA@CA composite scaffold provided a highly effective therapeutic strategy for rapid bone defects repair, making it a viable candidate for clinical applications in bone regeneration.

Key words: 3D printing, Bone immunomodulation, Angiogenesis, Osteogenic differentiation, Cerium hydroxyapatite nanowires