Immunomodulation and Oxidative Stress Mitigation Mediate Enhanced Bone Regeneration by a Platelet-Rich Plasma-Loaded...

Repairing critical-sized bone defects remains a clinical challenge. Nano-hydroxyapatite/polyamide 66 (n-HA/PA66) offers excellent biocompatibility and mechanics but limited bioactivity. This study aims to develop a multifunctional composite scaffold to overcome this limitation. A platelet-rich plasma (PRP)-loaded polydopamine/chitosan (PDA/CS) hydrogel was integrated with a 3D-printed porous n-HA/PA66 scaffold (PRP-PDA/CS-n-HA/PA66). Hydrogel's cytoprotective, immunomodulatory, and osteogenic effects were assessed in vitro using human osteoblasts, macrophages, and bone marrow mesenchymal stem cells (BMSCs) under hydrogen peroxide (H₂O₂)-induced oxidative stress or lipopolysaccharide (LPS)-induced inflammatory conditions. The osteogenic efficacy of the PRP-PDA/CS-n-HA/PA66 composite scaffold was further validated in a rabbit femoral condyle critical-sized defect model, assessed by micro-computed tomography, histological staining, and immunohistochemistry. The PRP-PDA/CS hydrogel demonstrated potent antioxidant activity, protected osteoblasts and BMSCs from H₂O₂-induced apoptosis and functional impairment, and promoted macrophage polarization toward the pro-healing M2 phenotype. It significantly enhanced BMSCs' proliferation, osteogenic differentiation, and mineralization. These effects were associated with the upregulation of the nuclear factor erythroid 2-related factor 2/heme oxygenase-1 antioxidant pathway and suppression of the nuclear factor kappa-B inflammatory pathway. In vivo, the PRP-PDA/CS-n-HA/PA66 composite scaffold markedly accelerated new bone formation, improved bone microarchitecture, including bone volume fraction (BV/TV), trabecular thickness (Tb.Th), trabecular number (Tb.N), and bone mineral density (BMD), upregulated osteogenic marker expression, and concurrently reduced local M1 macrophages, oxidative DNA damage, and pro-inflammatory cytokines. In conclusion, the PRP-PDA/CS-n-HA/PA66 composite scaffold synergizes structural support with multifaceted bioactivity, effectively promoting bone regeneration by mitigating oxidative stress, modulating immune responses, and enhancing osteogenesis, demonstrating significant translational potential.