Dual-functional 3D-printed polyetheretherketone scaffolds with immunomodulatory nano‑calcium silicate/interleukin-4 coating synergistically enhance osteogenesis through macrophage M2 polarization and NF-κB pathway suppression

Despite its biomechanical advantages, polyetheretherketone (PEEK) exhibits bioinertness and pro-inflammatory responses, which limit its efficacy in bone-defect repair. Therefore, we aimed to engineer three-dimensional-printed PEEK scaffolds functionalized with a polydopamine (PDA)-assisted nano‑calcium silicate (n-CS)/interleukin-4 (IL-4) coating (PEEK/PCS/IL-4) to synergistically modulate immune responses and osteogenesis to overcome the challenges associated with PEEK. The scaffolds were fabricated through fused deposition modeling under optimized conditions and subsequently subjected to sequential surface functionalization involving PDA-mediated n-CS immobilization followed by IL-4 impregnation. In vitro, the PEEK/PCS/IL-4 scaffolds significantly enhanced human bone marrow mesenchymal stem cell (hBMSCs) spreading, alkaline phosphatase activity, mineralization, and osteogenic gene expression compared with the controls. The scaffolds also promoted human umbilical vein endothelial cell migration and tube formation, suggesting potent pro-angiogenic effects. RAW264.7 macrophages cultured on PEEK/PCS/IL-4 exhibited M2 polarization, elevated transforming growth factor-β1 (TGFβ1), and suppressed tumor necrotic factor-α levels (TNFα), which correlated with the non-canonical NF-κB pathway inhibition. The conditioned medium from PEEK/PCS/IL-4-primed macrophages further amplified hBMSCs osteogenesis, confirming immune-osteogenic coupling. In vivo, PEEK/PCS/IL-4 scaffolds reduced fibrous encapsulation and enhanced M2 macrophage polarization in a rat subcutaneous implantation model. In rabbit femoral defects, micro-CT, confocal laser scanning microscopy, histology, and biomechanical testing results revealed that the PEEK/PCS/IL-4 scaffolds exhibited enhanced osteogenesis and superior osseointegration. This study pioneers an immuno-engineering approach to transform bioinert PEEK into a bioactive platform, leveraging n-CS/IL-4 coatings to harmonize immune homeostasis, bone regeneration, and revascularization, offering a transformative strategy for bone defect repair.