Effect of neurotrophin-3 on bone healing in distraction osteogenesis

Distraction osteogenesis (DO) is a surgical technique used to repair skeletal defects by promoting bone and soft tissue regeneration. However, prolonged use of external fixators often causes complications such as infections and joint contractures. Neurotrophin-3 (NT-3) enhances osteoblast differentiation via bone morphogenetic protein-2 (BMP-2) and Akt signaling and may also modulate the Wnt/β-catenin pathway. In this study, we investigated the role of NT-3 in bone regeneration during DO. A DO model was established in 9-week-old male ICR mice, in which tibiae were osteotomized and stabilized with external fixators under anesthesia. After a 5-day latency period, distraction was performed at 0.2 mm every 12 hours for 8 days. RNA was extracted at defined time points post-osteotomy for quantitative polymerase chain reaction (qPCR) analysis, and NT-3 was administered post-distraction. Bone regeneration was evaluated by radiographic, histological, and biomechanical analyses. The effects of NT-3 and Dickkopf-1 were further examined in ST2 bone marrow-derived mesenchymal stem cells. NT-3 significantly enhanced bone formation, increased bone mineral density within the DO gap, and improved the mechanical stiffness of regenerated bone. Histological examination revealed earlier bone continuity and greater callus formation in the NT-3 group. Gene expression analysis demonstrated upregulation of Wnt/β-catenin pathway components, including Wnt10b, β-catenin, and low-density lipoprotein receptor-related protein 5 (LRP5). In ST2 cells, NT3 increased the expression of LRP5, LRP6, β-catenin, and Runx2, while cotreatment with Dickkopf1 abolished these effects. These findings suggest that NT-3 promotes bone regeneration during DO by activating the Wnt/β-catenin signaling pathway, highlighting its therapeutic potential to improve DO outcomes.