Enhanced Bone Regeneration by Schwann Cells through Coupling of Osteogenesis and Angiogenesis via β-catenin signaling in a Preclinical Model of Distraction Osteogenesis.

Abstract

Background: The lengthy period of external fixation for bone consolidation increases the risk of complications during distraction osteogenesis (DO). Both pro-angiogenic and osteogenic potential of bone marrow mesenchymal stem cells (BMSCs) contribute to bone regeneration during DO. The underlying mechanism of Schwann cells (SCs) in promoting bone regeneration during DO remains poorly understood. Methods: The impacts of RSC-96 on the proliferation, migration, and osteogenic differentiation of BMSCs in the coculture system were investigated. The pro-angiogenic potential of BMSCs was evaluated by migration and tube formation assay. Quantitative real-time PCR was used to analyze angiogenic and osteogenic markers. ELISA was used to detect the secretion of various neurotrophins. Protein expressions of Activate protein kinase B (AKT)/β-catenin signaling were assessed by western blot. In vivo, dynamic expression levels of neurotrophic factors were detected in a preclinical rat DO model. Promotive effects of vascularization and mineralization provided by RSC-96 derived conditioned medium (CM) in a rat DO model were verified radiologically, biomechanically and histologically. Result: Coculture system with RSC-96 promoted osteogenic ability of BMSCs, with increased cell viability, alkaline phosphatase staining, mineralized nodule formation, and osteogenic gene expression. Additionally, increased angiogenic gene expression of BMSCs and angiogenic capacity of endothelial cells demonstrated enhanced pro-angiogenic potential of BMSCs. Secretion of angiogenic and neurotrophic factors were enhanced in the coculture system. These effects were accompanied by activation of AKT/GSK-3β/β-catenin signaling, as evidenced by western blot analysis and the inhibitory effect of AKT inhibitor. The mRNA expression of neurotrophic factors peaked at the end of the distraction phase during DO. Furthermore, RSC-96 derived CM accelerated bone regeneration, resulting in improved biomechanical parameters, radiological features and histological manifestations, along with increased vascularization in the distraction area. Conclusion: Through activation of AKT/GSK-3β/β-catenin signaling, SCs enhanced the coupled angio- and osteogenesis effects of BMSCs. The preclinical evidence demonstrates that SCs derived CM with increased neurotrophins secretion can be a promising treatment approach to accelerate bone regeneration in the DO process.