| Objective: The aim of this study is to investigate the role of Wnt/β-catenin signaling pathway and E-cadherin/β-catenin complex in ICMT–induced end plate cartilage degeneration.Methods: Intermittent cyclic mechanical strain at 0.5 Hz sinusoidal curve at 10% elongation was applied for 8 hours a day by using an FX-5000 Flexercell Tension Plus unit.Chondrocytes related gene expression were examined by Realtime RT-PCR and Western blot; Toluidine Blue and Alcian blue staining detect chondrocyte matrix after ICMT application; Alamar Blue assay,annexin V-PI staining and Red1-X phalloidin to detect the changes of cells viability, apoptosis and cytoskeleton.Multiple strategies to examine wnt/β-catenin signaling activation after ICMT application in vitro. Coimmunoprecipitation were performed to study the interaction between E-cadherin and β-catenin. Pathway-specific inhibitors and E-cadherin expression plasmid were used to regulate wnt/β-catenin signaling and E-cadherin expression.Results: The expression of β-catenin proteins was significant elevated, whereas the cartilaginous genes were down-regulated in the end plate chondrocytes after ICMT application. ICMT loading lead to Wnt/β-catenin signaling activation as well as end plate chondrocytes degeneration in vitro end plate chondrocytes culture system. Inhibiting Wnt/β-catenin signaling could relieve ICMT-induced end plate chondrocytes degeneration. Furthermore, E-cadherin expression was inhibited by ICMT stimulation, resulting in decreased interaction between E-cadherin and β-catenin protein. Over-expression of Ecadherin could rescue cartilaginous genes expression by enhance the interaction between of E-cadherin and β-catenin protein.Conclusion: Following results indicated that ICMT promotes end plate chondrocytes degeneration was via activating Wnt/β-catenin signaling pathway and suppressing physical protein-protein interaction between E-cadherin and β-catenin. |
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