Involvement Of Wnt/β-catenin Signaling In Tripchlorolide Protecting Against Oligomeric β-amyloid-_1-42-induced Apoptotic Neuronal Damage

Objective:To investigate the effect and the molecular mechanisms of oligomeric Aβ1-42-induced apoptosis in primary cultured rat cortical neurons, and to explore the underlied mechanism of protective action of tripchlorolide (T4) in oligomeric Aβ1-42-treated neurons.Methods:Primary cultures of cortical neurons were prepared from the embryonic day 16-18d Sprague-Dawley rats. The oligomeric Aβ1-42 (5μM for 24 h) was applied to induce apptotic neuronal death. Prior to treatment with Ap for 24 h, the cultured neurons were pre-incubated with T4 (2.5,10 and 40nM), Wnt3a (Wnt signaling agonists) and Dkkl (inhibitors) for indicated time. Then the cell viability, neuronal apoptosis, and protein levels of Wnt, glycogen synthase kinase 3β(GSK3β),β-catenin and phospho-β-catenin were measured by MTT assay, TUNEL staining and Western blot, respectively.Results:1. Oligomeric Aβ1-42 induced apoptotic neuronal cell death in a time-and dose-dependent manner. After treatment with 5μmol/1 oligomeric Aβ1-42 for 24 hours, the cortical neuronal survival was significantly decreased by 20% compared with control group by MTT assay. The neural body shrank, process shortened or dismissed and nucleus was stained black by TUNEL2. Treatment with oligomeric Aβ1-42 significantly increased the protein levels of phosphorylatedβ-catenin and GSK3β. In contrast, the non-phosphorylatedβ-catenin protein levels were decrease. Wnt signaling inhibitors Dkkl produced the familiar effect on the protein changes of P-catenin and GSK3β.3. Pretreatment with T4 significantly increased the neuronal cell survival and attenuated apoptotic neuronal death. Moreover, oligomeric Aβ1-42-induced the phosphorylation ofβ-catenin and GSK3βwas markedly inhibited by T4. Additionally, T4 stabilized cytoplasmicβ-catenin to activate P-catenin downsteam signaling. Wnt signaling agonists Wnt3a produced the familiar effect on the protein changes ofβ-catenin and GSK30.Conclusion:Tripchlorolide protects against the neurotoxicity of Aβby regulating Wnt/β-catenin signaling pathway. This will provide insight into the clinical application of tripchlorolide to Alzheimer's disease.