Fig. 3
Ex-4 promotes brain-derived insulin by activating Wnt/β-catenin pathway to antagonize tau hyperphosphorylation in db/db mice. A Workflow of animal study designed for db/db mice. Insulin levels B and c-peptide levels C in CSF were measured by ELISA kits (n = 4/ group). D Represented immunofluorescence staining of coronal brain sections obtained from mice of each group was performed for insulin (green) and DAPI (blue). Scale bar = 100 μm. E mRNA levels of Ins2 in the hippocampus were measured by RT-qPCR assay. β-actin was the internal control. n = 3/group. F The levels of the phosphorylated tau at Ser199, Ser202, Ser396 and Thr217 sites and total tau in each group; the activation of Wnt/β-catenin pathway as evidenced by the levels of the np-β-catenin to total β-catenin; insulin signalling activation as P-AKTS473 to total AKT and P-GSK-3βS9 to total GSK-3β were examined through Western blot analysis, (n = 4/group). G The gray density of E. H Represented immunohistochemical staining of coronal brain sections from each group of mice was performed with the antibody against phosphorylated tau at Thr231. For D and H, HIP: hippocampus, CA: cornue ammonis, DG: dentate gyrus. Results are representative of three independent experiments. Values are presented as mean ± SD. **P < 0.01, ***P < 0.001, ****P < 0.0001. vs. db/db; ††P < 0.01, †††P < 0.001. vs. Ex-4