Fig. 3

MW181 reduced the levels of pATF2 and pMK2, increased levels of synaptophysin, and blunted the expression of IFNγ and IL-1β in 20-month-old hTau mice. a Western blot analysis of hippocampal lysates shows no significant difference in p-p38α MAPK (pT180/pY182) levels, but reduced levels of phosphorylated ATF2 (pT71) and phosphorylated MK2 (pT334) and increased levels of synaptophysin were observed with MW181 treatment. b–e Quantification shows no difference for p-p38α MAPK, but statistically significant reduction (n = 3 for SB239063, n = 8 for MW181, and n = 5 for vehicle; mean ± SEM of IDV ratios; *p < 0.05; **p < 0.01; one-way ANOVA with Tukey’s multiple comparison test) in the pATF2/GAPDH and pMK2/GAPDH ratios in MW181-treated hTau mice compared with vehicle-treated hTau mice. None of these proteins were seen to be statistically altered with SB239063 treatment. f Quantitative real-time PCR analysis for various markers of classical activation or M1 state (IFNγ, IL-1β, TNFα, and IL-6) and alternative activation or M2 state (IL-4, YM1, and ARG1) shows a significant reduction (n = 3 for SB239063 and n = 6 for MW181 and vehicle; mean ± SEM of IDV ratios; *p < 0.05; two-way ANOVA with Tukey’s multiple comparison test) in the levels of IFNγ and IL-1β in the MW181-treated hTau mice compared with vehicle-treated hTau mice. g Immunohistochemical and immunofluorescence analysis shows Iba1⁺ microglia and CD45⁺ microglia/macrophages in the CA3 region of hippocampus, B220⁺ B cells and CD3⁺ T cells in the meninges, and CX3CR1⁺ microglia in the CA3 region of the hippocampus in MW181-treated, Veh-treated, or SB239063-treated 20-month-old hTau mice. Scale bars: 30 μm for Iba1, CD45, and B220; 10 μm for B220 and CD3. h Quantitative morphometry for Iba1, B220, CD3, and CX3CR1 for immunochemistry in g (n = 3–4 mice per genotype); four random fields in three sections per brain were quantified; mean area (± SEM) immunoreactive for Iba1, B220, and CD3 in the hippocampus (for Iba1) or in the meninges (for CD3 and B220). For CX3CR1, the number of CX3CR1⁺ microglia displaying activated morphology (reduced processes and swollen cell body) was counted manually in the CA1 region of hippocampus and expressed as number of cells per mm² (*p < 0.05; one-way ANOVA followed by Tukey multiple comparison post-hoc test). Note that B220 and CD3 showed a reduced trend in MW181-treated groups but were not statistically significant, unlike Iba1 and CX3CR1 which were significant. Veh vehicle