Figure 4.

TRIOL induces the nuclear translocation of Nrf2 in microglia. BV2 microglia were pretreated with/without 10 μM TRIOL or vehicle (HP-β-CD) 1 h before treatment of 1% O₂ hypoxia stimuli for 12 h, followed by reperfusion to normoxia (20% O₂) for 12 h. (A) Expression and nuclear localization of Nrf2 and HO-1 were measured using immunofluorescence staining with antibodies against Nrf2 (green) and HO-1 (red) in BV2 microglia. Nucleus was stained with Hoechst 33342 (blue); scale bar, 20 μm. (B) Relative HO-1 fluorescence intensity was measured by NIS-Elements AR. N = 4 independent ROIs for each group. (C) Nrf2 intensity accumulated in a single nucleus was also measured to evaluate the activation of Nrf2; N = 90 cells for each group. (D) Representative confocal imaging of Iba1 (red) and Nrf2 (green) in the optic nerves of wt mice in different groups. Nucleus was stained with Hoechst 33342 (blue). Fields in white boxes were enlarged in the panel below; scale bar, 25 μm. Statistical analysis of fluorescence intensity in B and C was performed using one-way ANOVA, followed by Dunnett’s post hoc test. N.s., no significance; *, p < 0.05; **, p < 0.01; ***, p < 0.001.