Fig 2: Effects of miR-126 overexpression on SIRT1 expression, Nrf2 protein distribution and HO-1 protein expression in OGD/R-exposed HUVECs. HUVECs were transfected with miR-126 m or miR-NC prior to OGD (4 h)/R (24 h) exposure. (A) Protein expression was determined via western blotting. Semi-quantification of (B) cytosolic SIRT1 and (C) Nrf2 normalized to GAPDH. (D) Representative western blots of nuclear Nrf2. (E) Quantification of nuclear Nrf2 normalized to histone H3. (F) Quantification of HO-1 normalized to GAPDH. Data are presented as the mean ± SD of at least three independent experiments. *P<0.05 and **P<0.01 vs. control group; #P<0.05 and ##P<0.01 vs. miR-NC + OGD/R group. miR-126 m, miR-126 mimics; miR-NC, miR-negative control; miR, microRNA; OGD, oxygen-glucose deprivation; R, reoxygenation; HUVEC, human umbilical vein endothelial cell; SIRT1, NAD-dependent protein deacetylase sirtuin-1; Nrf2, nuclear factor erythroid 2-related factor 2; HO-1, heme oxygenase 1.

Fig 3: Effects of LHE on (A) the expression of iNOS/COX-2, induction of HO-1/Nrf-2, activation of NF-κB, and (B) phosphorylation of MAPK in LPS/D-GalN-induced hepatitis mouse model. Mice were pretreated with LHE or vehicle once per day for 6 days. Then, LPS/D-GalN was injected 1 h after the last administration. After 6 h, mice were sacrificed, and liver tissue samples were collected. The expression of inflammatory synthetic enzyme, antioxidant molecules, and inflammatory pathways were determined via Western blot analysis. The histograms showed the expression levels of protein relative to those of β-actin. * p values of < 0.05, and *** < 0.001 (vs. LPS/D-GalN) were statistically significant.

Fig 4: Effects of LHE on (A) iNOS and COX-2 expression, (B) HO-1 induction, and (C) nuclear translocation of Nrf-2 in RAW 264.7 macrophages. (A) Cells were incubated with LHE for 1 h and then stimulated with LPS for 24 h (protein) or 12 h (mRNA). Cells were incubated with LHE alone for (B) 6 h or (C) 3 h. The quantified number and histograms showed the expression levels of protein and mRNA relative to those of β-actin or TBP. ** p < 0.01 and *** p < 0.001 were calculated by comparing LPS stimulation values.

Fig 5: DBZ ameliorated depressive behaviors, regulated M1/M2 polarization and affected Akt(Ser473)/GSK3β(Ser9)/Fyn-Nrf2 signal axis in the cerebral cortex of LPS-treated mice.(A) Experimental design is illustrated schematically. (B-E) Effects of DBZ (5, 20 mg·kg−1) on locomotor activity of mice administrated with LPS. The total distance, central time, and number of rearing were investigated by open field test. (F, G) DBZ reduced the increased number of Iba1+ cells and restored the microglia morphological alterations in neuroinflammatory mice induced by LPS. (F) The average number of Iba1+ cells in frontal cortex. (G) Representative images of labeling for Iba1 are shown. White arrows indicate the enlarged insert of a representative cell. The scale bar represents 100 μm. (H-M) DBZ inhibited LPS-induced pro-inflammatory M1 genes expression and increased anti-inflammatory M2 genes expression in mice cortex. The mRNA levels of IL-1β, IL-6, TNF-α, iNOS, CD206 and IL-10 were determined by qRT-PCR. (N, O) The protein levels of p-Akt, p-GSK3β, nuclear Fyn, nuclear Nrf2, HO-1 and NQO1 were determined by Western blotting assay and quantified by densitometry. Results are expressed as means ± SD (n = 12 mice per group for B-E; n = 5 mice per group for F and G; n = 4 mice per group for H-M; n = 3 mice per group for N and O). *p < 0.05 compared with the VEH + SAL group, #p < 0.05 compared with the VEH + LPS group. Statistical analyses were performed by one-way ANOVA followed by Bonferroni's post hoc test or Kruskal–Wallis test with Dunn's post hoc test. VEH: vehicle; SAL: saline.