Fig 2: Effects of DL0410 on oxidative damage and antioxidative enzyme activity in the hippocampus and cortex. (a) DL0410 decreased the production and accumulation AGEs and MDA and increased the activity of catalase and superoxide dismutase (SOD) in the hippocampus. (b) DL0410 decreased the levels of AGEs and MDA and strengthened the activity of catalase and SOD in the cortex. (c) The Western blot detection and quantitative analysis of SOD1 and SOD2 expression in the hippocampus. (d) The Western blot detection and quantitative analysis of SOD1 and SOD2 expression in cortex. Data are the mean ± SEM (n = 6 − 7). ###p < 0.001 vs. the control group, ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 vs. the model group.

Fig 3: Effect of HFI-419 treatment on expression of genes and proteins with pro-oxidant and antioxidant action and genes coding proteins involved in skeletal muscle and adipose tissue metabolism. Gene expression of superoxide dismutase [Cu-Zn] (Sod1), superoxide dismutase [Mn] (Sod2), extracellular superoxide dismutase [Cu-Zn] (Sod3), nuclear factor erythroid 2-related factor 2 (Nfe2l2), endothelial nitric oxid syntase (Nos3), NADPH oxidase (Nox4) and cytochrome b-245 light chain (Cyba), type-2 angiotensin II receptor (Agtr2), peroxisome proliferator-activated receptor gamma coactivator 1-α (Ppargc1a), forkhead box protein O1 (Foxo1), transforming growth factor β-1 (Tgfb1), fatty acid-binding protein 4 (Fabp4), mesoderm-specific transcript homolog protein (Mest), C-C chemokine receptor type 2 (Ccr2) and plasminogen activator inhibitor 1 (Pai1) in musculus quadriceps (A,E) and epididymal adipose tissue (B,F) of vehicle– and HFI-419–treated obese Zucker rats determined by real-time PCR. Data were normalized to the gene expression of 40S ribosomal protein S29 (Rps29) whose expression was not altered by the treatment. Protein expression of SOD1 and SOD2 and silent information regulator 1 (SirT1) (C) and SOD activity (D) in skeletal muscle of obese vehicle– and HFI-419–treated rats. Expression of proteins was evaluated by western blot and obtained data were normalized to loading control GAPDH whose expression was not altered by the treatment. Representative blots are shown at the panel (G). SOD activity is expressed as percentage of inhibition of tetrazolium salt WST-1 conversion to formazan dye and is related to 1 mg of tissue. Data are presented as mean ± SEM and were analyzed by Student’s t-test: *p < 0.05; **p < 0.01.

Fig 4: Mitochondria targeted antioxidant mitoTEMPO attenuates METH-induced autophagy in cardiomyocytes. (A) Left panel, representative Western blot membrane images demonstrate cellular antioxidant proteins, i.e., MnSOD, SOD1, GPx, and Catalase, in heart whole cell lysates of control and METH self-administered rats. GAPDH was used as the loading control. Right panel, box plots show densitometric quantifications of MnSOD, SOD1, GPx, and Catalase relative expressions as percent change to Control rat hearts. n = 3 male and 3 female rats per group. Data were subjected to nonparametric Kruskal-Wallis test to determine statistical significance. (B) Representative Western blot images demonstrating effects of vehicle, METH, and mitoTEMPO treatment on cellular antioxidant proteins, i.e., MnSOD, SOD1 and autophagy proteins LC3B-II, and p62 in primary neonatal rat cardiomyocytes (NRCs). NRCs were treated with saline (vehicle), METH (100 μM, dissolved in saline), and METH 100 μM in presence of 5 μM and 10 μM mitoTEMPO, respectively, for 24 h. MitoTEMPO (5 μM and 10 μM) were added to the culture medium to quench mitochondria-derived reactive oxygen species (ROS) under METH treatment. METH treatment significantly increased mitochondria localized MnSOD expression accompanied by increased accumulation of autophagy proteins LCB-II and p62 in NRCs compared to Vehicle-treated NRCs. Notably, mitoTEMPO (10 μM) treatment significantly attenuates MnSOD with a concomitant reduction in autophagy proteins LC3B-II and p62 levels compared to METH-only treated NRCs. GAPDH was used as the loading control. Boxes represent interquartile ranges, lines represent medians, and whiskers represent ranges. Data were subjected to One-Way ANOVA followed by Tukey's multiple comparisons test to determine statistical significance among the groups. A P value less than 0.05 between groups was accounted as statistically significant. *P < 0.05, **P < 0.01 and ***P < 0.001. Ctrl, Control; Veh, Vehicle; METH, Methamphetamine.

Fig 5: Proposed mechanisms of reduction in ROS in human coronary endothelial cells (HCAEC) by pequi extract. Hypoxia and H2O2 increase cytosolic and mitochondrial ROS in HCAEC. Polyphenol-rich pequi extract induces expression of antioxidant enzymes as a first line of defense against ROS, possibly by activating the Nrf2/ARE pathway and may reduce ROS and induce HCAEC proliferation. LDL: Low-density Lipoprotein; EC: Endothelial Cells; ARE: Antioxidant Responsive Element; Nrf-2: Nuclear factor-erythroid factor 2-related factor 2; HO-1: Heme Oxygenase-1; SOD-1: superoxide dismutase-1; SOD-2: superoxide dismutase-2; ROS: Reactive Oxygen Species.