Naringin Interferes Doxorubicin-Induced Myocardial Injury by Promoting the Expression of ECHS1

Figures

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  FIGURE 1. (A) Basic structure of Naringin (NR). (B) Schematic protocol for mice treatments showing models of exposure doxorubicin (DOX), which were performed in mice to test the efficacy of Naringin (NR).

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  FIGURE 2. Naringin inhibits the effect of DOX on myocardial atrophy in vivo. (A) Heart weight of four groups. (B) H&E staining of heart sections (Scale bar 1,500 μm). (C) Effect of NR on the ratios of heart weight to body weight (HW/BW). (D) Body weight of four groups. (E) Effect of NR on the ratios of heart weight to tibia length (HW/TL). (F) Heart cross-sections were stained with WGA, (left panel). Quantification of the relative myocyte cross-sectional area with quantification analysis (right panel). ***p < 0.001 DOX versus control group; # p < 0.05, ## p < 0.01, and ### p < 0.001 DOX versus NR + DOX.

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  FIGURE 3. Naringin inhibits the effect of DOX on apoptosis, inflammation and Oxidative stress in vivo (A) Naringin protects heart from vacuolization induced by DOX. H&E staining of heart sections (left panel), with quantification analysis (right panel). (B) The content of LDH (lactic acid dehydrogenase) activity in cardiac. (C) The content of CAT activity in cardiac. (D) The content of GSH-PX activity in cardiac. (E) The content of SOD activity in cardiac. (F) The content of MDA activity in cardiac. (G) Cardiac mRNA expression level of cybb was measured by qPCR analysis. (H) Cardiac mRNA expression level of nox4 was measured by qPCR analysis. (I) Mice heart sections were conducted TUNEL staining for determination of apoptotic cells (black arrows indicate TUNEL positive cells, left panel) with quantification of apoptotic cells with quantification analysis (right panel). (J) The expression of Bax, Bcl-2 and β-actin detected by Western Blot. (K–M) The level of IL-1β, IL-6 and TNF-α detected by ELISA. ***p < 0.001 DOX versus control group; ## p < 0.01, and ### p < 0.001 DOX versus NR + DOX.

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  FIGURE 4. Naringin inhibits the effect of DOX on apoptosis, inflammation and Oxidative stress in vitro (A) Effect of Naringin on H9C2 cells in the presence of DOX. Cell counting kit 8-based cell viability assays show Cell viability of H9C2 cells in the presence of DOX. (B) Effects of Naringin on the protein expression levels of Bcl-2, Bax and NOX2 in H9C2 cells (left panel) with quantification analysis (right panel). (C) Naringin treatment increases Dox-Induced Bcl-2 Secretion in NRVMS, which displays representative imaging of bcl-2 immunofluorescence staining. Bcl-2 cells are in green, DAPI in blue. (D) Representative Mito-SOX fluorescence images indicate the level of mitochondrial damage (left panel) with quantification of Mito-SOX florescence intensity. (E) Representation image of IF staining of the DNA damage markers γ-H2AX (left panel). The average number of γ-H2AX foci in γ-H2AX-positive NRVMS. (F–H) The level of IL-1β, IL-6 and TNF-α detected by ELISA. ***p < 0.001 DOX versus control group; ## p < 0.01, and ### p < 0.001 DOX versus NR + DOX.

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  FIGURE 5. Inhibition of ECHS1 interferes the effect of Naringin on DOX-induced on apoptosis, inflammation and Oxidative stress in vitro. (A) the protein expression levels of ECHS1 in heart tissue (left panel) with quantification analysis (right panel); (B) the protein expression levels of Bcl-2 in H9C2 cells (left panel) with quantification analysis (right panel); (C) representative imaging of ECHS1 immunofluorescence staining. ECHS1 cells are in green, DAPI in blue.; (D) the protein expression levels of ECHS1 after si-ECHS1 transfection; (E) the activity of caspase3 in different group; (F) the protein expression levels of Bcl-2, BAX, NOX2 and β-actin in heart tissue (left panel) with quantification analysis (right panel). (G) The content of SOD activity in H9C2. (H) The content of LDH activity in H9C2. (I–K) The level of IL-1β, IL-6 and TNF-α detected by ELISA. **p < 0.01, ***p < 0.001 DOX versus control group or si- ECHS1 versus control group; ## p < 0.01, and ### p < 0.001 DOX versus NR + DOX; $$ p < 0.01 NR + DOX versus NR + DOX + si- ECHS1.

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  FIGURE 6. Schematic diagram of the mechanism.