Astragaloside IV Reduces Sorafenib-Induced Cardiotoxicity by Inhibiting Apoptosis Through the STAT3/HIF-1α/Bcl-2 Signaling Pathway

Figures

• 

  Figure 1. Network pharmacology analysis reveals potential targets and pathways of AST against SOR-induced injury. (A) The Venn diagram illustrating the overlapping targets of AST- and SOR-induced cardiac injury. (B) PPI network. (C) Top 10 enriched GO terms in BP, CC, and MF categories. (D) Top 17 significantly enriched KEGG pathways.

• 

  Figure 2. Validation of STAT3 as a direct binding target of AST. (A) Venn diagram of potential AST targets, SOR injury−related genes, and genes screened from the human proteome microarray. (B) Core targets. (C) KEGG pathway. (D) Molecular docking result between AST and STAT3. (E) RMSD of the STAT3 backbone during molecular dynamics simulation. (F) Chemical structures of AST and its bioactive form. (G) Mass spectrum of Bio−AST−STAT3 binding complex. (H) Representative images of the protein array showed a positive control (marked with a green arrow), a negative control (marked with a blue arrow), and a STAT3 spot (marked with a yellow arrow) in a selected region of the microarrays. (I) Enlarged images and IMean_Ratio value for STAT3.

• 

  Figure 3. Astragaloside IV attenuates sorafenib-induced cardiotoxicity in mice. (A) Representative M-mode echocardiograms (parasternal short-axis view) and quantitative analysis of LVEF and LVFS, n = 6. (B) Masson’s representative trichrome-stained cardiac sections (scale bar = 200 μm), n = 5. (C) Representative HE-stained cardiac sections (scale bar = 50 μm), n = 6. (D) Representative TEM images (red arrows: damaged mitochondria; scale bar = 200 nm), n = 6. (E–I) Quantitative analysis of relative collagen content, CK-MB, BNP, SOD, MDA, and LDH levels. ## p < 0.01 vs. control; ** p < 0.01 vs. SOR.

• 

  Figure 4. AST attenuated SOR-induced cardiomyocyte injury and oxidative stress. (A,B) Representative illustrations of treated cardiomyocytes (scale bars = 20 µm) and quantitative analysis of TUNEL mean fluorescence intensity, n = 3. (C–F) Representative Western blot of Bcl-2, HIF-1α, and STAT3, n = 3. ## p < 0.01 vs. control; * p < 0.05 and ** p < 0.01 vs. SOR; && p < 0.01 vs. ML115.

• 

  Figure 5. AST protected against SOR−induced cardiomyocyte injury in HL−1 cells. (A) Cardiomyocyte viability assessed after treatment with SOR alone or in combination with different concentrations of AST (1.25−160 µM), n = 6. (B−D) Quantification of LDH release, SOD activity, and MDA content, n = 6 (AST−L, 10 µM), (AST−M, 20 µM), and (AST−H, 40 µM). (E,F) Hoechst 33258 staining in cardiomyocytes: mean fluorescence intensity and representative microscopy images. Scale bars = 50 µm, n = 6. ## p < 0.01 vs. control; * p < 0.05, ** p < 0.01 vs. SOR group.

• 

  Figure 6. AST alleviates SOR-induced mitochondrial oxidative stress and apoptosis through the STAT3/HIF-1α/Bcl-2 pathway. (A,C) Representative fluorescence images of MitoSOX staining, n = 3. Scale bars = 50 µm. (B,D) Representative images of TUNEL staining, n = 3. Scale bars = 50 µm (AST, 40 µM). (E–H) Representative Western blot images of STAT3, HIF-1α, and Bcl-2 protein expression, n = 3. ## p < 0.01 vs. control; * p < 0.05, ** p < 0.01 vs. SOR; && p < 0.01 vs. ML115.

• 

  Figure 7. Mechanism diagram of AST ameliorating sorafenib-induced cardiotoxicity.