Apocynum venetum leaf extract alleviated doxorubicin-induced cardiotoxicity by regulating organic acid metabolism in gut microbiota

Figures

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  FIGURE 1. Effects of AVLE on cardiac function after 5-week Dox treatment in mice. (A) Representative images of echocardiography (B) the ejection fraction (EF%) of mice in different groups (C) the fractional shortening (FS%) (D) left ventricular anterior diastolic wall thickness (LVAWd) (E) left ventricular anterior systolic wall thickness (LVAWs) (F) left ventricular end diastolic diameter (LVEDd) (G) left ventricular end systolic diameter (LVESd) (H) the ratios of heart weights to body weights were calculated. Data are presented as mean ± SD; n = 6, # p < 0.05, ## p < 0.01, ### p < 0.001 compared with model group; *p < 0.05, ** p < 0.01, *** p < 0.001 compared with A2 group.

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  FIGURE 2. Cardiac tissue staining and detection of cardiac injury indicators. (A) H&E and Masson staining of cardiac tissue after AVLE treatment (B–D) Serum brain natriuretic peptide (BNP), creatine kinase (CK) and lactate dehydrogenase (LDH) of Dox-induced cardiotoxicity mice after AVLE treatment. Data are presented as mean ± SD; n = 6, # p < 0.05, ## p < 0.01, ### p < 0.001 compared with model group; *** p < 0.001 compared with A2 group.

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  FIGURE 3. AVLE regulating organic acids generated by gut microbiota. (A–H) Detection of acetic acid (AA), propionic acid (PA), indole-3- propionic acid (IPA), butyric acid (BA), valeric acid (VA), isovaleric acid (iVA), hexanoic acid (HA) and 4-methylvaleric acid (4-MVA) in mice feaces after AVLE treatment (I) heatmap of organic acids concentration. N, normal group; M, model group; P, PGF group. Data are presented as mean ± SD; n = 6, *p < 0.05, *** p < 0.001 compared with model group.

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  FIGURE 4. Regulation of AVLE on mice gut microbiota. (A,B) The concentration of IPA and AA in gut microbiota culture system in vitro (C,D) IPA and AA in mice serum after treated with AVLE for 24 h (E) principal components analysis (PCA) score plot of three groups, N (normal group), M (model group), A (A2 group) (F) heatmap of top 50 genera in three groups, N (normal group), M (model group), A (A2 group). Data are presented as mean ± SD; n = 6, *p < 0.05, **p < 0.05, ***p < 0.001.

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  FIGURE 5. Serum brain natriuretic peptide (BNP), creatine kinase (CK) and lactate dehydrogenase (LDH) of Doxn-induced cardiotoxicity mice after treated with IPA and AA. Data are presented as mean ± SD; n = 6, *p < 0.05, **p < 0.05 compared with model group.

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  FIGURE 6. IPA improved cardiomyocyte apoptosis induced by Dox. (A) Flow cytometric scatterplot of Annexin-V-FLTC staining of HL-1 cell apoptosis (B) Flow cytometric histogram of Annexin-V-FLTC staining of HL-1 cell apoptosis (C–E) quantitative analysis of apoptotic HL-1 cells in different apoptosis stages, data are presented as mean ± SD; n = 3, ### p < 0.001 compared with Dox group, ***p < 0.001 compared with control group.

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  FIGURE 7. Cardioprotection function of AA by stimulating the secretion of Ghrelin. (A) Gh in mice plasma after Dox-induced cardiotoxicity mice treated with AA, data are presented as mean ± SD, n = 6, ***p < 0.001 compared with model group (B) Gh in mice plasma after normal mice operated with vagotomy, data are presented as mean ± SD, n = 6, ***p < 0.001 compared with sham group, ### p < 0.001 compared with sham AA group (C) BNP in mice serum after the 3 days AA treatment, data are presented as mean ± SD, n = 6, **p < 0.01, ## p < 0.01 (D–G) The mRNA levels of Collagen I, Collagen III, Activin A (Act A) and Follistatin (FS) in HL-1 cell after treated with Gh, data are presented as mean ± SD, n = 3, **p < 0.01 compared with control group, # p < 0.05, ## p < 0.01 compared with Dox group.

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  FIGURE 8. Potential mechanism of AVLE treated Dox-induced cardiotoxicity by regulating gut microbita.