Sodium Butyrate Attenuates Isoprenaline-Induced Myocardial Injury via Restoring the Gut–Heart Axis and Suppressing TLR4/NF-κB Signaling

Figures

• 

  Figure 1. Sodium butyrate attenuated isoprenaline-induced cardiac injury, oxidative stress, and fibrosis. Serum levels of cardiac injury markers (A) cardiac troponin (cTnI), (B) creatine kinase-MB (CK-MB), and (C) lactate dehydrogenase (LDH). Myocardial oxidative stress indicators: (D) malondialdehyde (MDA) content and (E) superoxide dismutase (SOD) activity. (F) Representative hematoxylin and eosin (H&E) staining of myocardial tissue (scale bar = 50 µm). (G) Semi-quantitative histological injury scoring. (H) Masson’s trichrome staining (scale bar = 50 µm). (I) Quantification of fibrotic area. Data are expressed as mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001; ns, not significant.

• 

  Figure 2. Sodium butyrate restored intestinal barrier structure and tight-junction integrity after ISO-induced myocardial injury. (A) Representative H&E staining of colonic tissue (scale bar = 100 µm). (B) Semi-quantitative histological injury scoring. Colonic mRNA expression of tight junction genes: (C) Tjp1 (ZO-1), (D) Cldn1 (claudin-1) determined by qRT-PCR. (E) Plasma endotoxin (lipopolysaccharide, LPS) levels. Representative immunofluorescence staining of tight junction proteins in colonic epithelium: (F) ZO-1, (G) claudin-1 (green fluorescence; nuclei counterstained with DAPI, blue) (scale bar = 50 µm). Quantification of fluorescence intensity for (H) ZO-1 and (I) claudin-1. Data are expressed as mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001; ns, not significant.

• 

  Figure 3. Sodium butyrate suppressed systemic inflammation induced by isoprenaline injury. Serum concentrations of pro-inflammatory cytokines: (A) tumor necrosis factor-alpha (TNF-α), (B) interleukin-6 (IL-6), and (C) interleukin-1β (IL-1β), measured by ELISA. Data are expressed as mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001; ns, not significant.

• 

  Figure 4. Sodium butyrate inhibited cardiac TLR4/NF-κB signaling activation after isoprenaline-induced injury. (A) Representative Western blot images showing protein expression of TLR4, IκBα, phosphorylated NF-κB p65 (p-p65), and β-actin (loading control) in cardiac tissue. Densitometric quantification of (B) TLR4, (C) IκBα, and (D) p-p65 protein expression, each normalized to the β-actin band from the same lane. Data are expressed as mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001; ns, not significant.

• 

  Figure 5. Sodium butyrate remodeled gut microbial community structure in isoprenaline-injured rats. Alpha diversity indices (Chao1, Shannon, observed species) (A–C); shared lowercase letter “a” above boxplots indicates groups not statistically different from one another (Kruskal–Wallis test, p > 0.05). LEfSe analysis showing discriminative taxa among groups (linear discriminant analysis score LDA ≥ 2.0, α = 0.05) (D). Beta diversity ordination by PCA, PCoA, and NMDS (E–G); sample labels C1-3, B1-3, M1-3, and T1-3 denote individual animals within the CON, CON-SB, MOD, and MOD-SB groups, respectively. Genus-level relative abundance composition (H) and relative abundance of selected genera significantly altered by ISO-induced myocardial injury and/or SB treatment (I–M). Data are expressed as mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; ns, not significant.

• 

  Figure 6. Serum metabolomic remodeling by sodium butyrate in isoprenaline-induced myocardial injury. (A) OPLS-DA score plots showing group separation across three pairwise comparisons; sample labels C1-3, B1-3, M1-3, and T1-3 denote individual animals within CON, CON-SB, MOD, and MOD-SB groups, respectively. (B) Heatmap of representative differential metabolites with hierarchical clustering. (C) Volcano plots of differential features for each pairwise comparison; sample labels C1-3, B1-3, M1-3, and T1-3 denote individual animals within the CON, CON-SB, MOD, and MOD-SB groups, respectively. (D) KEGG pathway enrichment analysis of SB-responsive metabolites. (E) Venn diagram summarizing overlaps of differential metabolites among key comparisons.

• 

  Figure 7. Microbiome–metabolome correlation heatmap shows SB-dependent reorganization of gut-systemic metabolic associations. Spearman correlation heatmap between differential bacterial genera and representative differential serum metabolites. Color intensity reflects Spearman’s correlation coefficient (ρ). Both genera and metabolites were hierarchically clustered to identify association modules. Asterisks denote correlations that remained significant after Benjamini–Hochberg false discovery rate correction (p < 0.05). * p < 0.05, ** p < 0.01, *** p < 0.001.