Multi-omics reveals the mechanism of vagus nerve stimulation in the treatment of chronic congestive heart failure

Figures

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  Fig. 1. Schematic of model construction and transthoracic echocardiography examination. (A) CHF and VNS model construction process. (B) LVEF measurements at different time points. (C) LVEF statistical analysis after 1 month of VNS stimulation. (ns: p > 0.05, *: p < 0.05, **: p < 0.01, ***: p < 0.001, ****: p < 0.0001).

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  Fig. 2. ELISA detection of changes in metabolites, ChAT, BNP, and inflammatory factors in the blood of CHF dogs. (ns: p > 0.05, *: p < 0.05, **: p < 0.01, ***: p < 0.001, ****: p < 0.0001)

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  Fig. 3. VNS reduces myocardial and mitochondrial damage in CHF dogs. (A) CHF resulted in reduced cardiomyocytes, disordered fibers, and extensive inflammatory infiltration, which were significantly improved after VNS treatment (scale bar: 25 μm. The red arrows indicate inflammatory cells, while the green arrows denote vacuolation). (B) The semi-quantitative scores for HE staining in each group. (C) Representative FACS plots showing ROS levels in myocardial tissue detected by flow cytometry. (D) Representative histogram plots of ROS levels in myocardial tissue across different groups measured by flow cytometry. (E) VNS significantly reduced ROS levels. (F) Representative FACS plots showing JC-1 levels in myocardial tissue detected by flow cytometry. (G) VNS significantly increased mitochondrial membrane potential.(ns: p > 0.05, *: p < 0.05, **: p < 0.01, ***: p < 0.001, ****: p < 0.0001).

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  Fig. 4. Metabolomics analysis to identify differential metabolites following VNS treatment. (A) Volcano plot illustrating the untargeted metabolomics data. (B) Heatmap of differential metabolites. Heatmap showing differentially expressed metabolites, constructed with the pheatmap package (v1.0.12; https://cran.r-project.org/package=pheatmap) in R 3.6.3. (C) Venn diagram showing the differential metabolites that align with the observed trends of change.

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  Fig. 5. Comparison of gut microbiota diversity and abundance between groups. LEfSe analysis reveals intergroup differences and Venn analysis. (A) Alpha diversity among the Control, CHF, and VNS groups. (ns: p > 0.05, *: p < 0.05, **: p < 0.01, ***: p < 0.001, ****: p < 0.0001) (B) PCA and PCoA plots illustrating differences in microbial composition between groups. (C,D) Evolutionary branch diagram and distribution histogram for Group B vs. Group A. (E,F) Evolutionary branch diagram and distribution histogram for Group C vs. Group B. (G) Venn diagram of genus-level microorganisms enriched in Group B.

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  Fig. 6. Results of multi-omics analysis, qPCR validation. (A) Venn diagram illustrating the results of transcriptomics. (B) Correlation analysis between differentially expressed metabolites and microorganisms. (C) Interaction network among microorganisms, metabolites, and genes: green circles denote genes, red hexagons represent microorganisms, pink diamonds indicate metabolites, brown lines signify positive correlations, and blue lines denote negative correlations. (D) VNS notably downregulated the mRNA expression levels of FSTL3, TNFRSF12A, and HBEGF. (ns: p > 0.05, *: p < 0.05, **: p < 0.01, ***: p < 0.001, ****: p < 0.0001)

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  Fig. 7. The potential therapeutic efficacy and treatment targets of VNS in CHF.