BMP6, a potential biomarker of inflammatory fibrosis and promising protective factor for dilated cardiomyopathy

Figures

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  Fig. 1. Differential expression analysis and WGCNA. A Normalization between samples. B Differential expression analysis of bulk RNA-seq sequencing results between DCM and NH. C Sample discrete dendrogram and heat map of disease trait distribution. D Scale independence and mean connectivity change curves and soft-threshold cutoff. E Gene module clustering dendrogram. F Correlation between DCM phenotypic characteristics and gene modules. G Correlation between yellow module membership and gene significance for DCM

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  Fig. 2. Assessment of cardiac immune response in DCM. A Differences in the abundance percentage of 22 immune cells across sample species. B Differences in the infiltration of different immune cells in each sample. C Differences in immune cell infiltration between DCM and NC

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  Fig. 3. Significant gene and assessment of biomarker sensitivity. A Heatmap of differential expression of immune-related genes between DCM and NC. B Heatmap of differential expression of senescene-related genes between DCM and NC. C Intersection of ImmDEGS, SenDEGS and gene sets in MEyellow. D Sensitivity and specificity analysis of ROC curves between BMP6 and DCM. E Correlation between BMP6 and 22 types of immune cells

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  Fig. 4. Cell clustering and annotation. A Elbow diagram obtained using the harmony method and we selected 0–22 as the number of dimensions. B Specific markers and expression levels in different cardiac cell populations. C–E Subdividing and annotating cardiac cell populations by TSNE method. F–H Subdividing and annotating cardiac cell populations by Umap method

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  Fig. 5. BMP6 expression in different cellular subpopulations. A Differential expression of BMP6 in various cell types of the heart. B Differential gene expression in fibroblasts between DCM and NH. C Differential gene expression in endothelial cells between DCM and NH. D Elbow diagram obtained using the harmony method and we selected 0–15 as the number of dimensions. E Specific markers and expression levels in fibroblasts and myofibroblasts. F–I subdividing and annotating fibroblast subpopulations by TSNE and Umap methods. J BMP6 expression between conventional fibroblasts and myofibroblasts. K differential expression of BMP6 in fibroblasts between DCM and NH. L Specific markers and expression levels of macrophage subpopulations. M–P Subdividing and annotating macrophage subpopulations by TSNE and Umap methods. Q Differences in the percentage of macrophage subpopulations abundance between DCM and NH

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  Fig. 6. Mendelian randomization analysis between BMP6 and DCM. A Scatter trend plot of effective SNPs. B The black line represents the nature of each SNP, and the red lines represent the combined nature of the SNPs analyzed by the IVW and MR Egger methods, respectively. C SNPs distribution map. D Leave-one-out analysis. The red line shows the results of the combined analysis

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  Fig. 7. Assessment of cardiac fibrosis and RNA-Seq analysis. A Comparison of cardiac ejection fraction between DCM and NC rats. B Comparison of left ventricular fractional shortening between DCM and NC rats. C and D Comparison of left ventricular internal diameters in systole and diastole between DCM and NC rats. E Echocardiography testing of cardiac function. F Morphology of longitudinal sections of the heart under HE staining. G HE staining of the lateral inner edge of the left ventricular wall of the heart at 50 × magnification. H Masson staining of the area located at the lateral inner edge of the left ventricular wall of the heart, viewed at 20 × magnification. I Comparison of fibrosis area by Masson staining (n = 3). J ELISA detection of changes in NT-proBNP in serum (n = 4). K and L Differential expression analysis of transcriptomics results between DCM and NC rats

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  Fig. 8. Immunofluorescence staining and cardiac tissue protein assay and BMP6 knockdown. A Immunofluorescence staining of markers specific to macrophages and type 2 macrophages. B and C Western blot was used to detect the changes in protein levels in the cardiac tissues between DCM and NC (n = 3). D and E The knockdown of BMP6 and the expression levels of SMAD6 and COL1A1 (n = 3). The siRNA group represents the knockdown group, si-NC represents the empty vector group, and blank represents the control group. The significance of *P in the above graph: *P < 0.05; **P < 0.01. F Potential interventional herbs based on BMP6 inverse prediction