Transcriptomic profiling of myocardial biopsies reveals metabolic suppression and autophagy activation in systemic sclerosis-associated primary heart involvement

Figures

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  Figure 1. Overview of myocardial transcriptomic differences between patients with systemic sclerosis (SSc) and diastolic myopathy (DCM). (A) The Minus versus Average (MA) and volcano plots of differentially expressed genes (DEGs) identified by DESeq2. Genes with adjusted p<0.05 and an absolute fold change ≥2 were considered significant. (B) Heatmap of the top 50 DEGs, shown as row-wise Z-scores of normalised expression. (C) Gene Set Enrichment Analysis of the top five positively and negatively enriched pathways across Kyoto Encyclopaedia of Genes and Genomes (KEGG), HALLMARK and Gene Ontology Biological Process (GOBP) databases, ranked by normalised enrichment score (NES).

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  Figure 2. Heatmap of pathway-specific gene expression. Z-score normalised expression levels of selected genes in systemic sclerosis (SSc) and dilated cardiomyopathy (DCM) samples. Genes were selected based on previous reports of their involvement in the pathogenesis of SSc and heart failure. Genes were grouped into the following categories: inflammation, fibrosis, ischaemia/hypoxia, vascular injury, energy metabolism, neurohumoral factors and autophagy, and subjected to hierarchical clustering.

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  Figure 3. Integrative pathway comparisons of systemic sclerosis (SSc) and non-failing (NF) myocardium samples relative to dilated cardiomyopathy (DCM). (A) Gene Set Enrichment Analysis bubble plot for the public cohort (NF vs DCM) showing the top five positively and negatively enriched pathways from the Kyoto Encyclopaedia of Genes and Genomes (KEGG), HALLMARK and Gene Ontology Biological Process (GOBP) analyses, ranked by normalised enrichment score (NES). (B) Bar chart of the top 10 discordantly enriched pathways between SSc and NF, selected for nominal p<0.05 in the local cohort and ranked by the largest absolute normalised enrichment score (NES) differences. (C) Summary of differential upstream regulators (UR), canonical pathways (CP) and downstream effects (DE) between SSc and NF in the ingenuity pathway analysis (IPA) with p<0.05 and ranked by the absolute Z-score difference; red and blue denote activation and inhibition, respectively.

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  Figure 4. LC3 expression by immunohistochemistry in the myocardial specimens obtained from patients with systemic sclerosis (SSc) and those with dilated cardiomyopathy (DCM). (A) Image of representative sections show 3,3’-diaminobenzidine-positive LC3 puncta in SSc-primary heart involvement (right) and DCM (left). (B) LC3-positive area proportions per region of interest were compared between SSc (n=7) and DCM (n=8). Samples SSc_1, SSc_2, SSc_3, SSc_4, DCM_1, DCM_2, DCM_3 and DCM_4 were used for RNA sequence analysis. PN: patient number.