Integrated single-cell RNA sequencing and mendelian randomization analysis identifies causal immune-related driver genes in the heart failure inflammatory microenvironment

Figures

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  Figure 1. Basic cell atlas of the GSE198784 and GSE247468 databases. (A) t-SNE-based cell clustering atlas of GSE198784. (B) t-SNE-based cell clustering atlas of GSE247468. (C) Manually annotated cell atlas of GSE198784 based on t-SNE. (D) Manually annotated cell atlas of GSE247468 based on t-SNE. (E) GSE198784 Markers. (F) GSE247468 markers. Ident, identity; t-SNE, t-distributed stochastic neighbor embedding.

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  Figure 2. Cell proportions in the GSE198784 and GSE247468 databases. (A) Stacked bar chart of intergroup cell proportions in GSE198784. (B) Stacked bar chart of intergroup cell proportions in GSE247468. (C) A heatmap of inter‑group differences in immune cell numbers. (D) Venn diagram of differential genes in GSE198784 and GSE247468. (E) Boxplots of the expression levels of highly expressed genes. (F) Boxplots of the expression levels of moderately-highly expressed genes. (G) Boxplots of the expression levels of moderately expressed genes. (H) Boxplots of the expression levels of lowly expressed genes. DEG, differentially expressed gene.

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  Figure 3. Mendelian randomization analysis. (A) Flowchart for screening 65 highly confident immune-related causal genes. (B,C) Scatter plots showing the causal relationship between key genes (TNNC1 and IFNGR1) and heart failure. In the graphs, different colors represent different statistical methods, and the slope of the line indicates the causal effect of each method. (D,E) Forest plots. Each forest plot represents an SNP corresponding to a key gene, showing the results of the leave-one-out analysis. Forest plot (D) shows the preliminary results of the causal relationship between TNNC1 and heart failure. Forest plot (E) shows the preliminary results of the causal relationship between IFNGR1 and heart failure. DEG, differentially expressed gene; FDR, false discovery rate; MR, Mendelian randomization; OR, odds ratio; SNP, single nucleotide polymorphism.

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  Figure 4. Enrichment analysis. (A,B) The GO enrichment analysis revealed that key genes causally associated with heart failure were primarily enriched in biological processes such as the regulation of lymphocyte, leukocyte, and T cell activation and tumor necrosis factor receptor activity. (C,D) KEGG enrichment analysis indicated that these key genes were mainly enriched in pathways including leukocyte transendothelial migration, natural killer cell-mediated cytotoxicity, and JAK-STAT, IgSF CAM, ErbB, and C-type lectin receptor signaling pathway. (E,F) GSEA further demonstrated enrichment of these genes in signaling pathways such as the IL-17 signaling pathway, neutrophil extracellular trap formation, the NF-κB signaling pathway, and natural killer cell-mediated cytotoxicity. BP, biological process; DEG, differentially expressed gene; GO, Gene Ontology; GSEA, Gene Set Enrichment Analysis; IL, interleukin; KEGG, Kyoto Encyclopedia of Genes and Genomes; MF, molecular function.

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  Figure 5. Association analysis between key genes and immune infiltration. (A) Violin plot showing the differential expression of TNNC1 in cardiomyocytes between groups in the GSE198784 dataset. (B) Violin plot showing the differential expression of IFNGR1 in T cells between groups in the GSE198784 dataset. (C) Violin plot showing the differential expression of TNNC1 in cardiomyocytes between groups in the GSE247468 dataset. (D) Violin plot showing the differential expression of IFNGR1 in neutrophils between groups in the GSE247468 dataset. (All comparisons, P<0.05).