T lymphocyte characteristics and immune repertoires in the epicardial adipose tissue of heart failure patients

Figures

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  Figure 1. Overall flowchart of this study.

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  Figure 2. Bioinformatics analyses reveal pro-inflammatory characteristics and key genes of EAT. (A) Volcano plot of DEGs between EAT and SAT in GSE64554 and GSE120774. (B) Top 10 up- and down-regulated DEGs identified by RRA method. (C) GO-BP functional enrichment analyses of up- and down-regulated DEGs in EAT compared to SAT. (D) Cluster dendrogram of WGCNA. (E) WGCNA key modules and EAT/SAT phenotype correlation. (F) GO-BP functional enrichment analyses of WGCNA-identified blue and greenyellow gene modules.

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  Figure 3. Immune cell infiltration and correlation analyses. (A) Violin charts of xCell immune infiltration score between EAT and SAT. (B) Correlation matrix of immune cell subtypes (Pearson correlation coefficients are displayed in the box). (C) Correlation matrix of immune cell infiltration scores with 19 identified key genes (Pearson correlation coefficients are displayed in the box). (D) Correlation matrix of immune cell infiltration scores with WGCNA key modules. *P < 0.05, **P < 0.01, ***P < 0.001 and ns refers to no significance.

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  Figure 4. Amplified lymphocyte activation features in EAT of HF patients. (A) GO-BP functional enrichment analyses of up-regulated DEGs in EAT of HF patients. (B) “Lymphocyte activation” and “myeloid leucocyte activation” GO term genes in DEGs of HF-EAT. (C) Immune cell infiltration analyses of HF-EAT and control EAT by CIBERSORT. (D) Differentially expressed key genes in HF-EAT. (E) Expression of T cell-inflamed GEPs in HF-EAT and control EAT. (F) Correlation of CCL5 and GZMK expression with T cell-inflamed GEPs score in GSE24425. (G) Top 10 potential key TFs of DEGs in HF-EAT identified by ChEA3 database. (H) PPI network of top 10 potential TFs. *P < 0.05, **P < 0.01.

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  Figure 5. Characteristics of TCR repertoires in EAT. (A) TCR clone counts and TCR clonotype counts in EAT and paired SAT. (B) Fraction of low (proportion>0.1%), middle (proportion>0.5%) and high (proportion>1%) frequency TCR clonotypes between EAT and paired SAT. (C, D) The ratio and difference of the top 100 TCR clonotypes in EAT and paired SAT. (E) Total TCR clonotypes sharing between paired EAT and SAT. (F) Total cardiac TCR clonotypes sharing in paired EAT and SAT. (G) Spearman’s correlation of cardiac TCR clonotypes with paired EAT and SAT. (H) Heat map of V-J usage between EAT, paired SAT and heart. (I) Spearman’s correlation of V-J combination (average usage>0.01) in EAT, SAT and heart. *P < 0.05 and ns refers to no significance.

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  Figure 6. Verification of T cells infiltration and key molecules in EAT. (A, B) CD3-specific immunohistochemical staining in EAT and SAT. (C, D) Gating strategy and representative flow cytometry results of IFN-γ+ T lymphocytes in EAT. (E, F) Representative flow cytometry results for proportion of T lymphocytes memory subtypes in EAT. (G) Representative fluorescent staining images of CCL5 and GZMK with CD3 from EAT of HF patients (scale: 50 μm). **P < 0.01, ***P < 0.001 and ns refers to no significance.