Multi-Omic Insight Into the Molecular Networks in the Pathogenesis of Coronary Artery Disease

Abstract

Background: Genome-wide association studies have revealed numerous loci associated with coronary artery disease (CAD). However, some potential causal/risk genes remain unidentified, and causal therapies are lacking.

Methods and results: We integrated multi-omics data from gene methylation, expression, and protein levels using summary data-based Mendelian randomization and colocalization analysis. Candidate genes were prioritized based on protein-level associations, colocalization probability, and links to methylation and expression. Single-cell RNA sequencing data were used to assess differential expression in the coronary arteries of patients with CAD. TAGLN2 (Transgelin 2), APOB (Apolipoprotein B), and GIP (Glucose-dependent insulinotropic polypeptide) were identified as the genes most strongly associated with CAD, with TAGLN2 exhibiting the most significant association. Higher methylation levels of TAGLN2 at specific Cytosine-phosphate-Guanine sites were negatively correlated with its gene expression and associated with a lower risk of CAD, whereas higher circulating TAGLN2 protein levels were positively associated with CAD risk (odds ratio,1.66 [95% CI, 1.32-2.08). These results suggest distinct regulatory mechanisms for TAGLN2. In contrast, APOB and GIP showed positive associations with CAD risk, whereas DHX58 (DExH-box helicase 58) and SWAP70 (Switch-associated protein 70) were associated with decreased risk.

Conclusions: Our findings provide multi-omics evidence suggesting that TAGLN2, APOB, GIP, DHX58, and SWAP70 genes are associated with CAD risk. This work provides novel insights into the molecular mechanisms of CAD and highlights the potential of integrating multi-omics data to uncover potential causal relationships that cannot be fully captured by traditional genome-wide association studies.

Keywords: Mendelian randomization; coronary artery disease; methylation; multi‐omics evidence.

Figure 1. Associations of genetically predicted gene methylation with coronary artery disease in summary‐based Mendelian randomization analysis.

OR indicates odds ratio; and PPH4, posterior probability of H4.

Figure 2. Associations of genetically predicted gene expression with coronary artery disease in summary‐based Mendelian randomization analysis.

OR indicates odds ratio; and PPH4, posterior probability of H4.

Figure 3. Associations of genetically predicted gene encoded protein with coronary artery disease in summary‐based Mendelian randomization analysis.

OR indicates odds ratio; and PPH4, posterior probability of H4.

Figure 4. A chromosome map of TAGLN2 illustrates the effect values of SNPs within 500 kbp upstream and downstream regions, identified as eQTLs and mQTLs, in the context of coronary artery disease GWAS data.

The x axis denotes the chromosomal position coordinates, whereas the y axis represents the negative logarithm of P values (including eQTLs, mQTLs, GWAS data, and SMR). The red dashed line indicates the P selection threshold of 1e‐5 for eQTLs analyzed with SMR, with genes exhibiting P<1e‐5 represented by red diamonds; the blue dashed line represents the P selection threshold of 0.001 for mQTLs analyzed with SMR, with methylated sites displaying P<0.001 denoted by blue solid circles. eQTL indicates expression quantitative trait loci; GWAS, genome‐wide association study; mQTL, methylation quantitative trait loci; SMR, summary‐data‐based Mendelian randomization; and SNPs, single‐nucleotide polymorphisms.

Figure 5. Associations of genetically predicted gene encoded protein with coronary artery disease in 2‐sample Mendelian randomization analysis.

OR indicates odds ratio.

Figure 6. Single‐cell type expression in atherosclerotic plaque for the 5 genes identified by Mendelian randomization.

A, A total of 17 cell clusters and 8 cell types based on UMAP visualization. B through C, Expression patterns of 4 genes (TAGLN2, SWAP70, DHX58, and APOB) across cell‐type clusters visualized using UMAP and dot plots. D, Differential expression of 3 genes (TAGLN2, SWAP70, and DHX58) between atherosclerotic plaques and controls, shown in 8 cell types at average Log2FC >0.25 and FDR <0.05 level. FDR indicates false discovery rate; Log2FC, log2 fold change; SMC, smooth muscle cells; T/NK, T or Natural Killer cells and UMAP, uniform manifold approximation and projection.

Figure 7. Validation of gene and protein expression in the aortas of AS and control mice.

A, Sequential panels show Oil Red O staining of the aorta, Oil Red O staining of the aortic valve, hematoxylin and eosin staining of the aortic valve, and Masson's trichrome staining of the aortic valve. B, Western blot DHX58, SWAP70, and TAGLN2 protein expression in the aortic tissues of AS and control mice, with GAPDH used as a loading control. C, Quantification of DHX58, SWAP70, and TAGLN2 protein levels in the aortic tissues of AS and control mice. Data are presented as mean±SE (n=6 per group, analyzed using the Mann‐Whitney U test). D, Quantification of DHX58, SWAP70, and TAGLN2 mRNA levels in the aortic tissues of AS and control mice. Data are presented as mean±SE (n=6 per group, analyzed using the Mann‐Whitney U test). AS indicates atherosclerotic. KD, kiloDalton.