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. 2022 Jun 30:9:890321.
doi: 10.3389/fcvm.2022.890321. eCollection 2022.

Macrophages-Related Genes Biomarkers in the Deterioration of Atherosclerosis

Yue Zheng  1   2   3   4   5 , Bingcai Qi  3   4   5   6 , Wenqing Gao  2   3   4   5   6 , Zhenchang Qi  3   4   5   6 , Yanwu Liu  3   4   5   6 , Yuchao Wang  1   3   4   5   6 , Jianyu Feng  3   4   5   6 , Xian Cheng  3   4   5   6 , Zhiqiang Luo  3   4   5   6 , Tong Li  1   2   3   4   5   6
Affiliations

Macrophages-Related Genes Biomarkers in the Deterioration of Atherosclerosis

Yue Zheng et al. Front Cardiovasc Med. .

Abstract

Background: The macrophages are involved in all stages of cardiovascular diseases, demonstrating the correlation between inflammation, atherosclerosis, and myocardial infarction (MI). Here, we aim to investigate macrophages-related genes in the deterioration of atherosclerosis.

Methods: GSE41571 was downloaded and the abundance of immune cells was estimated by utilizing the xCell. By utilizing the limma test and correlation analysis, differentially expressed macrophages-related genes (DEMRGs) were documented. The functional pathways and the protein-protein interaction (PPI) network were analyzed and the hub DEMRGs were obtained. The hub DEMRGs and their interactions were analyzed using NetworkAnalyst 3.0 and for validation, the expressions of hub DEMRGs were analyzed using the GSE135055 and GSE116250 datasets as well as atherosclerosis and MI mice model.

Results: A total of 509 differentially expressed genes (DEGs) were correlated with the abundance of macrophages and were identified as DEMRGs (Pearson correlation coefficients (PCC) > 0.6), which were mainly enriched in extracellular structure organization, lysosomal membrane, MHC protein complex binding, and so on. After screening out, 28 hub DEMRGs were obtained with degrees ≥20, including GNAI1 (degree = 113), MRPS2 (degree = 56), HCK (degree = 45), SOCS3 (degree = 40), NET1 (degree = 28), and so on. After validating using Gene Expression Omnibus (GEO) datasets and the atherosclerosis and MI mice model, eight proteins were validated using ApoE-/- and C57 mice. The expression levels of proteins, including SYNJ2, NET1, FZD7, LCP2, HCK, GNB2, and PPP4C were positively correlated to left ventricular ejection fraction (LVEF), while that of EIF4EBP1 was negatively correlated to LVEF.

Conclusion: The screened hub DEMRGs, SYNJ2, NET1, FZD7, LCP2, HCK, GNB2, EIF4EBP1, and PPP4C, may be therapeutic targets for treatment and prediction in the patients with plaque progression and MI recurrent events. The kit of the eight hub DEMRGs may test plaque progression and MI recurrent events and help in the diagnosis and treatment of MI-induced heart failure (HF), thus decreasing mortality and morbidity.

Keywords: GO/KEGG pathways analysis; GSEA; PPI; atherosclerosis; differentially expressed genes; immune infiltration; macrophages; progression.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Identification of differentially expressed genes (DEGs) in ruptured plaques and stable plaques. (A) Abundance of immune cells estimated by the xCell algorithm in GSE41571. (B) Upset diagram of plasm B cells, fibroblasts, activated myeloid dendritic cells, M0, M1, and M2 Macrophages-related DEGs with Pearson correlation coefficients >0.6. (C,D) Heatmap (C) and Volcano plot (D) of DEGs between ruptured plaques and stable plaques.
Figure 2
Figure 2
Enrichment pathway analysis of the DEMRGs. (A) Venn diagram showed 509 differentially expressed genes (DEGs) among M0, M1, and M2 macrophages were regarded as DEMRGs. (B) The significant Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways enriched by DEMRGs. (C) The gene sets associated with DEMRGs using Gene Set Enrichment Analysis (GSEA) analysis. DEMRGs, differentially expressed macrophages-related genes.
Figure 3
Figure 3
The hub differentially expressed macrophages-related genes (DEMRGs) and enrichment pathway analysis. (A) The hub DEMRGs were shown using NetworkAnalyst 3.0. (B) The significant GO and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways enriched by the hub DEMRGs. (C) The heatmap of the hub DEMRGs in GSE41571. (D) Left ventricle tissue-specific protein–protein interactions (PPIs) were applied, demonstrating UBC, FN1, PPP1CA, YAP1, and SOCS3 were core regulators in the network.
Figure 4
Figure 4
The hub differentially expressed macrophages-related genes (DEMRGs) and their interactions. (A) The hub DEMRGs-chemical interactions. (B–D) The hub DEMRGs, including FLT1 (B), GNAl1 (C), and HCK (D) and their drug interactions.
Figure 5
Figure 5
Expression patterns of the hub differentially expressed macrophages-related genes (DEMRGs) were validated in the GSE163154 dataset. (A,B) The principal components analysis (PCA) diagram (A) and Volcano plot (B) between the patients with intraplaque hemorrhage and non-intraplaque hemorrhage. (C) The expression level of the hub DEMRGs. (D) The receiver operation characteristic (ROC) curves of the hub DEMRGs validated in the patients with intraplaque hemorrhage compared to control. (E) The joint ROC curves of the hub validated DEMRGs. IPH, intraplaque hemorrhage. *p < 0.05; **p < 0.01; ***p < 0.001; ns, not significant.
Figure 6
Figure 6
Expression patterns of the hub differentially expressed macrophages-related genes (DEMRGs) were validated in the GSE116250 and GSE135055 datasets. (A) The expression level of the hub DEMRGs in patients with dilated cardiomyopathy compared to control. (B) The receiver operation characteristic (ROC) curves of the validated DEMRGs in patients with dilated cardiomyopathy. (C) The expression level of the hub DEMRGs in patients with ischemic cardiomyopathy compared to control. (D) The ROC curves of the validated DEMRGs in patients with ischemic cardiomyopathy. (E) The expression level of the hub DEMRGs in patients with ischemia-induced heart failure compared to control. (F) The ROC curves of the validated DEMRGs in patients ischemia-induced heart failure. DCM, dilated cardiomyopathy; ICM, ischemic cardiomyopathy; HF, heart failure. *p < 0.05; **p < 0.01; ***p < 0.001; ns, not significant.
Figure 7
Figure 7
The protein expression level of the nine screened hub differentially expressed macrophages-related genes (DEMRGs) in atherosclerosis and myocardial infarction (MI) mice model. (A) The expression level of the nine hub DEMRGs in atherosclerosis. (B) The expression level of the nine hub DEMRGs in MI and SHAM groups in ApoE-/- and C57Bl/6J mice. (C–J) The correlation between left ventricular ejection fraction (LVEF) and the nine DEMRGs expression level in the MI mice model, including SYNJ2 (C), NET1 (D), FZD7 (E), LCP2 (F), EIF4EBP1 (G), HCK (H), GNB2 (I), and PPP4C (J). *p < 0.05; **p < 0.01; ***p < 0.001; ns, not significant.
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