Identification of potential biomarkers of vascular calcification using bioinformatics analysis and validation in vivo

Chuanzhen Chen; Yinteng Wu; Hai-Lin Lu; Kai Liu; Xiao Qin

doi:10.7717/peerj.13138

Identification of potential biomarkers of vascular calcification using bioinformatics analysis and validation in vivo

PeerJ. 2022 Mar 16:10:e13138. doi: 10.7717/peerj.13138. eCollection 2022.

Authors

Chuanzhen Chen¹, Yinteng Wu², Hai-Lin Lu¹, Kai Liu¹, Xiao Qin¹

Affiliations

¹ Department of Vascular Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Province, China.
² Department of Orthopedic and Trauma Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Province, China.

Abstract

Background: Vascular calcification (VC) is the most widespread pathological change in diseases of the vascular system. However, we know poorly about the molecular mechanisms and effective therapeutic approaches of VC.

Methods: The VC dataset, GSE146638, was downloaded from the Gene Expression Omnibus (GEO) database. Using the edgeR package to screen Differentially expressed genes (DEGs). Gene set enrichment analysis (GSEA) and gene set variation analysis (GSVA) were used to find pathways affecting VC. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were performed on the DEGs. Meanwhile, using the String database and Cytoscape software to construct protein-protein interaction (PPI) networks and identify hub genes with the highest module scores. Correlation analysis was performed for hub genes. Receiver operating characteristic (ROC) curves, expression level analysis, GSEA, and subcellular localization were performed for each hub gene. Expression of hub genes in normal and calcified vascular tissues was verified by quantitative reverse transcription PCR (RT-qPCR) and immunohistochemistry (IHC) experiments. The hub gene-related miRNA-mRNA and TF-mRNA networks were constructed and functionally enriched for analysis. Finally, the DGIdb database was utilized to search for alternative drugs targeting VC hub genes.

Results: By comparing the genes with normal vessels, there were 64 DEGs in mildly calcified vessels and 650 DEGs in severely calcified vessels. Spp1, Sost, Col1a1, Fn1, and Ibsp were central in the progression of the entire VC by the MCODE plug-in. These hub genes are primarily enriched in ossification, extracellular matrix, and ECM-receptor interactions. Expression level results showed that Spp1, Sost, Ibsp, and Fn1 were significantly highly expressed in VC, and Col1a1 was incredibly low. RT-qPCR and IHC validation results were consistent with bioinformatic analysis. We found multiple pathways of hub genes acting in VC and identified 16 targeting drugs.

Conclusions: This study perfected the molecular regulatory mechanism of VC. Our results indicated that Spp1, Sost, Col1a1, Fn1, and Ibsp could be potential novel biomarkers for VC and promising therapeutic targets.

Keywords: Differentially expressed genes; Functional enrichment analysis; Gene set enrichment analysis; Gene set variation analysis; Protein–protein interaction; Vascular calcification.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Biomarkers, Tumor / genetics
Computational Biology / methods
Gene Expression Profiling* / methods
Humans
Protein Interaction Maps / genetics
Vascular Calcification* / genetics

Substances

Biomarkers, Tumor

Grants and funding

This work was supported by the National Natural Science Foundation of China (No: 81960091). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.