BACKGROUND: The molecular and genetic mechanisms underlying vascular calcification remain unclear. This study aimed to determine the differences in calcification marker-related gene expression in macrophages. METHODS: The expression profiling datasets GSE104140 and GSE235995 were analysed to identify differentially expressed genes (DEGs) between fibroatheroma with calcification and diffuse intimal thickening. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses, Weighted Gene Co-expression Network Analysis (WGCNA), and Gene Set Enrichment Analysis (GSEA) were performed to assess functional characteristics. Hub genes were identified through a protein-protein interaction (PPI) network and machine learning approaches. Single-cell RNA sequencing data (GSE159677) validated the expression of calcification-related genes in macrophages, while Mendelian randomization analysis explored their potential causal relationship with coronary calcification. Further validation was conducted using enzyme-linked immunosorbent assay (ELISA) on coronary calcification samples and immunohistochemistry in ApoE RESULTS AND CONCLUSIONS: Two key biomarkers, ITGAX and MYD88, were identified as diagnostic indicators of cardiovascular calcification. Both biomarkers were significantly upregulated in calcified samples and were strongly associated with immune processes. Single-cell RNA sequencing confirmed their high expression in multiple immune cell types. Additionally, molecular docking analysis revealed that retinoic acid interacted with both biomarkers, suggesting potential therapeutic relevance. Immunohistochemical and ELISA analyses further validated their elevated expression in calcified samples. These findings provide novel insights into the molecular mechanisms of vascular calcification and highlight potential diagnostic and therapeutic targets.