Decoding atherosclerosis through lactylation: multi-omics integration with experimental validation

Atherosclerosis (AS) remains a major cause of cardiovascular morbidity and mortality. Lactylation-a recently described post-translational modification linking cellular metabolism to gene regulation-has been implicated in vascular inflammation, yet its roles in AS are not fully defined.

We modelled AS by exposing HUVECs to oxidised LDL and performed data-independent proteomics. Public AS transcriptomes were integrated and batch-corrected; lactylation-related genes (LRGs) were profiled; consensus clustering and WGCNA defined subtypes/modules. Candidate biomarkers were prioritised by intersecting proteomic, differential and network features, then evaluated using an ensemble of 12 machine-learning algorithms with cross-validation and external validation. Immune infiltration (CIBERSORT) and single-cell data characterised immune contexts. Connectivity Map, molecular docking and molecular dynamics (MD) explored therapeutics. Core genes were validated by RT-qPCR, Western blotting and immunofluorescence in Apoe^-/- mice.

Proteomics identified 472 differentially expressed proteins; GEO analyses yielded 2,544 DEGs and WGCNA 2,059 module genes, converging on 25 candidates. The top ensemble (LASSO + GBM) achieved a mean AUC 0.979 across training and external sets, nominating UAP1, NRP1, QPRT and NDST1 as hub genes. These genes associated with immune-cell infiltration and showed prominent single-cell expression in macrophages and smooth muscle cells. RT-qPCR in vivo showed NRP1/NDST1/QPRT upregulated and UAP1 downregulated versus controls (all P < 0.05); WB/IF confirmed higher NRP1 and lower UAP1 protein abundance. CMap analysis suggested several candidate compounds. Among them, rivaroxaban was prioritised for further in silico evaluation because NRP1 emerged as a validated hub gene, and docking and molecular dynamics simulations supported a stable predicted interaction between rivaroxaban and NRP1.

UAP1, NRP1, QPRT and NDST1 represent lactylation-linked biomarkers of AS with diagnostic potential and plausible mechanistic relevance within immune-vascular pathways. Computational screening further prioritised a putative NRP1-rivaroxaban interaction as a hypothesis-generating lead for future experimental validation.