Unveiling macrophage-centric immunomodulatory mechanisms of radix salviae miltiorrhizae in abdominal aortic aneurysm through integrated single-cell transcriptomic, network pharmacological, and molecular docking analyses

Abstract

Background

Abdominal aortic aneurysm (AAA) represents a fatal vascular disease with prevalence rates of 4–8 % among individuals over 65 years, and rupture mortality rates reaching 60–80 %. With accelerating population aging, AAA incidence continues rising, creating urgent demands for novel therapeutic strategies. Current treatments including surgical intervention and endovascular therapy, while effective, demonstrate significant limitations, and the pathogenic mechanisms remain unclear. The inflammatory microenvironment and macrophages play crucial roles in AAA, yet their immune mechanisms remain controversial. The advancement of single-cell sequencing technology provides new opportunities for elucidating AAA immune mechanisms. This study employed single-cell RNA sequencing (scRNA-seq) and network pharmacology to characterize immune cell features and identify therapeutic targets, exploring the immunomodulatory effects of Radix Salviae miltiorrhizae (RS), aiming to reveal AAA pathogenic mechanisms and provide novel precision treatment strategies.

Method

Using scRNA-seq data from healthy and AAA mouse models in the GEO database (GSE239620), we performed preprocessing and quality control through Seurat, employed Harmony to correct batch effects, and combined clustering, pseudotemporal analysis (Monocle2), and cell communication analysis (CellChat) to reveal the characteristics and interactions of immune cells, particularly macrophages, within the AAA inflammatory microenvironment. Network pharmacology utilized TCMSP and SwissADME to screen active components and targets of RS, with molecular docking (AutoDock Vina) validating their binding to target proteins. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses (clusterProfiler) elucidated biological functions. This study integrates scRNA-seq and network pharmacology to explore AAA immune mechanisms and RS therapeutic potential.

Result

This study investigated the immune microenvironment of AAA via scRNA-seq, elucidating the pivotal role of macrophages in disease progression. UMAP clustering identified five distinct immune cell types, revealing a significant enrichment of macrophages in AAA samples (P < 0.01). Subsequent sub-clustering analysis distinguished five macrophage subtypes (Mac1–Mac5), each characterized by a unique marker gene expression profile (logFC > 0.5, P < 0.05). Pseudotime analysis using Monocle2 mapped their developmental trajectories and identified key gene modules associated with immune responses and inflammatory regulation (q < 0.1). Furthermore, cell communication analysis with CellChat indicated enhanced interactions between macrophages and T/NK cells, underscored by the robust activation of MHC-I signaling (communication probability > 0.5). GO and KEGG analyses showed that associated genes were primarily enriched in the “regulation of innate immune response” (P < 0.01) and the “Lipid and atherosclerosis” pathway (P < 0.05), network pharmacology screened the effective constituents of RS, and molecular docking revealed that luteolin possessed the strongest binding affinity for TNF (-8.6 kcal/mol), suggesting its therapeutic potential.

Conclusion

This study analyzed and predicted the potential pivotal role of macrophages in AAA through scRNA-seq and network pharmacology, suggesting their enhanced interactions with T/NK cells and enrichment in inflammatory pathways. The bioactive compound luteolin demonstrated computationally predicted strong binding affinity for TNF (-8.6 kcal/mol), suggesting its therapeutic potential.