Compound Danshen decoction identifies its mechanism of antagonizing atherosclerosis by network pharmacology and transcriptomics.

Abstract

Compound Danshen Decoction (CPDD), a widely used traditional Chinese medicine (TCM), is effective in treating atherosclerotic diseases. It significantly improves endothelial cell function and regulates inflammatory responses; however, its pharmacological mechanism remains unclear. Atherosclerosis is a common cardiovascular disease, and CPDD is often used clinically for its treatment. In this study, modern techniques such as transcriptomics and network pharmacology were applied to further explore the mechanism of action of CPDD. Quadrupole-Orbitrap LC-MS was used to identify the active chemical components of CPDD. The CCK-8 assay and phalloidin staining were employed to screen the active components of CPDD. Target genes related to inflammatory response in atherosclerosis were identified through database mining. By integrating transcriptomics and the WGCNA algorithm, the targets of the effective components Rg, R1, and PCAD in antagonizing endothelial cell injury were identified. The mechanism by which Rg-R1-PCAD (RRP) antagonizes atherosclerotic endothelial inflammation was further investigated using an ApoEmouse model. Oil red O staining was used to observe pathological changes in the vascular wall, such as lipid deposition, intimal thickening, and plaque formation. Hematoxylin-eosin (HE) staining, Masson staining, and picrosirius red staining were applied to examine pathological features, including plaque area and fibrous cap composition. Finally, the mRNA and protein expression levels of PIK3R1, Akt, and eNOS were detected using qRT-PCR and Western blot. We identified 23 chemical components in CPDD using the Quadrupole-Orbitrap LC-MS technique. Through database and Literature screening, we identified 9 active components and 531 potential targets of CPDD, along with 1037 inflammatory targets related to atherosclerosis. By integrating 18 potential targets from the network pharmacology screening with 14 differentially expressed genes from transcriptome analysis, we performed pathway enrichment analysis and found that the core targets may include HSP90AA1, PIK3R1, and HSP90B1. Based on the results of the CCK-8 assay and phalloidin staining, we ultimately selected RRP as the active ingredient combination of CPDD for further investigation. In vivo experiments showed that RRP reduced lipid plaque formation and plaque area, increased fiber content, improved aortic endothelial dysfunction, and enhanced endothelium-dependent relaxation of mouse aortic rings in response to acetylcholine (ACh). Western blot and qRT-PCR analyses revealed that, after intervention with RRP, the mRNA and protein expression of PIK3R1, Akt, and eNOS increased in the mouse aorta, while the expression of HSP90AA1 and HSP90B1 decreased. These results suggest that RRP acts through the regulation of the PI3K/AKT signaling pathway to treat atherosclerotic endothelial inflammation. The active ingredients of CPDD and RRP promote the expression of PIK3R1, Akt, and eNOS, and inhibit the expression of HSP90AA1 and HSP90B1, thereby antagonizing the PI3K/Akt signaling pathway and reducing inflammation in atherosclerotic endothelial cells.