Salvianic acid A ameliorates atherosclerosis through metabolic-dependent anti-EndMT pathway and repression of TGF-β/ALK5 signaling.

Background: Endothelial-to-mesenchymal transition (EndMT) has been identified as a key factor to the initiation and progression of the pathogenesis of atherosclerosis (AS). Salvianic acid A (SAAS) is the primary water-soluble bioactive ingredient found in Salvia miltiorrhiza, is renowned for its therapeutic effects on cardiovascular diseases. However, the efficacy and mechanisms of SAAS in treating EndMT-induced AS remain underexplored.

Purpose: This study aimed to investigate the role SAAS in reversing EndMT process to impede AS development.

Methods: We used a murine model of cholesterol-rich and high-fat diet-induced AS in ApoE mice to evaluate the effect of SAAS on EndMT during AS progression in vivo. The biological effects of SAAS on EndMT-induced HUVEC cells were also detected by transcriptome sequencing (RNA-seq). Mechanistic exploration was carried out using omics data mining and screening, gene knockout experiments, gene expression, protein expression, and localization of key gene expression in animal lesion areas.

Results: We found that SAAS treatment significantly alleviated EndMT injury in the AS mice model and also improved aortic root lesions and dyslipidemia. Furthermore, pre-treatment with SAAS effectively inhibited the EndMT in HUVEC cells, as evidenced by maintained endothelial cell morphology and reduced cell migration ability, as well as elevated CD31 and decreased α-SMA. RNA sequencing data indicated that key differentially expressed genes were mainly enriched in metabolism-related and TGF-β receptor signaling pathways. The metabolic regulator PDK4 and profibrotic TGF-β receptor ALK5 were identified specifically. Subsequently, RT-qPCR and western blot results demonstrated that SAAS notably increased metabolic regulator PDK4 and decreased profibrotic TGF-β receptor ALK5 in EndMT-induced HUVEC cells. Moreover, siRNA-directed PDK4 inhibition resulted in EndMT induction and SAAS mediated the suppression of EndMT in a PDK4-dependent manner. Additionally, SAAS partially reduced the TGF-β receptor ALK5 expression. Furthermore, ApoE AS mice with SAAS treatment displayed downregulation of ALK5 and upregulation of PDK4 with reduced EndMT during AS.

Conclusion: This investigation demonstrated that SAAS improved AS through metabolic-dependent anti-EndMT pathway and repression of profibrotic TGF-β receptor signaling, thereby providing SAAS as a promising therapeutic candidate for managing AS and EndMT-related disorders.