Macrophage membrane-functionalized biomimetic Yiqi Huoxue formula nanoparticles improve atherosclerosis by regulating smooth muscle cell phenotypic transition via the KLF4/NF-κB pathway

Abstract

This study aimed to analyze the active ingredients of the compound preparation of Yiqi Huoxue (YQHX) and evaluate the therapeutic effect of its nanoparticles (MM/YQHXF-NPs) on atherosclerosis (AS). First, the active ingredient in the YQHX formulation was identified by LC–MS analysis. Subsequently, transmission electron microscopy (TEM) tests particle size. Mapping tests nanoparticle surface elements. Dynamic light scattering (DLS) tests nanoparticle size and distribution. ZETA tests nanoparticle surface potential. HPLC tests drug release. The results showed that these nanoparticles were spherical, approximately 100 nm in size, and had good dispersion. The P element content of MM/YQHXF-NPs increased after cell membrane coating, and their hydrodynamic size also increased accordingly, but the Polymer dispersity index (PDI) value was low, indicating good monodispersity. In addition, the nanoparticle surface had a weak negative charge, the encapsulation efficiency of YQHXF was 59.4%, and the drug loading rate was 5.61%. In cell-based experiments, MM/YQHXF-NPs showed no cytotoxicity towards A7r5 cells at a concentration of 150 μg/mL. The study found that ox-LDL-induced A7r5 cell-to-foam cell transformation was significantly inhibited. Oil Red O staining revealed that MM/YQHXF-NPs reduced lipid accumulation. In addition, YQHXF and its active component, salvianolic acid B, can inhibit the foam cell formation of A7r5 cells. Furthermore, MM/YQHXF-NPs modulated the phenotype of smooth muscle cells, inhibiting the expression of genes such as Myh9, Icam-1, Vcam-1, Tnfrsf11b, Cd68, Lgals3, and Abca1, while promoting the expression of Myh11 and Smtn. Mechanistic studies revealed that MM/YQHXF-NPs exerted their effects by inhibiting the Krüppel-like factor 4 (KLF4) and NF-κB signaling pathways. In a high-fat diet, ApoE−/− mice model of AS, MM/YQHXF-NPs demonstrated significant therapeutic efficacy. H&E and Oil Red O staining revealed that MM/YQHXF-NPs mitigated pathological changes, reduced plaque size, and lowered serum TC, TG, LDL, and HDL levels. They also stabilized atherosclerotic plaques by increasing fiber area and promoting SM22α and SM-MHC expression. Consistent with the results from cell-based experiments, MM/YQHXF-NPs effectively inhibited the transformation of arterial smooth muscle cells (SMCs) into foam cells in vivo and suppressed the activation of KLF4 and NF-κB signaling pathways. In summary, MM/YQHXF-NPs can effectively prevent the transformation of SMCs into foam cells by inhibiting the KLF4 and NF-κB signaling pathways, thereby alleviating AS. These results provide a theoretical basis for MM/YQHXF-NPs as a potential therapeutic drug for AS.