Abstract
BACKGROUND: Impaired excretion of lipid deposits within vascular smooth muscle cell-derived foam cells (VSMC-FCs) contributes to the ongoing expansion of the plaque necrotic core. This study aims to explore the effects and underlying mechanisms of exosomes secreted by M2 macrophage (M2-exos) on lipid metabolism of VSMC-FCs and plaque stability. METHODS: First, immunofluorescence was used to detect the expression levels of CD45 (a recognized differentially-expressed molecule of myeloid and VSMC-FCs) and the key proteins of cholesterol efflux pathway, ABCA1 and ABCG1, in human early and late plaques. Next, an in vitro foam cell model was used to assess the effect and mechanism of M2-exos on lipid metabolism in vascular smooth muscle cells by western blot, Oil red O staining and cell total cholesterol assays. RNA-seq and quantitative real-time PCR were employed to characterize the miRNA profiles within M2-exos. The dual-luciferase reporting system and gene silencing approaches were utilized to assess the regulatory effect of candidate miRNA on target genes and signaling pathways. Subsequently, the effect of M2-exos on plaque progression and stability in ApoE(-/-) mice was evaluated using Oil Red O, H&E, Masson's trichrome, Movat's Pentachrome, and immunohistochemistry. RESULTS: Immunofluorescence revealed that compared to early plaques, VSMC-FCs (CD45(-)) were significantly increased in late plaques, and the expression levels of ABCG1 and ABCA1 were remarkably reduced compared to those in leukocyte-derived foam cells (CD45(+)). Purified M2-exos treatment significantly promoted the cholesterol efflux of VSMC-FCs in vitro. In high-fat-fed ApoE(-/-) mice, M2-exos significantly reduced the VSMC-FCs, delayed plaque progression, decreased the necrotic core area, and enhanced plaque stability. MiRNA profiling and analysis of signaling pathways identified miR-7683-3p as a key component in M2-exos, which modulated lipid metabolism in SMC-FCs lipid metabolism through the PPARγ-LXRα-ABCG1 pathway. Dual-luciferase reporting assay confirmed that miR-7683-3p could specifically bind to the promoter region of homeobox genes A1(HOXA1), an inhibitory factor of the PPARγ-LXRα-ABCG1 pathway. CONCLUSION: M2-exos exerted an obvious atherosclerotic protective effect, and the underlying mechanism was closely related to MiR-7683-3p, which targeted the 3'UTR of HOXA1 mRNA and activated the PPARγ-LXRα-ABCG1 mediated cholesterol efflux in VSMC-FCs.