Atorvastatin-preconditioned HUMSC attenuate atherosclerosis via exosomes transfer of hsa-miR-454-3p targeting IRF1/NF-kB to modulate macrophage polarization

Background Atherosclerosis (AS), a chronic inflammatory disorder, is a leading contributor to cardiovascular mortality through its role in coronary artery disease and ischemic stroke pathogenesis. Mesenchymal stem cell (MSC), multipotent stromal cells derived from bone marrow, adipose tissue, umbilical cord, and other sources, represent promising therapeutic agents due to their low immunogenicity, differentiation capacity, and immunomodulatory properties. While pharmacological preconditioning enhances MSC therapeutic potential, the efficacy of atorvastatin-preconditioned human umbilical cord MSC (HUMSC-ATV) in AS remains unexplored. Purpose To investigate the therapeutic efficacy and mechanisms of HUMSCATV and their paracrine exosomes in AS. Methods Six-week-old ApoE−/− mice were fed a Western diet for 12 weeks to establish AS (Fig. 1A). Systemic transplantation of HUMSC or HUMSC-ATV was performed in AS mice. Atherosclerotic plaque area was quantified by Oil Red O (ORO) staining. Macrophage polarization (M1/M2 phenotype) was further analyzed via immunofluorescence (IF) (Fig. 1E-F). Exosomes isolated from MSC supernatants via ultracentrifugation were intravenously administered. Plaque burden and macrophage polarization were similarly evaluated by ORO and IF. High-throughput miRNA sequencing and bioinformatics analysis identified candidate miRNAs, with dual-luciferase reporter assays validating target interactions. qPCR, Western blotting (WB), and IF were employed to assess macrophage polarization markers and downstream gene products. In vivo imaging was utilized to analyze HUMSC-ATV-derived exosomes (HUMSC-ATV-Exos) plaque-targeting capability. Results HUMSC-ATV transplantation significantly reduced atherosclerotic plaque area compared to HUMSC (Fig. 1B–D) and promoted macrophage polarization toward the M2 anti-inflammatory phenotype (Fig. 1E–F). HUMSC-ATV-Exos replicated these therapeutic effects. Notably, HUMSC-ATV-Exos demonstrated enhanced plaque retention compared to HUMSC-Exos, mediated by the CXCR4-SDF1 axis (Fig. 2A–C). miRNA sequencing identified hsa-miR-454-3p as a key mediator enriched in HUMSC-ATV-Exos. Mechanistically, miR-454-3p suppressed IRF1 expression and inhibited NF-κB nuclear translocation via the IRF1/NF-κB pathway, thereby reducing pro-inflammatory cytokine production and facilitating M2 polarization (Fig. 2D). Conclusions This study demonstrates that HUMSC-ATV attenuates AS progression through exosome transfer of miR-454-3p, which inhibits the IRF1/NF-κB pathway to exert anti-inflammatory and immunomodulatory effects. The enhanced plaque-targeting capability of HUMSC-ATV-Exos via the CXCR4-SDF1 axis provides a dual therapeutic mechanism. These findings highlight the translational potential of pharmacologically preconditioned MSC-derived exosomes as a novel strategy for alleviate advanced atherosclerotic plaques.