Platelet membrane-modified exosomes targeting plaques to activate autophagy in vascular smooth muscle cells for atherosclerotic therapy.

Abstract

Atherosclerosis is one of the leading causes of ischemic cardiovascular disease worldwide. Recent studies indicated that vascular smooth muscle cells (VSMCs) play an indispensable role in the progression of atherosclerosis. Exosomes derived from mesenchymal stem cells (MSCs) have demonstrated promising clinical applications in the treatment of atherosclerosis. However, there are still challenges and limitations persist in targeted therapy. This study aims to develop a bionic nano-delivery system by fusing platelet membranes with exosomes (MSC-Exo^P) and explore the anti-atherosclerosis effect of MSC-Exo^P by improving the targeting efficiency and participating in regulating the pathophysiological processes associated with VSMCs. The morphology, particle size, stability, and fusion efficiency of MSC-Exo^P were assessed using transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), immunofluorescence staining, and Western blotting, respectively. MSC-Exo^P was administered intravenously into ApoE^-/- mice via the tail vein. In vivo, immunofluorescence staining was used to assess the targeting efficacy of MSC-Exo^P. The ORO staining, H&E staining, Masson staining, aortic root immunofluorescence staining, and Western blot were utilized to evaluate the VSMC autophagy and anti-atherosclerosis effects of MSC-Exo^P. In vitro, the autophagy activation of MSC-Exo^P on VSMCs was further assessed by immunofluorescence staining and Western blotting. The effects of MSC-Exo^P on VSMCs proliferation, migration, and foam cell formation were detected by EdU experiment, Transwell experiment, wound healing experiment, ORO staining, and BODIPY staining. The TEM revealed that MSC-Exo^P retained a ring nanostructure, which was similar to MSC-Exo in morphology. NTA analysis indicated the MSC-Exo^P exhibited a slight increase after cell membrane fusion. Besides, the stability analysis of exosomes and MSC-Exo^P resulted in no significant changes in particle size. Western blot analysis confirmed that MSC-Exo^P simultaneously expressed platelet-specific markers (GPVI, GPIbα, CD62P) and exosome-specific markers (CD81, TSG101, and Alix). In ApoE^-/- mice, the immunofluorescence of aorta and its roots was significantly enhanced after injection of DiI-labeled MSC-Exo^P, indicating enhanced targeting of MSC-Exo to atherosclerotic plaques by platelets. In vivo experiments demonstrated that MSC-Exo^P could significantly suppress the progression of atherosclerosis and reduce the area of atherosclerotic plaques by reducing lipid deposition and necrotic nucleus area and increasing collagen content. In vitro experiments further revealed that the uptake of MSC-Exo^P by foam cells significantly increased, and their proliferation, migration, and foam formation were inhibited by autophagy activation. This study demonstrated successful fusion of platelet membranes with exosomes derived from MSCs. MSC-Exo^P could significantly improve the targeting efficiency of atherosclerosis and play an anti-atherosclerosis effect by activating VSMC autophagy.