Early Assessment of Atherosclerotic Lesions and Vulnerable Plaques in vivo by Targeting Apoptotic Macrophages with AV Nanobubbles
Xiaoju Ma, Jia Wang, Zhelong Li, Xueying Zhou, Xiao Liang, Junyan Wang, Yunyou Duan, Ping Zhao
Abstract
Background: The early detection of atherosclerotic lesions is particularly important for risk prediction of acute cardiovascular events. Macrophages apoptosis was significantly associated with the degree of AS lesions and especially contributed to plaque vulnerability. In this research, we mainly sought to explore the feasibility of a home-made AV-nanobubbles (NB AV ) for visualization of apoptotic macrophages and assessment of atherosclerosis (AS) lesions by contrast-enhanced ultrasound (CEUS) imaging. Methods: NB AV were prepared by “Optimized Thin-Film Hydration” and “Biotin-Avidin-Biotin” methods. Then, the characterization and echogenicity of NB AV were measured and analyzed in vitro. The targeting ability of NB AV to ox-LDL–induced apoptotic macrophages was observed by laser scanning confocal microscope. The ApoE−/− mice mode fed with high fat diet were observed by high-frequency ultrasound, microanatomy and oil red O staining. CEUS imaging in vivo was performed on AS plaques with NB AV and NB Ctrl injection through the tail vein in turn in ApoE−/− mice. After CEUS imaging, the plaques were confirmed and analyzed by histopathological and immunological assessment. Results: The prepared NB AV had a nano-scale size distribution with a low PDI and a negative zeta potential. Moreover, NB AV showed an excellent stability and exhibited a significantly echogenic signal than saline in vitro. In addition, we found that NB AV could target apoptotic macrophages induced by ox-LDL. Compared with NB Ctrl , CEUS imaging of NB AV showed strong and sustained echo enhancement in plaque area of aortic arch in vivo. Further research showed that NB AV sensitive plaques presented more significant pathological changes with several vulnerable plaque features and abundant TUNEL-positive area. Conclusion: NB AV displayed a sensitive indicator to evaluate apoptotic macrophages, indicating a promising CEUS molecular probe for AS lesions and vulnerable plaques identification. Graphical Abstract: Keywords: atherosclerosis, vulnerable plaques, macrophage, apoptosis, AV-nanobubbles