Dual-Functional Fluorescent Nanoprobe Tracks Lipid Droplet Accumulation and Superoxide Anion in Early Atherosclerotic Lesions

Abstract As the primary risk factor for cardiovascular events, early diagnosis of atherosclerosis (AS) is essential for reducing their incidence. Abnormal accumulation of lipid droplets (LDs) and excessive production of superoxide anion (O2•–) are key synergistic drivers that promote the early progression of AS. This study developed a dual-functional fluorescent nanoprobe, termed LDN-O, for simultaneous imaging and assessment of changes in LDs and O2•– levels during the onset and progression of AS. LDN-O was fabricated via nanoprecipitation to encapsulate two responsive molecules: LDB-O-ME enables high-fidelity targeted imaging of LDs, while LN-SF-O allows highly selective detection of O2•–. In vitro experiments confirmed that LDN-O exhibits excellent sensitivity and specificity for both targets. Dynamic monitoring with LDN-O revealed that during foam cell formation, the levels of LDs and O2•– were positively correlated with the duration of oxidized low-density lipoprotein (ox-LDL) stimulation, providing direct experimental evidence for understanding the intrinsic drivers driving early pathological progression in AS. Ex vivo imaging of aortas was further performed in mouse models of early-stage AS and in a combined model of AS with steatohepatitis. The results showed that LDs and O2•– levels in aortic tissue were significantly higher in the model groups than in the control group, with an even more pronounced increase in the group with concurrent steatohepatitis. Overall, LDN-O enables visualization of lipid deposition and oxidative stress in early AS, offering a new strategy for its early diagnosis.