ATO attenuates atherosclerotic plaque progression and destabilisation in ApoE-/- mice
To investigate the role of ATO in protecting against atherosclerosis, ApoE-/- mice were divided into two groups and fed a high-fat diet for 3 months. One group was treated with ATO (2.5 mg kg-1), and the other group was treated with vehicle (ATO and control groups, respectively; Figure 1A). ATO significantly reduced atherosclerosis development, as demonstrated by Oil Red O staining of both the aortic sinus and en face preparations of the entire aorta (Figure 1B, D, G, J). In addition, the lesion area and the proportion of necrotic cores in plaques significantly reduced after ATO treatment (Figure 1C, E, F), but the necrotic core area decreased slightly (Figure 1I). In addition, ATO increased the collagen content in plaques (Figure 1H, K). We further determined the effects of ATO on plaque composition. ATO reduced macrophage accumulation, as indicated by CD68 immunostaining (Figure 1L, M).
To determine whether the dose we used in this study was safe, we examined the histological features of various tissues and organs (Figure S1A, B). We also examined the levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), urea acid (UA), and creatine Kinase (CK) in ApoE-/- mouse plasma, apoptosis in peripheral lymphoid tissues, and the proportion of peripheral white blood cells (Figure S1D-G, S2). The results showed that the appropriate dose we used (2.5 mg kg-1) did not cause significant serological changes, histopathological changes, or extensive white blood cell apoptosis. Further, there were no significant changes in the body weight of ApoE-/- mice before and after ATO treatment (Figure S1C). These results indicate that the dose of ATO used in vivo (2.5 mg kg-1) attenuated atherosclerotic plaque progression but did not cause significant toxicity in ApoE-/- mice. Furthermore, our previous study has shown how the ATO dose affects the viability of macrophages (Fang et al. , 2021). Based on these results, we chose the dose of ATO as 2.5 mg kg-1in vivo and the doses of 2.5 and 5.0 μM in vitro in this experiment.