Celastrol ameliorates atherosclerosis by inhibiting TLR4/STAT3/NLRP3-mediated macrophage pyroptosis

Atherosclerosis is a chronic inflammatory disease characterized by plaque formation. Macrophage pyroptosis critically contributes to the formation and expansion of necrotic core as well as the instability of atherosclerotic plaque. Celastrol is a pentacyclic triterpene derived from the root of Tripterygium wilfordii Hook F, which has powerful anti-inflammatory properties. This study aimed to explore the effect and mechanism of celastrol in preventing atherosclerosis by regulating macrophage pyroptosis.

An atherosclerotic mouse model in vivo and a macrophage pyroptosis model induced by ox-LDL in vitro were established successfully. The cell viability was examined by CCK-8. The role of pyroptosis was detected by transmission electron microscopy, LDH assay, and PI staining. HE staining assay was applied to evaluate the morphology of vascular tissues. The degree of lipid accumulation in plaques was measured using Oil Red O staining. The collagen content of plaques was detected using Masson staining. Western blot analysis, RT-qPCR, immunofluorescence staining, and immunohistochemistry were used to determine the levels of key proteins related to pyroptosis in macrophages. The underlying target of celastrol was investigated by CETSA and SPR assays.

Ox-LDL effectively activated macrophage pyroptosis, which was markedly inhibited by celastrol in vitro. Consistently, celastrol suppressed atherosclerotic plaque progression and instability in apoE^-/- mice by reducing macrophage pyroptosis in vivo. Mechanistically, celastrol could reduce the phosphorylation and nuclear translocation of STAT3 by targeting and inhibiting TLR4, thereby reducing the activation of NLRP3 inflammasomes and preventing macrophage pyroptosis.

The present study demonstrates for the first time that celastrol prevents macrophage pyroptosis by inhibiting TLR4/STAT3/NLRP3 signaling pathway, ultimately alleviating atherosclerosis. These findings indicate that celastrol can serve as a potential therapeutic drug for atherosclerosis.