Stigmasterol attenuates atherosclerosis by inhibiting inflammatory signaling and foam cell formation

Foam cells derived from macrophages and smooth muscle cells (SMCs) play a pivotal role in the progression of atherosclerosis. While phytosterols (PS) have demonstrated cholesterol-lowering and anti-inflammatory properties, their impact on foam cells remains elusive. Here, we investigated the effects of PS on foam cell formation, inflammatory responses, and lipid metabolism using both single-cell RNA sequencing (scRNA-seq) and functional assays. scRNA-seq of aortic tissue from <i>ApoE</i> ^-/- mice revealed that PS supplementation reduced proinflammatory macrophages and SMC-derived intermediate cells. Among the PS components, stigmasterol most effectively attenuated foam cell formation by suppressing oxidized low-density lipoprotein (ox-LDL) uptake and enhancing cholesterol efflux. Mechanistically, stigmasterol inhibited the inflammatory CD86⁺ macrophage polarization by activating the Adenosine Monophosphate-Activated Protein Kinase (AMPK) pathway and inhibiting the NF-κB/NLRP3 signaling axis. In SMCs, stigmasterol upregulated ABCA1 and ABCG1 expression, reduced lipid accumulation, and suppressed CD68 expression, thereby limiting trans-differentiation into macrophage-like foam cells. In vivo, stigmasterol reduced plaque burden, promoted anti-inflammatory macrophage polarization, and inhibited SMC-to-macrophage transition in <i>ApoE</i> ^-/- mice. Collectively, these findings uncover a previously underexplored role of stigmasterol in modulating foam cell diversity and inflammation, providing mechanistic insight into the vascular protective effects of dietary PS.