Itaconate alleviates cholesterol burden via ABCA1 stabilization and cholesterol efflux

Background and aims: Itaconate (ITA) is a metabolite produced from the tricarboxylic acid cycle (TCA) that has been shown to regulate atherosclerotic plaque growth and induce stability via immunomodulation. However, lipid metabolism regulation by ITA is currently underexplored in atherosclerosis. Here, we take advantage of plaque-targeting ITA-conjugated nanoparticles (ITA-LNPs) to investigate the effects of ITA on regulating lipid metabolism in foam cells/macrophages in atherosclerosis via ABCA1 stabilization and increased triglyceride metabolism. Methods: Apoe−/− mice were fed a high-cholesterol/high-fat diet (HCHFD) for 12 weeks and injected once weekly with 50 mg/kg ITA-LNP or Ctrl-LNP. Aortas were tested for ITA-LNP biodistribution, followed by quantification of atherosclerotic plaque burden. Bone marrow-derived macrophages (BMDMs) or Raw 264.7 cells were treated with ITA-LNP or Ctrl-LNP in the presence of oxLDL, acLDL, or free cholesterol to investigate ITA’s actions on lipid metabolism and ABCA1 expression under a variety of conditions, including stable gene knockdown. Results: ABCA1 was significantly upregulated with ITA-LNP treatment compared to Ctrl-LNP both in vivo and in vitro at the protein level, but not at the transcriptional level. ITA-LNPs were shown to prevent ABCA1 decay via the HO-1-calpain axis, resulting in significantly increased cholesterol efflux in macrophages. This was further confirmed in Raw 264.7 cells with a stable HO-1 knockdown. Additionally, ITA decreased lipid burden in conjunction with increased expression of Slc25a1 in ITA-LNP-treated BMDMs, suggesting enhanced fatty acid-derived citrate shuttling and increased fatty acid metabolism. Conclusions: ITA-LNPs regulate lipid metabolism in atherosclerosis by inducing triglyceride catabolism and cholesterol efflux. RNA-seq was performed on whole abdominal aortas isolated from Apoe−/− male mice after 12 weeks of high fat diet and once weekly injection with Ctrl- or ITA-LNPs.