Fisetin Prevents Oxidized Low-density Lipoprotein–Induced Macrophage Foam Cell Formation

Abstract: Foam cell formation is an important event in atherosclerosis. Fisetin, a bioflavonoid, has been identified to possess anti-inflammatory, antilipidemic, and anticancerous properties; however, its role as a lipid homeostasis regulator in macrophages, specifically in the presence of metabolic stressors such as oxidized low-density lipoprotein (oxLDL) is not well understood. In this study, we have investigated the role of fisetin in preventing oxLDL-induced macrophage foam cell formation. U937-derived macrophages were stimulated with oxLDL with or without fisetin for varied time points, and various parameters were assessed including cell viability by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay; reactive oxygen species (ROS) by dichlorofluorescin diacetate assay; lipid accumulation by Oil Red O staining; and expression of NLR family pyrin domain containing 3 (NLRP3), sterol regulatory element-binding protein (SREBP)-1, and associated downstream proteins 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMGCR) and fatty acid synthase (FAS) by quantitative reverse transcription polymerase chain reaction (RT-qPCR) and immunoblotting. Functionality of FAS enzyme was determined using enzyme activity assay. Docking studies were performed to determine the in silico interaction between NLRP3 and fisetin. The results showed that fisetin up to the dose of 10 µM did not alter cell viability but at the same dose could decrease the accumulation of lipids in macrophages and prevented foam cell formation. Fisetin could also ameliorate and reduce oxLDL-induced upregulation of SREBP-1 and thereby the expression of its downstream lipid synthesis genes HMGCR and FAS and inhibited ROS-induced NLRP3 inflammasome activation. In conclusion, fisetin could inhibit foam cell formation by blocking oxLDL-induced ROS formation and subsequent NLRP3 activation, thereby inhibiting SREBP-1 and its downstream genes including FAS and HMGCR .