Fisetin, via CKIP-1/REGγ, limits oxidized LDL-induced lipid accumulation and senescence in RAW264.7 macrophage-derived foam cells

To test the hypothesis that the flavonoid compound, fisetin, protects macrophages from lipid accumulation and senescence through regulation of casein kinase 2-interacting protein-1 (CKIP-1)/REG gamma (11S regulatory particles, 28 kDa proteasome activator, proteasome activator subunit 3) signaling. RAW264.7 macrophage cells were exposed to 100 mu g/ml oxidized low-density lipoprotein (ox-LDL) with or without 20 mu g/ml fisetin for 24 h. Cell viability was detected by CCK-8 after 1 h. Intracellular lipid accumulation was measured using Oil Red O staining. Total cholesterol (TC) and free cholesterol (FC) contents were measured using assay kits, and cell senescence was inferred by beta-gal staining. Protein expression levels of CKIP-1, REG gamma, organic cation transporter 1 (Oct-1), lectin-like oxidized LDL receptor-1 (LOX-1), tumor suppressor protein p53 (p53), cell cycle regulatory protein p21 (p21), and multiple tumor suppressor-1 (p16) were detected by immunofluorescence and confirmed by Western blot. Stimulating RAW264.7 macrophage cells with 100 mu g/ml ox-LDL for 24 h induced the formation of foam cells, increased intracellular lipid accumulation, increased TC and FC content, and promoted cell senescence. Furthermore, cells induced with 100 mu g/ml ox-LDL for 24 h showed decreased CKIP-1 and REG gamma protein, while the expressions of Oct-1, LOX-1, p53, p21 and p16 were increased. In contrast, treatment with 20 mu g/ml fisetin reversed 100 mu g/ml ox-LDL effects to increase cell viability, and decrease beta-gal staining, intracellular lipid levels and TC and FC levels. These beneficial effects were associated with increased CKIP-1 and REG gamma and decreased Oct-1, LOX-1, p53, p21, and p16 protein expression. Results indicated that fisetin limited ox-LDL-mediated lipid accumulation and senescence in RAW264.7 macrophage-derived foam cells. The mechanism underlying these effects may involve regulation of CKIP-1/REG gamma signaling.