Testing the influence of lipid laden and iron-overloading on ceruloplasmin expression in RAW 264.7 cells

Abstract

To investigate whether the treatment of oxidized low density lipoprotein (ox-LDL) could reduce ceruloplasmin (Cp) expression and then, influence iron efflux in macrophages. RAW264.7 cells were treated by lipopolysaccharides (LPS), ferric ammonium citrate (FAC), deferoxamine (DFO) and ox-LDL. Biochemical analysis and histological assays were performed to detect the iron-related protein including ferritin (Ft), transferrin receptor (TfR), ferroportin 1 (FPN1) and Cp. And the cholesterol enzyme was used to evaluate cells' cholesterol content. Western blot showed that protein level of FPN1, Ft increased and protein level of TfR decreased after the treatment of FAC. The mRNA level (t=-5.995, P=0.019) and the oxidative activity (t=-9.875, P=0.001) of Cp increased significantly after the treatment of LPS+FAC compared to control group. The treatment of ox-LDL could reduce protein level of Cp compared to LPS treated cells and no different protein levels of Ft and Ferroportin was found after the treatment of ox-LDL as shown by Western blot. The protein level, ferroxidase activities (t=6.663, P=0.003) and mRNA levels (t=8.948, P=0.001) of Cp in LPS+ox-LDL+FAC group were reduced than LPS+FAC group. The protein levels of Ft and FPN1 in LPS+ox-LDL+FAC group was significant higher than LPS+FAC group as shown by Western blot. The findings suggest that the expression of Cp is reduced in foam cells and leads to obstructed iron efflux in iron overloaded foam cells.