Bilberries exert an anti-atherosclerotic effect in lipid-loaded macrophages

We hypothesized that the mechanism responsible for the anti-atherosclerotic action of bilberry extract (BE) is linked to its antioxidant and anti-inflammatory potential, and investigated its direct effect on the regulation of apolipoprotein E (apoE) and cholesteryl ester transfer protein (CETP) secretion from lipid-loaded macrophages. Human THP-1 macrophages were loaded with lipids by incubation with human copper-oxidized LDL (oxLDL) and then exposed to different concentrations of BE (1-5 A mu g mL(-1)) obtained from bilberries (mechanically homogenized and solubilized in ethanol). Cellular and secreted proteins, the phosphorylation level of NF-kappa B and protein kinase A (PKA) were quantified by Western blot and gene expression was evaluated by Real-time PCR. The results showed that BE induced in lipid-loaded macrophages has: (i) an antioxidant effect by reducing the expression of NADPH oxidase subunits, p22(phox), p47(phox) and NOX4, (ii) an anti-inflammatory effect by diminishing the secretion of CRP, MCP-1 and IL-1 beta and (iii) cholesterol efflux by increasing the secretion of apoE and CETP and by reducing cellular cholesterol content. BE exerted these effects by inhibition of NF-kappa B and activation of PKA signaling pathways. Our study supports BE therapeutic administration to decrease oxidative and inflammatory stress by a molecular mechanism regulated by NF-kappa B and PKA signaling pathways in lipid-loaded macrophages.