PHARMACOLOGICAL, ATP-BINDING CASSETTE TRANSPORTER A1 (ABCA1)-DEPENDENT MEMBRANE FREE CHOLESTEROL POOL REDUCTION LEADS TO ATHEROPROTECTIVE MODULATION OF MACROPHAGE FUNCTIONS

Introduction. Free cholesterol (FC) accumulates in macrophage-derived lipid-laden foam cells contributing to the progression of atherosclerotic plaque. Excess FC is transported to the plasma membrane where it induces a number of deleterious cellular responses including cell death. We have previously demonstrated that this accumulation leads to ruffles formation and impairs macrophage migration in an ABCA1-dependent manner. Probucol, an ABCA1 specific inhibitor, prevented FC deleterious effects. Aim. To investigate whether pharmacological modulation of the ABCA1-dependent membrane FC pool impacts on macrophage proatherogenic functions. Methods. Cells used were wild type (WT) and ABCA1 knock-out (KO) mouse peritoneal macrophages (MPMs). Pharmacological modulation of ABCA1 was evaluated as cholesterol efflux to apoA-I. Plasma membrane cholesterol was evaluated as oxidase-accessible pool. MCP-1 levels were quantified by standard ELISA assay. Results. Membrane cholesterol content positively correlates with ABCA1 gene expression in WT, heterozygous and homozygous ABCA1-KO MPMs. We have identified three structural probucol analogues, AGI-1067, compound A and compound B, all three more active than probucol in inhibiting ABCA1 activity (probucol IC50 =1.80µM and AGI1067, compound A, B IC50 <0.5µM). The two compounds specific inhibitors of ABCA1, compound A and B, inhibited macrophage release of MCP-1 with a higher efficiency than probucol (-32.2%, -33.6% and -22.8%, respectively) and a similar effect was obtained with AGI-1067, an inhibitor of both ABCA1 and ABCG1 (-38.8%). A similar inhibitory effect on ABCA1 activity and on FC-induced MCP-1 release and cytotoxicity in macrophages, was observed also with a non related structural compound berberine. Conclusions. Our results suggest that specific modulation of ABCA1-dependent FC pool may play a role in regulating macrophage functions involved in atherogenesis.