In vitro reverse cholesterol transport from THP-1-derived macrophage-like cells with synthetic HDL particles consisting of proapolipoprotein A1 or apolipoprotein A1 and phosphatidylcholine

Abstract

The human monocytic leukaemia cell line THP-1 was induced to differentiate to macrophage-like cells by the addition of phorbol myristoyl acetate (PMA). Subsequently, the cells were enriched in cholesterol and these cholesterol laden cells were used to study the capability of reconstituted discoidal complexes (RDCs), consisting of either human apolipoprotein Al (apo Al) or recombinant human proapolipoprotein Al (proapo Al) and phosphatidylcholine (PC), to promote cholesterol efflux. RDCs containing apo Al and proapo Al were both effective in the mobilization of intracellular cholesterol, whether this was measured by intracellular cholesterol mass or by the appearance of radiolabelled cholesterol in the supernatant. Using the radiolabelling technique, the activity was saturable and followed Michaelis-Menten kinetics. For both types of complexes and for native HDL the maximum rate of cholesterol removed was approximately 0.5 nmol h⁻¹ per 10⁶ cells. For RDCs of proapo Al and apo Al and for native HDL the K_m values were 3.7, 2.9 and 64.8 μg ml⁻¹ respectively. A significant in vitro cholesterol efflux could only be achieved with protein-lipid complexes; no significant export was observed with either free proapo Al or multilamellar PC liposomes without apolipoprotein. Both RDCs were found to be more active in the mobilization of intracellular cholesterol than HDL isolated from human plasma. The combined results demonstrate that synthetic complexes consisting either of apo Al or proapo Al and PC are both active in the in vitro reverse transport of cholesterol.

Key Words:

• cholesterol acceptor
• efflux
• egress
• foam cells
• recombinant
• reverse cholesterol transport