A simple method for the assessment of macrophage scavenger receptor-ligand interaction: adherence of erythrocytes coated with oxidized low density lipoprotein and modified albumin to macrophages

Abstract

A simple method is described for the assessment of the binding of macrophage scavenger receptor and its ligands such as oxidized low density lipoprotein (ox-LDL) and maleylated bovine serum albumin (m-BSA). In this method, the binding of ox-LDL or m-BSA to macrophages is observed as the adherence of erythrocytes precoated with ox-LDL or m-BSA under phase-contrast microscopy. Erythrocytes were coated with native LDL or ox-LDL simply by incubating mouse erythrocytes with LDL. The ox-LDL-coated erythrocytes attached to the monolayer of mouse peritoneal macrophages without phagocytosis at 37 degrees C, while native LDL-coated erythrocytes did not. The extent of the adherence of the ox-LDL-coated erythrocytes to the macrophages was conveniently expressed as the proportion of the macrophages binding the coated erythrocytes. An optimal concentration of ox-LDL for erythrocyte coating giving maximum erythrocyte adherence to macrophages was 10 micrograms/ml as protein at an erythrocyte concentration of 2%. The binding of the ox-LDL-coated erythrocytes to the macrophages could be inhibited by ligands for scavenger receptor, and reflected the extent of LDL oxidation. Mouse erythrocytes were successfully coated with m-BSA using chromium(III) ion as adsorbent. The m-BSA-coated erythrocytes attached to the macrophages, while native BSA-coated erythrocytes did not. The adherence was inhibitable with ligands for scavenger receptor, and reflected the extent of maleylation. The method would be useful particularly for measurement of the ligand-binding activity of the receptor, and the receptor-binding activity of the ligands.