Reduced uptake of cholesterol esterase-modified low density lipoprotein by macrophages

M Aviram,S Keidar,M Rosenblat,G.J Brook

doi:10.1016/s0021-9258(18)98994-5

Abstract

Changes in low density lipoprotein (LDL) lipid composition were shown to alter its interaction with the LDL receptor, thus affecting its cellular uptake. Upon incubation of LDL with 5 units/ml cholesterol esterase (CEase) for 1 h at 37 degrees C, there was a 33% reduction in lipoprotein cholesteryl ester content, paralleled by an increment in its unesterified cholesterol. CEase-LDL, in comparison to native LDL, was smaller in size, possessed fewer free lysine amino groups (by 14%), and demonstrated reduced binding to heparin (by 83%) and reduced immunoreactivity against monoclonal antibodies directed toward epitopes along the LDL apoB-100. Incubation of CEase-LDL with the J-774 macrophage-like cell line resulted in about a 30% reduction in lipoprotein binding and degradation in comparison to native LDL, and this was associated with a 20% reduction in macrophage cholesterol mass. Similarly, CEase-LDL degradation by mouse peritoneal macrophages, human monocyte-derived macrophages, and human skin fibroblasts was reduced by 20-44% in comparison to native LDL. CEase-LDL uptake by macrophages was mediated via the LDL receptor and not the scavenger receptor. CEase activity toward LDL was demonstrated in plasma and in cells of the arterial wall such as macrophages and endothelial cells. Thus, CEase modification of LDL may take place in vivo, and this phenomenon may have a role in atherosclerosis.

Highlights

From the Lipid Research Unit, Rambam Medical Center, Rappaport Family Institute forResearch in Medical Sciences and Technion Faculty of Medicine, Haifa, Israel
The results of the present study demonstrate that CEasemodified low density lipoprotein (LDL) undergoes compositional and conformational changes with modulation of the apolipoprotein B-100 epitope expressionwhich resulted inreducedcellular upt,ake of the lipoprotein
Incubation of content of unescholesterol esterase (CEase)-LDL with the 5-774 macrophage-like cell line resulted in about a 30%reduction in lipoprotein binding and degradation in comparison tonative LDL,and this was associated witha 20% reduction in macrophage cholesterol mass

Summary

The abbreviations usedare

LDL, low densitylipoprotein(s); MPM, mouse peritoneal macrophages; HUVEC,human umbilical in 5% CO,, 95% air, nonadherentcells were removed by three washes with serum-freemedium. After 5 days in cul- (four times) with PBS,cells were extracted by incubation with 0.1 N ture, the medium was changed to DMEM supplemented with 10% NaOH For1h at room temperature, and the bounraddiolabeled LDL human lipoprotein-deficient serum Cellular content of unescholesterol esterase (CEase) activity [11].Mouse peritoneal macro- terified cholesterol (UC) and cholesteryl ester (CE) mass was deterphages (MPM) and 5-774 A.l macrophage-like cells were harvested mined after lipid extraction (as described above) using a fluorescent by centrifugation a t 250 X g for 5 min and homogenized in 5 enzymatic method [26]. Beled with ["H]cholesteryl ester by incubation of HDL:I containing The columnwas saturated with bovine serum albumin (BSA) and ["H]cholesteryl linoleate with LDL in the presenceof partially puri- equilibrated to 0.05 M NaC1, 2 mM phosphate buffer (pH 7.4).

RESULTS

CEase-LDL 2

Findings

DISCUSSION