Involvement of the macrophage low density lipoprotein receptor-binding domains in the uptake of oxidized low density lipoprotein.
Macrophages, unlike most other cells, possess both low density lipoprotein (LDL) and scavenger receptors. The scavenger receptor has been shown to mediate the uptake of oxidized LDL (ox-LDL), which ultimately leads to cholesterol loading of the macrophages. The present study was undertaken to define epitopes on ox-LDL that are important for lipoprotein binding to macrophages and to ascertain whether ox-LDL can bind to the LDL receptor. Monoclonal antibodies (Mabs) directed against several epitopes along the apolipoprotein B-100 (apo B-100) molecule were used. LDL (300 micrograms/ml) was oxidized by incubation with 10 microM CuSO4 for 24 hours. Ox-LDL, as opposed to acetylated LDL (ac-LDL), reacted with Mabs directed against the LDL receptor-binding domains (Mabs B1B6 and B1B3). Similarly, uptake of ox-LDL but not ac-LDL by a murine J774 macrophage-like cell line was inhibited by as much as 40% after using Mab B1B6. The anti-LDL receptor antibody IgG-C7 also inhibited 125I-ox-LDL uptake by macrophages by 60%. Chromatography on heparin-Sepharose columns of LDL that was partially oxidized for only 3 hours resulted in two fractions: an unbound fraction with characteristics similar to those of ox-LDL and a bound fraction similar to native LDL. Macrophage degradation of the unbound fraction was inhibited by Mab IgG-C7 and Mab B1B6, which are directed toward the LDL receptor and the LDL receptor-binding domains on apo B-100, respectively. When incubated with three types of macrophages, J774 macrophage cells, mouse peritoneal macrophages, and human monocyte-derived macrophages, excess amounts of unlabeled ox-LDL, like native LDL but unlike ac-LDL, substantially suppressed the uptake and degradation of 125I-labeled LDL. Similar studies with fibroblasts, however, revealed that unlabeled LDL but not unlabeled ox-LDL or ac-LDL competed with 125I-LDL for cellular uptake and degradation. Mab directed against epitopes on the amino terminus domain of apo B-100 (C14) demonstrates a similar immunoreactivity with ox-LDL and native LDL but a much lower reactivity with ac-LDL. Mab C14 inhibited macrophage degradation of ox-LDL by 34% but had no inhibitory effect on the uptake of native LDL or ac-LDL. Thus, the ac-LDL and LDL receptor-binding domains as well as a unique epitope on the amino terminus of apo B-100 may be involved in macrophage binding of ox-LDL.(ABSTRACT TRUNCATED AT 400 WORDS)
- Copyright © 1992 by American Heart Association