Oxidation of high-density lipoprotein HDL3 leads to exposure of apo-AI and apo-AII epitopes and to formation of aldehyde protein adducts, and influences binding of oxidized low-density lipoprotein to type I and type III collagen in vitro

Abstract

The changes in the immunological properties of apolipoprotein AI (apo-AI) and AII (apo-AII) during the oxidation of the high-density lipoprotein HDL3 and its influence on the binding of heavily oxidized low-density lipoprotein (LDL) to type I and III collagen were investigated. Oxidation of HDL3 or Eu3+-labelled HDL3 was performed with CuSO4, varying the time of oxidation. Oxidation of HDL3 resulted in an increase in lipid hydroperoxides and enhanced the negative charge of this lipoprotein. Immunological studies with a solid-phase sandwich immunoassay revealed a strong increase in binding of Eu3+-labelled HDL3 to polyclonal antibodies against apo-AI and apo-AII within the first 4 h of oxidation. Neo-epitopes were also formed by interaction of the apolipoproteins with degradation products from the lipid peroxidation of polyunsaturated fatty acids, as evidenced by an immunoreaction of oxidized Eu3+-labelled HDL3 with antibodies to 4-hydroxynonenal (4-HNE)- and malondialdehyde (MDA)-protein adducts. Western blot analysis of oxidized HDL3 samples showed, as well as apo-AI and apo-AII bands, larger aggregated apolipoproteins, occurring after 0.5-2.5 h of oxidation. These aggregates were recognized by antibodies to apo-AI and apo-AII as well as by antibodies to 4-HNE- and MDA-protein adducts. Furthermore the original apo-AI monomers and apo-AII dimers decreased during the oxidation. The ability of native and oxidized HDL3 to prevent the binding of Eu3+-labelled 24 h-oxidized LDL to collagen on microtitration plates was estimated. Interestingly, 2 h-oxidized HDL3 competed most with the binding of 24 h-oxidized LDL on collagen type I and type III, followed by native HDL3. However, 24 h-oxidized HDL3 was a weaker competitor. Thus oxidative modification of HDL3 strongly alters the immunological properties of this lipoprotein and its binding affinity for collagen.

Effects of Protein Oxidation on the Structure and Stability of Model Discoidal High-Density Lipoproteins

“…located in the lipid-binding helical faces, which reduce the lipophilicity of these helices. In addition, apolipoprotein crosslinking, which reportedly increases solvent exposure of the α-helices (33,34) and is accompanied by partial α-helical unfolding on the disks (Fig. 4), may also contribute to the reduced lipophilicity.…”

“…In fact, cross-linking of lipid-bound apoA-I reduces its helical content (Fig. 4) and reportedly increases solvent exposure of specific protein α-helices (16,33,34), which may weaken protein-lipid interactions and promote protein dissociation and lipoprotein remodeling and fusion.…”

“…In fact, oxidation of Met 112 and Met 148 to MetO reportedly impairs apoA-I ability to activate LCAT (29); it also leads to partial unfolding and destabilization of the α-helices in lipid-free and in lipidbound apoA-I and apoA-II, as well as to reduced protein affinity for lipid that is attributed to reduced hydrophobicity of the Met-containing apolar helical faces (29,30,32). Oxidative crosslinking of apoA-I and apoA-II in HDL, which occurs via the tyrosyl radicals, reportedly alters apoA-I conformation, leading to increased solvent exposure of the protein α-helices (33,34) and to enhanced ability of HDL to promote cholesterol efflux from fibroblasts (35). Here, we analyze the effects of these oxidative modifications on thermal stability and remodeling of discoidal rHDLs.…”

Oxidized high-density lipoprotein inhibits platelet activation and aggregation via scavenger receptor BI

“…16 HDL is at least as susceptible to oxidation as LDL both in vivo and in vitro due to the depletion of antioxidants and decreased paraoxonase-1 (PON1) activity in HDL during hyperlipidemia. [17][18][19][20][21][22] Evidences for the presence of OxHDL in vivo are abundant. Lipids isolated from HDL and LDL found in atherosclerotic lesions have been reported to be oxidized to a comparable extent, and increases with increasing severity of disease.…”

“…17 Studies of animals and humans have indicated that OxHDL is also detected in plasma. 18,19,23 Scavenger receptor B type I (SR-BI), the principal HDL receptor, is expressed on the surface of various cell types, including hepatocytes and endothelial cells, and contributes to the antiatherogenic properties of HDL. 4,26-33 SR-BI, a multiligand transmembrane protein of the scavenger receptor CD36 superfamily, binds to a variety of ligands, including native and oxidized lipoproteins.…”