HUMAN LOW-DENSITY-LIPOPROTEIN AS A TARGET OF HYPOCHLORITE GENERATED BY MYELOPEROXIDASE

The aim of this study was to further clarify which parr of human low d ensity lipoprotein (LDL) is attacked by the MPO/H2O2/Cl--system and wh ich reactive oxygen species is responsible for the attack. Therefore t he influence of this system on the modification of the lipid and prote in moiety of LDL was studied in vitro. Using the monochlorodimedone as say it was found that HOCl is produced in micromolar quantities in the absence of LDL and is rapidly consumed by LDL in a concentration depe ndent manner. The consumption of HOCl was reflected in the formation o f HOCl-specific epitopes on apo B-100 as determined by an antibody rai sed against HOCl-modified LDL. The absorbency at 234 nm was applied to measure continuously the extent of modification of LDL, The general k inetic pattern of the absorbency measurement consisted of a lag phase where no LDL modification was observed, followed by a rapid increase o f absorbency and a plateau phase. Finally the absorbency decreased due to LDL precipitation. Time dependent absorption spectra indicated tha t this kinetic pattern is mainly caused by light scattering due to par ticle aggregation rather than by a specific absorption at 234 nm due t o conjugated diene formation. In agreement with this finding a low rat e of thiobarbituric acid reactive substances (TBArS) formation was obs erved after a lag phase. The aggregation of LDL occurs most likely by modification of apo B-100, which was determined fluorimetrically in te rms of LDL-tryptophan destruction in presence of the MPO/H2O2/Cl--syst em. The kinetic course of tryptophan fluorescence generally consisted of a rapid decrease leveling off into a low plateau phase. Gas chromat ographic determinations of linoleic acid in LDL in presence of the MPO system showed that this polyunsaturated fatty acid (PUFA) is easily a ttacked by HOCl. Consistent with this finding NMR spectra of HOCl modi fied LDL indicated a complete disappearance of bis-allylic methylene g roups. Since lipid peroxidation products only partially account for th is loss of PUFAs, other reactions of HOCl with unsaturated lipids-prob ably chlorohydrin formation-must be involved. Summarizing, although th e rate of lipid peroxidation is low, both the lipid and the protein mo iety of LDL are readily modified by the MPO system. It appears that th e immediate consequence of apo B-100 modification is its aggregation. It is concluded that MPO, which has been detected in atherosclerotic l esions, is able to contribute to the modification of LDL into a form r ecognizable for uncontrolled uptake by macrophages. (C) 1998 Elsevier Science Inc.