Paired samples of human Lipoprotein (a) (Lp(a)) and low density lipoprotein (LDL) were assayed for their platelet-activating factor (PAF) acetylhydrolase activity. Lp(a) displayed markedly enhanced PAF acetylhydrolase activity (approximately 7-fold based on equal particle concentrations) in comparison to LDL isolated from the same individual. Lp(a)-associated acetylhydrolase exhibited properties observed for LDL-associated acetylhydrolase as well as for the purified enzyme; significant inhibition was obtained by treatment with diisopropylfluorophosphate (1 mM, 90%) and phenylmethanesulfonyl fluoride (5 mM, 50%). Furthermore, the hydrolytic activity of both lipoproteins was abolished with paraoxon (6 mM, IC 50 0.9 mM) and with the fluorescent and active site-directed probe 4-hexyl-(6-O-butyl-(4-pyrenyl))-benzoic estersulfonyl fluoride (2) (K I(inact) ‫؍‬ 525 M), a novel irreversible inhibitor of PAF acetylhydrolase. Treatment with 2 and subsequent quantitation of protein-bound fluorescence suggests an increased concentration of enzyme associated to Lp(a) rather than alterations of kinetic constants due to the additional apolipoprotein apolipoprotein (a). Exposure of Lp(a) to Cu 2؉ (20 M, 37°C) was followed by a concomitant decrease of hydrolytic activity. A reduction of the basal activity by 91% was found after 15 h. Whereas immunoprecipitation with anti-apoB antiserum could remove enzymatic activity of Lp(a) regardless of a reductive treatment with dithiothreitol, precipitation with anti-apolipoprotein (a)-antibodies was accompanied by a minor reduction (approximately 30%) of the PAF-hydrolyzing ability. These results suggest that PAF acetylhydrolase exhibits an enhanced association with Lp(a) due to an increased affinity to Lp(a) apolipoprotein B. Lp(a)1 is a lipoprotein from human plasma differing from low density lipoprotein by apo(a) an additional unique glycoprotein (Ref. 1; for review see Ref.2). Apo(a) is linked to apoB, the main protein component of LDL, by a single disulfide bond (3, 4) and exhibits a homology to plasminogen (5). In clinical studies, elevated Lp(a) plasma concentrations have been shown to correlate with the incidence of stroke and coronary heart disease (6 -8), and an accumulation in atherosclerotic lesions has been demonstrated (9), features that provide evidence for elevated Lp(a) levels as a substantial risk factor. Moreover, an overexpression of apo(a) promotes the development of lesions in transgenic animals (10).

Addition of the phospholipids 1-O-hexadecyl-2-arachidonoyl-sn-glycero-3-phosphocholine (PLE) and 1-O-hexadecyl-2-desoxy-2-amino-ar achidonoyl-sn-glycero-3-phosphocholine (PLA) to [~2SIILDL and subsequent Cu2+-induced oxidation result in significant differences in protein modification and uptake by P388D~ macrophage-like cells. PLE-treated LDL is ingested at a 1.27-fold rate compared to PLE-treated LDL and displays enhanced electrophilic mobility. Similar results (1.43-fold enhanced uptake of LDL preloaded with PLE) are obtained when the uptake of phospholipid-enriched oxLDL particles are examined. The preference for ingestion as well as protein modification of both preparations is, however, reversed under experimental conditions allowing diffusion and inactivation of a fraction of the peroxidation products. These findings suggest that LDL-associated PAF-acetylhydrolase can exert a dual role and, to be protective to LDL, require an appropriate microenvironment, capable of binding certain species of oxidatively fragmented lipids.

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