AGGREGATION AND FUSION OF MODIFIED LOW-DENSITY-LIPOPROTEIN

In atherogenesis, low density lipoprotein (LDL, diameter 22 nm) accumu lates in the extracellular space of the arterial intima in the form of aggregates of lipid droplets (droplet diameter up to 400 nm). Here we studied the effects of various established in vitro LDL modifications on LDL aggregation and fusion. LDL was subjected to vortexing, oxidat ion by copper ions, proteolysis by alpha-chymotrypsin, lipolysis by sp hingomyelinase, and nonenzymatic glycosylation, and was induced to for m adducts with malondialdehyde or complexes with anti-apoB-100 antibod ies. To assess the amount of enlarged LDL-derived structures formed (d ue to aggregation or fusion), we measured the turbidity of solutions c ontaining modified LDL, and quantified the proportion of modified LDL that I) sedimented at low-speed centrifugation (14000 g), 2) floated a t an increased I-ate at high-speed centrifugation irate zonal flotatio n at 285000 g(max)), 3) were excluded in size-exclusion column chromat ography (exclusion limit 40 MDa), or 4) failed to enter into 0.5% Fast Lane agarose gel during electrophoresis. To detect whether particle fu sion had contributed to the formation of the enlarged LDL-derived stru ctures, particle morphology was examined using negative staining and t hin-section transmission electron microscopy. We found that 1) aggrega tion was induced by the formation of LDL-antibody complexes, malondial dehyde treatment, and glycosylation of LDL; 2) fusion of LDL was induc ed by proteolysis of LDL by alpha-chymotrypsin; and 3) aggregation and fusion of LDL were induced by vortexing, oxidation by copper ions, an d lipolysis by sphingomyelinase of LDL. The various modifications of L DL differed in their ability to induce aggregation and fusion.