Macrophage degradation of LDL extracted from human aortic plaques: effect of isolation conditions

Several laboratories have recently reported on the structural and functional characteristics of an LDL fraction isolated from atherosclerotic lesions, designated A-LDL. Given the wide variety of tissue sources and isolation conditions that have been employed, we have addressed whether several procedures currently used affect the interaction of A-LDL with macrophages, and, if so, by what mechanisms. We isolated A-LDL from human aortic plaques by ultracentrifugation and gel filtration chromatography. Although some differences in the chromatographic elution profiles on gel filtration were apparent between homogenized and nonhomogenized extracts, A-LDL isolated from the same pool of plaque minces with or without homogenization showed no differences in macrophage degradation or inhibition of this degradation by excess acetyl-LDL. A-LDL isolated from plaques obtained at surgery or at autopsy less than 12 hr after death also showed no major differences in macrophage recognition, suggesting that post-mortem changes were probably not affecting cell recognition. However, A-LDL particles underwent aggregation when subjected to concentration, when stored for periods of 2 weeks or more, or when subjected to vortexing. The aggregated A-LDL was degraded more readily by macrophages than unaggregated A-LDL, and inhibition of degradation of aggregated A-LDL by excess acetyl-LDL was less than for unaggregated A-LDL. Collectively, these studies show that although post-mortem changes and tissue homogenization do not appreciably affect the interaction of A-LDL with macrophages in culture, other isolation and preparation conditions have dramatic effects which could explain some of the diversity of A-LDL metabolism reported in the literature.