Macrophage uptake of oxidized LDL inhibits lysosomal sphingomyelinase, thus causing the accumulation of unesterified cholesterol-sphingomyelin-rich particles in the lysosomes. A possible role for 7-Ketocholesterol.
Macrophage uptake of oxidatively modified LDL (Ox-LDL), unlike the uptake of acetylated LDL (Ac-LDL), resulted in lysosomal accumulation of unesterified cholesterol (UC). As sphingomyelin (SM) binds UC with high affinity, we considered whether lysosomes also accumulate Ox-LDL-derived SM, and if such a phenomenon could be involved in the lysosomal trapping of Ox-LDL-derived UC. Incubation of J-774 A.1 macrophages with Ox-LDL increased the lysosomal accumulations of UC by 75% and SM by 63% compared with the effect of Ac-LDL. The addition of chlorpromazine, an inhibitor of lysosomal sphingomyelinase (SMase), to macrophages that were incubated with [3H]cholesteryl ester-labeled Ac-LDL also led to lysosomal accumulation of both SM and UC. 7-Ketocholesterol (7-KC), the major oxysterol in Ox-LDL, inhibited lysosomal SMase in a cell-free system. The addition of 7-KC to cells in the presence of [3H]choline- or [3H]cholesteryl ester-labeled Ac-LDL led to macrophage accumulation of SM or UC, respectively. Niemann-Pick type C disease (NP-C) is an inherited cholesterol-storage disease in which lysosomal SMase activity is attenuated after uptake of LDL. Incubation of monocyte-derived macrophages from two NP-C patients with Ac-LDL or Ox-LDL resulted in an accumulation of UC in the lysosomes, whereas normal monocyte-derived macrophages accumulate UC in their lysosomes after incubation with Ox-LDL but not Ac-LDL. These results suggest that inhibition of lysosomal SMase in NP-C cells or by 7-KC is required for lysosomal accumulation of UC. Analysis of the macrophage lysosomal extract (following cell incubation with Ox-LDL) by density-gradient ultracentrifugation and gel-filtration chromatography revealed the presence of a particle consisting of UC, SM, 7-KC, and apoB-100. We conclude that 7-KC in Ox-LDL can inhibit lysosomal SMase, thus leading to the accumulation of SM, which binds UC avidly and inhibits its further cellular processing out of the lysosome. As UC-SM particles of lysosomal origin exist in the atherosclerotic lesion, the formation of such particles may result from an impaired processing of Ox-LDL by arterial wall macrophages during early atherogenesis.
Maor
I
I
Lipid Research Laboratory, Rambam Medical Center, Haifa, Israel.
Mandel
H
H
Aviram
M
M
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002746:Chlorpromazine / Q000494:pharmacology; D002784:Cholesterol / Q000378:metabolism; D002788:Cholesterol Esters / Q000378:metabolism; D006801:Humans; D007653:Ketocholesterols / Q000494:pharmacology*; D008077:Lipoproteins, LDL / Q000378:metabolism*; D008247:Lysosomes / Q000201:enzymology*; D008264:Macrophages / Q000378:metabolism* / Q000648:ultrastructure; D051379:Mice; D009542:Niemann-Pick Diseases / Q000378:metabolism; D010084:Oxidation-Reduction; D013108:Sphingomyelin Phosphodiesterase / Q000037:antagonists & inhibitors*; D013109:Sphingomyelins / Q000378:metabolism*
D002788:Cholesterol Esters; D007653:Ketocholesterols; D008077:Lipoproteins, LDL; D013109:Sphingomyelins; C032629:acetyl-LDL; D002784:Cholesterol; D013108:Sphingomyelin Phosphodiesterase; C003001:7-ketocholesterol; D002746:Chlorpromazine