Essential involvement of 12-lipoxygenase in regiospecific andstereospecific oxidation of low density lipoprotein by macrophages.
To establish a role of the 12-lipoxygenase on the generation of oxidized low density lipoprotein (LDL) in macrophages that leads to foam cell formation in atherosclerosis, we overexpressed 12-lipoxygenases in a macrophage-like cell line, J774A.1, that does not show intrinsic enzyme activity. When the 12-lipoxygenase-expressing cells were incubated with 400 microg.mL-1 LDL in Dulbecco's modified Eagle's medium at 37 degrees C for 12 h, LDL oxidation, as determined by thiobarbituric acid reactive substance, was markedly increased compared with the mock-transfected cells. Oxygenated products in the modified LDL were examined by HPLC before and after alkaline hydrolysis. Most of the oxygenated derivatives were of an esterified form, and the major product was identified as 13S-hydroxyoctadeca-9Z,11E-dienoic acid. These results clearly demonstrate that esterified fatty acids in LDL are oxygenated by the 12-lipoxygenases expressed in the J774A.1 cells. Furthermore, the oxidized LDL generated by intracellular 12-lipoxygenases was recognized by a scavenger receptor as assessed by macrophage degradation assay.
Sakashita
T
T
Department of Pharmacology, Kanazawa University School of Medicine, Japan.
Takahashi
Y
Y
Kinoshita
T
T
Yoshimoto
T
T
D000818:Animals; D001092:Arachidonate 12-Lipoxygenase / Q000378:metabolism*; D001161:Arteriosclerosis / Q000378:metabolism; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D004952:Esters / Q000032:analysis; D005227:Fatty Acids / Q000378:metabolism; D005487:Foam Cells / Q000473:pathology; D008041:Linoleic Acids / Q000378:metabolism; D008077:Lipoproteins, LDL / Q000096:biosynthesis*; D008264:Macrophages / Q000378:metabolism*; D008565:Membrane Proteins; D051379:Mice; D008968:Molecular Conformation; D011971:Receptors, Immunologic / Q000378:metabolism; D018110:Receptors, Lipoprotein; D051116:Receptors, Scavenger; D051122:Scavenger Receptors, Class B; D017392:Thiobarbituric Acid Reactive Substances / Q000032:analysis; D013997:Time Factors; D014162:Transfection
D004952:Esters; D005227:Fatty Acids; D008041:Linoleic Acids; D008077:Lipoproteins, LDL; D008565:Membrane Proteins; D011971:Receptors, Immunologic; D018110:Receptors, Lipoprotein; D051116:Receptors, Scavenger; C454739:Scarb1 protein, mouse; D051122:Scavenger Receptors, Class B; D017392:Thiobarbituric Acid Reactive Substances; C099252:oxidized low density lipoprotein; C024348:13-hydroxy-9,11-octadecadienoic acid; D001092:Arachidonate 12-Lipoxygenase