Induction of acetyl-LDL receptor activity by phorbol ester in human monocyte cell line THP-1.
Human monocytic cell line THP-1 incubated with as little as 10 ng/ml of phorbol myristate acetate bound and metabolized 1-2 micrograms of Ac-LDL over a 5-h period. In the absence of phorbol treatment, no specific metabolism of Ac-LDL occurred. Optimal levels of receptor were reached after 72 h of exposure. Induction of receptor was dependent on protein and RNA synthesis and was partially reversed upon removal of the phorbol. Induction of receptor required activation of the protein kinase C pathway. Metabolism of Ac-LDL by THP-1 cells at 37 degrees C was saturated at 25 micrograms/ml. Binding at 4 degrees C was saturable with an average Kd of 8.0 x 10(-9) M. Cell population studies by fluorescent activated cell sorting indicated that approximately 87% of the THP-1 population was expressing scavenger receptor activity 96 h after phorbol treatment as compared to 99% for murine macrophage cell line P388D1. Uptake of Ac-LDL by THP-1 resulted in an 11-fold increase in the rate of cholesterol esterification which was saturable at 50 micrograms/ml. Incubation of cells for 48 h with 50 micrograms/ml of Ac-LDL resulted in a 60% increase in free cholesterol and a 10-fold increase in the cholesteryl ester content of the cells. Lipid accumulation in THP-1 cells after Ac-LDL uptake was readily visible by Oil Red-O staining. Solubilization of THP-1 cells, before and after phorbol treatment, followed by ligand blotting with Ac-LDL detected the presence of a 250-kDa protein only in cells treated with phorbol. The protein comigrated with the scavenger receptor derived from mouse macrophage cell line P388D1.
Via
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DP
Department of Medicine, Baylor College of Medicine, Houston, TX 77030.
Pons
L
L
Dennison
D K
DK
Fanslow
A E
AE
Bernini
F
F
D002454:Cell Differentiation; D002460:Cell Line; D002469:Cell Separation; D002784:Cholesterol / Q000378:metabolism; D004951:Esterification; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D006801:Humans; D008970:Molecular Weight; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism*; D014176:Protein Biosynthesis; D011493:Protein Kinase C / Q000378:metabolism; D012313:RNA / Q000096:biosynthesis; D011973:Receptors, LDL / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D013997:Time Factors
D005456:Fluorescent Dyes; D011973:Receptors, LDL; D012313:RNA; D002784:Cholesterol; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate