The secretion of apolipoprotein E by human monocyte-derived macrophages.
The secretion of newly synthesized apolipoprotein E (apo E) by human monocyte-derived macrophages (HMD macrophages) was measured in the medium of cells which had been incubated for 24 h with or without either native or acetylated low-density lipoproteins (LDL or AcLDL, respectively), and subsequently with [35S]methionine in the presence of high-density lipoproteins (HDL, 350 micrograms/ml) for 24 h, by isolating the lipoprotein fraction by centrifugation for 48 h at a density adjusted with KBr to 1.21 g/ml (d = 1.21). The d less than 1.21 medium was subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) or precipitation with trichloroacetic acid (TCA). Fluorography of the gels demonstrated that the d less than 1.21 fraction of the medium contained one major labeled band, which migrated with an apparent molecular weight of approximately 35,000. Immunoprecipitation of the d less than 1.21 fraction showed that the labeled band was precipitated by anti-apo E, but not by anti-HDL. As the apo E band appeared to be the only labeled band in the d less than 1.21 fraction, the amount of apo E secreted by the cells was quantitated by scintillation counting of the TCA-precipitable radioactivity in the d less than 1.21 fraction as compared with that in the whole medium. The proportion of secreted apo E to the total secreted protein was similar whether the cells had been in culture for 3 or 16 days, but was increased if the cells had been incubated with LDL or AcLDL. The proportion of apo E of the secreted proteins was always more than 6% and was as much as 16% when the cells were preincubated with lipoproteins, suggesting that the increased cholesterol influx induced apo E secretion.
H J
HJ
Kayden
F
F
Maschio
M G
MG
Traber
D000107:Acetylation; D001057:Apolipoproteins E / Q000378:metabolism*; D002478:Cells, Cultured; D002784:Cholesterol / Q000378:metabolism; D016264:Dextran Sulfate; D003911:Dextrans; D004591:Electrophoresis, Polyacrylamide Gel; D005470:Fluorometry; D006801:Humans; D007163:Immunosorbent Techniques; D007930:Leucine / Q000378:metabolism; D008075:Lipoproteins, HDL / Q000378:metabolism; D008077:Lipoproteins, LDL / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism*; D014461:Ultracentrifugation
D001057:Apolipoproteins E; D003911:Dextrans; D008075:Lipoproteins, HDL; D008077:Lipoproteins, LDL; D016264:Dextran Sulfate; D002784:Cholesterol; D007930:Leucine