Polarized secretion of newly synthesized lipoproteins by the Caco-2 human intestinal cell line.
Lipoprotein secretion by Caco-2 cells, a human intestinal cell line, was studied in cells grown on inserts containing a Millipore filter (0.45 micron), separating secretory products from the apical and basolateral membranes into separate chambers. Under these conditions, as observed by electron microscopy, the cells formed a monolayer of columnar epithelial cells with microvilli on the apical surface and tight junctions between cells. The electrical resistances of the cell monolayers were 250-500 ohms/cm2. Both 14C-labeled lipids and 35S-labeled proteins were used to assess lipoprotein secretion. After a 24-hr incubation with [14C]oleic acid, 60-80% of the secreted triglyceride (TG) was in the basolateral chamber; 40% of the TG was present in the d less than 1.006 g/ml (chylomicron + VLDL) fraction and 50% in the 1.006 less than d less than 1.063 g/ml (LDL) fraction. After a 4-hr incubation with [35S]methionine, apolipoproteins were found to be major secretory products with 75-100% secreted to the basolateral chamber. Apolipoproteins B-100, B-48, E, A-I, A-IV, and C-III were identified by immunoprecipitation. The d less than 1.006 g/ml fraction was found to contain all of the major apolipoproteins, while the LDL fraction contained primarily apoB-100 and apoE; the HDL (1.063 less than d less than 1.21 g/ml) fraction principally contained apoA-I and apoA-IV. Mn-heparin precipitated all of the [35S]methionine-labeled apoB-100 and B-48 and a majority of the other apolipoproteins, and 80% of the [14C]oleic acid-labeled triglyceride, but only 15% of the phospholipid, demonstrating that Caco-2 cells secrete triglyceride-rich lipoproteins containing apoB. Secretion of lipoproteins was dependent on the lipid content of the medium; prior incubation with lipoprotein-depleted serum specifically reduced the secretion of lipoproteins, while addition of both LDL and oleic acid to the medium maintained the level of apoB-100, B-48, and A-IV secretion to that observed in the control cultures.
Department of Medicine, New York University School of Medicine, NY 10016.
M G
MG
Traber
H J
HJ
Kayden
M J
MJ
Rindler
D001059:Apoproteins / Q000096:biosynthesis; D002460:Cell Line; D004553:Electric Conductivity; D004591:Electrophoresis, Polyacrylamide Gel; D006493:Heparin / Q000494:pharmacology; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D008055:Lipids / Q000096:biosynthesis; D008074:Lipoproteins / Q000096:biosynthesis*; D008715:Methionine / Q000378:metabolism; D008854:Microscopy, Electron; D009829:Oleic Acids / Q000378:metabolism; D011233:Precipitin Tests; D013462:Sulfur Radioisotopes; D014461:Ultracentrifugation
D001059:Apoproteins; D008055:Lipids; D008074:Lipoproteins; D009829:Oleic Acids; D013462:Sulfur Radioisotopes; D006493:Heparin; D008715:Methionine