Binding and intracellular trafficking of lipoprotein lipase and triacylglycerol-rich lipoproteins by liver cells.
The cellular mechanisms and pathways by which lipoprotein lipase (LPL) enhances the binding and uptake of lipoproteins remains unknown. Confocal and immunoelectron microscopy demonstrated that primary binding of bovine LPL (bLPL) occurs at the microvilli surface of HepG2 cells and hepatocytes. Internalized bLPL was associated with endocytic vesicles and multivesicular bodies. Quantitative immunofluorescence indicated that the presence of bLPL caused a marked increase in the cell-surface binding of DiI-conjugated triacylglycerol-rich lipoproteins (DiI-TRL). Confocal microscopy showed that when DiI-TRL was incubated with bLPL at 4 degrees C, the distributions of bound LPL and DiI-TRL were totally coincident, and covered the apical surface of both HepG2 cells and hepatocytes. When incubated separately, the time-courses of the internalization of fluorescence associated with DiI-TRL and bLPL were different: DiI-TRL was quickly internalized by both HepG2 cells and hepatocytes, and reached a plateau at 30 min, whereas intracellular LPL increased continuously, but more slowly in the same period. In the presence of bLPL, DiI-TRL was internalized progressively by HepG2 and by cultured hepatocytes for up to 1 h and no saturation was reached. At this time the intensity of labeling of bLPL was lower than of DiI-TRL and a higher number of DiI spots did not colocalize with bLPL immunofluorescence, suggesting that the ligands follow a different pathway after internalization. The data suggest that when lipoprotein lipase (LPL) is associated with the lipoproteins it directs them to specific endocytic pathways. A hypothetical model of the intracellular pathways followed by triacylglycerol-rich lipoproteins and LPL after internalization is proposed.
Casaroli-Marano
R P
RP
Department of Cellular Biology, University of Barcelona, Spain.
García
R
R
Vilella
E
E
Olivecrona
G
G
Reina
M
M
Vilaró
S
S
D000818:Animals; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006801:Humans; D008071:Lipoprotein Lipase / Q000378:metabolism*; D008074:Lipoproteins / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D018613:Microscopy, Confocal; D016253:Microscopy, Immunoelectron; D008954:Models, Biological; D011485:Protein Binding; D051381:Rats; D014280:Triglycerides / Q000378:metabolism*
D008074:Lipoproteins; D014280:Triglycerides; D008071:Lipoprotein Lipase