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. Conf ocal and immunoelectron microscopy demonstrated that primary binding o f bovine LPL (bLPL) occurs at the microvilli surface of HepG2 cells an d hepatocytes. Internalized bLPL was associated with endocytic vesicle s and multivesicular bodies. Quantitative immunofluorescence indicated that the presence of bLPL caused a marked increase in the cell-surfac e binding of DiI-conjugated triacylglycerol-rich lipoproteins (DiI-TRL ). Confocal microscopy showed that when DiI-TRL was incubated with bLP L at 4 degrees C, the distributions of bound LPL and DiI-TRL were tota lly coincident, and covered the apical surface of both HepG2 cells and hepatocytes. When incubated separately, the time-courses of the inter nalization of fluorescence associated with DiI-TRL and bLPL were diffe rent: DiI-TRL was quickly internalized by both HepG2 cells and hepatoc ytes, and reached a plateau at 30 min, whereas intracellular LPL incre ased continuously but more slowly in the same period. In the presence of bLPL, DiI-TRL was internalized progressively by HepG2 and by cultur ed hepatocytes for up to 1 h and no saturation was reached. At this ti me the intensity of labeling of bLPL was lower than of DiI-TRL and a h igher number of DiI spots did not colocalize with bLPL immunofluoresce nce, suggesting that the ligands follow a different pathway after inte rnalization. The data suggest that when lipoprotein lipase (LPL) is as sociated with the lipoproteins it directs them to specific endocytic p athways. A hypothetical model of the intracellular pathways followed b y triacylglycerol-rich lipoproteins and LPL after internalization is p roposed.