Abstract
Lipoprotein lipase (LPL), a key enzyme in lipoprotein triglyceride metabolism, produces a marked increase in the retention and uptake of all classes of lipoproteins by cultured cells. It was previously shown that two different receptors are involved in mediating the LPL effects: heparan sulfate proteoglycans (HSPG) and the low density lipoprotein (LDL) receptor-related protein/alpha 2 macroglobulin receptor (LRP). By immunofluorescence we show here that cell surface-bound LPL displays a pattern that corresponds to the previously described distribution of cell surface HSPG. No evident relation to the distribution of bound activated alpha 2-macroglobulin (alpha 2M*) or to LRP was observed. By immunoelectron microscopy we found that after 30 min at 37 degrees C most of the detected alpha 2M* (70% of the total gold particles) was inside the cells and associated with endosomal vesicles. However, at the same time, 76% of the LPL remained at the cell surface, suggesting that, LPL is internalized by a slow endocytic process. Binding of triglyceride-rich lipoproteins (TRL) or LDL together with LPL led to a spectacular increase in bound lipoproteins, which completely colocalized with LPL. After incubation at 37 degrees C, LPL and 1,1'-dioctadecyl-3,3,3,'3'-tetramethylindocarbocyanine (DiI)-TRL formed large clusters on the cell surface. Immunofluorescene and quantitative immunoelectron microscopy provided evidence of co-internalization of LPL and apoE-containing TRL by a slow endocytic process. In the absence of LPL, the fibroblasts rapidly internalized DiI-LDL and showed fluorescence in central, lysosome-like vesicles. In contrast, when LPL was present, internalization of DiI-LDL involved small, widely distributed vesicles. This pattern slowly changed to one consisting of large perinuclear vesicles. LDL receptor-deficient fibroblasts internalized DiI-LDL, either with or without LPL, into small widely distributed vesicles and no central vesicles were seen. Chloroquine-treated normal fibroblasts internalized DiI-LDL in a pattern similar to that of receptor-deficient fibroblasts. Taken together our results suggest an alternative receptor-independent endocytosis pathway for LDL. This pathway is potentiated by LPL and is characterized by a slow uptake involving small vesicles that gradually reach lysosomes. We suggest that, through its interaction with HSPG, LPL provides high capacity binding sites for lipoproteins and a independent internalization pathway.
Highlights
Lipoprotein lipase (LPL),a key enzyme in lipoprotein triglyceride metabolism, produces a marked increase in the retention and uptake of all classes of lipoproteins by cultured cells
We have previously shown that this characteristicpattern correspondsto the fibroblast cell surface distribution of heparan sulfate proteoglycans (HSPG), which are organizedby the actin cytoskeleton[39].When LRP was immunodetected in the same cells, no evident relation was found between LRP and LPL (Fig. lB), indicating that very little of the initial binding of LPL at the fibroblast cell surface could be explained by binding to plasma membrane LRP
The current results provide immunocytochemical evidence that human fibroblasts present a high binding capacity for LPL which is probably associated with cell surface HSPG
Summary
Lipoprotein lipase (LPL),a key enzyme in lipoprotein triglyceride metabolism, produces a marked increase in the retention and uptake of all classes of lipoproteins by cultured cells. The catabolism of chylomicrons and very low density lipoproteins (VLDL) is initiated by the hydrolysis of their triglycerides by the enzyme lipoprotein lipase (LPL) which is bound to the heparan sulfate proteoglycans (HSPG) on the luminal side Qf the vascular endothelium ( 1).Lipolysisby LPL causes a change in the lipid and protein composition of chylomicrons and VLDL, which become remnant particles and IDL-LDL, respectively [2]. Recent studies [11, 12] have confirmed previous observations by Felts, Abbrebiations: ACAT, acyl-Cokcholesterol acyltransferase; apo, apolipoprotein; azM*, activated awnacroglobulin; bLPL, bovine lipoprotein lipase; BSA, bovine serum albumin: DiI, 1,l'dioctadecylJ,3,3',3'-tetramethylindocarbocyanine;FH fibroblasts, LDL receptor-deficientfibroblasts;GPI, glycosyl phosphatidylinositol; IDL, intermediate densitylipoproteins;LDL, low densitylipoproteins; LRP, LDL receptor-related protein; HSPG, heparan sulfate proteoglycans; PBS, phosphate-buffered saline; RAP, receptorassociated protein; TRL, triglyceride-richlipoproteins: VLDL, very low density lipoproteins. It has been shown that LPL promotes binding of apoEcontaining lipoproteins to LRP [11, 20]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
