title: Uptake of hypertriglyceridemic very low density lipoproteins and their remnants by HepG2 cells: the role of lipoprotein lipase, hepatic triglyceride lipase, and cell surface proteoglycans.
abstract: Hypertriglyceridemic very low density lipoproteins (HTG-VLDL, S(f) 60-400) are not taken up by HepG2 cells. However, addition of bovine milk lipoprotein lipase (LPL) at physiological concentrations markedly stimulates uptake. In the present study, we determined whether: a) LPL catalytic activity is required for uptake, b) LPL functions as a ligand, and c) cell surface hepatic triglyceride lipase (HL) and/or proteoglycans are involved. Incubation of HepG2 cells with HTG-VLDL plus LPL (8 ng/ml) increased cellular cholesteryl ester (CE) 3.5-fold and triglyceride (TG) 6-fold. Heat-inactivation of LPL abolished the effect. Addition of tetrahydrolipstatin (THL, an LPL active-site inhibitor) to HTG-VLDL + LPL, inhibited the cellular increase in both CE and TG by greater than 90%. Co-incubation of HTG-VLDL + LPL with heparin, heparinase, or heparitinase, blocked CE accumulation by 70%, 48%, and 95%, respectively, but had no effect on the increase in cellular TG. Pre-treatment of cells with 1 mM 4-methylumbelliferyl-beta-D-xyloside, (beta-xyloside) to reduce cell surface proteoglycans inhibited the increase in CE induced by HTG-VLDL + LPL by 78%. HTG-VLDL remnants, prepared in vitro and isolated free of LPL activity, stimulated HepG2 cell CE 2.8-fold in the absence of added LPL, a process inhibited with THL by 66%. Addition of LPL (8 ng/ml) to remnants did not further enhance CE accumulation. HepG2 cell HL activity, released by heparin, was inhibited 95% by THL. The amount of HL activity and immunoreactive mass, released by heparin, was reduced 50-60% in beta-xyloside-treated cells. These results indicate that physiological concentrations of LPL promote HepG2 cell uptake of HTG-VLDL primarily due to remnant formation and that LPL does not play a major role as a ligand. HL activity and cell surface proteoglycans significantly enhance the subsequent uptake of VLDL remnants.
authors.affiliation: Department of Medicine and the Robarts Research Institute, The University of Western Ontario, London, Canada.
authors.forename: M W
authors.initials: MW
authors.lastname: Huff
authors.forename: D B
authors.initials: DB
authors.lastname: Miller
authors.forename: B M
authors.initials: BM
authors.lastname: Wolfe
authors.forename: P W
authors.initials: PW
authors.lastname: Connelly
authors.forename: C G
authors.initials: CG
authors.lastname: Sawyez
mesh_terms: D000818:Animals; D002417:Cattle; D002460:Cell Line; D002788:Cholesterol Esters / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006027:Glycosides / Q000494:pharmacology; D006493:Heparin / Q000494:pharmacology; D019763:Heparin Lyase; D006358:Hot Temperature; D006801:Humans; D015228:Hypertriglyceridemia / Q000097:blood*; D007783:Lactones / Q000494:pharmacology; D008049:Lipase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008071:Lipoprotein Lipase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008077:Lipoproteins, LDL / Q000378:metabolism; D008079:Lipoproteins, VLDL / Q000097:blood / Q000378:metabolism*; D008099:Liver / Q000201:enzymology / Q000378:metabolism*; D000077403:Orlistat; D011133:Polysaccharide-Lyases / Q000494:pharmacology; D011509:Proteoglycans / Q000378:metabolism*
chemical_list: D002788:Cholesterol Esters; D004791:Enzyme Inhibitors; D006027:Glycosides; D007783:Lactones; D008077:Lipoproteins, LDL; D008079:Lipoproteins, VLDL; D011509:Proteoglycans; C010807:xylosides; D006493:Heparin; D000077403:Orlistat; D008049:Lipase; D008071:Lipoprotein Lipase; D011133:Polysaccharide-Lyases; D019763:Heparin Lyase; C021349:heparitinsulfate lyase