Hepatic lipase mediates an increase in selective uptake of high-density lipoprotein-associated cholesteryl esters by human Hep 3B hepatoma cells in culture.
Selective uptake of high-density lipoprotein- (HDL-) associated cholesteryl esters (CE), i.e. lipid uptake independent from particle uptake, delivers CE to the liver and steroidogenic tissues in vivo. In vitro, besides hepatocytes and steroidogenic cells many other cell types selectively take up HDL CE. Hepatic lipase (HL) stimulates the internalisation of apoprotein (apo) B-containing lipoproteins by hepatocytes independent from lipolysis. In this study the role of HL in the hepatic metabolism of apo A-I-containing lipoproteins, i.e. HDL, was investigated. HDL3 (d = 1.125-1.21 g/ml) was radiolabeled in its protein (125I) and in its CE moiety ([3H]cholesteryl oleyl ether, ([3H]CEt)). HL originated from tissue culture media of hepatoma cells and from post-heparin plasma. Human Hep 3B hepatoma cells incubated in medium containing radiolabeled HDL3. In the absence of HL, the rate of apparent HDL3 particle uptake according to the lipid tracer ([3H]CEt) was in most cases in approximately 10-fold excess on that due to the protein label (125I), indicating selective CE uptake from HDL3. Addition of HL to these incubations increased the cellular uptake of [3H]CEt and of 125I from HDL3 and quantitatively the most prominent effect was an up to approximately 2.5-fold stimulation of apparent selective CE uptake ([3H]CEt-125I). This increase in selective CE uptake was observed in the presence of tetrahydrolipstatin, an inhibitor of the catalytically active site of HL, suggesting that this HL effect is independent from lipolysis. HL binds to cell surface heparan sulfate proteoglycans. To explore the role of these molecules for the HL effect on selective CE uptake, hepatoma cells were depleted of proteoglycans or Chinese hamster ovary (CHO) cells deficient in proteoglycan synthesis were used. Proteoglycan-deficiency reduced the HL-mediated increase in selective uptake by more than 80%. To investigate if low-density lipoprotein (LDL) receptors or the LDL receptor-related protein (LRP) are involved in the HL effect on selective CE uptake, murine embryonic fibroblasts (MEF) were used which are deficient in these receptors; alternatively, monensin, an inhibitor of endocytosis was present in the medium of Hep 3B cells during the uptake assay for labeled HDL3. These experiments yielded no evidence for a role of LDL receptors or LRP in the HL-mediated increase in selective CE uptake. In summary, HL mediates an increase in HDL3 selective CE uptake by human Hep 3B hepatoma cells. This HL effect is independent from lipolysis and independent from LRP and LDL receptors. However this HL effect is susceptible to cell surface proteoglycan deficiency. The potential physiologic implication is that HL modifies HDL selective CE uptake by the liver in vivo and such an effect could play a role in reverse cholesterol transport.
Rinninger
F
F
Universität Hamburg, Krankenhaus Eppendorf, Medizinische Kernklinik und Poliklinik, Germany.
Mann
W A
WA
Kaiser
T
T
Ahle
S
S
Meyer
N
N
Greten
H
H
D000818:Animals; D016466:CHO Cells; D006528:Carcinoma, Hepatocellular; D002788:Cholesterol Esters / Q000378:metabolism*; D006224:Cricetinae; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism; D019812:Heparan Sulfate Proteoglycans / Q000378:metabolism; D019763:Heparin Lyase / Q000494:pharmacology; D006801:Humans; D007783:Lactones / Q000494:pharmacology; D008049:Lipase / Q000494:pharmacology / Q000502:physiology*; D008075:Lipoproteins, HDL / Q000378:metabolism*; D053440:Lipoproteins, HDL3; D008099:Liver / Q000201:enzymology / Q000378:metabolism*; D051379:Mice; D008985:Monensin / Q000494:pharmacology; D000077403:Orlistat; D011973:Receptors, LDL / Q000172:deficiency / Q000502:physiology; D014407:Tumor Cells, Cultured
D002788:Cholesterol Esters; D004791:Enzyme Inhibitors; C059714:HDL cholesteryl ester; D019812:Heparan Sulfate Proteoglycans; D007783:Lactones; D008075:Lipoproteins, HDL; D053440:Lipoproteins, HDL3; D011973:Receptors, LDL; D008985:Monensin; D000077403:Orlistat; D008049:Lipase; D019763:Heparin Lyase