scholarly journals The human asialoglycoprotein receptor is a possible binding site for low-density lipoproteins and chylomicron remnants

1991 ◽  
Vol 276 (1) ◽  
pp. 79-87 ◽  
Author(s):  
E Windler ◽  
J Greeve ◽  
B Levkau ◽  
V Kolb-Bachofen ◽  
W Daerr ◽  
...  
Keyword(s):  
Cell Density ◽  
Hepg2 Cells ◽  
High Cell Density ◽  
Low Density Lipoprotein ◽  
Ldl Receptor ◽  
Density Lipoprotein ◽  
Asialoglycoprotein Receptor ◽  
Low Density ◽  
High Cell ◽  
Chylomicron Remnants

Binding and internalization of chylomicron remnants from rat mesenteric lymph by HepG2 cells was inhibited by both excess remnants and low-density lipoprotein (LDL) to the same extent. Ligand blots revealed binding of remnants and LDL to the LDL receptor. Measures regulating LDL receptor activity greatly influenced the binding of remnants: ethinyloestradiol, the hydroxymethylglutaryl-CoA reductase inhibitor pravastatin and the absence of LDL all increased binding, whereas high cell density or the presence of LDL decreased binding. Also, asialofetuin, asialomucin, the neoglycoprotein galactosyl-albumin and an antibody against the asialoglycoprotein receptor all decreased substantially the binding of remnants. At high cell density, binding internalization and degradation of chylomicron remnants was inhibited by up to 70-80%, yet binding of LDL was inhibited by no more than 20-30%. In cross-competition studies, the binding of 125I-asialofetuin was efficiently competed for by asialofetuin itself or by the antibody, and also by LDL and remnants, yet remnants displayed an approx. 100-fold higher affinity than LDL. Likewise, remnants of human triacylglycerol-rich lipoproteins and asialofetuin interfered with each others' binding to HepG2 cells or human liver membranes. It is concluded that the LDL receptor mediates the internalization of chylomicron remnants into hepatocytes depending on its activity, according to demand for cholesterol. Additionally, the asialoglycoprotein receptor may contribute to the endocytosis of LDL, but predominantly of chylomicron remnants.

scholarly journals Effect of reduced low-density lipoprotein receptor level on HepG2 cell cholesterol metabolism

1998 ◽  
Vol 329 (1) ◽  
pp. 81-89 ◽  
Author(s):  
Lahoucine IZEM ◽  
Eric RASSART ◽  
Lassana KAMATE ◽  
Louise FALSTRAULT ◽  
David RHAINDS ◽  
...  
Keyword(s):  
Hepg2 Cell ◽  
Hepg2 Cells ◽  
Cholesterol Metabolism ◽  
Low Density Lipoprotein ◽  
Ldl Receptor ◽  
Constitutive Expression ◽  
Density Lipoprotein ◽  
Low Density ◽  
Apo B ◽  
Acat Activity

Low-density lipoproteins (LDL) are taken up by both LDL receptor (LDLr)-dependent and -independent pathways. In order to determine the importance of these pathways in the activity of the various enzymes that are important in maintaining the cellular cholesterol level in hepatic cells, we created HepG2 cells expressing lower levels of LDLr. Thus HepG2 cells were transfected with a constitutive expression vector (pRc/CMV) containing a fragment of LDLr cDNA inserted in an antisense manner. Stable transformants were obtained that showed significant reductions of 42, 72 and 85% of LDLr protein levels compared with the control, as demonstrated by immunoblotting and confirmed by the LDL binding assay. The best inactivation was achieved with the construct containing the first 0.7 kb of LDLr cDNA. Incubating the different HepG2 cell subtypes with LDL showed similar association of apolipoprotein B (apo B) or cholesteryl esters from LDL with the cells, indicating that the LDLr deficiency did not significantly affect LDL uptake by the cell. However, apoB degradation was reduced significantly by 71-82% in the most LDLr-deficient HepG2 cells. We also found that 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCoA red) activity is significantly increased by 32-35% in HepG2 cells expressing very low levels of LDLr that also demonstrate a significant decrease of 20% in acyl-CoA:cholesterol acyltransferase (ACAT) activity. However, these effects are moderate compared with those observed when cells were incubated in lipoprotein-depleted medium, where a > 900% increase in HMGCoA red activity and a loss of 60% of ACAT activity was observed. Thus, in HepG2 cells, different levels of LDLr affect LDL-apoB degradation, but have very little effect on LDL association, HMGCoA red and ACAT activities, revealing that LDLr is more important in the clearance of LDL-apoB than in HepG2 cell cholesterol homoeostasis, a role that should be attributable to both LDLr-dependent and -independent pathways.

scholarly journals Role of the Low Density Lipoprotein (LDL) Receptor Pathway in the Metabolism of Chylomicron Remnants

1996 ◽  
Vol 271 (37) ◽  
pp. 22422-22427 ◽  
Author(s):  
Shun Ishibashi ◽  
Stéphane Perrey ◽  
Zhong Chen ◽  
Jun-ichi Osuga ◽  
Masako Shimada ◽  
...  
Keyword(s):  
Low Density Lipoprotein ◽  
Ldl Receptor ◽  
Density Lipoprotein ◽  
Low Density ◽  
Chylomicron Remnants

scholarly journals Co-ordinate regulation of low-density-lipoprotein receptor and 3-hydroxy-3-methylglutaryl-CoA reductase and synthase gene expression in HepG2 cells

1989 ◽  
Vol 260 (3) ◽  
pp. 731-736 ◽  
Author(s):  
D T Molowa ◽  
G M Cimis
Keyword(s):  
Hepg2 Cells ◽  
Cholesterol Biosynthesis ◽  
Hepatoma Cell ◽  
Low Density Lipoprotein ◽  
Ldl Receptor ◽  
Density Lipoprotein ◽  
Hepatoma Cell Line ◽  
Low Density ◽  
Hmg Coa Reductase ◽  
Coa Reductase

Cellular processes responsible for maintaining cholesterol homoeostasis are highly regulated. To determine whether two of these processes, cholesterol biosynthesis and receptor-mediated uptake of low-density lipoprotein (LDL), are co-ordinately regulated in human liver, we employed a human hepatoma cell line (HepG2) and measured the accumulation of mRNA for LDL receptor, 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase and HMG-CoA synthase under a variety of conditions. Genomic Southern-blot analysis demonstrated that the integrity of these genes is maintained in the transformed cell. Treatment of HepG2 cells with mevalonate, 25-hydroxycholesterol, LDL, lovastatin or miconazole resulted in a similar effect on the accumulation of all three mRNAs at the concentrations tested. The onset of the response to drug, whether repression or induction of mRNA accumulation, occurred after approximately the same period of exposure for each mRNA. We conclude that the expression of the LDL receptor, HMG-CoA reductase and HMG-CoA synthase is co-ordinately regulated in HepG2 cells.

scholarly journals Uptake of retinyl ester in HL-60 cells via the low-density-lipoprotein-receptor pathway

1989 ◽  
Vol 257 (1) ◽  
pp. 239-244 ◽  
Author(s):  
K O Wathne ◽  
B Carlander ◽  
K R Norum ◽  
R Blomhoff
Keyword(s):  
Low Density Lipoprotein ◽  
Cholesterol Acyltransferase ◽  
Ldl Receptor ◽  
Density Lipoprotein ◽  
Retinyl Palmitate ◽  
Low Density ◽  
Acyltransferase Activity ◽  
Retinyl Ester ◽  
Leukaemic Cells ◽  
Chylomicron Remnants

Newly absorbed retinol is transported in association with chylomicrons and their remnants. In addition, after intake of high doses of retinol, significant amounts are also found in low-density lipoprotein (LDL). As both chylomicron remnants and LDL may be taken up by cells via the LDL receptor, and retinoids inhibit proliferation of some leukaemic cells, we have studied the uptake of retinol in leukaemic cells via the LDL-receptor pathway. HL-60 cells contain saturable binding sites for LDL. The binding of LDL to its receptor has a dissociation constant of about 3.2 x 10(-9) M, and the number of receptors per cell was calculated to be about 2700. Uptake of 125I-LDL by HL-60 cells was increased 2-fold by preincubating the cells with mevinolin. The presence of specific receptors for LDL on HL-60 cells was further confirmed by the finding that exogenous LDL cholesterol was able to up-regulate the ACAT (acyl-CoA: cholesterol acyltransferase) activity of HL-60 cells. We then tested the uptake of retinyl ester in leukaemic cells via the LDL-receptor pathway. HL-60 cells were incubated with LDL or chylomicron remnants labelled with [3H]retinyl palmitate. Uptake of retinyl ester associated with both LDL and chylomicron remnants was observed. Furthermore, the presence of excess LDL decreased the uptake by 75-100%, supporting the hypothesis that the uptake of retinyl ester occurred via the LDL receptor in HL-60 cells.

scholarly journals Binding of rat chylomicrons and their remnants to the hepatic low-density-lipoprotein receptor and its role in remnant removal

1988 ◽  
Vol 252 (2) ◽  
pp. 553-561 ◽  
Author(s):  
E E T Windler ◽  
J Greeve ◽  
W H Daerr ◽  
H Greten
Keyword(s):  
Apolipoprotein E ◽  
Specific Binding ◽  
Low Density Lipoprotein ◽  
Ldl Receptor ◽  
Density Lipoprotein ◽  
Hepatic Uptake ◽  
Low Density ◽  
Pass Through ◽  
Chylomicron Remnants

Binding and uptake of rat chylomicrons of different metabolic stages by the hepatic low-density-lipoprotein (LDL) receptor were studied. Pure chylomicrons, characterized by apolipoprotein B-48 devoid of contaminating B-100, were labelled in their cholesteryl esters. Lymph chylomicrons and serum chylomicrons, enriched in apolipoprotein E and the C-apolipoproteins, bound poorly to rat hepatic membranes. In contrast, chylomicron remnants, containing the apolipoproteins B-48 and E, bound with high affinity. Specific binding of remnants was virtually completely competed for by LDL free of apolipoprotein E. In addition, in ligand blots both remnants and LDL associated with the same protein with an Mr characteristic of the LDL receptor. Uptake of remnants during a single pass through isolated perfused rat livers was decreased to about 50% by an excess of LDL. It is concluded that rat chylomicron remnants are a ligand of the hepatic LDL receptor. The much higher affinity as compared with LDL is mediated by apolipoprotein E but not B-48, and is inhibited by the C-apolipoproteins. This explains why serum chylomicrons are not taken up by the liver, whereas remnants are rapidly removed from the circulation. Results from experiments in vivo suggest that the LDL receptor makes an important contribution to the hepatic uptake of remnants and may be the principal binding site of the liver responsible for remnant removal.

scholarly journals Interaction of 4-hydroxynonenal-modified low-density lipoproteins with the fibroblast apolipoprotein B/E receptor

1986 ◽  
Vol 234 (1) ◽  
pp. 245-248 ◽  
Author(s):  
W Jessup ◽  
G Jurgens ◽  
J Lang ◽  
H Esterbauer ◽  
R T Dean
Keyword(s):  
Apolipoprotein B ◽  
Negative Charge ◽  
Low Density Lipoprotein ◽  
Ldl Receptor ◽  
Density Lipoprotein ◽  
Low Density ◽  
Lipid Peroxidation Product ◽  
Modified Low Density Lipoproteins ◽  
Peroxidation Product

The incorporation of the lipid peroxidation product 4-hydroxynonenal into low-density lipoprotein (LDL) increases the negative charge of the particle, and decreases its affinity for the fibroblast LDL receptor. It is suggested that this modification may occur in vivo, and might promote atherogenesis.

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