Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0019204 (hepatocellular carcinoma)
 71,386 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Insulin modulation of apolipoprotein B gene expression was studied at the translational level by the use of a cell-free translation system from a hepatoma cell-line, HepG2. Extracts of HepG2 cells lysed with lysolecithin were found to have high in vitro protein synthesizing activity utilizing endogenous mRNA. The level of peptide chain initiation was high, as suggested by a significant inhibition of translation by edeine. The translation products of endogenous mRNA in HepG2 cell-free lysate were probed with anti-apolipoprotein B antibodies to investigate its synthesis. A 550 kilodalton (kDa) polypeptide was selected by a polyclonal antibody, as well as a monoclonal antibody, against the C-terminal end of apolipoprotein B molecule. This in vitro synthesized polypeptide was also found to compare well in size with the in vivo product. The HepG2 lysate was also shown to efficiently synthesize in vitro a number of other proteins including albumin, apolipoprotein E, apolipoprotein A1, and actin. The in vitro synthesis of polypeptides as large as 500 kDa was unexpected and has not previously been demonstrated in a cell-free system. The HepG2 translation system was used to investigate the effect of insulin on the in vitro translation of apolipoprotein B. Lysates prepared from HepG2 cells treated with insulin were found to have lower translational activity (by an average of 52.3%) for apolipoprotein B compared with lysates from control untreated cells. In vitro synthesis of actin and apolipoprotein E were unaffected under these conditions. The insulin-stimulated decline in in vitro apolipoprotein B synthesis was not due to a change in apolipoprotein B mRNA levels as determined by slot- and Northern-blot analyses, suggesting that the inhibitory effect of insulin may be exerted partly at the level of apolipoprotein B mRNA translation.
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PMID:Insulin modulation of human apolipoprotein B mRNA translation: studies in an in vitro cell-free system from HepG2 cells. 133 69

We examined the effects of apolipoproteins A-IV and A-I on the catabolism of whole particles by hepatoma G2 cells and cultured primary hepatocytes. For this type of experiment, high density lipoprotein is unsuitable, because all of its lipid and protein components independently dissociate and exchange and hence poorly trace whole particle catabolism. We therefore used phosphatidylcholine liposomes with radioactive tracers entrapped within their aqueous cores. Apolipoproteins A-IV, A-I, or E added to liposomes became liposome-associated and produced no detectable release of encapsulated label. As a positive control, apolipoprotein E doubled the uptake of labeled liposomes by hepatoma cells, compared to apolipoprotein-free controls, and this increase could be blocked by the addition of excess unlabeled low density lipoprotein. Degradation of labeled liposomes by hepatoma cells was increased 6-fold by the addition of apolipoprotein E. In contrast, neither apolipoprotein A-IV nor A-I increased cellular uptake or degradation of the particles. Similar results were obtained with primary hepatocytes. In studies using apolipoprotein combinations, apolipoproteins A-IV and A-I were each able to displace apolipoprotein E from liposomes and thereby reduce cellular uptake. Our data indicate that apolipoproteins A-IV and A-I do not facilitate uptake or degradation of whole particles by liver-derived cells in vitro. However, these apolipoproteins may modulate receptor-mediated uptake of particles by reducing the amount of particle-bound apolipoprotein E.
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PMID:Effects of apolipoproteins A-IV and A-I on the uptake of phospholipid liposomes by hepatocytes. 249 20

Hepatic triglyceride lipase (H-TGL) is a key lipolytic enzyme in the metabolism of human plasma high density lipoproteins. The enzyme is bound to glycosaminoglycans on endothelial cells in the liver and is immediately released into the circulation after heparin administration. In addition to releasing H-TGL, heparin-like glycosaminoglycans have also been shown to suppress hepatocyte proliferation and to alter tissue-specific gene expression. In the present study, the effects of heparin exposure on the secretion of H-TGL were examined in a human parenchymal hepatoma (HepG2) cell line. The addition of heparin to serum-supplemented medium induced the secretion of H-TGL in a time- and concentration-dependent manner. At 5.4 micrograms/ml heparin, H-TGL levels, as determined by triacyglycerol hydrolase activity, increased 7-fold after a 44-h incubation. Heparin exposure decreased intracellular H-TGL activity from 21.3 to 4.8 nmol of oleic acids released/h/10(8) cells and increased enzyme activity in the medium from 16.2 to 165.3 nmol of oleic acids released/h/10(8) cells. The heparin-induced secretion of H-TGL was associated with increased levels of H-TGL-specific mRNA. The addition of actinomycin D or cycloheximide reversed the heparin-induced increase in H-TGL activity and mRNA. Heparin treatment did not increase the level of actin mRNA suggesting that elevated H-TGL mRNA is due to enhanced tissue-specific expression of H-TGL. Expression of apolipoprotein E, another protein involved in lipoprotein metabolism, also showed induced levels of mRNA by heparin but to a lesser extent than that for H-TGL. We conclude that heparin stimulates the de novo synthesis of H-TGL in liver parenchymal cells in vivo by influencing both transcriptional and post-transcriptional events.
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PMID:Heparin induces the expression of hepatic triglyceride lipase in a human hepatoma (HepG2) cell line. 254 13

Characterization of the membrane receptor for the low density lipoproteins (LDL) has led to insights into cellular receptor physiology as well as mammalian lipid transport. Result with LDL have stimulated the search for specific receptors for other plasma lipoproteins. Receptors for high density lipoproteins (HDL) have been identified in human fibroblasts and smooth muscle cells. Specificity for this receptor has been difficult to define since normal HDL contains several apolipoproteins, and particles containing apolipoproteins B and E have been shown to compete for HDL binding. In the present study, we demonstrate that HDL isolated from a patient devoid of apolipoprotein E was bound specifically by human hepatic membranes. This binding reached saturation within 2 hours and was EDTA-resistant. Assuming a single receptor model, we found that 2.9 x 10(15) receptors/mg membrane protein bound with an affinity KD = 3.5 x 10(-7) M at 0 to 4 degrees C and KD = 1.9 x 10(-7) M at 37 degrees C. The binding was effectively competed with intact HDL3, with HDL3 that had undergone selective arginine and lysine residue modification, and with antibodies to apolipoproteins A-I and A-II. However, LDL, asialofetuin, and HDL3 which had undergone tyrosine modification by nitration, and anti-apolipoprotein B did not compete with apo A-I HDL binding. In contrast to LDL binding, the human hepatoma cell line, HEPG2, increased HDL binding with cholesterol loading that was specific for HDL3. Thus, hepatic tissue can modulate its recognition of HDL. Finally, hepatic membranes from a patient lacking normal hepatic LDL receptors bound apo A-I HDL normally. These data indicate that a saturable, specific regulatable receptor for apo E-free HDL is present in human liver.
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PMID:Characterization of a human hepatic receptor for high density lipoproteins. 298 87

We studied the effect of the plant alkaloid castanospermine on the biosynthesis and secretion of human hepatoma glycoproteins. The HepG-2 cells, grown in the presence or absence of the alkaloid, were labelled with [2-3H]mannose and then the labelled glycopeptides were prepared by Pronase digestion. This material was analysed by gel filtration on Bio-Gel P-4 before and after treatment with endo-beta-N-acetylglucosaminidase H. Castanospermine caused an accumulation of high-mannose oligosaccharides, by 70-75% over control. The major accumulated product, which could also be labelled with [3H]galactose and was only partially susceptible to alpha-mannosidase digestion, was identified by h.p.l.c. as a Glc3Man9GlcNAc. Thus the alkaloid inhibits glucosidase I in the human hepatoma cells. Analysis of total glycoproteins secreted by the cells into the medium revealed the presence of only complex oligosaccharides in both control and treated cultures, and the amount of the oligosaccharides labelled with radioactive mannose, galactose or N-acetylmannosamine, secreted by treated cells, was decreased by about 60%. The rate of secretion of total protein labelled with [35S]methionine and precipitated from the medium with trichloroacetic acid was inhibited by up to 40% in the presence of castanospermine. Pulse-chase studies utilizing [35S]methionine labelling were performed to study the effect of the alkaloid on secretion of individual plasma proteins. Immunoprecipitation at different chase times with monospecific antisera showed that castanospermine markedly decreased the secretion rates of alpha 1-antitrypsin, caeruloplasmin and, to a lesser extent, that of antithrombin-III. Secretions of apolipoprotein E, a glycoprotein containing only O-linked oligosaccharide(s), and albumin, a non-glycosylated protein, were not affected by the drug. It is suggested that castanospermine inhibits secretion of at least some glycoproteins containing N-linked oligosaccharides, owing to the inhibition of oligosaccharide processing.
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PMID:Castanospermine inhibits glucosidase I and glycoprotein secretion in human hepatoma cells. 300 19

The regulation of the hepatic catabolism of normal human very-low-density lipoproteins (VLDL) was studied in human-derived hepatoma cell line HepG2. Concentration-dependent binding, uptake and degradation of 125I-labeled VLDL demonstrated that the hepatic removal of these particles proceeds through both the saturable and non-saturable processes. In the presence of excess unlabeled VLDL, the specific binding of 125-labeled VLDL accounted for 72% of the total binding. The preincubation of cells with unlabeled VLDL had little effect on the expression of receptors, but reductive methylation of VLDL particles reduced their binding capacity. Chloroquine and colchicine inhibited the degradation of 125I-labeled VLDL and increased their accumulation in the cell, indicating the involvement of lysosomes and microtubuli in this process. Receptor-mediated degradation was associated with a slight (13%) reduction in de novo sterol synthesis and had no significant effect on the cellular cholesterol esterification. Competition studies demonstrated the ability of unlabeled VLDL, low-density lipoproteins (LDL) and high-density lipoproteins (HDL) to effectively compete with 125I-labeled VLDL for binding to cells. No correlation was observed between the concentrations of apolipoproteins A-I, A-II, C-I, C-II and C-III of unlabeled lipoproteins and their inhibitory effect on 125I-labeled VLDL binding. When unlabeled VLDL, LDL and HDL were added at equal contents of either apolipoprotein B or apolipoprotein E, their inhibitory effect on the binding and uptake of 125I-labeled VLDL only correlated with apolipoprotein E. Under similar conditions, the ability of unlabeled VLDL, LDL and HDL to compete with 125I-labeled LDL for binding was a direct function of only their apolipoprotein B. These results demonstrate that in HepG2 cells, apolipoprotein E is the main recognition signal for receptor-mediated binding and degradation of VLDL particles, while apolipoprotein B functions as the sole recognition signal for the catabolism of LDL. Furthermore, the lack of any substantial regulation of beta-hydroxy-beta-methylglutaryl-CoA reductase and acyl-CoA:cholesterol acyltransferase activities subsequent to VLDL degradation, in contrast to that observed for LDL catabolism, suggests that, in HepG2 cells, the receptor-mediated removal of VLDL proceeds through processes independent of those involved in LDL catabolism.
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PMID:Studies on the binding and degradation of human very-low-density lipoproteins by human hepatoma cell line HepG2. 300 89

The human liver cell line HepG2 was investigated for its synthesis and secretion of lecithin-cholesterol acyltransferase. The cells were grown to confluency in Eagle's minimal essential medium plus 10% fetal bovine serum. At the onset of the study, fetal bovine serum was removed and cells were grown in minimal essential medium only. At 6, 12, 24, and 48 h the cells were harvested, and the culture medium collected at each time point was assayed for lecithin-cholesterol acyltransferase mass and activity, cholesterol esterification rate, and apolipoprotein A-I mass. The rate of the enzyme secretion measured by both mass and activity was linear over 24 h of culture. The enzyme mass by radioimmunoassay was 1.7, 4.1, 7.9 and 13.7 ng/ml culture medium (or 8.3, 19.9, 38.5 and 66.7 ng/mg cell protein), respectively, and enzyme activity using an exogenous source of phosphatidylcholine/cholesterol liposomes containing apolipoprotein A-I as substrate was 85, 170, 315, and 402 pmol cholesterol esterified/h per ml culture medium (or 414, 828, 1534 and 1957 pmol cholesterol esterified/h per mg cell protein) for 6, 12, 24, and 48 h of culture, respectively. The endogenous cholesterol esterification rate of the culture medium was 47, 104, 224 and 330 pmol/h per ml and apolipoprotein A-I mass was 305, 720, 2400 and 3940 ng/ml culture medium over the same time frame. In contrast to culture medium, low levels of enzyme activity (approximately 10% of that in culture medium at 24 and 48 h) were observed in the extracts of HepG2 cells. The enzyme secreted by HepG2 was found to be similarly activated by apolipoprotein A-I, apolipoprotein E, or apolipoprotein A-IV, and was similarly inhibited by phenylmethylsulfonyl fluoride, dithiobisnitrobenzoate, p-hydroxymercuribenzoate, or iodoacetate as compared to human plasma enzyme. High-performance gel filtration of the culture medium revealed that the HepG2-secreted enzyme was associated with a fraction having a mean apparent molecular weight of approximately 200,000. We concluded that human hepatoma HepG2 cells synthesize and secrete lecithin-cholesterol acyltransferase, which is functionally homologous to the human plasma enzyme.
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PMID:Synthesis and secretion of lecithin-cholesterol acyltransferase by the human hepatoma cell line HepG2. 301 21

The binding and metabolism of [3H]vitamin A-containing chylomicron (CM) remnants by the human hepatoma cell line HepG2 were studied. Mesenteric lymph chylomicrons were collected from [3H]retinol-fed rats and incubated with lipoprotein lipase to obtain CM remnants. At 4 degrees C, specific CM remnant binding was inhibited by an excess of unlabeled CM remnants. Specific binding predominated at low concentrations and approached saturation while total binding continued to increase over an extensive concentration range (0.45-32 microgram triglyceride/ml). CM remnant uptake at 37 degrees C was greater than that of CM and at least 70 times more efficient than the pinocytosis of sucrose. CM remnant binding increased with the extent of lipolysis. Addition of human apolipoprotein E enhanced both CM remnant and CM binding. After internalization, HepG2 cells hydrolyzed CM remnant-[3H]retinyl esters, and radiolabeled metabolites accumulated. As a function of the concentration of [3H]retinoid initially bound to cells, retinol and retinyl esters accumulated as the major cell-associated metabolites. In contrast, retinol was the major metabolite in the medium only at low retinoid concentrations; other more polar metabolites accumulated at higher concentrations (greater than 110 pmol retinoid/mg cell protein). The accumulation in the medium of labeled metabolites derived from CM remnant-retinoid was reduced when cells were preincubated in unlabeled retinol-supplemented media. The specific activity of retinol in the medium indicated that CM remnant-vitamin A had mixed with the cellular store prior to its secretion as retinol. These results indicate that HepG2 cells internalize CM remnants in part by specific binding sites, and that the metabolism of CM remnant-retinoids by the HepG2 cell involves retinyl ester hydrolysis and the secretion of retinol and other more polar metabolites. These processes were regulated in part by the concentration of retinoid delivered by the CM remnant and by the initial retinoid content of the cell.
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PMID:Chylomicron remnant-vitamin A metabolism by the human hepatoma cell line HepG2. 303 18

The present study was designed to investigate whether plasma lipoproteins and albumin can affect the basal synthetic rate of apolipoproteins in differentiated rat hepatoma cells (Fao) incubated in serum-free medium. The synthesis of apolipoproteins was measured by the incorporation of [35S]methionine into medium lipoproteins isolated by density gradient ultracentrifugation. Under all the experimental conditions used, Fao cells synthesized almost exclusively apolipoprotein E. When cells were incubated in the presence of 5-10% rat plasma the synthesis of apolipoprotein E increased 2-3-fold; lipoprotein-deficient serum had a negligible effect. Fatty acid-poor bovine serum albumin (BSA), which had been found to reduce very-low-density lipoprotein secretion in isolated rat hepatocytes, did not modify the synthesis of apolipoprotein E. When Fao cells were incubated in medium containing rat plasma lipoprotein fractions, the synthesis of apolipoprotein E increased. The d less than 1.090 g/ml plasma lipoprotein fraction had the major stimulatory effect. Increased apolipoprotein E synthesis was observed when cells were incubated in the presence of lipids extracted from rat plasma lipoproteins. These results suggest that the intracellular accumulation of lipoprotein-lipids plays an important role in regulating apolipoprotein E synthesis in Fao cells.
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PMID:Modulation of the synthesis of apolipoproteins in rat hepatoma cells. 319 28

The human hepatoma cell line, Hep G2, has been used to compare the metabolism by isolated liver cells of purified isoforms of human apolipoprotein E (apo E). Complexes of [125I]apo E-3/3, 2/2, 3/2 and 4/3 with dimyristoyl phosphatidylcholine (DMPC) were prepared by a detergent-dialysis method: discoidal, bilayer complexes with a stoichiometry of 125 +/- 15 mol DMPC/mol apo E resulted. The predominant phenotype apo E-3/3, and the phenotype apo E-2/2 characteristic of patients with Type III hyperlipoproteinemia, interact similarly with DMPC and adopt the same conformation with 60-70% alpha-helix, as monitored by circular dichroism spectroscopy. The uptake and degradation at 37 degrees C, and binding at 4 degrees C by Hep G2 cells, of [125I]apo E-3/3/DMPC and [125I]apo E-2/2/DMPC complexes were compared. Apo E-3/3 was degraded more rapidly than apo E-2/2 suggesting that the diminished catabolism of the latter phenotype by intact livers is due to lack of recognition by the hepatocytes. The observed degradation of apo E was 3-4 times greater than that which could be attributed to fluid phase endocytosis and low-affinity adsorptive endocytosis. The degradation of [125I]apo A-I by Hep G2 cells can be accounted for by the above endocytotic mechanisms. The distinction between apo E-3/3 and apo E-2/2 isoforms is attributed to the presence of a cell-surface receptor on Hep G2 cells which binds apo E-3/3 with a higher affinity than apo E-2/2.
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PMID:The conformation of apolipoprotein E isoforms in phospholipid complexes and their interaction with human Hep G2 cells. 608 44


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