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Query: UMLS:C0019204 (hepatocellular carcinoma)
 71,386 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effect of 12-O-tetradecanoylphorbol-13-acetate (TPA) on the function of the insulin receptor was examined in intact hepatoma cells (Fao) and in solubilized extracts purified by wheat germ agglutinin chromatography. Incubation of ortho[32P]phosphate-labeled Fao cells with TPA increased the phosphorylation of the insulin receptor 2-fold after 30 min. Analysis of tryptic phosphopeptides from the beta-subunit of the receptor by reverse-phase high performance liquid chromatography and determination of their phosphoamino acid composition suggested that TPA predominantly stimulated phosphorylation of serine residues in a single tryptic peptide. Incubation of the Fao cells with insulin (100 nM) for 1 min stimulated 4-fold the phosphorylation of the beta-subunit of the insulin receptor. Prior treatment of the cells with TPA inhibited the insulin-stimulated tyrosine phosphorylation by 50%. The receptors extracted with Triton X-100 from TPA-treated Fao cells and purified on immobilized wheat germ agglutinin retained the alteration in kinase activity and exhibited a 50% decrease in insulin-stimulated tyrosine autophosphorylation and phosphotransferase activity toward exogenous substrates. This was due primarily to a decrease in the Vmax for these reactions. TPA treatment also decreased the Km of the insulin receptor for ATP. Incubation of the insulin receptor purified from TPA-treated cells with alkaline phosphatase decreased the phosphate content of the beta-subunit to the control level and reversed the inhibition, suggesting that the serine phosphorylation of the beta-subunit was responsible for the decreased tyrosine kinase activity. Our results support the notion that the insulin receptor is a substrate for protein kinase C in the Fao cell and that the increase in serine phosphorylation of the beta-subunit of the receptor produced by TPA treatment inhibited tyrosine kinase activity in vivo and in vitro. These data suggest that protein kinase C may regulate the function of the insulin receptor.
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PMID:Phorbol ester-induced serine phosphorylation of the insulin receptor decreases its tyrosine kinase activity. 312 81

Using the particulate fraction of tissue homogenate, plasma membrane-associated sialidase was assayed at pH 4.5 with bovine brain mixed gangliosides as the substrate. The activity was lower in rat hepatoma induced by 3'-methyl-4-dimethylaminoazobenzene (MeDAB) and transplantable AH-109A rat hepatoma than in normal rat liver. The enzyme was almost quantitatively solubilized from liver particulate fraction by using 0.5% (w/v) sodium deoxycholate plus 0.2% (w/v) Triton X-100. When chromatographed on DEAE-cellulose, the solubilized activity emerged as a single peak. The enzyme thus obtained was maximally active at pH 4.5, and readily hydrolyzed mixed gangliosides but was less active toward 4-methylumbelliferyl-alpha-N-acetylneuraminic acid, 3'-sialyllactose and fetuin. The corresponding enzyme from MeDAB-induced hepatoma was indistinguishable from the liver enzyme in terms of ease of solubilization, pH-activity relationship, chromatographic behavior and substrate preference. It therefore appears that the plasma membrane-associated sialidase of hepatomas differs from that of liver only in the tissue level of activity.
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PMID:Membrane-associated sialidase of rat liver and its decrease in hepatomas. 312 7

By method of light and electron microscopy the influence of heparin (0.1%) sodium heparinate) on Zajdela ascites hepatoma cells was studied. To provide penetration of heparin into cells, the latter were treated with low-concentration detergent--Triton X-100. Heparin causes increase of cells observable by phase-contrast method, and homogenization of the karyoplasm. When examining stained preparations light-optically, chromatin is distributed evenly and structurelessly over the whole cell. Streaks of DNA-containing material protrude into the cytoplasm. Electronograms show lowering of electron density of cells, transformation of chromatin fibrils into a thin-fibrillar network, vanishing of nucleoli, and appearance of structureless globules about the size of 100 nm. The observed changes are discussed in the light of data on the anti-cancer action of heparin.
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PMID:Structural changes induced by heparin in Zajdela ascites hepatoma cells. 351 12

Glucosidase II is regarded as a resident protein of the endoplasmatic reticulum. The enzyme removes alpha-1-3-linked glucose from high mannose oligosaccharides N-linked to asparagine residues of glycoproteins. Monospecific antibodies raised against the pig kidney enzyme are used to study the metabolism of the enzyme in a rat hepatoma cell line. These antiglucosidase II antibodies specifically immune precipitate glucosidase II as a 100,000-Da species from [35S]methionine-labeled cells. In addition, protein blotting and immune staining of cell extracts from both rat liver and human and rat hepatoma cell lines show identity in apparent Mr (100,000). Glucosidase II synthesized in the presence of tunicamycin is approximately 94,000 Da, indicating the presence of one or more N-linked oligosaccharide chains. Cell-free protein synthesis of rat hepatoma total RNA demonstrates that glucosidase II is synthesized as a slightly higher molecular weight species as compared to the polypeptide synthesized in whole cells in the presence of tunicamycin, indicating that the enzyme has a cleavable signal sequence. Using a pulse-chase protocol, the apparent molecular weight does not change upon longer chase periods. In addition, the 100,000-Da protein remains sensitive to endo-beta-N-acetylglucosaminidase H regardless of prolonged chase periods. The cells incorporate [3H]mannose into the enzyme; after release with endo-beta-N-acetylglucosaminidase H, most of the radioactivity comigrates with Glc1-Man9-GlcNAc on a gel filtration column. Phase separation in Triton X-114 shows a partition between the aqueous and the Triton phase, the major portion being separated in the aqueous phase. In rat hepatoma cells glucosidase II has a half-life of 50 min. This value is not altered if the cells are grown in the presence of monensin nor of methyl-deoxynoijirimycin. However, tunicamycin and low concentrations or primaquine (raising the pH of acidic compartments) causes a 100% increase in half-life of glucosidase II. We conclude that glucosidase II is a hydrophilic, probably not a transmembrane membrane, protein with a short half-life. It is the first example of an oligosaccharide-processing enzyme not being an integral membrane protein.
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PMID:Glucosidase II, a protein of the endoplasmic reticulum with high mannose oligosaccharide chains and a rapid turnover. 354 12

Using a monospecific, monoclonal antibody against the glucocorticoid receptor (GR), an immunocytochemical study was performed to investigate the intracellular localization of GR both in the presence or absence of ligand. With all fixation methods tested (paraformaldehyde, acetic acid in ethanol, Bouin's fixative, and bensochinone in PBS), it was possible to obtain specific GR staining. Fixation with paraformaldehyde was chosen for further studies on the effect of permeabilization, using several concentrations of Triton X-100 or saponin. A rat Rueber hepatoma (H-4-II-E) and a human uterus carcinoma (NHIK 3025) cell line were used as well as cultured hepatocytes from normal rat. The accessibility of the different cell compartments after fixation and permeabilization was tested for by using antibodies against cellular constituents with known locations (i.e. core-nucleosome proteins and tubulin), in combination with the anti-GR antibody in double immunofluorescence staining experiments. The specific GR stain obtained with the indirect peroxidase antiperoxidase technique or with fluorescein isothiocyanate-labeled second antibodies was shown to be present both in the cytoplasm and in the nucleus. Staining of all cellular compartments was abolished (peroxidase antiperoxidase) or diminished (fluorescein isothiocyanate) if the monoclonal antibody was preincubated with a 90% pure GR preparation. These findings are in contrast to recently reported immunocytochemical studies, where a strict nuclear existence of the estrogen and progestin receptors has been reported. Consequently, generalizations with regard to steroid receptor localization cannot be made. Furthermore, an in vitro model is described, where the effect of dexamethasone administration upon the localization of receptor staining in H-4-II-E cells can be studied.
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PMID:Intracellular localization of the glucocorticoid receptor: evidence for cytoplasmic and nuclear localization. 354 55

We investigated the biosynthesis of the human insulin receptor in IM-9 lymphocytes and HEP-G2 hepatoma cells. Cells were first pulse labeled for 15 min with [35S]methionine and then chased for up to 4 h. At each time, the cells were solubilized in 1% Triton X-100; the insulin receptor was immunoprecipitated and then analyzed with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (6%) and fluorography. At 15 min, a major precursor protein of 190,000 Mr was precipitated. During the chase period, two smaller proteins became apparent, which evolved into two major species of 130,000 and 95,000 Mr, the mature alpha- and beta-subunits, respectively. When IM-9 cells were trypsinized after pulse chase, the alpha- and beta-subunits were completely digested, whereas the 190,000-Mr precursor was unaffected. 125I-surface labeling of cells, followed by immunoprecipitation, revealed the presence of only the alpha- and beta-subunits, indicating that only these two species were on the cell surface. To study this biosynthetic pathway, several inhibitors were used (tunicamycin, monensin, and swainsonine). These inhibitors revealed the following. The receptor is first synthesized as a 170,000-Mr protein that is cotranslationally N-glycosylated to yield a high-mannose 190,000-Mr precursor. This precursor is rapidly transported from the endoplasmic reticulum to the Golgi apparatus where it is cleaved into two subunits of 120,000 Mr (alpha) and 90,000 Mr (beta). These subunits then increase in molecular weight by processing of the high-mannose oligosaccharides to the low-mannose complex type. The two subunits then migrate to the cell surface where they function to transmit the insulin signal.
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PMID:Biosynthesis and processing of the human insulin receptor. 372 Oct 67

Transglutaminase activity was reduced in malignant hepatoma, virus-transformed human and hamster cells, and chemically transformed mouse cells when compared to normal counterparts. The reduction in enzyme activity reflected the presence of fewer transglutaminase molecules in transformed cells. Greater amounts of the enzyme activity were particulate-associated in confluent and arrested normal human cells. Indirect immunofluorescence studies with antibody to cellular transglutaminase demonstrated the presence of transglutaminase in Triton X-100-insoluble material. A parallel between pericellular fibronectin and transglutaminase (TGase) was demonstrated. Normal human and mouse cells that elicited contact inhibition of growth and had the high TGase activity also had more epsilon-(gamma-glutamyl) lysine isopeptide bonds than transformed counterparts. Similarly nonproliferating human cells had higher transglutaminase activity and isopeptide levels than did proliferating populations. These results suggest that isopeptide bond formation stabilizes the cell membrane and contributes to a nonproliferating state. Inhibition of isopeptide formation should therefore lead to a mitogenic response. Preliminary results support such a relationship. A model depicting control of isopeptide formation at either enzyme or substrate level is presented.
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PMID:Transglutaminase and epsilon-(gamma-glutamyl) lysine isopeptide bonds in eukaryotic cells. 610 9

Cultured rat hepatoma cells were homogenized and subjected to subcellular fractionation by analytical sucrose density centrifugation to determine the localization of gamma-glutamyltransferase ((5-glutamyl-)-peptide: amino acid 5-glutamyltransferase, EC 2.3.2.2). The activity was exclusively localized to the plasma membrane. Diazotized sulphanilic acid was used as a non-penetrant membrane reagent which inactivates ectoenzymes. With both intact and sonicated cells, only 70-75% inhibition of gamma-glutamyltransferase activity was observed. At least 12% of the total cell complement of gamma-glutamyltransferase activity is highly resistant to inactivation by diazotized sulphanilic acid even after Triton X-100 solubilization. The enzyme was purified from hepatoma cells and its properties compared with enzyme from normal liver. Apart from the striking increase in Vapp there were only minor differences between the enzymes from the two sources. In contrast to the complete abolition of transpeptidase activity of the purified hepatoma enzyme by diazotized sulphanilic acid, the hydrolytic activity of this preparation was only slightly inhibited.
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PMID:Subcellular localization and isolation of gamma-glutamyltransferase from rat hepatoma cells. 611 21

Purified rat liver nuclei were labelled in vitro in the presence of (32P) ATP and submitted to sequential extraction with DNAse, 0.4 or 2.0 M NaCl and Triton X-100. The residual or matrix structures contained 8-10 phosphoproteins between 76 and 260 kd including a triplet of major bands with 110, 117 and 128 kd. The 110 kd species was purified by chromatography on oligo(dT)-cellulose. It was shown to be identical with the 110 kd phosphoprotein of rat liver or Morris hepatoma free polyribosomes using the technique of limited digestion with S. aureus protease V 8.
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PMID:High molecular mass phosphoproteins in the rat liver nuclear matrix identification of a prominent 110,000 Dalton species. 622 86

The effects of several detergents commonly used to solubilize membrane glycoproteins have been investigated on the binding of hepatoma cell surface [3H]-galactoglycoproteins to, and their elution from, concanavalin A or Ricinus communis lectins conjugated to Sepharose 4B. The optimum conditions (pH, ionic strength) in the presence of ionic [sodium deoxycholate (DOC) and sodium dodecyl sulphate (SDS)] and non-ionic detergents (Triton X-100) at a constant concentration were determined in order to ascertain which would yield the better efficiency. The effects of different detergent concentrations on binding and elution were then studied. The range of concentrations for each detergent to be used without modifying efficiency was determined. Triton X-100 and DOC (0.1-1%) did not change the efficiency on Ricinus lectin-Sepharose, whereas SDS, at a concentration greater than 0.05%, caused a dramatic decrease in efficiency. On concanavalin A-Sepharose, by contrast, the non-ionic detergent had no effect on the efficiency at all the concentrations tested (0.1-1%), while concentrations of more than 0.5% DOC and 0.1% SDS significantly decreased both binding and elution.
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PMID:Interactions of insolubilized lectins with membrane glycoproteins in presence of detergents. 625 Nov 1


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