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)

Co-secretion of plasminogen activator inhibitor type 1 (PAI-1) and urokinase-type plasminogen activator was identified in short-term cultures of primary type II pneumocytes isolated from adult rats. After separation by sodium dodecyl sulfate (SDS)-PAGE and reverse fibrin autography (reverse FA) of serum-free conditioned medium (SFCM), cellular lysate, and extracellular matrix (ECM), the inhibitor was seen as a zone of spared lysis at an apparent molecular mass of 46 to 48 kD. The plasminogen activator (PA) activity could only be visualized when human instead of bovine fibrin was used in the indicator gel. It presented as a single band of lysis at an apparent molecular mass of 45 kD when tested by regular FA and was found adjacent to PAI-1 when examined by reverse FA. Immunoblot analysis of type II pneumocyte SFCM, cellular lysate, and ECM revealed two bands at 46 and 48 kD, consistent with the apparent molecular masses (Mr) reported for rat PAI-1 from HTC hepatoma cells. Type II pneumocyte PAI-1 formed SDS-resistant complexes with tissue-type and urokinase-type plasminogen activator and was found to be stable to acid, to short-term exposure to heat, and to the denaturants guanidine HCl and SDS, while being sensitive to treatment with alkali and urea. When levels of type II pneumocyte PAI-1 activity were monitored over time during short-term culture conditions, the level of PAI-1 in SFCM remained stable, whereas activity in the lysate accumulated and activity in the ECM declined.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Plasminogen activator inhibitor type 1 production by rat type II pneumocytes in culture. 154 Mar 77

Human interleukin-6 or B-cell stimulatory factor-2 is a cytokine involved in acute phase and immune response. Cloning of cDNA for human interleukin-6 in the pT7.7 expression plasmid under the control of a bacteriophage T7 RNA polymerase promoter system allows rapid production of the cytokine in Escherichia coli. Upon cell induction with isopropyl thiogalactopyranoside, recombinant human interleukin-6 is overexpressed and forms insoluble inclusion bodies. Solubilization of the protein with 6 M guanidine hydrochloride and refolding in the presence of a reduction/oxidation system results in a quantitative recovery of recombinant human interleukin-6. This material is already 70% pure and can be further purified to homogeneity with a single passage over a weak anionic-exchange column. Extended structural characterization of the purified protein by electrospray mass spectrometry, automated Edman degradation and peptide mapping by high-pressure liquid chromatography/fast-atom-bombardment mass spectrometry demonstrates that recombinant human interleukin-6 is identical to the natural protein both in amino acid sequence and S-S bridge content. However, it contains a minor component accounting for about 20% of the entire translated protein which exhibits a Met-Ala dipeptide extension at the N-terminus. Purified recombinant human interleukin-6 is biologically active because it is able to induce at least 70-fold the human C-reactive promoter transfected in human hepatoma Hep 3B cells and is stable for several months in 10% glycerol at 4 degrees C. The expression system described in the present work has the main advantage of producing a high yield of recombinant human interleukin-6 (about 25 mg/l) combined with a very simple purification scheme.
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PMID:Single-step purification and structural characterization of human interleukin-6 produced in Escherichia coli from a T7 RNA polymerase expression vector. 205 Jan 35

Previous studies in this laboratory suggested that in adult liver, either the gene for the tumor-type poly(A) polymerase is poorly transcribed or the mRNA for this enzyme is largely not expressed. To test these possibilities, total RNA from rat liver and Morris hepatoma 3924A RNA were isolated by using a guanidine thiocyanate method; poly(A+) RNA and poly(A-) RNA were separated by oligo(dT)-cellulose chromatography and used for translation in a rabbit reticulocyte lysate system. After in vitro translation, the products were immunoprecipitated with either purified anti-tumor poly(A) polymerase antibodies or control immunoglobulins. When the polypeptides translated from poly(A+) or poly(A-) hepatoma RNA were precipitated with immune sera, a unique [35S]methionine-labeled 35-kilodalton (kDa) protein was observed. This band was not apparent when control serum was used for the immunoprecipitation. The radiolabeled 35-kDa polypeptide was not evident when the products were incubated with highly purified tumor nuclear poly(A) polymerase prior to immunoprecipitation. Prior incubation of the translation products with bovine serum albumin instead of poly(A) polymerase had no effect on the immunoprecipitation. This 35-kDa protein was not apparent when liver poly(A+) RNA was used to direct translation. These data demonstrate that (a) the tumor enzyme is not synthesized as a precursor, (b) tumor mRNA, but not normal liver mRNA, contains detectable sequences coding for tumor-type poly(A) polymerase, and (c) poly(A) polymerase mRNA also exists as a poly(A-) population.
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PMID:Cell-free synthesis of tumor-type poly(A) polymerase. 242 13

A protein preparation that specifically binds insulin-like growth factors (IGFs) I and II was purified from medium conditioned by rat liver BRL-3A cells using molecular sieve chromatography in 1 M acetic acid followed by affinity chromatography on IGF-II-agarose. The affinity-purified IGF-binding protein exhibits a single major band with apparent Mr = 36,300 under reducing conditions on sodium dodecyl sulfate-polyacrylamide gels. The IGF-binding protein is efficiently and specifically cross-linked to either 125I-IGF-I (human) or 125I-IGF-II (rat) using disuccinimidyl suberate. An IGF-binding protein of similar apparent molecular weight was also affinity purified from rat hepatoma H-35 cell conditioned medium and found to differ from the BRL-3A protein such that potent polyclonal antisera prepared in rabbits against the purified BRL-3A IGF-binding protein exhibited a much lower titer for the H-35 protein in an enzyme-linked immunosorbent assay and upon immunoblotting. In order to determine whether a single BRL-3A IGF-binding protein is present in the affinity-purified preparation, the protein was prepared for sequencing on a Sephacryl S-300 column in 6 M guanidine HCl after reduction and alkylation. The amino acid composition (expressed in percentages) of this IGF-binding protein was determined to be: Cys = 5.5, Lys = 4.8, His = 2.8, Arg = 7.8, Asx = 10.2, Thr = 5.1, Ser = 3.9, Glx = 15.7, Gly = 17.4, Ala = 7.3, Val = 4.6, Met = 1.4, Ile = 2.4, Leu = 8.3, Tyr = 1.0, Phe = 1.9. Sequencing of the NH2-terminal portion of this protein led to the identification of 31 amino acids in the following order: Phe-Arg-Cys-Pro-Pro-Cys-Thr-Pro-Glu-Arg-Leu-Ala-Ala-Cys-Gly-Pro-Pro-Pro- Asp-Ala-Pro-Cys-Ala-Glu-Leu-Val-Arg-Glu-Pro-Gly-Cys. We conclude that rat liver BRL-3A cells secrete a single major IGF-binding protein capable of binding both IGF-I and IGF-II.
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PMID:Purification and amino-terminal sequence of an insulin-like growth factor-binding protein secreted by rat liver BRL-3A cells. 242 67

The ability of cultured normal human fetal liver and kidney epithelial cells to repair the premutagenic and precarcinogenic O6-methylguanine (O6-MeGua) DNA adduct was determined by directly monitoring its loss in cellular DNA and quantitating the number of O6-MeGua-DNA-methyltransferase (O6-MT) molecules per cell. Following treatment of the epithelial cells with the direct acting carcinogen N-methyl-N-nitrosourea (MNU), the loss of the O6-MeGua adduct was biphasic, exhibiting a half-life of 2.0 and 1.5 h in the liver and kidney cells, respectively. The activity of O6-MT in the liver and kidney epithelial cells in culture was 0.19 pmol/mg protein or 18,500 molecules/cell. The activity of O6-MT was maintained throughout the life of the cultures, i.e., 20 subpassages or 50 cumulative population doublings for the liver and kidney. In order to ascertain whether human fetal epithelial cells exhibit an induction of O6-MT, the cell cultures were treated with single and multiple conditioning doses of N-methyl-N-nitro-N-nitroso-guanidine (MNNG) or gamma-irradiated and assayed for the amount of O6-MT. A 1 h exposure of cells to 2, 4, and 8 microM MNNG resulted in an 80-100% decrease of the initial O6-MT activity which was restored to the constitutive levels within 48 and 72 h post-treatment. Rat hepatoma cells, used as a positive control, increased their levels of O6-MT to 2.8-fold the constitutive levels following treatment with MNNG. Treatment of the human liver and kidney epithelial cells with chronic low doses of MNNG exhibited O6-MT levels identical to untreated cells. The O6-MT activity in epithelial cells remained unaffected upon pre-irradiation with 1.2 or 2.5 Gy of gamma-irradiation.
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PMID:Absence of DNA damage-mediated induction of human methyltransferase specific for precarcinogenic O6-methylguanine. 257 88

We have constructed and analyzed amino terminally deleted analogs of IL-6. Progressively shortened variants of mature IL-6 were constructed at the cDNA level and expressed in Escherichia coli. Mutant proteins were recovered from refractile bodies by solubilizing in 6 M guanidine-HCl. The mutant protein concentration in these preparations was estimated by Western blotting by using an IL-6-specific mAb and the biologic activity was measured in the B9 (hybridoma growth factor) assay. The first 28 amino acids of mature IL-6 could be removed without significantly affecting biologic activity. A further removal of amino acids 29 and 30 resulted in an approximately 50-fold decrease, whereas removal of amino acids 31 to 34 virtually abolished the activity. The mutants showed the same reaction pattern in three other IL-6 assays: induction of murine thymocyte proliferation, induction of fibrinogen synthesis by a human hepatoma cell line (HepG2), and the induction of IgM synthesis by an EBV-transformed B cell line. This suggests that a single functional domain might be responsible for all four activities of IL-6.
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PMID:Analysis of human IL-6 mutants expressed in Escherichia coli. Biologic activities are not affected by deletion of amino acids 1-28. 266 92

Incubation of HTC rat hepatoma cells with the synthetic glucocorticoid dexamethasone rapidly inhibits tissue-type plasminogen activator activity by inducing a specific plasminogen activator-inhibitor (PAI-1). Using immobilized polyclonal antibodies raised against HT-1080 human fibrosarcoma PAI-1, we have purified HTC PAI-1 from serum-free medium conditioned by dexamethasone-treated HTC hepatoma cells and shown it to be antigenically related to human PAI-1. Greater than 100-fold purification with greater than 75% yield was achieved in a single step. The purified PAI-1 migrates on SDS-polyacrylamide gels as a single major band of 49 kDa with a minor band of 46 kDa. Digestion of PAI-1 with endoglycosidase F causes a shift toward faster migrating species which retain inhibitory activity. The purified PAI-1 was stable at pH 2.5, lost 50% of its activity after 15 min at 45 degrees C, and showed marked activation after treatment with SDS or guanidine-HCl. Purified PAI-1 rapidly inhibited and formed complexes with both tissue-type and urokinase-type plasminogen activators. Polyclonal rabbit antirat PAI-1 antibodies were raised which immunoprecipitate both free and complexed PAI-1.
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PMID:Immunoaffinity purification of HTC rat hepatoma cell plasminogen activator-inhibitor-1. 312 13

In primary monolayer cultures of mature rat hepatocytes, cell growth and hepatocyte-specific functions were mediated by a cell surface component (named the cell surface modulator) via cell-cell contact. The modulator activity was heat-labile and trypsin-sensitive. Activity was also found in plasma membranes from kidney, brain, lung, and erythrocytes. The modulator was solubilized by 4% octylglucoside plus 4M guanidine HCl from liver membranes. The molecular weight of the modulator was 670KD determined by Sephacryl S-400 gel filtration. Hepatoma cells established from Reuber and Morris hepatoma did not show any cell density-dependency on either cell growth or hepatocyte-specific function. However, these hepatoma cells had strong cell surface modulator activity. These results suggest that hepatoma cells have lost their cell density-dependent regulation because they have lost the ability to respond to the cell surface modulator. Characterization of the cell surface modulator and its mechanism of transmitting a signal for gene regulation would be helpful in understanding the process by which cells assemble into tissues in vivo and the mechanism of changes in gene expression in tissue during development, regeneration and carcinogenesis.
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PMID:[Reciprocal regulation of growth and differentiation in hepatocytes by cell surface modulator and loss of regulation during carcinogenesis]. 398 39

The nuclear matrix of Zajdela hepatoma cells, in which DNA synthesis was blocked by novobiocin, contained 2.5-3.0 times more DNA and protein not dissociating in 2 M NaCl than the nuclear matrix of control cells. Chromatography of nuclear matrix preparations on Sepharose 2B-CL resulted in isolation of tightly bound DNA-protein complexes which did not dissociate in 8 M urea or 0.1% SDS. Subsequent elution of DNA-protein complexes on a hydroxylapatite column with a buffer containing 4 M guanidine hydrochloride and 5 M urea caused partial dissociation of the complexes. Electrophoretic analysis revealed essential changes in the composition of proteins DNA-protein complexes of hepatoma cells nuclear matrix during inhibition of DNA synthesis.
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PMID:[Reorganization of the protein composition of the nuclear matrix of hepatoma cells after inhibition of DNA synthesis with novobiocin]. 407 85

Replication of hepatitis A virus (HAV) in the human hepatoma-derived PLC/PRF/5 cell line was neither inhibited in the presence of various concentrations of guanidine or D-2-(alpha-hydroxybenzyl)benzimidazole (D-HBB), nor were the two chemicals effective in combination. Under identical conditions, however, replication of poliovirus type 1 was inhibited. Tracer experiments with radiolabelled guanidine and D-HBB also furnished no evidence that the two antiviral substances were metabolized gradually to inactive derivatives in PLC/PRF/5 cells. Therefore, it is concluded that resistance to the action of guanidine and D-HBB is an inherent characteristic of HAV. However, the insensitivity of HAV to these drugs does not exclude the virus from the family of picornaviruses.
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PMID:Failure of guanidine and 2-(alpha-hydroxybenzyl)benzimidazole to inhibit replication of hepatitis A virus in vitro. 628 46


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