amanitins and Carcinoma--Hepatocellular

The enzyme glutamine synthetase (GS) ranks as one of the most remarkable glucocorticoid-inducible vertebrate genes. However, little is known about the responsible DNA elements and the mode of glucocorticoid action. This is especially the case for the induction of GS in hepatoma cells. In the work presented, the rat hepatoma cell line FAO was used as a model to study the induction of GS under the influence of glucocorticoids. FAO cells do not show GS activity in the absence of glucocorticoids and are strongly responding to their presence. Analyzing sequences of several thousand base pairs upstream and downstream from the transcriptional start point of the GS gene, a glucocorticoid responsible element was identified within the first intron of the gene. However, evidence is presented that aside from a primary effect on transcription glucocorticoids mediate their effect on the expression of GS also at the posttranscriptional level.

Topics: Amanitins; Animals; Blotting, Northern; Carcinoma, Hepatocellular; Cytoplasm; Dexamethasone; DNA; Genes, Reporter; Glucocorticoids; Glutamate-Ammonia Ligase; Humans; Introns; Liver Neoplasms; Male; Plasmids; Rats; Rats, Sprague-Dawley; RNA Processing, Post-Transcriptional; RNA, Messenger; Time Factors; Transcription, Genetic; Transfection; Tumor Cells, Cultured

To examine the role of hepatitis delta virus antigen in the replication of hepatitis delta virus RNA, we have transfected a stable HDAg-positive cell line (A3) and the parental HDAg-negative line (HepG2) with HDV RNA produced in vitro; synthesis of complementary HDV RNA was only detected in HDAg-positive cultures. In contrast, nuclear homogenates from both HDAg-positive and -negative cells synthesized comparable levels of complementary RNA from exogenous HDV RNA. These findings indicate that HDAg is not a necessary component of the transcriptional complex and suggest with other evidence, that a major role for HDAg is likely to be transport of HDV RNA from cytoplasm to nucleus. Transcription of HDV RNA by intact nuclei was sensitive to 1 microgram/ml alpha-amanatin providing firm evidence that, like viroids, this function is performed by host RNA polymerase II.

Topics: Amanitins; Antigens, Viral; Carcinoma, Hepatocellular; Cell Line; Cell Transformation, Viral; Hepatitis delta Antigens; Hepatitis Delta Virus; Humans; Liver Neoplasms; RNA Polymerase II; RNA, Viral; Transcription, Genetic; Transfection

The hormone-responsive enzymes tyrosine aminotransferase and glycerol-3-phosphate dehydrogenase were studied with respect to current models of the mechanism of glucocorticoid/cAMP interaction during the induction of enzyme activity in responsive cell hybrids between rat C6 glioma cells and rat FU5AH hepatoma cells. The results of experiments involving protein and mRNA synthesis inhibitors, sequential addition of inducers, and the assay of cyclic-AMP-dependent protein kinase could not be adequately explained by any one model of inducer interaction. Comparison of the hybrid clones revealed the presence of factors that may modify induction but that are not essential for synergistic induction.

Topics: Amanitins; Animals; Bucladesine; Carcinoma, Hepatocellular; Cycloheximide; Drug Synergism; Enzyme Induction; Glioma; Glycerolphosphate Dehydrogenase; Hybrid Cells; Hydrocortisone; Kinetics; Liver Neoplasms; Rats; Tyrosine Transaminase

The effect of Ca2+ antagonists (CA) on the receptor-mediated low density lipoprotein pathway has been investigated "in vitro" in human skin fibroblasts (HSF) and in human hepatoma cell line Hep G2. The specific binding and internalization of human 125I-labeled LDL are dose-dependently increased in HSF by CA of the verapamil series (verapamil, anipamil, gallopamil, ronipamil, and diltiazem), but neither by CA of the dihydropyridine series (nifedipine, nitrendipine) nor by flunarizine. BAY K 8644, a Ca2+ agonist, elicited an opposite effect. In the presence of the tested CA, LDL degradation is either unaffected (lower concentrations) or inhibited (higher concentrations). 125I-LDL uptake is stimulated also in fibroblasts from type IIa hypercholesterolemic patients, heterozygous for defective expression of LDL receptor. The enhanced cellular uptake of 125I-LDL was prevented by cycloheximide and by alpha-amanitin. CA of the verapamil series including diltiazem retained their effect in human hepatoma cell line Hep G2, a model proposed for hepatic metabolism of LDL. Our studies show that a) CA stimulate the high affinity binding and internalization of LDL in HSF and in human hepatoma cell line Hep G2; b) this stimulation involves DNA transcription and new protein synthesis; c) this effect is specific to one subgroup of Ca2+ antagonists (the verapamil class only).

Topics: Amanitins; Biotransformation; Calcium Channel Blockers; Carcinoma, Hepatocellular; Cell Line; Cycloheximide; Diltiazem; Fibroblasts; Humans; Lipoproteins; Lipoproteins, LDL; Liver Neoplasms; Papaverine; Receptors, LDL; Verapamil

Exposure of cultured human hepatoma cells (Hep G2) to medium containing 2% (v/v) dimethyl sulphoxide resulted in an approximate doubling in the activity of delta-aminolaevulinate dehydratase, an increase in the haem content and a decreased growth rate; induced enzyme activity was decrease by 50% after treatment with alpha-amanitin. The findings are strikingly similar to those seen in murine Friend-virus-transformed erythroleukaemia cells.

Topics: Amanitins; Carcinoma, Hepatocellular; Cell Line; Dimethyl Sulfoxide; Heme; Humans; Liver; Liver Neoplasms; Porphobilinogen Synthase; Transcription, Genetic

Topics: Amanitins; Animals; Carcinoma 256, Walker; Carcinoma, Hepatocellular; DNA; DNA-Directed RNA Polymerases; Kidney; Kinetics; Liver Neoplasms; Magnesium; Manganese; Multiple Myeloma; Neoplasms, Experimental; Organ Specificity; Rats; Sarcoma, Yoshida; Spleen

Studies were made on the effects of alpha-amanitin, cycloheximide, and thioacetamide on synthesis and content of low molecular weight nuclear RNA. Cycloheximide, an inhibitor of protein synthesis and the synthesis of 45S pre-rRNA and 5S RNA, also inhibited synthesis of nuclear U1 and U3 RNAs. alpha-Amanitin, an inhibited the synthesis of U1 and U2 low molecular weight nuclear RNA. Thioacetamide, which induces nucleolar hypertrophy and increased nucleolar RNA polymerase activity, markedly increased synthesis of 5.8S RNA and U3 RNA. These results show that syntheses of individual low molecular weight nuclear (LMWN) RNAs are controlled by different regulatory mechanisms. In particular, there appears to be a specific relationship between U3 RNA and functional states of the nucleolus.

Topics: Acetamides; Amanitins; Animals; Carcinoma, Hepatocellular; Cell Nucleus; Cycloheximide; Dose-Response Relationship, Drug; Liver; Liver Neoplasms; Male; Molecular Weight; Neoplasms, Experimental; Phosphates; Rats; RNA; RNA, Neoplasm; Thioacetamide

Rat hepatocytes and AS-30 D hepatoma cells were treated in vitro with phallolysin, the toxic haemolysin from the death cap fungus Amanita phalloides. Scanning electron microscopy revealed dose-dependent formation of bulky protrusions on the cell surface, which later on burst, expelling intracellular material into the surrounding medium, while the cells became stainable with trypan blue. After 1 haemolytic unit (HU)/ml the protrusions burst within 10 min; after 3 HU/ml all cells were destroyed within 3-5 min. Susceptibility of hepatoma cells did not differ from that of hepatocytes.

Topics: Amanitins; Animals; Carcinoma, Hepatocellular; Female; In Vitro Techniques; Liver; Liver Neoplasms; Microscopy, Electron, Scanning; Neoplasms, Experimental; Rats