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S S Thorgeirsson

Publications and source records attributed to S S Thorgeirsson.

At least 19 recordsLinked to original sources
PubMed1992

Regulation of P-glycoprotein gene expression in hepatocyte cultures and liver cell lines by a trans-acting transcriptional repressor.

Previously we have demonstrated that expression of the multidrug resistance (mdr) genes in rat liver and primary rat hepatocyte cultures is induced by exposure to 2-acetylaminofluorene and 3-methylcholanthrene. The mdr expression induced by both of these compounds occurs primarily via increased gene transcription. To determine the nature of possible regulatory proteins involved in mdr gene regulation we inhibited protein synthesis using cycloheximide or emetine in primary rat hepatocyte cultures, mouse (HePa 1), human (Hep G2) and rat (H4-II-E) cell lines. Each cell type responded by strongly increasing its steady state mdr1 mRNA levels. In hepatocytes increased mdr expression was observed after greater than 50% inhibition of protein synthesis, and was first detected after 2h of protein synthesis inhibition with maximal induction occurring by 24h. Nuclear run-on analysis showed that the increased steady state mRNA level was due to increased gene transcription without alteration of the transcription start site. Combined these data indicate that one regulatory mechanism by which mdr gene expression is controlled is via a trans-acting transcriptional repressor.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1992

Metabolic activation and genotoxicity of heterocyclic arylamines.

Because of the potential for human exposure to mutagenic and carcinogenic heterocyclic arylamines in the diet, the carcinogenicity of three HAAs, 2-amino-3-methylimidazo[4,5-f]quinoline, 2-amino-3,8-dimethyl-imidazo[4,5-f]quinoxaline, and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine, is being evaluated in nonhuman primates, especially cynomolgus monkeys. Concomitant with the carcinogenicity studies, the metabolic processing, disposition, and DNA-adduct formation of these compounds are being examined in these monkeys. This report highlights the results from studies in monkeys and from in vitro models examining metabolic activation and genotoxicity of HAAs. The extent of in vivo activation of HAAs in monkeys was assessed by measuring DNA adducts in various tissues. Both 2-amino-3-methylimidazo[4,5-f]quinoline and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine form high levels of DNA adducts in a number of organs, particularly the liver, kidney, and heart. The implications of metabolic activation and DNA-adduct formation to the carcinogenicity of HAAs are discussed.

Animals↗
PubMed1992

Stable expression of human cytochrome P450IIE1 in mammalian cells: metabolic activation of nitrosodimethylamine and formation of adducts with cellular DNA.

To introduce cytochrome P450IIE1 DNA stably into the chromosomal DNA of mammalian cells, we constructed recombinant retroviruses containing the full-length complementary DNA for human cytochrome P450IIE1 and a selectable neo gene. Rat and mouse cells were infected with these viruses, and clones expressing the neo marker gene product were selected in G418-containing medium. Analysis of the DNA of the clones by Southern blotting showed that the viral DNA was integrated into the cellular DNA. Enzymatic analysis of the clones showed that the transduced DNA directed the expression of enzymatically active cytochrome P450IIE1. Treatment of the cells with the carcinogen [14C]-nitrosodimethylamine and analysis of the cellular DNA by CsCl equilibrium density gradients showed incorporation of the label into DNA, indicating the formation of covalent adducts with the cell DNA. Construction of recombinant cell lines constitutively expressing cytochrome P450IIE1 provides a permanent source for this enzyme and can aid in the analysis of its catalytic properties, such as the metabolic activation of chemical mutagens/carcinogens, and the consequent cytotoxic and genotoxic effects of these compounds.

3T3 Cells↗
PubMed1992

Low frequency of p53 gene mutation in tumors induced by aflatoxin B1 in nonhuman primates.

Aflatoxin B1 has been suggested as a causative agent for a G to T mutation at codon 249 in the p53 gene in human hepatocellular carcinomas from southern Africa and Qidong in China. To test this hypothesis, nine tumors induced by aflatoxin B1 in nonhuman primates were analyzed for mutations in the p53 gene. These included four hepatocellular carcinomas, two cholangiocarcinomas, a spindle cell carcinoma of the bile duct, a hemangioendothelial sarcoma of the liver, and an osteogenic sarcoma of the tibia. None of the tumors showed changes at the third position of codon 249 by cleavage analysis of the HaeIII enzyme site at codon 249. A point mutation was identified in one hepatocellular carcinoma at the second position of codon 175 (G to T transversion) by sequencing analysis of the four conserved domains (II to V) in the p53 gene. These data suggest that mutations in the p53 gene are not necessary in aflatoxin B1 induced hepatocarcinogenesis in nonhuman primates. The occurrence of mutation in codon 249 of the p53 gene in selective samples of human hepatocellular cancers may indicate involvement of environmental carcinogens other than aflatoxin B1 or that hepatitis B virus-related hepatitis is a prerequisite for aflatoxin B1 induction of G to T transversion in codon 249.

Adenoma, Bile Duct↗
PubMed1992

Occurrence of oval-type cells in hepatitis B virus-associated human hepatocarcinogenesis.

Proliferation of a new population of epithelial cells with distinct structure, as well as cytokeratin and alpha-fetoprotein expression, was observed in nonneoplastic liver tissues from 14 cases (13 hepatitis B virus-positive) of human hepatocellular carcinoma. These cells were characterized by oval nuclei; scant, pale cytoplasm; small cell size; and cross-reaction with a monoclonal antibody against rat oval cells. These putative human oval cells were strongly positive for cytokeratin 19 and displayed considerable heterogeneity in alpha-fetoprotein and albumin expression. The oval cells were most prominent in actively regenerating nodules and in liver tissue surrounding the cancer. Oval cells and transitional types of cells appear to be the principal producers of alpha-fetoprotein in the regenerating liver. Cancer cells positive for cytokeratins 8, 18 and 19 were observed in half the hepatocellular carcinomas studied. The data suggest that a new cell population structurally similar to oval cells seen in early stages of chemical hepatocarcinogenesis in rats is consistently present in regenerating liver lesions associated with human hepatocellular carcinoma. Furthermore, it is possible that the proliferation of these oval-type cells may partly account for the elevation of serum alpha-fetoprotein frequently seen in precancerous stages of hepatitis B virus-associated human hepatocellular carcinoma.

Adult↗
PubMed1992

Expression of transforming growth factor-alpha in regenerating liver and during hepatic differentiation.

Both the level of expression and cellular distribution of transcripts for transforming growth factor-alpha (TGF-alpha) were studied in adult rat liver after partial hepatectomy and during hepatic differentiation in fetal, neonatal, and adult livers by northern blot analysis and in situ hybridization. A marked increase in the expression of TGF-alpha was observed in neonatal livers and in adult livers after partial hepatectomy and during hepatic regeneration following modification of the Solt-Farber protocol. Quantitation of silver grains after in situ hybridization with a TGF-alpha riboprobe revealed a sixfold to eightfold increase in fetal and neonatal hepatocytes. Moreover, the expression of TGF-alpha in the liver 3 wk after birth was still fourfold higher than that of the adult quiescent liver. Both proliferating oval cells and basophilic foci of hepatocytes generated by modification of the Solt-Farber protocol were positive for TGF-alpha transcripts. The level of TGF-alpha transcripts was sixfold higher in the basophilic foci than in the surrounding liver. High concentrations of TGF-alpha transcripts were observed in the oval cells that lined pseudoducts and in the transitional cells proliferating within the ducts. The combination of in situ hybridization and immunocytochemistry using cell-specific antibodies revealed the presence of TGF-alpha transcripts in both oval cells and in perisinusoidal stellate cells. The observation that TGF-alpha transcripts were found both in primitive liver epithelial cells and perisinusoidal stellate cells suggests that this growth factor, in addition to its mitogenic action, may also have other important functions in the liver.

Animals↗
PubMed1992

Homologous and heterologous gap-junctional intercellular communication in v-raf-, v-myc-, and v-raf/v-myc-transduced rat liver epithelial cell lines.

We examined gap-junctional intercellular communication (GJIC) in a series of normal and v-raf-, v-myc-, and v-raf/v-myc-transduced rat liver epithelial (RLE) cell lines using the scrape loading-dye transfer and fluorescence-recovery-after-photobleaching (FRAP) assays. Whereas the normal RLE cell line, the control helper virus-transduced cell line, and the v-myc-transduced cell line all showed excellent GJIC, the v-raf-transduced cell lines displayed decreasing levels of GJIC associated with their increasing tumorigenicity. The v-raf/v-myc-transformed cell lines showed the lowest levels of GJIC and were also the most tumorigenic. Heterologous GJIC of these oncogene-transduced cell lines was also compared with that in the normal RLE cells. A modified FRAP assay, using fluorescent-microbead labelling to identify the oncogene-transduced cell from surrounding normal cells, was used to quantify the heterologous GJIC. The v-raf/v-myc-transformed RLE cells had no heterologous communication with the normal RLE cells, whereas v-raf- and v-myc-transduced cell lines maintained heterologous GJIC. Northern analysis showed that connexin 43 was the only gap-junction protein message expressed in these cell lines; connexin 32 and connexin 26 were not expressed. The levels of connexin 43 mRNA expression were relatively unchanged in all cell lines, suggesting that the reduction in GJIC was primarily at the posttranslational level. These findings suggest that reduction of homologous GJIC in v-raf- and v-raf/v-myc-transformed RLE cells is linked to their tumorigenic potential. Furthermore, the loss of heterologous GJIC, which we observed only in the v-raf/v-myc-transformed cells, might release such cells from the growth-regulating effects of surrounding normal cells, possibly contributing to their enhanced tumorigenic potential.

Actins↗
PubMed1992

Cellular pattern of multidrug-resistance gene expression during chemical hepatocarcinogenesis in the rat.

Increased expression of multidrug-resistance (mdr) gene transcripts and of the encoded protein, P-glycoprotein, is found in many types of tumors. The biological significance of mdr overexpression during the stepwise process of neoplastic development, however, is not well understood. To assess the possible significance of mdr overexpression in carcinogenesis, we examined the cellular distributions of both mdr gene transcripts and P-glycoprotein during hepatocarcinogenesis induced in rats by the Solt-Farber protocol and then compared them to the distributions of the placental form of glutathione S-transferase (GST-P), a known marker of preneoplastic and neoplastic lesions in the liver. In situ hybridization and immunohistochemical techniques were employed. Neither mdr transcripts nor P-glycoprotein was expressed in oval cells that appeared early in the carcinogenic process. GST-P was strongly expressed in the early focal lesions, whereas the levels of mdr transcripts and P-glycoprotein expressed were low and heterogeneous. Expression of mdr transcripts and P-glycoprotein was increased and became more uniform in hyperplastic nodules and carcinomas, although considerable heterogeneity of expression was still found, particularly at the nodular stage. These data suggest that increased expression of mdr is associated with later stages of neoplastic development in the liver. Furthermore, that no chemical treatment of the animals was employed when the expression of mdr was increasing in the preneoplastic and neoplastic lesions suggests that the enhanced mdr expression is intrinsic to the carcinogenic process.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1992

Adenovirus-mediated in vivo gene transfer and expression in normal rat liver.

Replication deficient, recombinant adenovirus (Ad) vectors do not require target cell replication for transfer and expression of exogenous genes and thus may be useful for in vivo gene therapy in hepatocytes. In vitro, primary cultures of rat hepatocytes infected with a recombinant Ad containing a human alpha 1-antitrypsin cDNA (Ad-alpha 1AT) synthesized and secreted human alpha 1AT for 4 weeks. In rats, in vivo intraportal administration of a recombinant Ad containing the E. coli lacZ gene, was followed by expression of beta-galactosidase in hepatocytes 3 days after infection. Intraportal infusion of Ad-alpha 1AT produced detectable serum levels of human alpha 1AT for 4 weeks. Thus, targeted gene expression has been achieved in the liver, albeit at low levels, suggesting that adenovirus vectors may be a useful means for in vivo gene therapy in liver disorders.

Adenoviruses, Human↗
PubMed1992

Expression of acidic fibroblast growth factor in regenerating liver and during hepatic differentiation.

BACKGROUND: Acidic fibroblast growth factor belongs to a family of growth factors that show a high affinity for heparin sulfate proteoglycans. In vitro, it participates in various cellular functions including proliferation, differentiation, angiogenesis, and cell migration, but in vivo, the physiologic role of this growth factor is still not clearly defined. EXPERIMENTAL DESIGN: The level of expression and also cellular distribution of transcripts for acidic fibroblast growth factor (aFGF) were studied in adult rat liver after partial hepatectomy and during hepatic differentiation in fetal, neonatal, and adult livers by Northern analysis and in situ hybridization techniques. RESULTS: After partial hepatectomy a significant increase in the transcripts for aFGF was observed at 24 hours, whereas at 4 and 12 hours after the operation, the level of transcripts were similar to those of sham-operated animals. In the postnatal liver a high level of aFGF expression was present when the most evident transition from 2 to 3 cell thick hepatic cords to normal hepatic structure is taking place (Ogawa K, Medine A, Farber E. Br J Cancer 1979;40: 782-90). In contrast during the prenatal period, when the liver is still a hemopoietic organ and only a small number of sinusoids are present, low level of aFGF transcripts could be found. Animals treated with 2-acetylaminofluorene and partial hepatectomy (Evarts RP, Nagy P, Marsden E, Thorgeirsson SS. Carcinogenesis 1987;8:1737-40) displayed a marked increase in hepatic aFGF transcripts at the peak of proliferation of primitive liver epithelial cells (oval cells) and perisinusoidal stellate cells (Ito cells) in addition to hepatocytes. In situ hybridization combined with immunocytochemistry using oval and Ito cell specific antibodies revealed the presence of transcripts both in oval cells and Ito cells. Basophilic areas composed of small hepatocytes had a 3-fold increase in the level of transcripts as compared with the surrounding hepatocytes. CONCLUSIONS: These experiments demonstrate that the expression of aFGF is highest during the late stages of hepatic morphogenesis in newborn animals as well as during hepatic differentiation in adult liver.

Animals↗
PubMed1992

Pathology of spontaneous and oncogene transformed rat liver epithelial cells and derived tumours in nude mice.

The histological and ultrastructural features of oncogene transformed rat liver epithelial (RLE) cells in culture and spontaneously transformed RLE cells were studied. The tumours produced in nude mice by all the transformed cells were either moderately well differentiated carcinomas or sarcomatous tumours. Epithelial tumours were produced in the RLE cells after transduction of both v-raf and v-myc oncogenes whereas sarcomatous tumours were produced after transduction of v-raf alone. These data suggested that transformation of RLE cells with a single oncogene, v-raf, produced malignant tumours which were consistent with sarcomas while a combination of two oncogenes, v-raf and v-myc, produced an epithelial tumour, consistent with a carcinoma. The effects of these oncogenes on RLE cells indicate that they were able to differentiate and were capable of producing two morphologically distinct tumour types. The possible role of v-myc in switching the sarcomatous lineage to an epithelial tumour lineage is considered to be significant and worthy of further studies. The epithelial tumour produced in RLE cells by combination of v-raf and v-myc is consistent with an embryonal type of hepatoblastoma. The trabecular type of liver cell carcinoma which resulted from spontaneous transformation of RLE cells illustrates the inherent potential of the RLE cell to undergo malignant change and strongly suggests that the RLE cells may be the precursor cells in the development of hepatocellular carcinoma in the rat.

Animals↗
PubMed1979

Genetic aspects of metabolic processing of chemical substances.

The Ah locus in the mouse controls the induction of cytochrome P1-450 and at least 10 associated monooxygenase activities. These enzyme systems metabolically potentiate and detoxify chemical carcinogens, environmental pollutants, drugs, and other chemicals, as well as numerous endogenous substrates. With certain substrates cytochrome P1-450 is known to produce a predominance of reactive intermediates and products that differ from those formed by other cytochrome P-450 species. Numerous conditions in the mouse, including cancer, drug toxicity, and birth defects, are directly associated with the Ah locus, which is considered to be a single gene or a small number of genes.

Acetaminophen↗
PubMed1979

Metabolism and activation of 2-acetylaminofluorene in isolated rat hepatocytes.

The metabolism of 2-acetylaminofluorene (AAF) as well as the activation of AAF to covalently bound and mutagenic intermediates were studied in isolated rat hepatocytes. The cell system readily formed oxidized, deacetylated, and conjugated AAF metabolites. Pretreatments of animals with the inducer beta-naphthoflavone led to increases in phenolic and conjugated as well as covalently protein-bound products. Addition of 4-nitrophenol, a substrate for conjugation, increased the levels of free phenols and inhibited the formation of water-soluble metabolites. At the same time, the rates of covalent protein binding were decreased. Formation of 9-hydroxy-2-acetylaminofluorene could also be demonstrated. The pathway leading to this alicyclic hydroxylated AAF metabolite was not induced by prior beta-naphthoflavone treatment, nor was it inhibited by 4-nitrophenol addition. The cells converted AAF as well as aminofluorene and 2,4-diaminoanisole to mutagenic intermediates which were released into the incubation medium. 2-Aminofluorene was considerably more mutagenic than was AAF in this system. Addition of microsomes increased the mutagenicity of AAF, but not that of 2-aminofluorene or 2,4-diaminoanisole, presumably by deacetylation of N-hydroxy-2-acetylaminofluorene to N-hydroxy-2-aminofluorene.

2-Acetylaminofluorene↗