bluuebunny/crossref_metadata_2025_split huggingface dataset:
The molecular progression of plutonium‐239–induced rat lung carcinogenesis: Ki‐ras expression and activation
Authors: Stegelmeier Bryan L. , Gillett Nancy A. , Rebar Alan H. , Kelly Gregory 
Published: February 2007

AbstractSpecific, transforming point mutations of ras and alterations in ras expression have been associated with many neoplastic processes, and their presence may be pivotal in neoplastic transformation. Our objectives were to evaluate the molecular and genetic alterations of Ki‐ras in preneoplastic foci and neoplasms in the lungs of rats that inhaled 239PuO2 aerosols. Histologically classified pulmonary lesions were evaluated by in vitro nucleic acid amplification, oligonucleotide hybridization, and direct nucleic acid sequencing for activating Ki‐ras point mutations. We evaluated ras expression in similar lesions using immunohistochemistry and in situ hybridization. Specific Ki‐ras point mutations were present in 46% of the radiation‐induced malignant neoplasms. Spontaneous pulmonary neoplasms, which are rare in rats, had similar activating mutations and frequencies (40%). We found similar mutation frequencies in radiation‐induced adenomas and foci of alveolar epithelial hyperplasia. No mutations were identified in normal lung tissue. Ras expression in hyperplastic lesions and neoplasms was similar to that observed in normal pulmonary epithelia. These findings suggest that Ki‐ras activation, not alterations in expression, is an early lesion associated with many radiation‐induced, proliferative pulmonary lesions and that this molecular alteration may be an important component of both radiation‐induced and spontaneous pulmonary carcinogenesis in the rat.
An arylamine acetyltransferase (AT‐I) from syrian golden hamster liver: Cloning, complete nucleotide sequence, and expression in mammalian cells
Authors: Abu‐Zeid Medhat , Nagata Kiyoshi , Miyata Masaaki , Ozawa Shogo , Fukuhara Morio , Yamazoe Yasushi , Kato Ryuichi 
Published: February 2007

AbstractA cDNA clone (designated hamAT101) encoding an arylamine acetyltransferase, AT‐I, was isolated from a hamster liver λgt11 cDNA library using a specific polyclonal antibody raised against AT‐I. The cloned cDNA insert consisted of 1181 nucleotides, including an open reading frame of 870 nucleotides encoding 290 amino acids (Mr 33,503). The isolated cDNA displayed high sequence similarity to those of chicken, rabbit, and human acetyltransferases. In Northern blots, the hamAT101 cDNA probe hybridized to an RNA band of 18S in the livers of both slow and rapid acetylator phenotypes. To confirm that hamAT101 cDNA encodes the monomorphic but not the polymorphic protein, the isolated cDNA was expressed in monkey kidney cells (COS‐1 cells) using the vector p91023(B). A protein with a molecular weight similar to that of AT‐I was detected upon Western blotting in the 9000 × g supernatant from the transfected cells. The activity toward four different substrates of the 9000 × g supernatant was also examined. In agreement with the results of purified AT‐I, the cDNA‐expressed protein exhibited a high capacity for N‐acetylation of 4‐aminoazobenzene and 2‐aminofluorene, and O‐acetylation of 2‐hydroxyamino‐6‐methyldipyrido[1,2‐a:3′,2′‐d]imidazole, whereas no activity was found for the N‐acetylation of p‐aminobenzoic acid. These results, in addition to the RNA blot hybridization, indicate that hamAT101 encodes the hamster acetyltransferase AT‐I.
Analysis of ras gene mutations in rainbow trout liver tumors initiated by aflatoxin B1
Authors: Chang Yung‐Jin , Mathews Catherine , Mangold Kathy , Marien Koenraad , Hendricks Jerry , Bailey George 
Published: February 2007

AbstractThe suspect human hepatocarcinogen aflatoxin B1 (AFB1) is a well‐known potent initiator of hepatic tumors in rainbow trout (Oncorhynchus mykiss). Both hepatocellular carcinomas and mixed hepatocellular/cholangio‐cellular carcinomas are induced by AFB1 in trout, with the mixed form predominating. We previously isolated two c‐ras genes from trout liver cDNA, and in the present study we analyzed DNA from 14 AFB1‐induced trout liver tumors for point mutations in exon 1 of both genes. Using the polymerase chain reaction (PCR) and oligo‐nucleotide hybridization methods, a high proportion (10/14) of the AFB1‐initiated tumor DNAs showed evidence of activating point mutations in the trout c‐Ki‐ras gene. Of the 10 mutant ras genotypes, seven were codon 12 GGA→GTA transversions, two were codon 13 GGT→GTT transversions, and one was a codon 12 GGA→A transition. Nucleotide sequence analysis of cloned PCR products from four of these tumor DNAs provided definitive evidence for two codon 12 GGA→GTA mutations, one codon 12 GGA→A mutation, and one codon 13 GGT→GTT mutation, in complete agreement with the oligonucleotide hybridization results. No mutations were detected in exon 1 of a second trout ras gene also expressed in liver, nor in DNA from control livers. This is the first report of experimentally induced ras gene point mutations in a lower vertebrate fish model. The results indicate that the hepatocarcinogen AFB1 induces c‐Ki‐ras gene mutations in trout similar to those in rat liver tumors.
TGF‐β1 and skin carcinogenesis: Antiproliferative effect in vitro and TGF‐β1 mRNA expression during epidermal hyperproliferation and multistage tumorigenesis
Authors: Krieg Peter , Schnapke Ruben , Fürstenberger Gerhard , Vogt Ingeborg , Marks Friedrich 
Published: February 2007

AbstractThe role of transforming growth factor‐β1 (TGF‐β1) in multistage carcinogenesis in mouse skin was assessed by studying its growth inhibitory effects on nontumorigenic and tumorigenic keratinocytes and by examining its mRNA expression in vitro and during epidermal hyperproliferation and multistage carcinogenesis. While growth of primary basal keratinocytes was inhibited by TGF‐β1 in doses as low as 0.1 ng/mL, the immortal keratinocyte line MCA/3D (“putatively initiated” cells) responded to TGF‐β1 with slightly reduced sensitivity, and the papilloma‐producing keratinocyte line 308 was considerably less sensitive. In contrast, the squamous carcinoma cell line Carc B was completely nonresponsive, and two other tumorigenic cell lines (PDV and PDVC57) were sensitive to growth inhibition by TGF‐β1. Steady‐state levels of TGF‐β1 mRNA were high in all the malignant cell lines and in line 308 papilloma cells, but low in primary basal cells and in the nontumorigenic keratinocyte lines V2, Reb1, and MCA/3D. Our in vivo studies showed that tumor promoters, but not mitogenic or weak hyperplasiogenic agents, were able to induce transient expression of TGF‐β1 mRNA in mouse epidermis. A constitutive overexpression of TGF‐β1 mRNA was observed in malignant carcinomas but not in the benign premalignant lesions, indicating that overexpression may be associated with malignant progression.
Mutational specificities of environmental carcinogens in the lacl gene of Escherichia coli H. V: DNA sequence analysis of mutations in bacteria recovered from the liver of swiss mice exposed to 1,2‐dimethylhydrazine, azoxymethane, and methylazoxymethanolacetate
Authors: Zeilmaker Marco J. , Horsfall Michael J. , van Helten José B. M. , Glickman Barry W. , Mohn Georges R. 
Published: February 2007

AbstractThe host‐mediated assay (HMA) was used to determine the spectra of mutations induced in the lacl gene of Escherichia coli cells recovered from the livers of Swiss mice exposed to the carcinogens 1,2‐dimethylhydrazine (SDMH), azoxymethane (AOM), and methylazoxymethanolacetate (MAMA). These spectra were further compared with changes induced by dimethylnitrosamine (DMNA) in the HMA methodology. A total of 177 independent lacl mutations arising in the HMA following exposure to SDMH, AOM, and MAMA were analyzed. Single‐base substitutions accounted for 97% of all mutations analyzed. The vast majority of the single‐base substitutions consisted of G:C→ A:T transitions (94% of all mutations). The remaining mutations consisted of A:T →G:C transitions (3% of all mutations) while non‐base substitutions accounted for only 3% of the total mutagenesis. The latter mutations consisted of one frameshift mutation and four lacO deletions. The distribution of G:C→ A:T transitions induced by the three chemicals in the first 200 bp of the lacO gene was not random, but rather clustered at sites where a target guanine was flanked at the 5′ site by a purine residue.
Altered Regulation of TGF‐β1 and TGF‐α in Primary Keratinocytes and Papillomas Expressing v‐Ha‐ras
Authors: Glick Adam B. , Sporn Michael B. , Yuspa Stuart H. 
Published: February 2007

AbstractThe influence of an oncogenic v‐Ha‐ras gene on the expression of TGF‐β and TGF‐α by mouse keratinocytes and derived tumors has been investigated. Normal mouse keratinocytes cultured as basal cells in 0.05 mM Ca2+ secreted low levels of TGF‐β2 peptide, and this increased markedly following culture in 1.4 mM Ca2+, retinoic acid, or phorbol esters. In contrast, introduction of a v‐Ha‐ras gene into normal keratinocytes increased basal expression and secretion of TGF‐β1 (rather than TGF‐β2) in response to all three agents. The selective secretion of TGF‐β1 in v‐Ha‐ras keratinocytes in response to 1.4 mM Ca2+ occurred even though the four TGF‐β2 transcripts were induced and the TGF‐β1 transcript decreased, suggesting that the activated v‐Ha‐ras gene product regulates expression of the TGF‐β isoforms at the posttranscriptional level. Immunohistochemical analysis of papillomas formed following skin grafting of v‐Ha‐ras keratinocytes onto nude mice indicated that TGF‐β1 was abundant in the basal and spinous layers, while there was no expression of TGF‐β1 in normal skin. In contrast, both normal and neoplastic tissues expressed TGF‐β2 and TGF‐β3 in the granular layers. Furthermore, TGF‐α mRNA expression was also elevated fivefold in cultured v‐Ha‐ras keratinocytes, and TGF‐α protein was overexpressed in the grafted papillomas, but there was no detectable expression in normal skin. Elevated expression of both TGF‐β1 and TGF‐α in the basal and spinous layers of benign tumors may be important for the high proliferation rate in these tumors as well as for increased proliferation in the suprabasal layer.
Morphological transformation of 10T1/2 mouse embryo cells can be initiated by DNA double‐strand breaks alone
Authors: Borek Carmia , Ong Augustus , Morgan William F. , Cleaver James E. 
Published: February 2007

AbstractMalignant transformation of mouse fibroblasts was produced by electroporation with restriction enzymes. Similar transformation frequencies were observed with Pstl, Pvull, and Xbal, which cut genomic DNA at similar overall frequencies but have different termini, i.e., a 3′ overhang, a blunt end, and a 5′ overhang, respectively. The dose‐response curve for restriction enzyme transformation shows a marked plateau in frequencies of transformed foci per surviving cell, whereas x‐irradiation of the same cells gives a linear dose‐response curve. Evidently, transformation can be caused by DNA double‐strand breaks alone at a limited number of sites, but the evidence from x rays suggests that other kinds of DNA damage can cause transformation independently.
The use of rabbit polyclonal antibodies for the isolation of carcinogen‐adducted DNA by immunoprecipitation
Authors: Brylawski Bruna P. , Cordeiro‐Stone Marila , Kaufman David G. 
Published: February 2007

AbstractPolyclonal rabbit antibodies elicited against DNA with high levels of (±) 7r, 8t‐dihydroxy‐9t,10t‐epoxy‐7,8,9,10‐tetrahydrobenzo[a]pyrene (BPDE‐I) adducts were used to isolate DNA fragments modified by this carcinogen. DNA treated in vitro with different concentrations of BPDE‐I was used as substrate in double‐antibody immuno‐precipitation reactions. The IgG fraction from immune rabbit serum (primary antibody) was reacted with single‐stranded plasmid DNA bearing BPDE‐I adducts, and the complexes were immunoprecipitated using goat antirabbit‐IgG as secondary antibody. DNA was isolated from the immunoprecipitated pellet, blotted onto nitrocellulose or nylon, and hybridized with 32P‐labeled sequences homologous to a fragment of the plasmid DNA used in the assay. The recovery of both DNA and adducts in the immunoprecipitated pellet increased with the level of carcinogen adduction of the DNA. The immunoprecipitation reaction appeared to be more efficient for fragments of DNA containing a high number of adducts. The amount of 32P‐hybridizing material recovered by immunoprecipitation was virtually identical to the amount added to the reaction in DNA samples that contained three adducts per 103 nucleotides.
Low‐level β1 protein kinase C expression in cloned rat embryo fibroblast cells enhances transformation induced by the adenovirus type 5 E1A gene
Authors: Su Zao‐Zhong , Duigou Gregory J. , Fisher Paul B. 
Published: February 2007

AbstractExpression of the E1A gene of adenovirus type 5 (Ad5) in a cloned rat embryo fibroblast (CREF) cell line results in morphological transformation. The efficiency of E1A‐mediated transformation of CREF cells is increased if a wild‐type Ad5 E1A gene is cotransfected with a rat β1 protein kinase C (β1 PKC) gene. A direct demonstration of complementation between a functional‐transforming Ad5 E1A gene and β1 in inducing transformation was demonstrated using Ad5 E1A cold‐sensitive mutant (E1ACS) genes. The E1Acs gene enhanced transformation only at the transformation‐permissive temperature of 37° C and not at the nonpermissive transforming temperature of 32° C. CREF cells constitutively expressing low levels of β1, PKC mRNA were transformed at a higher frequency than parental CREF cells after transfection with an Ad5 E1A gene or infection with wild‐type Ad5 or the Ad5 host‐range cold‐sensitive mutant H5hr1. There was no enhancement of transformation in low‐level β1 PKC‐expressing CREF cells when cultures were grown continuously in the presence of the PKC‐inhibitor 1‐(5‐isoquinolynsulfonyl)‐2‐methylpiperazine dihydrochloride. Transfected CREF cells expressing low levels of β1 PKC mRNA displayed CREF‐like morphology and did not form colonies when grown in agar. In contrast, retroviral vector‐transformed CREF cells expressing high levels of β1 PKC mRNA and β1 PKC enzyme activity were morphologically transformed and grew efficiently in agar. These findings indicate that the β1, PKC gene, when expressed at low levels, can cooperate with the Ad5 E1A gene in the initiation of viral oncogene‐mediated transformation.
Chromatin and cell surface receptors mediate melanoma cell growth response to nerve growth factor
Authors: Rakowicz‐Szulczynska Ewa M. , Reddy Usha , Vorbrodt Andrzej , Herlyn Dorothee , Koprowski Hilary 
Published: February 2007

AbstractGrowth response to nerve growth factor (NGF) was tested in the primary melanoma cell line WM 164, which expressed a low level of NGF cell‐surface receptor, and in WM 164 cells transfected with cDNA for the cell‐surface receptor (TrWM 164 cells), which expressed a higher level of the cell‐surface receptor. Neither cell line expressed the chromatin receptor for NGF or internalized NGF. Both cell lines were stimulated to growth by NGF. After 10 d of exposure to NGF, a 230 000 Mr chromatin protein (receptor) was induced in both cell lines; as a result, NGF bound to the chromatin, and ribosomal RNA synthesis and cell proliferation were inhibited. We suggest that the cell‐surface and chromatin receptors each mediate a different function of NGF.
Spectrum of cis‐diamminedichloroplatinum(II)‐induced mutations in a shuttle vector propagated in human cells
Authors: Bubley Glenn J. , Ashburner Brian P. , Teicher Beverly A. 
Published: February 2007

AbstractThe supF gene of the shuttle vector pZ189 was used as a target for the study of mutations induced by cis‐diamminedichloroplatinum(II) (cis‐DDP). Normal human repair‐proficient fibroblasts and cis‐DDP repair‐deficient xeroderma pigmentosum (XP) cells were used as host cells to study the effect of cis‐DDP on the inhibition of shuttle vector replication and mutagenesis. Transfection of cis‐DDP‐treated pZ189 into normal and XP cell lines resulted in a marked increase in the mutation frequency and a decrease in the replication efficiency of the vector. However, these effects were much greater for the plasmid propagated in XP cells. Atomic absorption spectroscopy showed that six to eight Pt‐DNA adducts per plasmid were necessary to inhibit plasmid replication by 50% in normal cells. In contrast, only one to two Pt‐DNA adducts were necessary to inhibit replication of the plasmid by 50% in XP cells. Analysis of mutation sites demonstrated that cis‐DDP treatment resulted primarily in single and double mutations separated by one base and limited to a few locations within the 85‐bp mature tRNA. Propagation of the cis‐DDP‐treated vector in either normal or XP cells led to predominantly transversion mutations at AGA, AGG, and GAG sites and a cis‐DDP‐associated deletion of 174 bp. Although mutations occurred at target sites for cis‐DDP adduct formation, there was no correlation between sites of mutation and the most frequent sites of adduct formation.
Cytochrome P4501A2 constitutively expressed from transduced DNA mediates metabolic activation and DNA‐adduct formation of aromatic amine carcinogens in NIH 3T3 cells
Authors: Battula Narayana , Schut Herman A. J. , Thorgeirsson Snorri S. 
Published: February 2007

AbstractWe transduced mouse cytochrome P4501A2 DNA into NIH 3T3 cells by retrovirus‐mediated gene transfer. The capacity of the transduced cytochrome P4501A2 for metabolic activation and DNA‐carcinogen adduct formation of aromatic amine carcinogens was investigated. Clones of NIH 3T3 cells that constitutively express cytochrome P4501A2 and controls were exposed to a prototype food‐derived carcinogenic heterocyclic amine, 2‐amino‐3‐methylimidazo[4,5‐f]quinoline (IQ), and an aromatic amine, 2‐acetylaminofluorene (AAF), and their genomic DNAs were analyzed for adducts by 32P‐postlabeling assays. Kinetic analysis of DNA‐carcinogen adducts indicated that adduct formation was dependent on the level of the enzyme, the dose of carcinogen, and the duration of exposure. Addition of 7,8‐benzoflavone, an inhibitor of P4501A2, blocked both the enzyme activity and DNA‐adduct formation, indicating the specific role of P4501A2 in metabolic activation and adduct formation. Three specific IQ‐DNA adducts were detected in cells expressing P4501A2. Fingerprints of the in situ DNA adducts were similar to those of the in vivo adducts in rodent hepatic DNA after the administration of IQ. A single AAF‐DNA adduct was observed in cells exposed to AAF, but other minor adducts were also detected in vivo. These results show that cells expressing constitutive levels of single cytochrome P450s provide an excellent in situ model system for analyzing the catalytic specificity, metabolic activation, and genotoxicity of putative toxic, mutagenic, and carcinogenic substances.
Activation of Ki‐ras gene by point mutation in human liver angiosarcoma associated with vinyl chloride exposure
Authors: Marion Marie‐Jeanne , Froment Olivier , Trépo Christian 
Published: February 2007

AbstractPoint mutations of c‐ras genes were investigated in human angiosarcomas of the liver associated with occupational exposure to vinyl chloride. DNA prepared from either frozen or paraffin‐embedded tissues was amplified by the polymerase chain reaction, and putative point mutations at codons 12, 13, and 61 of c‐Ha‐ras, c‐Ki‐ras, and N‐ras were analyzed by dot‐blot hybridization with allele‐specific oligonucleotides. A G·C→A·T transition in the second nucleotide at codon 13 of the c‐Ki‐ras‐2 gene was detected in 5 of 6 tumors. This mutation is likely a consequence of vinyl chloride‐DNA adduct formation. It leads to the substitution of glycine by aspartic acid in the resulting p21 protein, a consistent amino acid substitution found so far in all types of human cancer exhibiting a codon 13‐mutated Ki‐ras gene.
Involvement of fos in spontaneous and ultraviolet light‐induced genetic changes
Authors: van den Berg Susanne , Kaina Bernd , Rahmsdorf Hans J. , Ponta Helmut , Herrlich Peter 
Published: February 2007

AbstractTransient overexpression of ras, mos, or fos transcribed from various inducible promoters in NIH 3T3 cells causes significant increases in the frequency of chromosomal aberrations and, as shown for fos, in gene mutations. Under the experimental conditions of exponential growth and full serum supply, overexpression of the oncogenes does not increase the proliferation rate of cells. The generation of ras‐ and mos‐induced chromosomal aberrations was suppressed in cells that had been deprived of fos protein by antisense c‐fos oligodeoxynucleotides. The induction of chromosomal aberrations by ultraviolet irradiation is also suppressed by antisense c‐fos oligodeoxynucleotides. The data suggest that fos protein alone, or a transcription factor that contains fos protein as a subunit, activates or induces the synthesis of one or several mutator functions. Oncogenedriven mutagenesis could account for the accumulation of additional mutations after the activation of an oncogene, which may furnish a mechanistic basis for tumor promotion and tumor progression.
Expression of mouse cytochrome P4501A1 cDNA in repair‐deficient and repair‐proficient CHO cells
Authors: Trinidad Alma C. , Wu Rebekah W. , Thompson Larry H. , Felton James S. 
Published: February 2007

AbstractRecombinant DNA techniques have been used to develop Chinese hamster ovary cell lines for studying chemically induced genotoxicity. These cell lines express a specific cytochrome P450 isozyme responsible for the metabolism of polycyclic aromatic hydrocarbons and exhibit defined differences in DNA repair capacity. A bacterial gene (neo) conferring resistance to gentamicin was inserted into the pcD expression vector containing the mouse cytochrome P1450 (P450IA1) cDNA to facilitate the selection of transformed cells. This plasmid was introduced into the nucleotide‐excision‐repair‐deficient UV5 cell line by electroporation. Transformed clonal isolates expressing the P1450 cDNA were identified by differential cytotoxicity assays using benzo[a]pyrene (B[a]P). One such clone, termed UV5P1, was mutagenized with ethyl methanesulfonate and selected for resistance to killing by UV radiation to derive a repair‐competent clone that expresses similar P1450 activity to that of the parental cell line. Two repair‐competent clones were selected and called 5P1R1 and 5P1R3. The resulting cell lines (UV5P1, 5P1R1, and 5P1R3) expressed arylhydrocarbon hydroxylase activity. UV5P1 and 5P1R3 were compared in terms of cytotoxicity and mutagenicity after exposure to B[a]P. Induced mutations were measured at the adenine phosphoribosyltransferase (aprt) and hypoxanthine guanine phosphoribosyltransferase (hprt) loci. Repair‐deficient UV5P1 cells were killed by B[a]P at concentrations below 0.1 μM, while the repair‐proficient 5P1R3 cells showed no toxicity up to 60 μM. Mutation induction at both loci was also much more efficient in UV5P1 cells. These new cell lines provide a more sensitive system that can be used in a battery of assays to evaluate the genotoxicity of chemicals requiring P450IA1 metabolic activation and to assess the role of DNA repair in modulating the biological effects of DNA damage.