bluuebunny/crossref_metadata_2025 huggingface dataset:
Assessment of 1,2‐dimethylhydrazine in bone marrow micronucleus assay: Variations in protocol and response
Authors: Goldberg M. T. , Schop R. N. , Reidy J. A.
Published: February 2007

The effect of variations in experimental protocol on the assessment of the genotoxicity of 1,2‐dimethylhydrazine (DMH) in the bone marrow micronucleus assay was determined. The incidence of micronuclei (MN) in the bone marrow of CBA mice treated with DMH (either intraperitoneally (i.p.) or orally) was found to be significantly greater than that observed in C57B1/6J mice using the same dose and dosing regimen. With i.p. injection, DMH, at doses of 20 and 50 mg/kg, was found to be positive in the bone marrow MN test in CBA mice only. In C57B1/6J mice, DMH (i.p.) was found to be positive at only the 50 mg/kg dose. With oral administration, DMH was positive in the MN test only at the 50 mg/kg dose and only in CBA mice. No significant difference in the percentage of MN was observed when 300,500, or 1,000 polychromatic erythrocytes (PCEs) were scored following a single treatment of DMH. Cyclophosphamide (CY) was found to induce a dose‐dependent increase in the percentage of MN observed in the bone marrow of C57B1/6J mice.

DMH tested positive in the colon nuclear aberration (NA) assay in both strains of mice using both i.p. and oral routes of administration, although C57B1/6J mice were found to be more sensitive than CBA mice. No significant difference was observed regarding the percentage of NAs observed in the colon between mice injected i.p. or orally gavaged.
Replication index in cultured human lymphocytes: Methods for statistical analysis and possible role in genetic toxicology
Authors: Lazutka J. R. , Margolin B. H.
Published: February 2007

The replication index (RI) is an index of cytokinetics in cultured cells. This paper presents statistical methods for the analysis of RI data. It also proposes formulas for the calculation of RI and its standard error. The proposed statistical analysis may be applied only in cases when replicate cultures are used. If duplicate or repeated cultures are used, the interculture variation may provide a more suitable error term for RI. Determining the RI in human blood cultures may be useful in genotoxicity testing, because inhibition of DNA synthesis will result in a decrease of the RI. Monte Carlo methods are used to show that the sensivity of the proposed method is roughly equivalent to that of a χ2 test. Power studies indicate that for the study sample, no less than 200 cells should be analyzed when assessing the RI.
Responses of the L5178Y mouse lymphoma cell forward mutation assay. V: 27 coded chemicals
Authors: McGregor Douglas B. , Brown Alison G. , Howgate Susan , McBride Douglas , Riach Colin , Caspary William J. , Carver J. H.
Published: February 2007

Twenty‐seven chemicals were tested for their mutagenic potential in the L5178Y tk+/tk− mouse lymphoma cell forward mutation assay using procedures based upon those described by McGregor et al. (McGregor DB, Martin R, Cattanach P, Edwards I, McBride D, Caspary WJ (1987): Environ Mol Mutagen 9:143–160). Cultures were exposed to the chemicals for 4 hr, then cultured for 2 days before plating in soft agar with or without trifluorothymidine (TFT), 3 μg/ml. The chemicals were tested at least twice. Statistically significant responses were obtained with acid orange 10, aniline, benzaldehyde, o‐chloroaniline, chlorodibromomethane, cytembena, 1,2‐dibromo‐4‐(1,2‐dibromomethyl) cyclohexane, dieldrin, lithocholic acid, oxytetracycline, phenazopyridine HCl, 1‐phenyl‐3‐methyl‐5‐pyrazolone, sodium diethyldithiocarbamate, solvent yellow 14, tetraethylthiuram disulfide (disulfiram), 2,4‐toluene diisocyanate, and 2,6‐toluene diisocyanate. Apart from phenazopyridine HCl, acid orange 10, and solvent yellow 14, rat liver S9 mix was not a requirement for the mutagenic activity of these compounds. Chemicals not identified as mutagens were N‐4‐acetylaminofluorene, chlorpheniramine maleate, chlorpropamide, 1,4‐dioxane, endrin, ethylene glycol, iron dextran, methapyrilene, sodium(2‐ethylhexyl)alcohol sulfate, and 2,3,7,8‐tetra‐chlorodibenzo‐p‐dioxin.
Analysis of bleomycin‐induced mutagenic functions related to the PSO4 (= XS9) gene of saccharomyces cerevisiae
Authors: Severgnini A. , Lillo O. , Nunes E. , Brockman H. E.
Published: February 2007

In the present work we analyse some properties of the pso4‐1 (= xs9) mutant of the yeast Sac‐charomyces cerevisiae, which is blocked in both reverse mutation and recombination. We have applied the “apparent survival test” as a method to characterize putative inducible errorprone repair activities related to the PSO4 gene. As the mutagenic agent we have used bleomycin, which is an antitumour radiomimetic antibiotic. Survival, reverse mutation yield, and reverse mutation frequency were determined as a function of bleomycin concentrations (1‐60 μg/ml) in the logarithmic phase of growth. It is shown that the PSO4 gene product is poorly involved in bleomycin sensitivity and that reverse mutation induced by this mutagen is dramatically reduced in the pso4‐1 lys2 mutant strain, as compared with the isogenic SC7K (lys2) strain. M(x) functions exhibit linear‐quadratic courses for both the SC7K (lys2) and the pso4‐1 lys2 strains. The apparent survival functions display “humps” in both strains, corresponding to the resistant components of the survival functions. These facts suggest that the mutagenicity of bleomycin depends on at least one inducible error‐prone repair pathway and that the PSO4 gene product could act as a mutation triggering factor.
Sex and strain differences in the hepatocyte primary culture/DNA repair test
Authors: McQueen Charlene A. , Way Barbara M. , Casciano D. A.
Published: February 2007

The hepatocyte primary culture (HPC)/DNA repair test was developed using hepatocytes isolated from male F‐344 rats. A number of genetic polymorphisms have been shown to occur in inbred strains of rats, which may lead to variation in biotransformation of xenobiotics resulting in differences in susceptibility to genotoxins. The effect of the strain utilized as a source of hepatocytes was investigated with female Lewis, F‐344, and DA rats. Variation was observed when hepatocytes from the three strains were exposed to aflatoxin B1 (AFB1). Hepatocytes from female Lewis rats were more sensitive to AFB1 genotoxicity than cells from F‐344 or DA rats. DNA repair was induced by a concentration as low as 108 M AFB 1 in the Lewis strain while 104 M was needed for hepatocytes from female F‐344 or DA rats. Sex‐related differences were also observed with AFB1; DNA repair was induced at 108 M in hepatocytes from male F‐344 rats. No clearcut strain differences were seen when cells were exposed to diethylnitrosamine (DEN) or 2‐acetylaminofluorene, although 102 M DEN was only cytotoxic to hepatocytes from DA rats. These results demonstrate that both the strain and the sex of the animal used as a source of hepatocytes can affect the HPC/DNA repair test.
Frequency of sister chromatid exchange in peripheral lymphocytes of male pesticide applicators
Authors: Rupa D. S. , Reddy P. P. , Sreemannarayana K. , Reddi O. S. , Galloway S. M.
Published: February 2007

In the present study 61 male pesticide applicators who worked in cotton fields and regularly sprayed pesticides such as DDT, BHC, endosulfan, malathion, methyl parathion, phosphamidon, dimethoate, monocrotophos, quinalphos fenvelrate, and cypermethrin were analyzed for sister chromatid exchanges, mitotic index, and cell cycle kinetics in peripheral lymphocytes. Subjects who handled pesticides were non‐smokers and teetotalers and the data were compared with the matched control group. Statistical analysis revealed that the frequency of sister chromatid exchanges was significantly higher among the pesticide applicators at all the durations of exposure when compared to controls. Subjects exposed to pesticides also showed cell cycle delay and decrease in mitotic index when compared to the control group.
A molecular Geneticist's view of complex genomic lesions
Authors: Hozier John C.
Published: February 2007

Molecular genetic techniques have been used effectively to study the DNA lesions, many of which are large and complex, that are responsible for human genetic disease. In this workshop we explore some of these techniques, how they are applied to study genetic lesions, and how they may be applied to the needs of genetic toxicology.
Molecular analysis of mutations affecting hprt mRNA splicing in human T‐lymphocytes in vivo
Authors: Rossi Anna M. , Tates Ad D. , Van Zeeland Albert A. , Vrieling Harry
Published: February 2007

Molecular analysis of hypoxanthine‐guanine phosphoribosyltransferase (hprt) cDNA from 6‐thioguanine‐resistant T‐lymphocytes cloned from smoking and non‐smoking adult donors showed that 35% of these mutants were defective in splicing of hprt mRNA. Among a set of 42 hprt splice mutants, we observed i) complete loss of one or more exons, ii) partial loss of one exon, or iii) inclusion of part of an intron sequence between adjacent exons. Loss of exon 4 was significantly more frequent than of the other exons, suggesting that the sequences that regulate splicing of this exon are either larger than those of the other exons or especially prone to mutation. In order to identify the molecular nature of DNA alterations causing aberrant splicing of hprt mRNA, 17 splice mutants were analyzed in more detail by sequencing the genomic regions flanking the mis‐spliced exon. Base pair substitutions or small deletions causing defective splicing were either detected in exon sequences or in splice site consensus sequences of introns. Furthermore, genomic deletions encompassing entire exons were found. In some mutants, the alteration responsible for incorrect splicing could not be identified, suggesting that the target sequence for splice mutations is larger than merely the splice junctions. Molecular characterization of hprt splice mutations will lead to the identification of specific sequences regulating splicing of hprt mRNA and will reveal whether the mutational spectrum in splice mutants is similar to that found in the hprt coding region.
Induction of 6‐thioguanine‐resistant mutants in human diploid fibroblasts in vitro with ethylene oxide
Authors: Kolman Ada , Bohušová Tatiana , Lambert B. , Simons J. W. I. M.
Published: February 2007

Human diploid fibroblasts (strain VH‐10) were exposed to the direct‐acting alkylating agent, ethylene oxide (EtO), in vitro, and the frequency of HPRT mutants was evaluated by selection in medium containing 6‐thioguanine. A dose‐dependent increase of the mutant frequency was found in the dose range of 2.5–10 mMh of EtO. The EtO‐induced mutant frequency increased 5–19 times the background frequency at low or moderately toxic doses, which indicates that EtO is a strong mutagen in human fibroblasts in vitro. The mutagenic potency was 9.8 × 10−6 per mMh.
Further evidence for the aneuploidogenic properties of chelating agents: Induction of micronuclei in mouse male germ cells by EDTA
Authors: Russo A. , Levis A. G.
Published: February 2007

A micronucleus assay based on cytogenetic analysis of early spermatids (Tates et al.; Mutation Research 121:131–138, 1983) was applied to determine if the chelating agent ethylenedinitrilotetraacetic acid (EDTA) may induce aneuploidy in mouse meiotic cells. Previous results indicated aneuploidogenic activity of EDTA in Drosophila female germ cells (induction of chromosome loss; Zordan et al.: Environ Mol Mutagen 15:205–213, 1990). In the same study, a standard aneuploidy test based on chromosome counting in mouse secondary spermatocytes failed however to show aneuploidogenic properties of FDTA in mouse somatic and germ cells. In the present study the effects of two clastogens, adriamycin (ADM) and mitomycin C (MMC), and of the aneuploidogenic agent chloral hydrate (CH) were also evaluated. All compounds were tested at a single dose level and at two time intervals corresponding to the treatment of diakinesis/metaphase I/metaphase II spermatocytes. The clastogenic potential of the compounds under study was also evaluated, by chromosomal aberration analysis in mouse spermatogonia, in an independent set of experiments. The results obtained indicate that ADM, CH and EDTA are able to induce micronuclei at meiosis. On the contrary, only ADM and MMC induced chromosomal aberrations in mouse spermatogonia. Therefore, the most probable origin of micronuclei produced by CH and EDTA is whole chromosome lagging. These results provide further evidence for the aneuploidogenic properties of these chelating agents.
Interspecies cytogenetic comparisons: Studies with x‐radiation and bleomycin sulfate
Authors: Kligerman Andrew D. , Bryant Miriam F. , Doerr Carolyn L. , Halperin Edward C. , Kwanyuen Puntipa , Sontag Marc R. , Erexson Gregory L.
Published: February 2007

A series of in vitro experiments were conducted to determine if there are innate differences in the sensitivity of peripheral blood lymphocytes (PBLs) from different mammalian species to clastogens. Mouse, rat, and human whole blood samples were exposed to either 0, 0.38, 0.75, 1.5, or 3.0 Gy x‐radiation or 0, 5, 10, 20, 40, or 80 (μg/ml bleomycin for 4 hr. Bromodeoxyuridine‐containing cultures were initiated and the PBLs stimulated to divide with phytohemagglutinin. All cultures were harvested following a 3‐hr colcemid treatment. Slides were made and differentially stained, and first‐division metaphases were scored for chromosome aberrations. In the x‐radiation studies human PBLs were significantly more sensitive than mouse PBLs which were in turn more sensitive than rat PBLs as measured by either the total percent aberrant cells or the number of dicentrics. Data from all three species could be fitted to a linear‐quadratic model. Results with bleomycin suggest that the mouse and human PBLs are equally sensitive to the clastogenic effects of bleomycin. Both appeared to be more sensitive than the rat PBLs, but the variation between experiments was such that the results among species were not significantly different. These results indicate that there may be inherent differences in sensitivity among PBLs of mammalian species; however, more studies are needed to determine if the differences presented here hold for other agents.
Mutation spectrum of spontaneous frameshift revertants in yeast using double‐strand gap repair
Authors: Plewa Michael J. , Kalinowski Douglas P. , Larimer Frank W.
Published: February 2007

A mutation spectrum was constructed from a series of randomly isolated spontaneous His+ revertants of the frameshift mutant his4–38 in Saccharomyces cerevisiae. For each true revertant, a 438 bp region encompassing his4–38 on chromosome III was recovered into a shuttle vector by double‐strand gap repair. Of the 45 independent His+ revertants sequenced, 44 were −1 base deletions and one revertant was a +2 base insertion. The −1 deletions exhibited a bimodal distribution. Of the bases encompassing the his4–38 region from +153–181, 45% were not involved in a reversion event, although a −1 frameshift within this region will result in a viable His+ revertant. Approximately 49% of −1 events occurred within runs of 3 repeated bases. At these sites the strand‐slippage model for frameshift mutation is supported. However, the −1 events occurring at sites of 2 repeated bases and the low frequency (2%) of +2 base insertions suggest that the transiently misaligned template model is a significant mechanism in reversion of his4–38. When the distribution of −1 events at repeated bases was discounted, a hotspot involving a ‐T at position +163 was resolved. © 1992 Wiley‐Liss, Inc.
DNA sequence analysis of spontaneous and n‐ethyl‐n‐nitrosourea‐lnduced hprt mutations arising in vivo in cynomolgus monkey t‐lymphocytes
Authors: Harbach P. R. , Filipunas A. L. , Wang Y. , Aaron C. S.
Published: February 2007

The study of hprt mutations in cynomolgus monkey T‐lymphocytes is part of our effort to understand the mechanisms of mutagenesis in vivo. This primate model allows us to study mutations and their kinetics at the molecular level under well‐controlled conditions using recently developed techniques for selection of mutant T‐cells and polymerase chain reaction (PCR) amplification of hprt cDNA, which is directly sequenced. This is the first report of the sequence of the coding region of the cynomolgus monkey hprt gene and PCR/DNA sequence analysis of seven spontaneous mutant T‐cell clones, as well as 23 mutant clones isolated 63 and 601 days after treatment with ethylnitrosourea (ENU, 77 mg/ kg, intraperitoneal). cDNA was reverse transcribed from hprt mRNA directly from a lysate of about 2–4 × 103 cells, and a 700 bp fragment including the coding region was amplified by PCR and sequenced. Of the seven spontaneous mutants, only one point mutation (GC↠ transition) was detected, and the other six failed to amplify by PCR, possibly due to functional deletions. Of the 14 mutant clones isolated 63 days after ENU treatment, nine base substitutions were detected in ten clones: four transitions (three AT↠GC and one GC↠AT) and five transversions (four AT↠TA and one AT↠CG). Of the nine mutants isolated 601 days after ENU treatment, six single base substitutions were detected in six clones (five AT↠TA and one AT↠CG transversions), and one mutant had a large deletion or insertion. No changes were detected in three clones (one Day 63 and two Day 601 clones). In summary, only one of 15 single base substitutions isolated after ENU treatment was a GC↠AT transition mutation and the rest were transitions and transversions at AT sites. © 1992 Wiley‐Liss, Inc.
structure‐activity relationship of genotoxic polycyclic aromatic nitro compounds: Further evidence for the importance of hydrophobicity and molecular orbital energies in genetic toxicity
Authors: Debnath Asim Kumar , Hansch Corwin
Published: February 2007

A quantitative structure‐activity relationship (QSAR) has been formulated for 15 polycyclic aromatic nitro compounds acting on E. coli PQ37. Upon damage of DNA by these substances β‐galactosidase is induced and can be easily assayed colorimetrically, hence, this is a short‐term test for mutagenicity. The QSAR (log SOSIP = 1.07 log P ‐ 1.57 ± ‐ 6.41) is strikingly similar to that found earlier with nitroaromatics acting in the Ames test (TA100) and differs significantly for that found using TA98 organisms. The QSAR brings out in a unique manner the underlying similarity in the two test systems. © 1992 Wiley‐Liss, Inc.
Tissue‐specific genotoxic effects of acrylamide and acrylonitrile
Authors: Butterworth Byron E. , Eldridge Sandra R. , Sprankle Catherine S. , Working Peter K. , Bentley Karin S. , Hurtt Mark E.
Published: February 2007

Acrylamide (AA) has been reported to induce dominant lethal mutations in male rat germ cells and tumors in a variety of organs, including the scrotum, thyroid and mammary glands, but not the liver of rats. The structurally similar vinyl monomer acrylonitrile (ACN) does not induce dominant lethal mutations but does induce tumors of the brain, Zymbal gland, forestomach and mammary gland, but not the liver of rats. Several in vitro and/or in vivo unscheduled DNA synthesis (UDS) assays were employed to examine the potential tissue‐specific genotoxic activity of these compounds. Neither AA nor ACN induced DNA repair in either the in vitro or in vivo hepatocyte DNA repair assays. Glycidamide (GA), a mutagenic metabolite of AA, induced DNA repair in the in vitro hepatocyte DNA repair assay. Cyanoethylene oxide (CEO), a mutagenic metabolite of ACN, did not yield a DNA repair response in the in vitro hepatocyte DNA repair assay, but was highly toxic and could not be tested at doses equivalent to GA. AA, but not ACN, produced a DNA repair response in the in vivo spermatocyte DNA repair assay. AA produced a slight response in the in vitro human mammary epithelial cell (HMEC) DNA repair assay in normal cells derived from discarded surgical samples from five different women. GA produced a strong UDS response in all cases in the same assay. CEO, but not its parent compound ACN, produced a response in the HMEC DNA repair assay. These results show a highly tissue‐specific pattern of genotoxic activity for AA and ACN that correlates, to the extent that it has been examined, with the tissue‐specific pattern of carcinogenic and dominant lethal activity. The induction of DNA repair by GA and CEO confirms the genotoxic potential of these metabolites. While the observation of genotoxic activity of AA in the HMEC DNA repair assay suggests that mammary cells might be a target for carcinogenic activity of this compound in humans, other factors such as pharmacokinetics and epidemiology must be evaluated to establish that effect. © 1992 Wiley‐Liss, inc.
Chromosomal breakage following treatment of CHO‐K1 Cells in Vitro With U‐68,553B is due to Induction of Undercondensation of Heterochromatin
Authors: Yu R. L. , Aaron C. S. , Ulrich R. G. , Thilagar A. , Kumaroo P. V. , Wang Y.
Published: February 2007

Preferential breakage of chromosomes af specific sites (so‐called “fragile sites”) has been observed to occur spontaneously, and has been induced by some metal salts and chemicals. Furthermore, a heterochromatic region of the long arm of the Chinese hamster ovary (CHO) M‐chromosome is known to be suceptible to a disproportionately high frequency of spontaneous breakage; unless there is physical displacement of chromatin the resulting achromatic lesions are not scored as structural aberrations. We have encountered such anomalous breakage associated with C‐band positive regions of the chromosomes of a CHO‐K1 cell line following exposure of the cells to toxic doses of U‐68,553B and in this report present evidence that the apparent breaks are due to undercondensed heterochromatin (UH) and evidence that the phenomenon appears to occur at higher frequency in a particular cell line of Chinese hamster. This finding has important implications on the assessment of potential risk due to exposure to the drug. Such apparent breaks at sites of UH in chromosome 1 was not observed in an alternate CHO cell line (CHO‐WBL) which supports the notion that the UH associated achromatic lesions in the CHO‐K1 line may be a cell line specific phenomenon. Furthermaore, careful electron microscopy of the chromosomes revaled chromatin fibers connecting the apparently broken chromosomes. The UH was not observed in the presence of the phenomenon in risk assessment is further reduced. The kata presented here provide evidence that sites of UH accur preferentially at locations of C‐band positive constitutive heterochromatin in CHO cells; we believe that this is the first report of induced fragile sites in rodent cells in vitro documented in this way. In addition, evidence is presented that U‐68,553B lacks the ability to induce breakage in vivo in rodents and locks the ability to induce chromosome breakage in human peripheral lympocytes in vitro. Therefore, it is concluded that the positive results with CHO‐k1 cells treated with U‐68,553B are unlikely to be predictive of a genotoxic hazard. This is a specific example of the importance of careful followup to an in vitro result in risk assessment. © 1992 Wiley‐Liss, Inc.
Development of a specific‐locus assay in the ad‐3 region of two‐component heterokaryons of neurospora: A review
Authors: De Serres Frederick J.
Published: February 2007

In recognition of the need for a more comprehensive data base for genetic risk assessment of human exposure to mutagenic agents in the environment, a model system was developed for specific‐locus studies in Neurospora crassa. This lower eukaryotic organism permits the utilization of microbial techniques for recovery of large numbers of specific‐locus mutations at two closely linked loci as well as their subsequent genetic analysis. In particular, this assay makes possible exploratory experiments with different environmental mutagens to obtain data on a wide variety of experimental conditions. Such data make it possible to study induction kinetics and mutational spectra in a manner that is not as yet feasible in higher eukaryotic organisms. The adenine‐3 (ad‐3) specific‐locus assay was modeled after the 2–gene, morphological specific‐locus assay in the dilute‐short‐ear region of the mouse, and it also detects forward‐mutations at two closely linked loci, namely, ad‐3A and ad‐3B. Because ad‐3 mutations are recovered by a direct method, based on the accumulation of a reddish‐purple pigment in the vacuoles of the mycelium rather than their requirement for adenine, this system is both a morphological and biochemical specific‐locus assay. The use of the ad‐3 assay system in experiments with different environmental mutagens has provided precise dose‐response curves not only for inactivation, but also the overall induction of ad‐3 mutations. Genetic characterization of these ad‐3 mutations by a series of 3 rapid and simple genetic tests permits the identification of 18 subclasses of gene/point mutations, and 12 subclasses of multilocus deletion mutations. These subclasses also include 3 different classes of multiple‐locus mutations with separate sites of recessive lethal damage either in the immediately adjacent regions or elsewhere in the genome. In summary, this specific‐locus assay provides a capability that is unique among eukaryotic organisms for the recovery and analysis of genetic damage at 2 closely linked loci. © 1992 Wiley‐Liss, Inc.
Certain tryptophan photoproducts are inhibitors of cytochrome p450‐dependent mtuagenicity
Authors: Rannug U. , Agurell E. , Rannug A. , Cederberg H.
Published: February 2007

Two photoproducts, derived from UV‐irradiation of the amino acid L‐tryptophan and with high Ah (TCDD) receptor binding affinity, were tested for genotoxic and antimutagenic effects. The two indolo[3,2‐b]carbazole derivatives, with the molecular weights of 284 and 312, respectively, were tested in Saccharomyces cerevisiae strain D7 for mitotic gene conversion and reverse mutation and in strain RS112 for sister chromatid conversion and gene conversion. No significant (P > 0.05) genotoxic effects were found in strain D7, while strain RS112 showed a small but significant increase in the frequency of sister chromatid conversions. In Chinese hamster ovary (CHO) cells the two compounds induced a statistically significant but less than twofold increase in the frequency of sister chromatid exchanges (SCE). No mutations were detected when the compounds were tested in Salmonella typhimurium strains TA98 and TA100. However, both 284 and 312 acted as antimutagens on strain TA100+S9 in the presence of benzo(a)pyrene. The decrease in mutagenicity by the most potent compound 284 was 20 revertants/nmol. This effect could be explained by an inhibitory effect on the cytochrome P450‐dependent ethoxyresorufin O‐deethylase (EROD) activity as seen in rat hepatocytes. The two compounds were also tested with hamster cells expressing rat cytochrome [P‐450IA]. The results support the conclusion that this cytochrome P‐450 isozyme is inhibited by the tryptophan photoproducts. Similar results were also seen with two other high affinity Ah receptor ligands the quinazolinocarboline alkaloids rutaecarpine and dehydrorutaecarpine. © 1992 Wiley‐Liss, Inc.
Assessment of oxidative DNA damage in the oxyR‐deficient sos chromotest strain escherichia coli PQ300
Authors: Müller Jürgen , Janz Siegfried
Published: February 2007

The SOS chromotest is a simple short‐term genotoxicity assay measuring the induction of gene sfiA in Escherichia coli K‐12. The recent availability of SOS tester strains with additional mutations in DNA repair or protection systems allows testing of DNA damaging compounds for genotoxic specificity. E. coli PQ300 differs from the standard SOS tester strain PQ37 in that it contains an additional mutation in gene oxyR that renders it more sensitive to oxidative genotoxins. The generation of reactive oxygen intermediates (ROI) by hydroperoxides (H2O2, t‐butyl hydroperoxide, cumene hydroperoxide), γ‐radiation, glucose oxidase, and xanthine oxidase resulted in a more vigorous SOS response in strain PQ300 compared to strain PQ37. PQ300 was also more sensitive than PQ37 for the detection of reducing agents such as ascorbic acid, cysteine, and glutathione, which also alter the redox status of the bacterial cells. However, intercalating agents (adriamycin, bleomycin, and mitomycin C) and the UV‐and radiomimetic compound 4‐nitroquinoline‐1‐oxide whose DNA damaging potential are known also to involve ROI did not show significant differences between strains PQ37 and PQ300. It is concluded that the oxyR‐deficient strain PQ300 is useful for detecting certain classes of genotoxins that change the oxidative/antioxidative balance of tester bacteria in the SOS chromotest. © 1992 Wiley‐Liss, Inc.
Mutagenicity test schemes and guidelines: U.S. EPA office of pollution prevention and toxics and office of pesticide programs
Authors: Auletta Angela E. , Dearfield Kerry L. , Cimino Michael C.
Published: February 2007

New requirement for chemicals subject to mutagenicity testing from the U.S. Environmental Protection Agency (USEPA) are discussed. Also detailed are two categories in the 1986 Mutagenicity Risk Assessment Guidelines. © 1993 Wiley‐Liss, Inc.
Mutagenicity of peroxyacetyl nitrate (PAN) in vivo: Tests for somatic mutations and chromosomal aberrations
Authors: Heddle J. A. , Shepson P. B. , Gingerich J. D. , So K. W.
Published: February 2007

A series of experiments was conducted in which Chinese hamsters inhaled PAN, an ubiquitous pollutant that is present in the atmosphere at concentrations that are as high as, or higher than, other known genotoxic agents. The animals were exposed to PAN in air at concentrations of = 3 ppm for up to 1 month and then examined for somatic mutations and chromosomal aberrations. Mutations were assayed by measuring the frequency of thioguanine‐resistant lung fibroblasts (isolated de novo and cultured). Chromosomal aberrations were assayed by measuring the frequency of micronuclei in either the bone marrow (polychromatic erythrocytes) or the lungs (binucleate lung fibroblasts cultured in the presence of cytochalasin B). The results for the test animals were compared to those from animals exposed similarly, but without PAN. Although in each experiment the mutation frequencies for the test animals were higher than the corresponding controls, the mutation frequencies were not significantly different from the concurrent negative controls (P > .05) or the historical controls, except for experiment C. In experiment C, there was a significant regression of mutation frequency versus dose (P < 0.001) if all of the historical controls for pooled animals are included at zero dose. No reproducible evidence of chromosomal breakage was found in either lung or bone marrow. Thus, although PAN has been found to be a bacterial mutagen, we did not find statistically significant evidence of mutagenicity in vivo. The toxicity of PAN limited the exposure concentration that could be used. When all of the PAN data were used, the best estimate of the mutagenic potency proved to be comparable to that of ethylene dibromide, a carcinogenic atmospheric pollutant. © 1993 Wiley‐Liss, Inc.
Incorporation of a micronucleus study into a developmental toxicology and pharmacokinetic study of L‐selenomethionine in nonhuman primates
Authors: Choy Wai Nang , Henika Philip R. , Willhite Calvin C. , Tarantal Alice F.
Published: February 2007

Concomitant to a developmental toxicology study of selenium in long‐tailed macaques (Macaca fascicularis), a transplacental bone marrow micronucleus assay was conducted in the fetuses of treated animals. Selenium was administered as L‐selenomethionine by nasogastric intubation at 0, 150 or 300 μg/kg‐day to pregnant macaques daily throughout organogenesis (gestation days 20–50). Pregnancy was terminated on gestation day 100 ± 2 and fetuses were obtained by hysterotomy. Selenium concentrations in maternal blood were monitored throughout pregnancy and selenium concentrations in fetal blood were measured at hysterotomy. Maternal circulating selenium did not exceed 4 ppm in plasma or 3.7 ppm in erythrocytes. Selenium in cord blood was ≤ 0.1 ppm in plasma and ≤ 1.1 ppm in erythrocytes at 300 μg/kg‐day. Fetal bone marrow smears were prepared from the humerus and micronucleated polychromatic erythrocytes were scored. No increase of micronucleus frequency was detected in any dose group, although signs of maternal selenosis were obvious. This finding is compared to the previous observation that micronuclei were induced in the bone marrow of adult nonpregnant macaques treated at 600 μg/kg‐day, a lethal dose yielding blood selenium levels to 7.3 ppm in plasma and 5.7 ppm in erythrocytes after 15 days of daily treatment, when death occurred. These data demonstrate that measurement of circulating xenobiotics can be useful for the interpretation of genetic toxicology results. © 1993 Wiley‐Liss, Inc.
Mutagenicity of the human carcinogen treosulphan, and its hydrolysis product, dl‐1,2:3,4‐diepoxybutane in mammalian cells
Authors: Zhu Songyun , Zeiger Errol
Published: February 2007

The cytotoxicity and mutagenicity of the human carcinogen, treosulphan, and its hydrolysis product, dl‐1,2:3,4‐diepoxybutane (DEB), were studied in Chinese hamster ovary, AS52, cells. Treosulphan (0.1–1.0 mM) is toxic and mutagenic at the gpt locus. A strong pH dependence was noted. DEB is cytotoxic and mutagenic at a much lower dose (0.025 mM), but these effects were not affected by pH. The results suggest that the toxic and mutagenic effects of treosulphan are mediated by its hydrolysis product DEB, and that the conversion of treosulphan to DEB is highly pH‐dependent. © 1993 Wiley‐Liss, Inc.
Analysis of the six additional chemicals for in vitro assays of the european economic communities' EEC aneuploidy programme using saccharomyces cerevisiae D61.M and the in vitro porcine brain tubulin assembly assay
Authors: Albertini Silvio , Brunner Martin , Würgler Friederich E.
Published: February 2007

We tested six additional chemicals (acetaldehyde, benomyl, diethylstilboestrol, diethylstilboestrol dipropionate, griseofulvin, and mercaptoethanol) for in vitro systems of the coordinated programme to study aneuploidy induction sponsored by the Commission of the European Communities in two in vitro test systems.

Using Saccharomyces cerevisiae D61.M (mitotic chromosomal malsegregation assay), benomyl showed a dose‐dependent increase in the frequency of chromosomal malsegregation with a lowest effective dose tested (LEDT) of 30 μg/ml (0.1 mM). Diethylstilboestrol (DES) showed solvent‐dependent effects. DES dissolved in ethanol induced an increase in chromosomal malsegregation as well as in the frequency of total resistant colonies (mutations and recombinations) with a LEDT arround 13 μg/ml (0.048 mM). Using dimethylsulfoxide as the solvent, no increases were observed with DES up to 333 μg/ml (1.24 mM). Acetaldehyde induced an increase in chromosomal malsegregation with the cold treatment protocol (LEDT: 1.25 μl/ml (21 mM) and 0.75 μ/ml (13 mM), respectively) but no increase with the overnight protocol (highest dose tested (HDT): 1.75 μ/ml; 30 mM). Concerning the frequency of total cycloheximide‐resistant colonies (mutations and recombinations) increases were obtained with both protocols. The other three compounds were negative when tested up to toxic doses (survival below 10%), up to the maximum solubility in the solvent used or up to heavy precipitation in the incubation mix. The HDT were 333 μ/ml (0.88 mM) for diethylstilboestrol dipropionate, 1,600 μg/ml (4.5 mM) for griseofulvin and 0.5 μl/ml (7 mM) for mercaptoethanol.

Concerning effects on porcine brain tubulin assembly in vitro, diethylstilboestrol and griseofulvin inhibited the assembly process. The IC30% (30% inhibition concentration) values were 12.5 μM and 100 μM for DES and griseofulvin, respectively. Mercaptoethanol showed no effects up to 50 mM. © 1993 Wiley‐Liss, Inc.
In vitro system for detecting nongenotoxic carcinogens
Authors: Dertinger Stephen D. , Torous Dorothea K. , Tometsko Andrew M.
Published: February 2007

Chemical risk assessment has been limited by the inability of in vitro short‐term assays to identify the true carcinogenic potential of many substances. Numerous methods exist for identifying mutagenic and clastogenic agents, but a practical means of identifying nongenotoxic carcinogens has remained elusive. Experiments described here suggest that some chemicals may participate in carcinogenesis by modulating the enzymatic processes of drug metabolism. The tumor promoters butylated hydroxyanisole, butylated hydroxytoluene, deoxycholic acid, reserpine, trypan blue, and 12,‐O‐tetradecanoyl phorbol‐13‐acetate were chosen as model non‐genotoxic carcinogens. The enzyme‐modulating action of these chemicals was measured using a modified Ames plate incorporation assay whereby the known tumor promoters were plated with a promutagen in the presence of a mammalian metabolic activation system (S9). Each of the nongenotoxic carcinogens significantly increased the mutagenic response of metabolically activated promutagen(s). These experiments suggest that the carcinogenic role of some chemicals may be attributed to their ability to modify the biochemical pathways of drug metabolism. By enhancing or inhibiting the activity of various enzymes, some tumor promoters may create an environment that increases a cell's mutational burden, thereby contributing to neoplastic transformation.
Benzene metabolite, 1,2,4‐benzenetriol, induces micronuclei and oxidative DNA damage in human lymphocytes and HL60 cells
Authors: Zhang Luoping , Robertson Moire L. , Kolachana Prema , Davison Allan J. , Smith Martyn T.
Published: February 2007

The triphenolic metabolite of benzene, 1,2,4‐benzenetriol (BT), is readily oxidized to its corresponding quinone via a semiquinone radical. During this process, active oxygen species are formed that may damage DNA and other cellular macromolecules. The ability of BT to induce micronuclei (MN) and oxidative DNA damage has been investigated in both human lymphocytes and HL60 cells. An antikinetochore antibody based micronucleus assay was used to distinguish MN containing kinetochores and potentially entire chromosomes (kinetochore‐positive, K+) from those containing acentric chromosome fragments (kinetochore‐negative, K−). BT increased the frequency of MN formation twofold in lymphocytes and eightfold in HL60 cells with the MN being 62% and 82% K+, respectively. A linear dose‐related increase in total MN, mainly in K+‐MN, was observed in both HL60 cells and lymphocytes. Addition of copper ions (Cu2+) potentiated the effect of BT on MN induction threefold in HL60 cells and altered the pattern of MN formation from predominantly K+ to K−. BT also increased the level of 8‐hydroxy‐2′‐deoxyguanosine (8‐OH‐dG), a marker of active oxygen‐induced DNA damage. Cu2+ again enhanced this effect. Thus, BT has the potential to cause both numerical and structural chromosomal changes in human cells. Further, it may cause point mutations indirectly by generating oxygen radicals. BT may therefore play an important role in benzene‐induced leukemia. © 1993 Wiley‐Liss, Inc.
Genotoxicity analysis of N,N‐dimethylaniline and N,N‐dimethyl‐p‐toluidine
Authors: Taningher Maurizio , Pasquini Rossana , Bonatti Stefania
Published: February 2007

N,N‐Dimethylaniline (DMA, CAS No. 121—69—7) and N,N‐dimethyl‐p‐toluidine (DMPT, CAS No. 99—97—8) belong to the N‐dialkylaminoaromatics, a chemical class structurally alerting to DNA reactivity. Their applications may be industrial (dye and pesticide intermediates, polymerizing agents) and surgical (polymerization accelerators for the manufacture of bone cements and prosthetic devices), thus implying heterogeneous types of human exposure. Findings of carcinogenicity in rodents and some nonexhaustive genotoxicity data are available for DMA, but to our knowledge no information is available on DMPT concerning either carcinogenicity or any kind of genetic toxicity. To investigate their mechanism cf action and mutagenic/carcinogenic potential, DMA and DMPT were analyzed for complementary genotoxicity end‐points, namely, gene mutation in Salmonella (Ames test), structural and numerical chromosome aberrations in hamster V79 cells (micronucleus test, matched with an immunofluorescent staining for kinetochore proteins), and in vivo DNA damage in mouse and rat liver (alkaline DNA elution test). The results essentially indicate that both chemicals are chromosome damaging agents. Indeed, at the maximum nontoxic doses, they proved nonmutagenic in Salmonella (although their toxicity did not allow concentrations > 70 μg/plate to be tested) and weakly positive in inducing DNA damage (increases in DNA elution rates at most ∼2.4 times control value). Conversely, they proved clearly positive in inducing numerical chromosome alterations, with dose‐dependent increases up to more than five times the control value for DMPT. At the highest dose tested, both chemicals also showed a significant clastogenic effect. © 1993 Wiley‐Liss, Inc.
Mutagenicity of soluble and insoluble nickel compounds at the gpt locus in G12 chinese hamster cells
Authors: Lee Y. W. , Pons C. , Tummolo D. M. , Klein C. B. , Rossman T. G. , Christie N. T.
Published: February 2007

Nickel is an established human and animal carcinogen, but efforts to demonstrate its mutagenicity in a number of cell types have not been successful. In this report we describe the mutational response to nickel compounds in the G12 cell line, an hprt deficient V79 cell line containing a single copy of the E. coli gpt gene. This cell line has a low spontaneous background, making it suitable for assessment of mutagenic responses to environmental contaminants. When G12 cells were treated with insoluble particles of crystalline nickel sulfide < 5 μm in diameter, a strong, dose‐dependent mutagenic response was observed up to 80 times the spontaneous background. Of 48 mutant gpt(−) clones isolated that were induced by insoluble nickel, all were capable of DNA amplification of the gpt sequences by polymerase chain reaction (PCR). The ability to produce full‐length PCR products is an indication that large deletions of gene sequences have not occurred. When G12 cells were treated with soluble nickel sulfate, the mutational response was not significantly increased over the spontaneous background. This difference in mutagenic response reflects a large difference in the mutagenic potential of soluble and insoluble nickel compounds, which reflects the carcinogenic potential of these forms of nickel. © 1993 Wiley‐Liss, Inc.
Genetic toxicity of malathion: A review
Authors: Flessel Peter , Quintana P. J. E. , Hooper Kim
Published: February 2007

Mammalian in vivo and in vitro studies of technical or commercial grade malathion and its metabolite malaoxon show a pattern of induction of chromosome damage, as measured by chromosome aberrations, sister chromatid exchanges, and micronuclei. Experiments with purified (<99%) malathion gave weak or negative results. In contrast to the cytogenetic effects of technical grade malathion, responses in gene mutation assays were generally negative except for malaoxon, which was positive for mammalian gene mutations in both tested instances. This result also could be a consequence of chromosome level changes, however. Dermal exposure, a common human route, caused cytogenetic damage in test animals at doses near those producing positive results by intraperitoneal injection. Workers who apply technical grade malathion and other pesticides have higher levels of chromosomal damage than unexposed individuals. Because of the inactivity of malathion mixtures in gene mutation assays, malathion has been thought to be of little genotoxic concern. However, the pattern of chromosome damage in animals and mammalian cells in culture (including human) indicates that technical grade malathion and its components have not been adequately studied for genotoxic potential in humans. © 1993 Wiley‐Liss, Inc.
In vitro and in vivo assays of isopropanol for mutagenicity
Authors: Kapp Robert W. , Marino Dale J. , Gardiner Thomas H. , Masten Lawrence W. , McKee Richard H. , Tyler Tipton R. , Ivett James L. , Young Robert R.
Published: February 2007

To assess the mutagenic potential of isopropanol, an in vitro Chinese hamster ovary (CHO) cell/HGPRT gene mutation assay and a bone marrow micronucleus study in mice were conducted. In the CHO/HGPRT assay, concentration levels ranged from 0.5 to 5.0 mg/ml. No elevated mutant frequencies attributable to treatment were observed in the test under either activated or non‐activated conditions. In the micronucleus assay, mice were injected intraperitoneally (IP) with either 350, 1,173, or 2,500 mg/kg of isopropanol at constant volumes of 10 ml/kg. No increased incidence of micronuclei was observed in bone marrow polychromatic erythrocytes (PCEs) harvested at 24, 48, or 72 hr post‐dosing. In both assays, negative and positive control mutant frequencies were within historical control ranges. These results, in conjunction with previously published data, clearly demonstrate that isopropanol is not a mutagen. © 1993 Wiley‐Liss, Inc.
Use of fluorescence in situ hybridization to detect chromosome‐specific changes in exfoliated human bladder and oral mucosa cells
Authors: Moore Lee E. , Titenko‐Holland Nina , Smith Martyn T.
Published: February 2007

Change in chromosome number, numerical aneuploidy, has been consistently linked with cancer development. Since 90% of cancers arise in epithelial tissues, techniques that measure aneuploidy in these tissues would be very useful. Here we describe methods of optimization and suggest use of fluorescent in situ hybridization (FISH) to detect aneuploidy in exfoliated epithelial cells collected from the mouth and bladder. A total of 10,383 urothelial cells and 4,691 buccal cells were scored in order to determine a baseline frequency of aneuploidy in human volunteers using a classical satellite probe for chromosome 9. Protein digestion with pepsin was found to be more efficient at removing the keratinized cell membrane and optimizing probe penetration than acid washes, detergent washes, or hypotonic treatments. A 20 min cellular digestion with 200 μg/ml and a 30 min digestion with 300 μg/ml of pepsin in 0.01 M HCI optimized probe penetration in urothelial and buccal cells, respectively. Average frequencies for 0, 1, 2, 3, and 4 hybridization regions were 10.3, 10.1, 78.4, 1.0, and 0.3% for urothelial cells and 8.8, 9.8, 79.4, 1.3, and 0.3% for buccal cells, respectively. These results are very similar to those previously described in lymphocytes. The urothelial cells of males had a lower frequency of diploid cells and a higher frequency of cells without hybridization regions than females (P < 0.02). No statistically significant variability was found between individuals or sex groups in buccal cells. Our data show that FISH is a useful tool to detect changes in frequency of aneuploidy in exfoliated epithelial cells and has good potential for monitoring human populations exposed to genotoxic agents. © 1993 Wiley‐Liss, Inc.
Spectrum of spontaneous HPRT mutations in TK6 human lymphoblasts
Authors: Giver Cynthia R. , Nelson Stephen L. , Grosovsky Andrew J.
Published: February 2007

The occurrence of deletions, coding sequence alterations, and intronic changes leading to aberrant splicing has been characterized among 33 spontaneous HPRT− mutants in TK6 human lymphoblasts. Deletions detectable by multiplex PCR amplification accounted for 45% (15/33) of the mutant collection. Base substitutions represented 30% (10/33) of the total, and were predominated by changes at G:C base pairs. The remaining mutants were distributed among frameshifts (9%, 3/33), small deletions (6%, 2/33), and compound alterations (9%, 3/33). Five mutants (15%) demonstrated aberrant splicing of the hprt transcript. A cluster of 4 deletion/insertion events was identified in hprt exon 6. A nearly perfect 13 bp duplication differed from the original sequence only by an A:T to G:C transition, which was observed as a unique alteration in another HPRT− mutant. A model involving correction of a mismatch in a secondary structure formed by the duplicated sequence may account for these results. © 1993 Wiley‐Liss, Inc.