bluuebunny/crossref_metadata_2025 huggingface dataset:
The relationship between lethal mutation yield and intake of ethylnitrosourea (ENU) in Drosophila melanogaster
Authors: Ayaki Toshikazu , Ohshima Kayoko , Okumura Yutaka , Yoshikawa Isao , Shiomi Toshio
Published: February 2007

To estimate the absorbed dose of N‐ethyl‐N‐nitrosourea (ENU) ingested in Drosophila melanogaster, males were fed with sucrose solutions containing various concentrations of ENU plus 3H‐labeled sucrose for 24 hr. Flies showed decreasing intakes with increase in ENU concentration when monitored by intake 3H radioactivity. Absorbed dose, D, per male can be estimated by the following formula: D = vμ C, where v is intake volume per male of sucrose solution at concentration C. Estimating the v value for each ENU solution from 3H radioactivities in male flies, and using the above formula, we reached the conclusion that average absorbed doses of ENU were 0.064, 0.221, and 0.302 nmol, respectively, for the ENU concentrations of 0.03, 0.3, and 1.0 mM. Sex‐linked recessive lethals were measured for males exposed to these sucrose solutions at three different ENU concentrations. Their frequencies increased nonlinearly with increasing exposure doses—ie, ENU concentration C—but linearly with increasing absorbed doses estimated in the above‐mentioned way.
Microbial mutagenicity of isomeric two‐, three‐, and four‐ring amino polycyclic aromatic hydrocarbons
Authors: Later Douglas W. , Pelroy Richard A. , Stewart Dorothy L. , McFall Ten , Booth Gary M. , Lee Milton L. , Tedjamulia Marvin , Castle Raymond N.
Published: February 2007

The isomers of various two‐, three‐, and four‐ring amino polycyclic aromatic hydrocarbons were tested for mutagenic activity using a microbial plate incorporation test with four Salmonella typhimurium strains (TA98, TA 100, TA1535, and TA 1537). All compounds were assayed with an S9 metabolic activating enzyme system. The two‐ring compounds were tested only with TA98. All were weakly mutagenic (1–10 rev/μg) except 2‐aminobiphenyl, which was not mutagenic under these test conditions. All except two of the 13 fused three‐ring compounds (aminofluorenes, aminoanthracenes, and aminophenanthrenes) were active frame shift mutagens; only the aminophenanthrenes were active base‐pair mutagens. The potency of this group of isomeric compounds ranged from moderately (˜ 20 rev/μg) to strongly (> 5,000 rev/μg) mutagenic. As a group, the pericondensed fourring amino compounds were the most mutagenic of the three groups tested. All of the aminofluoranthene and aminopyrene isomers showed significant mutagenic activity with TA98, TA100, and TA1537. In general, the mutagenic potency of the amino polycyclic aromatic compounds tested was highly dependent on the structural position of the amino group.
In vivo persistence of sister chromatid exchanges (SCE) induced by gamma rays in mouse bone marrow cells
Authors: Morales‐Ramírez Pedro , Vallarino‐Kelly Teresita , Rodriguez‐Reyes Regina
Published: February 2007

The sister chromatid exchange (SCE) frequencies induced in bone marrow cells by in vivo irradiation with gamma rays before or after bromodeoxyuridine (Brdurd) incorporation were compared. The frequency of SCE at different postir‐radiation times was also measured in bone marrow cells in vivo, irradiated before Brdurd incorporation. Increased sensitivity to SCE induction by radiation was found in cells after Brdurd incorporation for one cycle when compared with cells irradiated before Brdurd incorporation. The increased SCE frequency persisted for at least 72 hr after the initial irradiation, implying that the gamma ray‐induced lesion(s) capable of eliciting an SCE are persistent and cannot be easily repaired.
Reduced mutant yield at high doses in the Salmonella/activation assay: The cause is not always toxicity
Authors: McGregor Douglas , Prentice Robert D. , McConville Malcolm , Lee Young Jack , Caspary William J.
Published: February 2007

In the Salmonella/activation assay developed by Ames et al [1973, 1975] toxicity is not measured, though it is recognized by the loss of a cloudy appearance on the plate. One approach to the measurement of toxicity is described here and uses a microscope‐linked automated colony counter to estimate the number of micro‐colonies formed by histidine auxotrophs that stop growing after the depletion of histidine. This technique was used to evaluate the effect of toxicity on the revertant count for 16 mutagens, most of which were chosen because, from previous experience, their dose‐response curves manifested a maximum at an intermediate dose tested.

One of the sixteen, 2‐nitrofluorene, was not toxic up to the maximum dose tested. The relationship between mutation and toxicity for the remaining fifteen allowed them to be grouped into two categories: (1) compounds that induced decreases in survival at the same dose at which the number of mutants decreased, and (2) compounds that induced toxicity, but survival was reduced at dose levels higher than those required to reduce the number of mutants. Possible explanations for this reduction of mutant counts occurring with little apparent concomitant increase in toxicity are examined. These results may be significant for attempts to estimate mutagenic potency and, to a lesser extent, construct mathematical models of the Ames test.
Enhancing activity of rat tissue extracts for induction of lambda prophage by L‐azaserine
Authors: Suit J. L. , Cruz B. Miranda‐Da , Sito L. , Rogers A. E.
Published: February 2007

We studied the effect of rat tissue extracts on induction of lambda prophage in Escherichia coli (λ) by L‐azaserine. Hepatic and pancreatic extracts, primarily the cytosolic fraction, markedly increased the rate of induction. Hepatic extracts from lipotrope‐deficient rats were somewhat more active than extracts from normal rats. The enhancing activity in normal rat hepatic cytosol was partially characterized. It reduced by about one‐half the dose of azaserine required for a given purpose. The enhancement was increased by preincubating the bacterial cells with cytosol; cells retained the effect after cytosol was removed. Enhancing activity was inhibited strongly by the amino acids phenylalanine, tryptophan, and tyrosine; to lesser extents by leucine, methionine, and serine; and not at all by proline or glutamine. It was eliminated by dialysis of the cytosol and reduced by omission of nicotinamide adenine dinucleotide phosphate (NADP) from the reaction mixture. Heating the cytosol to 60°C or 80°C or varying the pH of the reaction mixture from 6 to 8 had no significant effect. Treating the cytosol with trypsin appeared to release an inhibitor of the activity. Glutathione, cysteine, and β‐mercaptoethanol also enhanced X induction by azaserine, but the cytosolic activity was not affected by the thiol‐inactivating compound diethylmaleate (DEM). The results suggest that factors in cytosol interact with bacterial cells to facilitate transport of azaserine into the cells, primarily through the aromatic amino acid transport system. A small molecule, not a free thiol compound, appears to be involved. It may serve to establish reducing conditions protective for azaserine, the probable mechanism of action of sulthydryl compounds.
Lack of effect of dibromochloropropane on the mouse testis
Authors: Oakberg E. F. , Cummings C. C.
Published: February 2007

A single intraperitoneal injection of (101 x C3Hf)F male mice with 110 mg/kg of 1,2‐dibromo‐3‐chloropropane (DBCP) showed no effect on spermatogonial stem cells or on differentiating spermatogonia as measured by cell counts made 3, 5, and 8 days after injection.
A comparison of genotoxicity of automotive exhaust particles from laboratory and environmental sources
Authors: Brooks A. L. , Li A. P. , Dutcher J. S. , Clark C. R. , Rothenberg S. J. , Kiyoura R. , Bechtold W. E. , McClellan R. O.
Published: February 2007

This research (1) ranked the genotoxicity of methylene chloride extracts of laboratory and environmentally collected particles and (2) evaluated the role of collection location and sample composition on genotoxic potency. Samples of exhaust from a spark‐ignition automobile, light‐duty diesel automobile, and a heavy‐duty diesel engine operated in a laboratory on a dynamometer were studied, as well as samples taken in a highway tunnel and outside the same tunnel. The tunnel samples were collected 30 m inside or 56 m outside the exit portal at times when between 70%‐95% of the traffic consisted of diesel trucks. In the Ames Salmonella mutagenicity assay, each extract produced a dose‐dependent increase in mutagenicity in strain TA‐98 without addition of liver S‐9 fraction. Extracts from two tunnel samples collected 1 yr apart, and extracts of particles collected outside the tunnel had similar mutagenic activity. The order of mutagenic activity per μg of extract in TA‐98 without S‐9 from the lowest to the highest was environmental sample ⩽ tunnel < heavy‐duty diesel < light‐duty diesel < spark ignition. Addition of S‐9 or testing in Salmonella strains resistant to the mutagenicity of nitroaromatic compounds (TA‐98 NR and TA‐98 1,8‐DNP6) decreased the mutagenic response. With cell killing, sister chromatid exchanges, and mutations as endpoints in Chinese hamster ovary cells (CHO), the order of potency was tunnel < light‐duty < spark‐ignition samples. All three extracts induced a similar amount of mitotic delay per μg with or without S‐9. Enhanced chromosome aberration frequency was detected only in cells exposed to extracts from sparkignition exhaust. The data indicated that genotoxic activity was detected in each particle extract, that the potency ranking was similar using different genetic endpoints, and that the magnitude of the genotoxic potency was similar.
Mutagenicity of azo dyes following metabolism by different reductive/oxidative systems
Authors: Reid Thomas M. , Morton Kenneth C. , Wang Ching Y. , King Charles M.
Published: February 2007

The mutagenic activity of a group of diazo dyes based on benzidine and its congeners was compared following metabolic activation of the dyes through sequential reduction and oxidation. The dyes were reduced by incubating them with either a suspension of rat cecal flora or a hamster S9 mix supplemented with flavin mononucleotide. The products of dye reduction were then subjected to oxidative metabolism by either Aroclor‐induced rat liver S9 or by hamster liver S9; the resultant mutagenic activity was assayed with Salmonella typhimurium TA1538. Fifteen of the 17 compounds tested were mutagenic, and the degree of mutagenicity was affected by the activity of both the reduction and oxidation systems used. Purified dyes required a reductive step to become mutagenic, but several of the crude dyes did not. All the positive compounds, however, were more mutagenic when the reduction step was included. The mutagenicity of the purified dyes was equal to or greater than that of an equimolar amount of benzidine or appropriate benzidine congener. For the crude dyes, there were no consistent quantitative relationships between the mutagenicity of the dye and that expected from the benzidine moiety.
Calibration of the maize yg2 assay using gamma radiation and ethylmethanesulfonate
Authors: Plewa Michael J. , Dowd Patrick A. , Wagner Elizabeth D.
Published: February 2007

A standard protocol for the yellow‐green‐2 (yg2) forward mutation assay in Zea mays is proposed. A detailed calibration of the assay using 137Cs gamma rays and ethylmethanesulfonate (EMS) was conducted. Gamma ray‐induced mutant sectors in leaves 4 and 5 exhibited one‐hit kinetics. The radiation doses ranged from 25 to 500 rads. The mean induced mutation rate per rad of gamma radiation was 4.54 × 10−6. This value was constant for the primordial cells of leaves 4 or 5. The induction of forward mutation by EMS also exhibited one‐hit kinetics in the concentration range 0.25–20 mM (0.33–23.54 μmol EMS/kernel). The mean induced mutation rate per mM EMS was 1.79 × 10−4, and the mean induced mutation rate per μmol of EMS per kernel was 1.52 × 10−4. Using the induced mutation rates for gamma radiation and EMS, the rad equivalent was calculated. One rad of gamma radiation is equivalent to the exposure of a 2.53 × 10−5 M EMS solution or to 2.99 × 10−8 mol of EMS per kernel.
Evaluation of the genotoxicity of process stream extracts from a coal gasification system
Authors: Shimizu R. W. , Benson J. M. , Li A. P. , Henderson R. F. , Brooks A. L.
Published: February 2007

Extracts of three complex organic environmental mixtures, two from an experimental coal gasifier (a raw gas and a clean gas sample) and one from a coke oven main, were examined for genotoxicity. Three short‐term genotoxicity assay systems were used: Ames Salmonella typhimurium reverse mutation assay, Chinese hamster ovary cell/hypoxanthine‐guanine phosphoribosyl transferase (CHO/ HGPRT) gene locus mutation assay, and the Chinese hamster lung primary culture/sister chromatid exchange (CHL/SCE) assay. Aroclor‐1254‐induced rat liver homogenate fraction (S‐9) was required to observe genotoxicity in both gene locus mutation assays (CHO/HGPRT and Ames). The relative survival of CHO cells exposed to extracts was highest in cells exposed to clean gas samples, with the raw gas sample being the most cytotoxic either with or without the addition of S‐9. All three complex mixtures induced sister chromatid exchanges in primary lung cell cultures without the addition of S‐9. The relative genotoxicity ranking of the samples varied between the mammalian and prokaryotic assay systems. Coke oven main extract produced fewer revertants in bacteria than the raw gas sample. However, the coke oven main extract was more genotoxic in the two eukaryotic systems (CHL/SCE and CHO/HGPRT) than was the raw gas sample. The results of all three assays indicate that the cleanup process used in the experimental gasifier was effective in decreasing the genotoxic materials in the process stream. These data also reemphasize the necessity of evaluating genotoxicity of complex mixtures in a variety of short‐term systems.
Detoxification of nitrosamides and nitrosocarbamates in blood plasma and tissue homogenates
Authors: Aukerman Sharon Lea , Brundrett Robert B. , Hartman Philip E.
Published: February 2007

Nitrosamides and nitrosocarbamates exhibit relatively high mutagenic activity in Salmonella when compared with nitrosoureas. This high activity can be accounted for by activation of nitrosamides and nitrosocarbamates by cellular thiols, predominantly reduced glutathione, that are present intracellularly at concentrations in the millimolar range. In striking contrast to the in vitro mutagenicity tests, a number of studies have indicated that nitrosamides and nitrosocarbamates are less potent than nitrosoureas when tested in vivo in model systems such as the mouse. We extend here previous studies [Aukerman et al, 1983] that demonstrate striking chemical decomposition and inactivation of mutagenic activity of nitrosamides and nitrosocarbamates during exposure to murine blood plasma. Plasma glutathione concentrations are inadequate to account for the rapid inactivations noted. Furthermore, the predominant inactivating species is heat‐sensitive, nondialyzable, and is greater than 25,000 daltons in size as judged by ultrafiltration experiments. Serum albumin has some inactivating capacity at the concentration found in undiluted plasma and could account for the very low but significant inactivating capacity of human plasma. On the other hand, serum albumin lacks the potency necessary to account for the extremely high levels of inactivating activity observed in rodent and rabbit plasma. Elsewhere we present evidence that carboxylesterase activity is the predominant inactivating species in mouse plasma [Aukerman et al, 1983; Aukerman, 1983; Brundrett and Aukerman, 1984]. Mouse liver, large intestine, kidney, and stomach have more activity per milligram protein under the assay conditions used than plasma itself. Rat liver S9 is also active at enhancing the decomposition of nitrosamides and nitrosocarbamates; most of this inactivating capacity resides in the microsomal fraction. The relatively rapid detoxification of these N‐nitroso compounds by plasma and other tissues of rodents has important implications regarding the utility of rodents in assessment of tumorigenicity and/ or antitumor activity of these classes of drugs in other animal species. Tests with Salmonella may be of use in estimating relative levels of protection that vary widely among mammalian species.
Chemicals showing no evidence of carcinogenicity in long‐term, two‐species rodent studies: The need for short‐term test data
Authors: Shelby Michael D. , Stasiewicz Stanley
Published: February 2007

Studies to determine the performance of short‐term tests (STT) in predicting chemical carcinogenicity are frequently characterized by a poor representation of noncarcinogens among the test chemicals. As a result, the performance of several STT is well characterized with carcinogenic chemicals, but the ability of the tests to discriminate between carcinogens and noncarcinogens is largely unknown because of the shortage of test results for noncarcinogens.

A list of 70 chemicals that have been tested by the National Cancer Institute or the National Toxicology Program in long‐term rodent carcinogenicity studies and that have yielded no evidence of cancer induction in male and female rats and mice is presented. The list helps document the shortage of STT results of noncarcinogens and provides a potential source of chemicals for consideration in future studies designed to assess the ability of STT to predict chemical carcinogenicity.
Chemical mutagenesis testing in Drosophila. II. Results of 20 coded compounds tested for the national toxicology program
Authors: Zimmering S. , Mason J. M. , Valencia R. , Woodruff R. C.
Published: February 2007

Results are presented from mutagenesis testing in Drosophila of 20 coded compounds. Two compounds were positive in the sex‐linked recessive lethal test, and two were inconclusive. The positive compounds were calcium chromate in adult feeding experiments and ferrocene after adult injection. The two inconclusive compounds were 2,4‐diaminophenol dihydrochloride and hexachiorocyclopentadiene. Compounds that produced a positive response were assayed for chromosome breakage using the conventional translocation test. Calcium chromate was negative in the translocation test, and ferrocene was positive. Many of the test compounds were poorly soluble in water, raising questions regarding the effective concentration to which the flies were exposed.
Effects of some DNA‐ligands and of some mutagenic compounds on the induction of meiotic disomic or diploid yeast products
Authors: Sora Silvio
Published: February 2007

Data were collected following the administration of various compounds to meiotic yeast cultures to investigate their effects on disomic or diploid induction. Epi chlorohydrin, actinomycin D, and 9‐aminoacridine were unable to induce either diploidy or disomy. Ethyl methanesulphonate was active in inducing diploids, while hycanthone induced disomics. β‐Propiolactone, N‐methyl‐N′‐nitro‐N‐nitrosoguanidine, and β‐naphtylamine were able to increase both disomy and diploidy.
Evaluation of chemicals used for drinking water disinfection for production of chromosomal damage and sperm‐head abnormalities in mice
Authors: Meier John R. , Bull Richard J. , Stober Judy A. , Cimino Michael C.
Published: February 2007

Chemical oxidants are commonly added during water treatment for disinfection purposes. These chemicals have not been tested previously for their ability to induce genetic damage in vivo. Chlorine (hypochlorite and hypochlorous acid), monochloramine, chlorine dioxide, sodium chlorite, and sodium chlorate were evaluated for induction of chromosomal aberrations and micronuclei in bone marrow of CD‐I mice, and for induction of sperm‐head abnormalities in B6C3F1 mice. Oral administration of chlorine at pH 8.5 (where hypochlorite predominates) at dose levels equivalent to approximately 4 and 8 mg/kg/day induced significant increases in the level of sperm‐head abnormalities. There was no evidence of other effects with any of the disinfectants. Halogenated acetonitriles, which have previously been shown to form in the stomach following oral dosing of sodium hypochlorite to rats, were also tested in the sperm‐head abnormality assay but gave no indication of an effect.
Mutagenicity testing of Di(2‐ethylhexyl)phthalate and related chemicals in Salmonella
Authors: Zeiger Errol , Haworth Steve , Mortelmans Kristien , Speck William
Published: February 2007

Di(2‐ethylhexyl)phthalate and 33 other phthalates, ethylhexanol derivatives, and related chemicals were tested for mutagenicity in Salmonella typhimurium strains TA98, TA 100, TA 1535, and TA 1537 without metabolic activation and in the presence of rat and hamster liver S‐9 metabolic activation systems. No mutagenic activity was seen with any of the chemicals tested.
Nick translation—a new assay for monitoring dna damage and repair in cultured human fibroblasts
Authors: Snyder Ronald D. , Matheson Dale W.
Published: February 2007

An in vitro assay has been developed to detect DNA damage and repair following chemical treatment of human diploid fibroblasts. DNA damage is measured by following the Escherichia coli DNA polymerase I‐catalyzed incorporation of radiolabeled deoxycytidine triphosphate (dCTP) into the DNA of lysolecithin‐permeabilized cells. DNA strand breaks with free 3′ OH termini serve as template sites for incorporation, and decrease of this incorporation with time, following removal of the test chemical, indicates loss (repair) of initial damage. Inhibition of the DNA excision repair process by the addition of the repair inhibitors arabinofuranosyl cytosine (ara‐C) and hydroxyurea (HU) during the incubation period gives rise to an increased number of template sites, manifesting itself in increased incorporation and indicating the induction of long‐patch excision repair. This nick translation assay, originally proposed by Nose and Okamoto [1983], is very sensitive, allows detection and quantitation of both DNA damage and repair, distinguishes between various types of induced damage, does not require radioactive prelabeling of cells, and circumvents some of the problems inherent in unscheduled DNA synthesis (UDS) assays. The assay is also useful in detecting those agents that inhibit replicative DNA synthesis and/or the excision repair process.

Results presented demonstrate that all 14 direct‐acting carcinogens tested and 8 of 14 carcinogens requiring metabolic activation give positive indication of DNA damage, repair, or both. Eleven of 14 noncarcinogens tested were scored as negative, the other 3 having previously been shown to interact with cellular DNA. This assay is shown to have predictive capability at least equal to that of UDS assays but to allow a broader spectrum of genotoxic effects to be analyzed.
Induction of sister chromatid exchanges by coal dust and tobacco snuff extracts in human peripheral lymphocytes
Authors: Tucker James D. , Ong Tong‐Man
Published: February 2007

The organic solvent extracts of sub‐bituminous coal dust and tobacco snuff, both together and separately, were tested for the induction of sister chromatid exchanges (SCEs) in human peripheral lymphocytes. The results indicate that these extracts induced SCEs, and that when tested together synergistically induced SCEs in two of three donors. Studies with the organic solvent extracts of all five ranks of coal indicate that the extracts of bituminous, lignite, and peat, but not anthracite, induced SCEs. Similar experiments conducted with water extracts show that bituminous, lignite, and peat, but not sub‐bituminous extracts, induced SCEs, and that anthracite was equivocal. To determine whether individuals differed in their SCE responses to coal dust extracts, lymphocytes from five donors were tested with organic solvent extracts of bituminous and sub‐bituminous coal. An analysis of variance indicates that the SCE response was significantly influenced by the donor (p < 0.0001) and each of the two coal extracts (p < 0.0001). From studies of workers occupationally exposed to coal dust, it is known that inhaled coal dust is cleared from the lungs by mucociliary action and introduced into the stomach by swallowing. Coal dust, or coal dust plus snuff, may be responsible for the increased frequency of gastric cancer observed in coal miners. The findings presented here suggest that coal dust, with or without tobacco snuff, may play a role in the elevated incidence of gastric cancer in coal miners. Because water extracts of some ranks of coal induced SCEs, there exists the possibility of adverse environmental effects due to coal leachates.
Chemical mutagenesis testing in Drosophila. III. Results of 48 coded compounds tested for the national toxicology program
Authors: Valencia R. , Mason J. M. , Woodruff R. C. , Zimmering S.
Published: February 2007

Forty‐eight chemicals were tested for mutagenicity in Drosophila melanogaster. Each compound was tested first for the induction of sex‐linked recessive lethals (SLRLs) by feeding the chemical in a solution of 5% aqueous sucrose. If it was not mutagenic by this route, the chemical was tested by injection in a solution of 0.7% aqueous NaCl. If found inactive, the compound was not tested further. If it was mutagenic, the compound was tested for the induction of reciprocal translocations (RTs) using the exposure protocol and germ cell stage that yielded positive results in the SLRL assay. Eight chemicals (diglycidylresorcinol ether, AF‐2, alloxan monohydrate, 1‐aziridine ethanol, p,p'‐DDE, dichloroacetonitrile, 1,3‐dichloropropene, and 3,3′‐dimethylbenzidine dihydrochloride) were found to induce SLRLs. Only diglycidylresorcinol ether also induced RTs.
Autoradiographic detection of diphtheria toxin resistant mutants in human diploid fibroblasts
Authors: Gupta Radhey S. , Singh Bhag
Published: February 2007

An autoradiographic procedure for the detection of diphtheria toxin (DT) resistant (DipR) mutants in human diploid fibroblast (HDF) cells has been developed. The assay is based on the observation that when HDFs from confluent cultures are seeded in medium containing 0.01 flocculating units/ml or higher concentration of DT, protein synthesis in sensitive cells is severely inhibited by 4–6 hr. If at this or later time, a radiolabeled protein precursor (eg, 3H‐leucine) is added to the culture, it is almost exclusively incorporated into the resistant cells, which are then readily identified by autoradiography. The DipR cells can also be identified by labeling in the presence of 3H‐thymidine, although a higher background is observed in these experiments. Reconstruction experiments using DipS and DipR HDFs show that the frequency of heavily labeled cells that are detected by autoradiography show an excellent correlation with the number of DipR cells added and to the number of DipR cells as detected by conventional colony forming assay. These studies provide strong evidence that the labeled cells identified by autoradiography are bona fide DipR mutants. The detection of DipR cells by autoradiography is apparently not affected by the presence of the sensitive cells in the mixtures. The spontaneous frequency of DipR cells in HDFs has been found to be in the range of 1–5 × 10−6, and this increases in a dose dependent manner upon treatment with the mutagen ethyl methanesulfonate. These results indicate that the autoradiographic assay could be used for quantitative mutagenesis. Since the autoradiographic assay does not depend on cell division, it may prove useful in estimating the incidence of pre‐existing mutations in cell populations that either do not divide or have very limited growth potential (eg, lymphocytes, muscle cells, neurons, senescent fibroblasts, etc).
The role of DNA sequence and structure of the electrophile on the mutagenicity of nitroarenes and arylamine derivatives
Authors: Rosenkranz Herbert S. , McCoy Elena C. , Frierson Manton , Klopman Gilles
Published: February 2007

The mutagenicities of a series of nitroarenes and related electrophilic metabolites of aromatic amines were studied in Salmonella typhimurium strains TA98 and TA97, which have, respectively, (GCGC/CGCG)2 and (CCC/GGG)2 as their mutational sites. For electrophiles, primarily bicyclic species, that form adducts with nucleophilic sites other than the C8 of guanine (G), the ratios of activities in the two strains is near unity. On the other hand, for electrophiles consisting of cyclic structures with more than two rings and that are reported to react solely with the C8 of G, the ratio of activities (TA98/TA97) is in excess of 1.6.

Quantum mechanical calculations indicate that (1) in the plane of G, the N7. N3, and 06 of G are more likely nucleophilic targets than C8 and (2) adduct formation at the C8 of G requires electrophilic attack from above or below the plane of G. It is hypothesized that because of steric hindrances, nitroarenes and derivatives containing in excess of two rings cannot react with the preferred nucleophilic sites and, therefore, form adducts with the more accessible C8. However, such an out‐of‐the‐plane attack is influenced by the nature of the nearest neighbor, as evidenced by the difference in mutagenic responses of TA98 and TA97, alternating GCs being preferred over other combinations. Further quantum mechanical calculations indicate that the nature of the nearest neighbor does indeed affect the nucleophilicity of the C8 of G but that the expression of this effect is apparently masked by the overriding steric effects.
Embryonic exposure to aflatoxin‐B1: Mutagenicity and influence on development and immunity
Authors: Dietert R. R. , Qureshi M. A. , Nanna U. C. , Bloom S. E.
Published: February 2007

Chick embryos were mutagenized in ovo in order to study developmentally related alterations in immune functions in survivors of this prenatal toxicant insult. In this experimental system, a single exposure of 6‐day chick embryos to 0.1 μg aflatoxin‐B1 (AF‐B1) in 10 μ1 of acetone was employed, and the control embryos received 10 μ1 of solvent alone. This dosage of AF‐B1 administered to 6‐day embryos was found to increase the incidence of sister chromatid exchanges in blood cells approximately fivefold above the baseline observed in solvent controls. A second sham control, where no solvent was administered, was included in some experiments. The cell cycle times in blood increased slightly during the initial exposure to AF‐B1. However, a majority of the AF‐B1 and acetone exposed embryos survived and hatched without incident. Losses occurred mainly in the latter part of embryogenesis. After hatching, no significant differences were observed in body weight between different treatment groups up to 26 weeks of age and no change in primary humoral immunity was detected. In contrast, two parameters of cell‐mediated immunity, graft vs host (GvH), and cutaneous basophil hypersensitivity (CBH) reactions were both depressed as a result of exposure to AF‐B1. The AF‐B1 treatment group was significantly reduced in the GvH reaction compared with sham‐treated controls. In the CBH assay, AF‐B1‐exposed chicks showed reduced immunity compared with acetone controls. These results suggest that long‐term selective immune depression can occur following embryonic exposure to AF‐B1.
Investigations into the reported ability of cimetidine to initiate UDS in rat hepatocyte primary cultures
Authors: Lefevre P. A. , Ashby John
Published: February 2007

Cimetidine is widely prescribed as a palliative or cure for gastric disorders in man. It is known to be noncarcinogenic to rodents and has been shown to be inactive in a wide range of in vitro and in vivo genotoxicity assays. It was therefore of concern when an injectable formulation of this drug (Cimetex) was reported to initiate UDS in cultured primary rat hepatocytes [Martelli et al, 1983]. We have confirmed the ability of Cimetex to elicit UDS in primary hepatocytes and established that cimetidine itself is inactive. These data indicate that it is not cimetidine per se, but rather its protonated formulation which is responsible for the activity. These observations pose fundamental questions regarding the validity of the UDS end‐point when testing salts in vitro, rather than presenting a challenge to the nongenotoxic status of cimetidine.
Aroclor 1254 pretreatment effects on DNA repair in rat hepatocytes elicited by in vivo or in vitro exposure to various chemicals
Authors: Kornbrust Douglas , Dietz Dennis
Published: February 2007

Inducers of liver mixed function oxidase (MFO) activities have profound effects on the genotoxicity of substances that undergo metabolic activation by the MFO system. The polychlorinated biphenyl mixture Aroclor 1254 is a broad‐spectrum inducer of liver MFO activities that has been employed as a pretreatment to augment the metabolic activation capabilities of rat liver fractions used in a number of short‐term tests for genotoxicity, including the Ames Salmonella/bacterial mutagenicity assay. The present study was designed to characterize the effects of Aroclor pretreatment of rats on the DNA repair responses elicited by various chemicals in the in vitro hepatocyte primary culture/DNA repair (HPC/DR) assay as well as the in vivo/in vitro HPC/DR assay. The amount of DNA repair produced in vitro by diethylnitrosamine (DEN), benzo(a)pyrene (B(a)P), 3‐meth‐ylcholanthrene (3‐MC), 2‐acetylaminofluorene (2‐AAF), o‐aminoazotoluene (o‐AT), and aflatoxin B1 (AFB1) was significantly greater in hepatocytes derived from Aroclor‐pretreated rats than in control rat hepatocytes; in vitro responses to dimethylnitrosamine (DMN), 7,12‐dimethylbenzanthracene (DMBA), benzidine (BZ), and 2‐naphthylamine (2‐NA) were not significantly affected by Aroclor pretreatment. DNA repair elicited by the direct‐acting alkylating agents methyl methanesulfonate and N‐methyl‐N′‐nitro‐N‐nitrosoguanidine was also not increased by Aroclor pretreatment, which indicated that Aroclor does not exert a general stimulatory effect on the hepatocellular DNA repair capacity. Therefore, the pretreatment‐related potentiation of DNA repair observed for 6 out of 12 compounds tested in vitro was considered to be due to enhanced metabolic activation. These results suggested that pretreatment with Aroclor may increase the sensitivity of the in vitro HPC/DR assay to certain compounds. In contrast, Aroclor pretreatment had little effect on the amount of hepatocellular DNA repair elicited by in vivo administration of DMN, DEN, o‐AT, 2‐AAF, 3‐MC, or AFB1, which indicated that this pretreatment regimen may have little utility for improving the sensitivity of the in vivo/in vitro HPC/DR assay.
Rapid analysis of DNA strand breaks in soft tissues
Authors: Morris Stanton R. , Shertzer Howard G.
Published: February 2007

A technique was developed to measure rapidly DNA strand breaks in soft tissues. This method measured the rate of alkaline unwinding of DNA, which was proportional to strand breakage. Alkaline unwinding of DNA was done by treating tissue homogenates with NaOH. Single‐stranded DNA was removed by extraction with aqueous phenol. DNA unwinding was quantitated by measuring the remaining double‐stranded DNA. Using the described technique, a dose‐effect relationship was observed between N‐nitrosodimethylamine (NDMA) and alkaline unwinding of mouse liver DNA.
Mutagenicity of chili extract and capsaicin in short‐term tests
Authors: Nagabhushan M. , Bhide S. V.
Published: February 2007

Vanillin, capsaicin and chili extracts were tested for mutagenicity in Salmonella typhimurium histidine‐deficient tester strains TA 98, TA 100, TA 1535. TA 1537, and TA 1538. Vanillin was nonmutagenic, whereas chili extract and capsaicin were mutagenic with metabolic activation. Capsaicin, an active component of chili extract, was the more potent mutagen. The positive samples were also tested in two mammalian test systems: the micronucleus test and the 8‐azaguanine‐resistant mutagenesis assay that used V79 Chinese hamster cells. It was observed that both were negative for the latter test at the dose level tested, whereas in the micronucleus test, only capsaicin was positive near the LD50 dose. Capsaicin also inhibited DNA synthesis in the testes of Swiss mice injected at two dose levels.
Classifying mutagens as to their specificity in causing the six possible transitions and transversions: A simple analysis using the salmonella mutagenicity assay
Authors: Levin David E. , Ames Bruce N.
Published: February 2007

The standard Salmonella tester strains used to detect base substitution mutations carry the hisG428 ochre mutation (TA102 and TA104) and the hisG46 missense mutation (TA100). These mutations can be reverted by base changes at their mutant his loci or at extragenic suppressor loci. The base changes resulting in each class of revertants of these mutations have been identified, and simple phenotypic screens have been developed to distinguish among them. Revertants at extragenic suppressor loci are distinguished from those at the his loci by their sensitivity to inhibitory histidine analogs. The four ochre suppressor loci of hisG428 are distinguished by their ability to support growth of nonsense mutants of phage P22. These screens are the basis for a rapid and simple system for determining the base substitution specificity of mutagens using hisG428‐ and hisG46‐containing tester strains. Diagnostic mutagens specific for each of the six possible base changes (transitions and transversions) have been identified. Using these diagnostic mutagens, two additional strains, each specifically reverted by a single base substitution mutation, have been developed to provide a minimum of two loci at which to detect each type of base change. The ability of this system to provide detailed information about mutational specificity in a variety of DNA repair backgrounds will allow further elucidation of the mechanisms of mutagenesis and DNA repair.
Sodium arsenite potentiates the clastogenicity and mutagenicity of dna crosslinking agents
Authors: Lee T. ‐C. , Lee K. C. , Tzeng Y. J. , Huang R. Y. , Jan K. Y.
Published: February 2007

To see if sodium arsenite enhances the clastogenicity and the mutagenicity of DNA crosslinking agents, Chinese hamster ovary (CHO) cells and human skin fibroblasts were exposed to cis‐diamminedichloroplatinum (II) (cis‐Pt(II)) or 8‐methoxypsoralen (8‐MOP) plus long‐wave ultraviolet light (UVA) and then to sodium arsenite. The results indicate that the clastogenicity of cis‐Pt(II) and 8‐MOP plus UVA are enhanced by the post‐treatment with sodium arsenite. Chromatid breaks and exchanges are predominantly increased in doubly treated cells. Furthermore, the mutagenicity of cis‐Pt(II) at the hypoxanthine‐guanine phosphoribosyl transferase locus is also potentiated by sodium arsenite in CHO cells.
Chemically induced aneuploidy in mammalian cells: Mechanisms and biological significance in cancer
Authors: Oshimura Mitsuo , Barrett J. Carl
Published: February 2007

A growing body of evidence from human and animal cancer cytogenetics indicates that aneuploidy is an important chromosome change in carcinogenesis. Aneuploidy may be associated with a primary event of carcinogenesis in some cancers and a later change in other tumors. Evidence from in vitro cell transformation studies supports the idea that aneuploidy has a direct effect on the conversion of a normal cell to a preneoplastic or malignant cell. Induction of an aneuploid state in a preneoplastic or neoplastic cell could have any of the following four biological effects: a change in gene dosage, a change in gene balance, expression of a recessive mutation, or a change in genetic instability (which could secondarily lead to neoplasia). To understand the role of aneuploidy in carcinogenesis, cellular and molecular studies coupled with the cytogenetic studies will be required.

There are a number of possible mechanisms by which chemicals might induce aneuploidy, including effects on microtubules, damage to essential elements for chromosome function (ie, centromeres, origins of replication, and telomeres), reduction in chromosome condensation or pairing, induction of chromosome interchanges, unresolved recombination structures, increased chromosome stickiness, damage to centrioles, impairment of chromosome alignment, ionic alterations during mitosis, damage to the nuclear membrane, and a physical disruption of chromosome segregation. Therefore, a number of different targets exist for chemically induced aneuploidy. Because the ability of certain chemicals to induce aneuploidy differs between mammalian cells and lower eukaryotic cells, it is important to study the mechanisms of aneuploidy induction in mammalian cells and to use mammalian cells in assays for potential aneuploidogens (chemicals that induce aneuploidy). Despite the wide use of mammalian cells for studying chemically induced mutagenesis and chromosome breakage, aneuploidy studies with mammalian cells are limited. The lack of a genetic assay with mammalian cells for aneuploidy is a serious limitation in these studies.
Chromosome analysis of trifluorothymidine‐resistant L5178y mouse lymphoma cell colonies
Authors: Blazak W. F. , Stewart B. E. , Galperin I. , Allen K. L. , Rudd C. J. , Mitchell A. D. , Caspary W. J.
Published: February 2007

Cells from small (σ) and large (σ) trifluorothymidine‐resistant (TFTr) colonies induced by chemical mutagen treatment of TK+/‐ L5178Y mouse lymphoma cells were examined for chromosomal abnormalities. Analysis of G‐banded metaphase chromosomes from 34 σ‐TFTr colonies revealed that cells from 20 (59%) possessed one or more chromosomal abnormalities. The most frequent (16/20 colonies) abnormality observed in cells from σ‐TFTr colonies involved the addition of extra chromatin to the distal region of one chromosome number 11. In 13 of these 16 colonies, the origin of the chromatin translocated to chromosome number 11 could not be identified; the chromatin was not missing elsewhere in the genome. The remaining three σ‐TFTr colonies with an abnormal chromosome number 11 had apparently whole chromosomes translocated, in tandem, to the distal region of chromosome number 11. Chromosomal abnormalities observed in cells from σ‐TFTr colonies with normal number 11 chromosomes included 2N/4N and 2N/4N/ 8N mosaicism (two colonies), a Robertsonian translocation involving chromosome 10 and a marker chromosome (one colony), and trisomy 7 (one colony). In most (14/16) σ‐TFTr colonies with structural damage to chromosome number 11, the cells within a colony were heterogeneous in that some possessed chromosomal damage whereas others were apparently normal. Analysis of chromosomes in cells from eight λ‐TFTr colonies revealed one colony in which all cells had a Robertsonian translocation involving chromosomes 1 and 16 plus other structural abnormalities. The chromosomes of cells from the remaining λ‐TFTr colonies were apparently normal.
A comparison of mutagenic and snbryotoxic effects of cyclophosphamide on the chick embryo
Authors: Novotná Boẑena , Jelinek Ríchard
Published: February 2007

The cytogenetic and embryotoxic effects of cyclophosphamide (CP) after injection on day 3 of single doses ranging from 0.1 to 30 μg were investigated in the chick embryo. The occurrence of embryotoxicity was checked on incubation day 8; clastogenic effects as well as mitotic indices were evaluated in primitive blood cells 6, 12, 18, 24 and 30 hr after the administration of CP. Teratogenic doses of CP induced high percentages of aberrant cells with many unstable aberrations. mitotic activity appeared to be inhibited at the early collection intervals. Non‐embryotoxic doses of CP also increased the incidence of aberrant cells without any substantial effect upon mitotic activity. Although the clastogenic effects of CP occur at lower doses than the teratogenic effects, the thresholds for both effects lie in the lower part of the embryotoxicity dose range, as estimated by the Chick embryotoxicity Screening Test (CHEST).
Chromosome aberrations in mouse bone marrow cells following treatment in vivo with vinblastine and colcemid
Authors: Satya‐Prakash K. L. , Liang Jan C. , Hsu T. C. , Johnston Dennis A.
Published: February 2007

The present study was undertaken to determine if chemicals that are capable of inducing mitotic arrest and aneuploidy can also induce chromosomal breakage. Two chemicals, vinblastine and Colcemid, were selected to be studied. Their potentially clastogenic effects were investigated in mouse bone marrow cells in vivo. Two doses of vinblastine, 10−5 M (9 mg/kg) and 10−6 M (0.9 mg/kg), and one dose of Colcemid 10−4 M (37 mg/kg), were administered to mice as single intraperitoneal injections. Bone marrow preparations were made at multiple time periods after injections, ie, 17, 24, 48, 72, and 96 hr. Both these agents, at the concentrations tested, induced mitotic arrest of bone marrow cells within 5 hr after injection. In recovering bone marrow cell populations, 2–3 days after drug administration, significantly elevated levels of chromosomal breakage (mainly the chromatid type) were observed. Vinblastine was found to be much more potent than Colcemid. Since both of these agents affect mainly microtubules, their actions to cause chromosomal breakage are likely to be indirect. Several possible clastogenic mechanisms such as interference with DNA synthesis, active metabolites, and cytoplasmic endonucleases are discussed.
Tests for mutagenic effects of ammoniated glycyrrhizin, butylated hydroxytoluene, and gum arabic in rodent germ cells
Authors: Sheu C. W. , Cain K. T. , Rushbrook C. J. , Jorgenson T. A. , Generoso W. M.
Published: February 2007

Ammoniated glycyrrhizin, butylated hydroxytoluene, and gum Arabic are “generally recognized as safe” (GRAS) substances that are used primarily as additives in foods. These substances were incorporated into rodent diets and fed to male rats and mice for 10 and 8 wk, respectively. The treated male mice and rats were then tested for dominant lethal effects. The mice were also tested for induced heritable translocation. Results of the rat studies indicated a statistically significant dominant lethal effect of each of the compounds tested; however, the biological significance of this response is not known. Results of the mouse dominant lethal and heritable translocation studies, on the other hand, indicated no adverse effects of the compounds tested.
In vitro induction of sister chromatid exchanges and chromosomal aberrations in peripheral lymphocytes of the oyster toadfish and american eel
Authors: Ellingham Tara J. , Christensen Elizabeth A. , Maddock Michael B.
Published: February 2007

A series of experiments was conducted to characterize the proliferation of oyster toadfish lymphocytes in medium containing 5‐bromodeoxyuridine (BrdUrd) and to determine the effectiveness of cytogenetic endpoints for assessing the genotoxic effects of in vitro exposure of toadfish and eel lymphocytes to known mammalian clastogens. Although the rate of proliferation of toadfish lymphocytes was low compared to that of mammalian lymphocytes, the effects of increasing BrdUrd concentrations were similar, in that proliferation exhibited a concentration‐dependent inhibition for concentrations above 10 μM BrdUrd, and sister chromatid exchange (SCE) frequencies exhibited a concentration‐dependent increase for concentrations above 100 μM BrdUrd. Mitomycin C (MMC) and ethylene dibromide (EDB) induced concentration‐dependent increases in chromatid‐type exchange and SCE frequencies with least effective concentrations (control SCE frequency divided by the slope of the least‐squares line) for SCE induction by MMC (6.8 × 10−9 M) and EDB (2.6 × 10−4 M) that were comparable to or slightly lower than those that have been obtained with mammalian in vitro systems. In vitro exposure of toadfish lymphocytes to dimethoate (DIM) induced a concentration‐dependent increase in SCE frequency with a least effective concentration of 2.8 × 10−3 M that was much higher than that observed with mammalian in vitro systems. In vitro exposure of American eel lymphocytes to MMC also induced a concentration‐dependent increase in the frequency of chromosomal aberrations and SCEs with a least effective concentration for SCE induction of 2.0 × 10−9 M. These results indicate that cytogenetic endpoints can be effectively scored with cultured lymphocytes from these and perhaps other fish species with comparable karyotypes that contain an average of at least 0.07 pg DNA/chromosome.