bluuebunny/crossref_metadata_2025 huggingface dataset:
Mechanism of chromosome exchange formation in human fibroblasts: Insights from “chromosome painting”
Authors: Brown J. M. , Evans J. W. , Kovacs M. S.
Published: February 2007

We have used the techniques of premature chromosome condensation (PCC) and fluorescence in situ hybridization (FISH) with a library for human chromosome 4 to analyze the rate of rejoining of chromosome breaks and development of exchange aberrations in AG1522 human fibroblasts. AG1522 cells were irradiated in plateau phase with 10 Gy and fused with mitotic HeLa cells either immediately after irradiation or at intervals up to eight days later. The slides were then hybridized with the chromosome 4 library and unrejoined breaks and exchange events (visualized as bicolor chromosomes) scored in these cells. At the earliest time point after irradiation, the number of exchange events in the irradiated cells was low, but increased with kinetics similar to that of the joining of the breaks. Furthermore, when we analyzed those cells which had exchange events for their distribution, almost all of the cells initially contained one exchange event (1 bicolor chromosome). As time progressed, the number of cells containing exchanges with two exchange events per cell increased as the number with one exchange event per cell decreased. Extrapolation of the number of exchange events to zero time (with an estimate of 20 min for the fusion and condensation times) gave a value consistent with zero exchanges at zero time after irradiation. In a separate experiment, we also scored AG 1522 cells at the first metaphase after a dose of 6 Gy and were able to show that as many as 50% of the complete exchanges were non‐reciprocal in nature, that is, the two broken ends of a single break in chromosome 4 joined to two different chromosomes. These data support the classical breakage‐and‐reunion model rather than the Revell Exchange Theory of exchange formation. © 1993 Wiley‐Liss, Inc.
Histone shuttle driven by the automodification cycle of poly(ADP‐ribose) polymerase
Authors: Althaus Felix R. , Höfferer Liane , Kleczkowska Hanna E. , Malanga Maria , Naegeli Hanspeter , Panzeter Phyllis , Realini Claudio
Published: February 2007

In mammalian cells, the incision step of DNA excision repair triggers a dramatic metabolic response in chromatin. The reaction starts with the binding of a zinc‐finger protein, i.e. poly‐(ADP‐ribose)polymerase to DNA nicks, activation of four resident catalytic activities leading to poly(ADP‐ribose) synthesis, conversion of the polymerase into a protein modified with up to 28 variably sized ADP‐ribose polymers, and rapid degradation of polymerase‐bound polymers by poly(ADP‐ribose)glycohydrolase. This automodification cycle catalyzes a transient and reversible dissociation of histones from DNA. Shuttling of histones on the DNA allows selected other proteins, such as DNA helicase A and topoisomerase I, to gain access to DNA. Histone shuttling in vitro mimics nucleosomal unfolding/refolding in vivo that accompanies the postincisional steps of DNA excision repair. Suppression of the automodification cycle in mammalian cells prevents nucleosomal unfolding and nucleotide excision repair. © 1993 Wiley‐Liss, Inc.
Assessing the risk of heritable gene mutation in mammals: Drosophila sex‐linked recessive lethal test and tests measuring DNA damage and repair in mammalian germ cells
Authors: Bentley Karin S. , Sarrify Awni M. , Cimino Michael C. , Auletta Angela E.
Published: February 2007

The former U.S. EPA OPPT tiered test scheme for heritable gene mutations included the Drosophila sex‐linked recessive lethal (SLRL) test in which positive results triggered the mouse specific locus (MSL) test. However, review of available literature indicated that the evaluation of mutations in the germ cells of this insect is not a good predictor of the risk of heritable gene mutations in mammals. The database contained 29 compounds for which there were conclusive MSL test results in either spermatogonial and/or postspermatogonial cells. Results in the SLRL test were available for 27 of those compounds. Of the 24 SLRL‐positive chemicals, only 13 (54%) induced heritable mutations in mice; the three SLRL‐negative compounds were nonmutagenic in mouse germ cells. The overall concordance between the two tests was 59%. In contrast, results of unscheduled DNA synthesis (UDS: 18 chemicals) and alkaline elution (AE: 14 chemicals) assays in rodent testicular cells following in vivo exposure correlated well with results in the MSL test (83% and 86%, respectively). MSL test results in spermatogonia and postspermatogonia were also compared separately to the SLRL, UDS, and AE assays. The concordances for the two cell types in the SLRL relative to the MSL test were 36% and 79%, respectively, indicating that the SLRL test is extremely poor in predicting heritable gene mutations in mammalian spermatogonia. Concordances for UDS and AE assays relative to MSL test results in spermatogonia (53% and 54%, respectively) and postspermatogonia (91% and 100%, respectively) were greater. Based on these analyses, the U.S. EPA OPPT has revised its tiered test scheme using assays for interaction with gonadal DNA (e.g., UDS and AE) in place of the SLRL test. © 1994 Wiley‐Liss, Inc.
Overview of the workshops on statistical analysis of mutation data from transgenic mice
Authors: Gorelick N. J. , Thompson E. D.
Published: February 2007

Early results from two transgenic mouse mutation assays, Big Blue™ [Kohler SW et al. (1991): Proc Natl Acad Sci USA 88:7958‐7962] and Muta™ Mouse [Myhr BC (1991): Environ Mol Mutagen 18:308‐315], raised questions about appropriate study design and methods for statistical analysis. First, there were a number of potential sources of variability in the technical aspects of the assay. These are ‘how we do it in our laboratory’ differences, which, if not controlled, ultimately make inter‐laboratory comparison of data difficult. Second, separate from the technical sources of variability were a number of study design issues, e.g., how many animals are needed in each treatment group, how many times should the animal be dosed, what is the appropriate expression period for a particular tissue, how many plaques need to be collected from each animal, and so on.

To address these questions and to identify and understand the sources of variability in mutation data from these systems, a workshop was held in June, 1992 in Cincinnati, Ohio, USA. A core group of biologists and statisticians discussed possible sources of bias (tissue sampling, phage recovery and transgene recovery), possible sources of variability in mutant frequency (between animals, protocol‐based sources, and design‐based sources) and assay sensitivity. Following two days of discussion on protocol design and assay procedures, three action steps were recommended: (1) compile a data base of existing mutation data in transgenic mice to study its statistical features, (2) develop standard protocols for the mutation assays; and (3) use the standard protocol to generate a large data base of mutant frequencies in liver DNA from untreated mice for statistical study and analysis. This report summarizes the proceedings and recommendations of the workshop. The progress made toward these recommendations was reviewed in a second workshop, held in April, 1993, in Norfolk, Virginia, part of which is the subject of the accompanying paper by Piegorsch et al. To date, a standard protocol has been developed for the Big BlueTM mutagenesis assay and a data base of over 90 million plaques from seven labs using either the Big BlueTM or MutaTM Mouse system has been assembled, including a large data set of spontaneous liver mutant frequencies in the Big BlueTM system. © 1994 Wiley‐Liss, Inc.
Sources of variability in data from alacltransgenic mouse mutation assay
Authors: Piegorsch Walter W. , Lockhart Ann‐Marie C. , Margolin Barry H. , Tindall Kenneth R. , Gorelick Nancy J. , Short Jay M. , Carr Gregory J. , Thompson Edward D. , Shelby Michael D.
Published: February 2007

Experimental features of a transgenic mouse mutation assay based on a lacl target transgene from Escherichia coli are considered in detail. Sources of variability in the experimental protocol that can affect the statistical nature of the observations are examined with the goal of identifying sources of excess variation in the observed mutant fractions. The sources include plate‐to‐plate (within packages), package‐to‐package (within animals), and animal‐to‐animal (within study) variability. Data from two laboratories are evaluated, using various statistical methods to identify excess variability. Results suggest only scattered patterns of excess variability, except possibly in those cases where genomic DNA from test animals is stored for extended periods (e.g., > 90 days) after isolation from tissues. Further study is encouraged to examine the validity and implications of this time/storage‐related effect.
Chemical mutagenesis testing in Drosophila. IX. Results of 50 coded compounds tested for the national toxicology program
Authors: Foureman P. , Mason J. M. , Valencia R. , Zimmering S.
Published: February 2007

Fifty chemicals were tested for mutagenic activity in post‐meiotic and meiotic germ cells of male Drosophila melanogaster using the sex‐linked recessive lethal (SLRL) assay. As in the previous studies in this series, feeding was chosen as the first route of administration. If the compound failed to induce mutations by this route, injection exposure was used. One gaseous chemical (1,3‐butadiene) was tested only by inhalation. Those chemicals that were mutagenic in the sex‐linked recessive lethal assay were further tested for the ability to induce reciprocal translocations. Eleven of the 50 chemicals tested were mutagenic in the SLRL assay. These included bis(2‐chloroethyl) ether, 1,4‐butanediol diglycidyl ether, 1‐chloro‐2‐propanol, dimethyl methylphosphonate, dimethyl morpholinophosphoramidate, dimethyloldihydroxy‐ethylene urea, 2,2‐dimethyl vinyl chloride, hexamethylphosphoramide, isatin‐5‐sulfonic acid (Na salt), isopropyl glycidyl ether, and urethane. Five of these, including 1,4‐butanediol diglycidyl ether, 2,2‐dimethyl vinyl chloride, hexamethylphosphoramide, isopropyl glycidyl ether, and urethane, also induced reciprocal translocations. © 1994 Wiley‐Liss, Inc.
Mutational spectrum of ICR‐191 at the hprt locus in human lymphoblastoid cells
Authors: Taft Sharon A. , Liber Howard L. , Skopek Thomas R.
Published: February 2007

Human TK6 lymphoblasts were treated with the acridine derivative ICR‐191, and mutants at the hprt locus were isolated. Mutant hprt cDNA was reverse‐transcribed from mRNA, amplified by polymerase chain reaction (PCR), and sequenced. Additions of single G:C base pairs (+1 frameshift mutations) in repetitive G:C sequences were found in 82% (32/39) of the mutants. Sixteen of the +1 frameshifts analyzed were located in a single sequence of six consecutive guanine bases in exon 3. The remaining +1 frameshifts occurred at six different GGG sequences (14 mutants) and a single GGGG sequence (2 mutants) in other hprt exons. The repetitive guanine sequences that underwent frameshift mutagenesis were located in both the transcribed and nontranscribed strands of hprt. No single base deletions (‐1 frameshift mutations) were observed. Base substitutions were observed in 13% (5/39) of the clones analyzed and occurred at both G:C and A:T bases. Loss of exon 4 from the cDNA was also observed in 5% (2/39) of the mutants. Hprt mutants containing seven consecutive guanines (produced from a +1 frameshift in a GGGGGG sequence) were treated with ICR‐191 and wild‐type revertants selected in CHAT medium. Revertants were recovered at a frequency of approximately 10−7 and contained the wild‐type sequence (GGGGGG) in all clones analyzed. The observed frequency of ICR‐191‐induced ‐1 frameshift reversion in the GGGGGGG sequence was ∼500‐fold lower than the estimated frequency of +1 frameshifts observed in the wild‐type GGGGGG sequence following the same ICR‐191 treatment. These results suggest that ICR‐191 produces predominantly +1 frameshift mutations at the hprt locus in human cells. © 1994 Wiley‐Liss, Inc.
Polymerase chain reaction‐directed DNA sequencing of bleomycin‐induced “nondeletion”‐Type, 6‐thioguanine‐resistant mutants in chinese hamster ovary cell derivative AS52: Effects of an inhibitor and a mimic of superoxide dismutase
Authors: An Jie , Hsie Abraham W.
Published: February 2007

Bleomycin‐induced, 6‐thioguanine‐resistant, “non deletion” mutants pretreated with or without either TRIEN (triethylenetetramine), a superoxide dismutase (SOD) inhibitor, or TEMPOL (4‐hydroxy‐2,2,6,6‐tetramethylpiperidine‐1‐oxyl), a SOD mimic, were analyzed by polymerase chain reaction (PCR)‐directed DNA sequencing in a Chinese hamster ovary (CHO) cell derivative, AS52. Among the 23 bleomycin‐induced mutants, six have 3‐bp 5′‐TGA‐3′ deletions in the region of 366‐371, five have single‐base deletions, seven have base substitutions, three have insertions, and two have possible translocations. Among the 16 bleomycin‐induced mutants pretreated with TRIEN, six have the 5′‐TGA‐3′ deletion (366‐371), two have single‐base deletions, one has a 13‐bp deletion, four have single‐base substitutions, one has a double‐base substitution, and two have insertions. Among the 17 bleomycin‐induced mutants pretreated with TEMPOL, six have the same TGA deletions, two have single‐base deletions, two have single‐base insertions, four have single‐base substitutions, one mutant has a 12‐bp deletion, one has a 13‐bp deletion, and one mutant shows no detectable change in its coding region in the DNA sequence. A possible shift from a ROS‐mediated mutational spectrum to a spontaneous mutational spectrum by TRIEN further indicates that reactive oxygen species play an important role in bleomycin mutagenesis in mammalian cells. © 1994 Wiley‐Liss, Inc.

This article is a US Government work and, as such, is in the public domain in the United States of America.
Differential effects of luminol, nickel, and arsenite on the rejoining of ultraviolet light and alkylation‐induced DNA breaks
Authors: Lee‐Chen S. F. , Yu C. T. , Wu D. R. , Jan K. Y.
Published: February 2007

When Chinese hamster ovary cells were treated with ultraviolet (UV) light or methyl methanesulfonate (MMS), a large number of DNA strand breaks could be detected by alkaline elution. These strand breaks gradually disappeared if the treated cells were allowed to recover in a drug‐free medium. The presence of nickel or arsenite during the recovery incubation retarded the disappearance of UV‐induced strand breaks, whereas the disappearance of MMS‐induced strand breaks was retarded by the presence of arsenite or of luminol, a new inhibitor for poly(ADP‐ribose) synthetase. Luminol, however, had no apparent effect on the repair of UV‐induced DNA strand breaks, and nickel had no effect on the repair of MMS‐induced DNA strand breaks. When UV‐ or MMS‐treated cells were incubated in cytosine arabinofuranoside (AraC) plus hydroxyurea (HU), a large amount of low molecular weight DNA was detected by alkaline sucrose sedimentation. The molecular weight of these DNAs increased if the cells were further incubated in a drug‐free medium. This rejoining of breaks in cells pretreated with UV plus AraC and HU was inhibited by nickel and by arsenite, but not by luminol. The rejoining of breaks in cells pretreated with MMS plus AraC and HU was inhibited by luminol and by arsenite, but not by nickel. These results suggest that different enzymes may be used in DNA resynthesis and/or ligation during the repairing of UV‐ and MMS‐induced DNA strand breaks, and that nickel, luminol, and arsenite may have differential inhibitory effects on these enzymes. © 1994 Wiley‐Liss, Inc.
32P‐postlabelling analysis of DNA adducts in white blood cells of humans exposed to residential wood combustion particulate matter
Authors: Heussen G. A. H. , Bouman H. G. M. , Roggeband R. , Baan R. A. , Alink G. M.
Published: February 2007

Residential wood combustion (RWC) in open fireplaces poses a possible health risk because of the emission into the indoor air of mutagenic and carcinogenic compounds. In the present report it was investigated whether this emission leads to enhanced levels of DNA adducts in white blood cells (WBC) of exposed subjects. Under conditions that most likely reflect the Dutch pattern of use of open fireplaces, RWC increased both indoor air mutagenicity and levels of benzo(a)pyrene (B(a)P) and pyrene. The indirect mutagenicity showed a stronger increase than the direct mutagenicity. The increase in indirect mutagenicity was not directly correlated with the increase in the levels of B(a)P and pyrene. 32P‐postlabelling analysis of DNA adducts following nuclease P1 enrichment or butanol extraction revealed low adduct levels. No combustion‐related increase in the amount of adducts was observed. Possible explanations for the lack of correlation between air monitoring data and WBC DNA adduct levels are discussed. © 1994 Wiley‐Liss, Inc.
Comparison of effects of direct‐acting DNA methylating and ethylating agents on inducible gene expression in vivo
Authors: McCaffrey Jennifer , Hamilton Joshua W.
Published: February 2007

Our laboratory is interested in whether chemical carcinogen‐induced DNA damage is non‐ran‐domly distributed in the genome, i.e., “targeted,” at the level of individual genes. As one means of investigating this, we have examined whether carcinogen treatment differentially alters the expression of specific genes in vivo. In this study, we have compared the effects of four direct‐acting simple alkylating agents (methyl methanesulfonate, ethyl methanesulfonate, methylnitrosourea, and ethylnitrosourea) on the steady‐state mRNA expression of a model in‐ducible gene, phosphoenolpyruvate carboxyki‐nase (PEPCK), using the chick embryo as a simple in vivo test system. We observed no effect of any of these four carcinogens on the steady‐state mRNA expression of the constitutively expressed β‐actin, transferrin, or albumin genes in chick embryo liver following a single dose of carcinogen. In contrast, these same treatments significantly altered both the basal and inducible expression of the glucocorticoid‐inducible PEPCK gene. These results support the hypothesis that inducible gene expression is a target for the effects of chemical carcinogens in vivo. In addition, the direction, magnitude, and time course of these effects were agent‐specific. Qualitative and quantitative differences in effects between the methylating and ethylating agents and between the methanesulfonates and nitrosoureas were correlated with differences in their specific patterns of DNA adduct formation, suggesting that different DNA lesions have different effects on inducible gene expression.
Variation of mouse oocyte sensitivity to griseofulvin‐induced aneuploidy and meiotic delay during the first meiotic division
Authors: Marchetti Francesco , Mailhes John B.
Published: February 2007

The effects of varying the time of chemical treatment on the induction of areuploidy and meiotic delay in metaphase II (Mll) oocytes were studied by administering 1,500 mg/kg griseofulvin (GF) at 0, 2, 4, 6, or 8 hr after on injection of human chorionic gonadotrophin (HCG). The results show that the oocytes have a different sensitivity to GF‐induced aneuploidy and meiotic delay during the course of meiotic maturation. Although not restricted to a particular period of meiotic maturation, the frequency of aneuploidy was highest (P < 0.05) when GF was given at 2, 4, or 6 hr after HCG. The maximum frequency of hyperploidy (42.4%) occurred at the 4‐hr treatment time. Also, GF treatment resulted in the induction of meiotic delay as demonstrated by ovulated metaphase I (Ml) and polyploid Mll oocytes. The meiotic delay data depict a period of relative resistance between two periods of sensitivity in that the percentages of ovulated Ml oocytes were 53.3, 21.3, 3.5, 6.7, and 25.7 when GF was given at 0, 2, 4, 6, and 8 hr after HCG, respectively. Also, at these treatment times the percentages of polyploid oocytes were 0.6, 1.7, 7.7, 20.1, and 15.4, respectively. Therefore, the oocytes seem to be more sensitive to GF‐induced meiotic delay during the periods preceding and following meiotic spindle assembly. In conclusion, the results demonstrate that the time of chemical treatment influences the frequency of aneuploidy and the degree of meiotic delay. Also, the results emphasize that to thoroughly characterize the aneugenic potential of a specific chemical several treatment times may be needed. © 1994 Wiley‐Liss, Inc.
Etoposide (VP‐16): Cytogenetic studies in mice
Authors: Agarwal Kalpana , Mukherjee Anita , Sen Soumitra
Published: February 2007

Etoposide (VP 16‐213), the epipodophyllotoxin derivative that is widely used in the treatment of cancer, forms complexes with DNA‐topoisomerase type llα to exert its cytotoxicity. The drug was evaluated in vivo in Swiss albino mouse bone marrow cells for its ability to induce clastogenicity and sister chromatid exchanges (SCEs). Doses of 5, 10, 15, and 20 mg/kg body weight etoposide given intraperitoneally induced a dose‐dependent significant increase of clastogenicity (Trend test, α ≦ 0.05). The aberrations induced were predominanty chromatid types. The drug shows specificity for S‐phase cells: cells harvested 6 and 12 hr posttreatment showed a significantly increased number of damaged cells and aberrations per cell. Doses of 0.5, 1.0, 2.5, 5.0, and 10.0 mg etoposide/kg body weight induced a dose‐dependent significant induction of SCEs (Trend test, α ≦ 0.05). The minimal effective concentration was 0.5 mg/kg body weight. Etoposide significantly prolonged the cell cycle time at all concentrations tested: 12‐13 hr in treated animals vs. 11 hr in control. The results confirm in vivo cell cycle phase specificity of the drug and further designate etoposide as a potent clastogen and a genotoxic agent in mice. © 1994 Wiley‐Liss, Inc.
Automatic scoring of sister chromatid exchanges by image analysis in a dose response experiment
Authors: García‐Sagredo J. M. , Piper J. , Rutovitz D. , Vaquero J. J. , Vázquez‐Mazariego Yolanda
Published: February 2007

A system which automatically selects second division metaphases and then, automatically scores the number of SCEs of each cell is described. In an initial set of experiments, the performance of the components of the system was measured using a data set in which metaphases had been visually classified as either 2nd division or other; and in 2nd division metaphases, every SCE had been marked on a hard copy. SCE scoring had a true positive rate of about 75% and a false positive rate of about 1.5 false SCEs per metaphase analyzed. Second division detection had a true positive rate of 80% and a false positive rate of about 10% of the non‐2nd division cells. Next, the overall system was compared to human visual scoring in a dose‐response experiment by analyzing the effect of mitomycin C on human chromosomes scored visually by two observers and by the fully automatic scoring. Human visual scoring and machine analysis showed similar dose responses, but the variability between them was considerable. © 1994 Wiley‐Liss, Inc.
Correlation between meteorological conditions and mutagenicity of airborne particupate samples in a tropical monsoon climate area from Kaohsiung City, Taiwan
Authors: Lee H. , Su S. Y. , Liu K. S. , Chou M. C.
Published: February 2007

Kaohsiung is a city of 1.5 million located in the southern part of Taiwan. It has a serious air pollution problem mainly attributable to much industrial and commercial activity. In order to estimate the effects of traffic, season, and meteorological conditions on the mutagenicity of Kaohsiung City's urban ambient particulate matter, 624 airborne particulate samples were collected on a weekly basis from 12 locations for an entire year. The mutagenic potential of acetone extracts of air samples was evaluated by the Salmonella/microsomal test with S. typhimurium TA98 in the presence and absence of S9 mixtures. The air samples from November 1990 showed the highest direct and indirect mutagenicity among the 12 months, whereas those from June and July 1991 had the lowest direct and indirect mutagenic activity, respectively. The mutagenicity showed a good correlation with amounts of the acetone extractable matter of airborne particulates. The meteorological conditions, monthly mean precipitation, and wind speed also showed a good correspondence with mutagenicity. Wind direction and temperature had a moderate relationship. The major mutagenic fractions of air samples that had the highest mutagenic activity in a month were purified using Sephadex LH‐20 column chromatography, and the contents of PAHs, 1‐NP, and DNPs were analyzed by HPLC. The characteristic concentration ratios of PAHs indicated that, for the main pollution sources of airborne particulates from Kaohsiung city, the mobile sources were more important than the stationary ones. The total amounts of 1‐NP and DNPs in airborne particulates seemed to correspond to their mutagenicity. Although the total amounts of 1‐NP and DNPs in the air samples correlated with their mutagenicity, the major mutagenic chemicals in the airborne particulate samples from Kaohsiung City need further investigation. © 1994 Wiley‐Liss, Inc.
Effects of spermine on formation of HGPRT− mutants induced by ethylmethanesulfonate, methylmethanesulfonate, and mitomycin C in V79 chinese hamster cells
Authors: Fiorio R. , Vellosi R. , Bronzetti G.
Published: February 2007

Spermine is a polyamine found in bacteria, animal, and plant tissues. It is involved in a variety of biological processes, and its interaction with DMA stabilizes the secondary structure of the double helix. Spermine is one of the first reported antimutagens, reducing the mutation rate in several prokaryotic test systems, while in eukaryotic organisms conflicting results have been obtained. In light of the significant antimutagenic effect of spermine, it is important to evaluate its activity in mammalian cells in culture. The present study was undertaken to evaluate the ability of spermine to suppress the level of HGPRT mutants induced by ethylmethanesulfonate, methylmethanesulfonate, and mitomycin C. Spermine reduced the mutation frequency induced by ethylmethanesulfonate and methylmethanesulfonate but did not affect survival; with mitomycin C survival was reduced but mutation rate was not influenced.
Genotoxic effects of the carbamate insecticide methomyl. I. In vitro studies with pure compound and the technical formulation “Lannate 25”
Authors: Bonatti Stefania , Bolognesi Claudia , Degan Paolo , Abbondandolo Angelo
Published: February 2007

The carbamate insecticide methomyl and the methomyl‐containing technical formulation “Lannate 25” were tested on whole blood human lymphocyte cultures. Both products induced dose‐dependent increases in chromosome aberrations and micronuclei. Lannate 25 induced DNA damage as measured by the alkaline elution assay and hydroxylation of guanine at the C8 position. Sister chromatid exchanges were not increased significantly with either product. Overall, the technical formulation was more active than the pure compound, when compared at similar concentrations of active principle. Moreover, a different ratio of CREST‐positive/CREST‐negative micronuclei was observed with the two products, pure methomyl being relatively more active than Lannate 25 in the induction of CREST‐positive micronuclei. On the basis of these results, previous evaluations of methomyl as a nongenotoxic compound should be reconsidered. © 1994 Wiley‐Liss, Inc.
Assessment of the in vivo genotoxicity of 2‐hydroxy 4‐methoxybenzophenone
Authors: Robison Steven H. , Odio Mauricio R. , Thompson Edward D. , Aardema Marilyn J. , Kraus Albert L.
Published: February 2007

The genotoxic potential of 2‐hydroxy 4‐methoxy‐benzophenone (benzophenone‐3, Bz‐3), a commonly used sunscreen, has been evaluated previously with in vitro systems. Data from Salmonella studies (with and without activation) have been predominantly negative, but two reports have shown weakly positive results in a single bacterial strain under conditions of metabolic activation. In addition, Bz‐3 has been reported to induce chromosome aberrations and equivocal results for sister chromatid exchange in Chinese hamster ovary (CHO) cells. We used the Drosopha somatic mutation and recombination test (SMART) and in vivo cytogenetics in rat bone marrow to define the potential for in vivo expression of this in vitro activity. For the SMART assay, larva from a mating of “multiple wing hair” (mwh) females with heterozygous “flare” (fir) males were exposed to 0,3000, or 3500 ppm Bz‐3 or 25 ppm dimethylnitrosamine (DMN, positive control) for 72 hr. A recombination between the mwh and flr genes produces twin wing spots, while events such as deletions produce single spots. None of the Bz‐3‐treated larva produced flies with significantly more single or multiple wing spots than controls. In contrast, DMN‐treated larva produced flies with significantly more single or multiple wing spots than controls. The in vivo cytogenetic assay in rat bone marrow cells was conducted to evaluate the clastogenicity of Bz‐3. Sprague‐Dawley rats were treated by oral gavage with a single administration of 0.0, 0.5, 1.67, or 5 gm/kg Bz‐3 or a single dose of 5 gm/kg/day Bz‐3 for 5 consecutive days. Cyclophosphamide (CP) was the positive control and was administered at 20 mg/kg with both treatment regimens. Colchicine growth‐arrested bone marrow cells were collected 8 and 12 hr after the single treatment and 12 hr after the last daily treatment. Under either treatment protocol none of the Bz‐3 concentrations caused any significant increase in chromosomal aberrations. Results from these two studies strongly support the conclusion that Bz‐3 is not genotoxic in vivo. © 1994 Wiley‐Liss, Inc.
Response of fibroblast cultures from ataxia‐telangiectasia patients to reactive oxygen species generated during inflammatory reactions
Authors: Ward A. J. , Olive P. L. , Burr A. H. , Rosin M. P.
Published: February 2007

Cells from patients with ataxia‐telangiectasia (AT) are more sensitive than cells from normal individuals to a number of compounds which induce DNA damage via oxygen‐derived free radical attack. We tested the hypothesis that AT cells would show a sensitivity to reactive oxygen species (ROS) generated by activated inflammatory cells. AT cells were exposed to neutro‐phils activated with 12‐O‐tetradecanoyl‐phor‐bol‐13‐acetate (TPA) or to xanthine/xanthine oxidase (X/XO), an enzyme system which generates superoxide and hydrogen peroxide. Induced micronuclei (MN) frequencies (corrected for spontaneous MN frequencies) were significantly higher in AT cell cultures than in cultures from normal individuals (comparison of MN frequencies of AT vs. normal cultures: for treatment with activated neutrophils, P = 0.003; for X/XO, P = 0.05). The comet assay was used to determine whether the elevated chromosomal damage in the treated AT cells was due to a difference in strand breakage or its rejoining. X/XO treatment was used in studies of single‐stranded (SS) DNA breakage, and X‐ray treatment for double‐stranded (DS) DNA damage. AT and normal cells showed no significant differences in the initial levels of SS (P = 0.29) or DS (P = 0.91) DNA damage. Likewise, they exhibited similar rejoining kinetics (rejoining half‐time for SS = 10 min, for DS = 30 min). These data support the involvement of the AT loci in determining a cell's ability to deal with oxidative stress, although the mechanism underlying this effect has yet to be resolved. The data also suggest that AT patients are at elevated risk of sustaining DNA damage in tissues undergoing inflammatory reactions. © 1994 Wiley‐Liss, Inc.
further studies with the somatic white‐ivory system of Drosophila melanogaster: Genotoxicity testing of ten carcinogens
Authors: Batiste‐Alentorn M. , Xamena N. , Creus A. , Marcos R.
Published: February 2007

To provide further background data for the white‐ivory somatic mutation Drosophila assay, ten selected carcinogens (acetamide, acrylamide, benzo(α)pyrene, cyclophosphamide, diethylstilbestrol, 4‐nitroquinoline N‐oxide, propyleneimine, safrole, thiourea, and o‐toluidine) have been tested in this system. Seventy‐two hours after egg laying, larvae were fed with different concentrations of each carcinogen during the rest of their development until pupation, and the genotoxic effects were measured as significant increases in the appearance of visible mutant clones of ommatidia in the eyes of the emerging adult flies. Our results indicate that three of the ten carcinogens tested (cyclophosphamide, 4‐nitroquinoline N‐oxide, and propyleneimine) were strong genotoxic agents, two (diethylstilbestrol and acrylamide) induced significant positive results but without a dose‐response relationship, and safrole was weakly positive. On the other hand, acetamide, benzo(α)pyrene, thiourea, and o‐toluidine were unable to increase the frequency of mutant clones. © 1994 Wiley‐Liss, Inc.
Induction of DNA strand breaks by ethylene oxide in human diploid fibroblasts
Authors: Nygren Jonas , Cedervall Björn , Eriksson Stefan , Duškinská Mária , Kolman Ada
Published: February 2007

In vitro exposure of normal human diploid fibro‐blasts (strain VH‐10) to ethylene oxide (EtO) induced DNA strand breaks in the dose range of 2.5–30 mMh of EtO. Alkaline DNA unwinding (ADU), neutral filter elution (NFE), pulsed field gel electrophoresis (PFGE), and the comet assay were used to measure DNA single (SSBs) and double strand breaks (DSBs).

Different induction rates of SSBs and DSBs, depending on applied method and also on treatment conditions (cells in monolayer or in suspension were used), were found. A dose‐dependent increase of DNA strand breaks was found by the ADU method in the dose range of 2.5–20 mMh of EtO when treatment was performed in monolayer and in suspension. DSBs were detected by NFE only when the cells were treated with EtO in suspension (doses 10–30 mMh). The highest induction rate of DSBs (about 4 DSBs per 100 Mbp per 1 mMh of EtO) was detected in suspension with PFGE applied. We have shown that heat‐labile sites are formed by EtO. Presumably, the different DSB levels detected by PFGE and NFE result from the conversion of these sites to DSBs during cell lysis at elevated temperature in the PFGE method. The results of the comet assay confirmed that apoptotic processes are not involved in the formation of DSBs in our experimental conditions (less than 1% of apoptotic cells were observed at all doses studied). Possible mechanisms for the induction of DNA strand breaks by EtO‐treatment are discussed. The capacity to repair DSBs in EtO‐exposed (5–7.5 mMh) cells was studied, and it was found that a considerable part of the damage (about 50%) could be repaired during 18 hr of incubation. © 1994 Wiley‐Liss, Inc.
Evidence that SCEs induced by mutagens do not occur at the same locus in successive cell cycles: Lack of cancellation in three‐way stained CHO chromosomes
Authors: Cortés F. , Daza P. , Piñero J. , Escalza P.
Published: February 2007

An approach based on the synchronization of CHO cells after a first cell cycle incorporating a relatively low amount of bromodeoxyuridine (BrdUrd) into DNA, followed by mutagenic treatment and subsequent culture for second and third generations of BrdUrd incorporation for the scoring of sister chromatid exchanges (SCEs) per cell cycle in three‐way differentially (TWD) stained chromosomes, has been used to investigate the possible concellation of SCEs. Cancellation is expected to occur if two mutagen‐induced SCEs occur at exactly the same site in subsequent rounds of replication. Lesions in DNA seem to persist and are able to induce SCE throughout two cell cycles after treatment with the three mutagens tested—mitomycin C (MMC), ethyl methanesulfonate (EMS) and ultraviolet (UV) light—though this latter agent was shown as only moderately persistent. Our results seem to indicate that SCEs induced by these mutagens do not take place at the same locus in successive cell generations, as assessed by a lack of SCE cancellation. © 1994 Wiley‐Liss, Inc.
Ascorbic acid (vitamin C) modulates the mutagenic effects produced by an alkylating agent in vivo
Authors: Aidoo Anane , Lyn‐Cook Lascelles E. , Lensing Shelly , Wamer Wayne
Published: February 2007

Recent reports suggest that ascorbic acid (vitamin C) inhibits tumorigenesis as well as exerts a protective effect against mutagenesis in vitro; however, there is no information on its ability to affect gene mutations induced in vivo. In this study, we have investigated the antimutagenic effects of ascorbic acid on the frequency of 6‐thioguanine‐resistant (6‐TGr) T‐lymphocytes produced in Fischer 344 rats dosed with the direct‐acting alkylating agent, N‐ethyl‐N‐nitrosourea (ENU). The freqeuncy of 6‐TGr T‐lymphocytes from the spleen measured five weeks after ENU treatment indicated that ENU produced a substantial mutagenic response. Pretreatment and/or post‐treatment of rats with ascorbic acid administered in the drinking water appeared to inhibit the response, but the inhibition was statistically significant only when data from the various dosing schedules were pooled. In addition, there was no clear dose‐dependency to the inhibitory effect of ascorbic acid. To further evaluate the time effects of the vitamin supplement on ENU mutagenicity, rats were exposed to the mutagen together with ascorbic acid, which was given continuously for the entire duration of the experiment. At specific times after ENU treatment, the frequency of 6‐TGr T‐cells was determined in lymphocytes isolated from the spleen and the thymus. Time‐dependent increases in the frequency of 6‐TGr T‐cells were observed with ENU treatment; ascorbic acid significantly reduced the ENU‐mediated mutagenic responses, most dramatically in the spleen at weeks 6 and 8 (P < 0.0001), and to a lesser extent in the thymus (P < 0.01 at week 6 and P < 0.006 at week 8). Our data suggest that ascorbic acid intake affects the in vivo mutagenicity of ENU, a direct‐acting mutagen/carcinogen, and that the reported inhibitory effects of the antioxidant on carcinogenesis may be partially mediated by its effects on mutagenesis. Although it is difficult to extrapolate from rodent studies to humans, the results presented suggest an explanation for epidemiological data that link vitamin C ingestion with decreased cancer risk. © 1994 Wiley‐Liss, Inc.
mutation spectra in salmonella of complex mixtures: Comparison of urban air to benzo[a]pyrene
Authors: Demarini David M. , Shelton Melissa L. , Bell Douglas A.
Published: February 2007

We used an ion‐exchange procedure coupled to the Salmonella assay to fractionate the dichlo‐romethane‐extractable particulate organics from an urban air sample collected in Boise, ldaho. A resulting base/neutral fraction contained 81% of the mutagenic activity but only 36% of the mass of the unfractionated sample. Chemical analysis showed that polycyclic aromatic hydrocarbons (PAHs) accounted for much of the mutagenic activity of the air sample. Colony probe hybridization, PCR, and DNA sequence analysis were then used to determine the mutations induced by the complex mixtures and a model PAH, benzo[a]pyrene (BAP) in ∼900 revertants of the frameshift hisD3052 allele and ∼400 revertants of the base‐substitution hisG46 allele. The majority (93–94%) of the mutations induced at the frameshift allele in strain TA98 by the whole or base/neutral fraction of the urban air sample was a hotspot 2‐base deletion of a CG or GC within the sequence CGCGCGCG. The remaining mutations were complex frame‐shifts that consisted of −2 or +1 frameshifts associated with a flanking base substitution. BAP induced a somewhat similar pattern of mutations, with 70% being the hotspot mutation, 23% being complex frameshifts, and the remaining being deletions. The inferred base‐substitution specificity associated with the complex frame‐shifts at the hisD3052 allele (primarily G · C→T · A transversions) was consistent with the observation that this same transversion was the primary mutation induced by the whole urban air sample and BAP at the base‐substitution allele in strain TA100. At the frameshift allele, adducts that promote correct incorporation/slippage could account for hotspot mutations, whereas those that promote misincorporation/slippage could account for complex frameshifts. At the base‐substitution allele, a mixture of adducts or of adducts with multiple conformations could account for the observed proportion of transitions and transversions. Combined with the bioassay‐directed chemical analysis, these results from the first mutation spectra of a complex mixture suggest that such spectra reflect the dominance of particular classes of chemical mutagens within the mixture. © 1994 Wiley‐Liss, Inc.
cytogenetic and germ cell effects of phosphine inhalation by rodents: II. subacute exposures to rats and mice
Authors: Kligerman A. D. , Bishop J. B. , Erexson G. L. , Price H. C. , O'Connor R. W. , Morgan D. L. , Zeiger E.
Published: February 2007

Phosphine (PH3) is a highly toxic grain fumigant to which there is significant human workplace exposure. To determine the in vivo cytogenetic effects of inhalation of PH3, male F344/N rats and B6C3F1 mice were exposed to target concentrations of 0, 1.25, 2.5, or 5 ppm PH3 for 6 hr/day for 9 days over an 11 ‐day period. Approximately 20 hr after the termination of exposures, blood was removed from the mice and rats by cardiac puncture and the lymphocytes cultured for analyses of sister chromatid exchanges and chromosome aberrations in rats and mice, and micronuclei (MN) in cytochalasin B‐induced binucleated lymphocytes from mice. In addition, bone marrow (rats) and peripheral blood (mice) smears were made for the analysis of MN in polychromatic and normochromatic erythrocytes. No significant increase in any of the cytogenetic endpoints was found at any of the concentrations examined. These results indicate that concentrations of PH3 up to 5 ppm are not genotoxic to rodents when administered by inhalation for 9 days during an 11 ‐day period as measured by several cytogenetic assays. To evaluate the effects of PH3 on male germ cells, a dominant lethal test was conducted in male mice exposed to 5 ppm PH3 for 10 days over a 12‐day period and mated to groups of untreated females (2 females/male) on each of 6 consecutive 4‐day mating intervals. None of the 6 groups of females exhibited a significant increase in percent resorptions. These results indicate that exposure to 5 ppm PH3 by inhalation does not induce dominant lethality in male mouse germ cells at steps in spermatogenesis ranging from late differentiating spermatogonia/early primary spermatocytes through mature sperm. © 1994 Wiley‐Liss, Inc.
Reciprocal mitotic recombination is the predominant mechanism for the loss of a heterozygous gene in saccharomyces cerevisiae
Authors: Acuña Gonzalo , Würgler Friedrich E. , Sengstag Christian
Published: February 2007

The loss of a functional copy of a heterozygous tumor suppressor gene represents an important step during neoplastic transformation. In order to learn more about the genetic events that lead to spontaneous and drug‐induced loss of heterozygosity, a diploid Saccharomyces cerevisiae strain was constructed that allows the detection of the loss of a heterozygous gene by means of direct selection. The strain contains a single functional URA3 gene copy inserted at the ADE2 locus located on the right arm of chromosome 15. In addition, the chromosome contains two other phenotypic marker genes, HIS3 which is located distal from URA3, and PHO80 which is closely linked to the centromere. The homologous chromosome lacks all three marker genes. Loss of the heterozygous copy of URA3 can easily be detected by 5‐fluoro‐orotic acid resistance of the resulting clones. Simple phenotypic tests of the resistant clones further allows one to distinguish whether the loss of the URA3 gene copy occurred by crossing over, chromosomal loss, or point mutation and gene conversion. Loss of heterozygosity was found to be induced in a dose‐dependent fashion by UV radiation and by several chemical agents. All the tested mutagens induced loss of heterozygosity predominantly by crossing over. © 1994 Wiley‐Liss, Inc.
Excision repair and gene orientation modulate the strand specificity of UV mutagenesis in a plasmid‐borne yeast tRNA gene
Authors: Armstrong John D. , Kunz Bernard A.
Published: February 2007

Ultraviolet (UV) mutagenesis in a plasmid‐borne Saccharomyces cerevisiae tRNA gene (SUP4‐o) occurs preferentially at sites where the pyrimidine in the base pair is part of a dipyrimidine sequence on the transcribed strand. In this study, we examined whether excision repair, or strand identity with respect to DNA replication, influences this strand bias. The specificity of UV mutagenesis was determined for a wild type (RAD) strain and an isogenic excision repair‐deficient (rad1) derivative, each carrying SUP4‐O on the vector YCpMP2, or another vector (YCpJA1) that differed only in the orientation of SUP4‐O with respect to a unique origin of replication. Most (≧ 90%) of the SUP4‐O mutations induced by UV in these strains were single base pair substitutions, predominantly (> 87%) transitions. The rad1 defect and inversion of SUP4‐O in the RAD strain eliminated the strand preference, whereas inversion of SUP4‐O in the rad1 strain caused it to reappear. Both conditions also altered the distribution of frequently mutated sites and the relative fraction of transitions at TT sequences. These results suggest that excision repair and gene orientation can be important determinants for the strand and site specificities of UV mutagenesis in SUP4‐O on YCpMP2 and YCpJA1. We consider several possible explanations for our observations, including potential roles for transcription by RNA polymerase II, sequence context effects on the efficiency of excision repair, and inherent differences in strand mutability or translesion synthesis by the leading and lagging strand DNA replication complexes. © 1995 Wiley‐Liss, Inc.
Effects of X‐irradiation on mouse testicular cells and sperm chromatin structure
Authors: Sailer B. L. , Jost L. K. , Erickson K. R. , Tajiran M. A. , Evenson D. P.
Published: February 2007

The testicular regions of male mice were exposed to x‐ray doses ranging from 0 to 400 rads. Forty days after exposure the mice were killed and the testes and cauda epididymal sperm removed surgically. Flow cytometric measurements of acridine orange stained testicular samples indicated a repopulation of testicular cell types following x‐ray killing of stem cells. Cauda epididymal sperm were analyzed by the sperm chromatin structure assay (SCSA), a flow cytometric measurement of the susceptibility of the sperm nuclear DNA to in situ acid denaturation. The SCSA detected increased susceptibility to DNA denaturation in situ after 12.5 rods of x‐ray exposure, with significant increases following 25 rads. Abnormal sperm head morphology was not significantly increased until the testes were exposed to 60 rads of x‐rays. These data suggest that the SCSA is currently the most sensitive, non‐invasive method of detecting x‐ray damage to testicular stem spermatogonia. © 1995 Wiley‐Liss, Inc.