bluuebunny/crossref_metadata_2025 huggingface dataset:
Statistical analyses for in vitro cytogenetic assays using chinese hamster ovary cells
Authors: Margolin B. H. , Resnick M. A. , Rimpo J. Y. , Archer P. , Galloway S. M. , Bloom A. D. , Zeiger E.
Published: February 2007

It is a widely held view that objective statistical criteria are needed for the evaluation of genetic toxicity assays. This paper presents statistical methods for the analysis of data from in vitro sister chromatid exchange (SCE) and chromosome aberration tests that use Chinese hamster ovary cells. For SCEs, an extensive study of solvent control results demonstrated that there is a substantial interday component of variability in the data, and that a Poisson sampling model is applicable to data generated via the protocol of Galloway et al [1985]. Consequently, a trend test for evidence of a dose response is proposed for such SCE data. As an illustration of this statistical method, analysis of data previously considered to be negative [Gulati et al, 1985] indicates that di(2‐ethyl‐hexyl)phthalate induces a weak, but reproducible, SCE dose response in CHO cells. Monte Carlo methods are used to show that the trend test is more sensitive than four other statistical procedures considered for the analysis of Poisson‐distributed SCEs. A similar trend test for dose response in proportions is proposed for chromosome aberration data, where the percent of cells with chromosome aberrations is the response of interest. Sensitivity (or power) studies indicate that three doses and a control with 50 cells/dose point is a reasonable design for an in vitro SCE study that uses the Galloway et al protocol. For in vitro chromosome aberrations, however, three doses and a control with 100 cells/dose point appears to produce too insensitive an assay; an increase to 200 cells/dose point in the Galloway et al protocol seems worthy of serious consideration.
Comparison of multiple parameters of rodent carcinogenicity and in vitro genetic toxicity
Authors: Tennant Raymond W. , Stasiewicz Stanley , Spalding Judson W.
Published: February 2007

The relationship between chemically induced patterns of tumorigenesis in rodents and of in vitro genetic toxicity was evaluated for 73 substances. Tumorigenicity patterns were defined according to sex and species effects, the induction of common or uncommon tumors, and benign or malignant tumors. These results and the genetic toxicity results derived from the testing of chemicals under code were compared. Chemicals that induced tumors in both sexes of both rodent species (trans‐sex/species carcinogens) were divided into those that showed multiple responses in genetic toxicity assays and those that showed little or no response. Some of the nongenotoxic trans‐sex/species carcinogens exhibit properties that do not necessarily fit classification as only tumor promoters and may involve some other mode(s) of action. Those chemicals showing tumorigenicity in only one of the four groups exposed (uni‐sex/species carcinogens) generally showed little or no response in genetic toxicity assays. Uni‐sex/species carcinogens may be difficult to identify by in vitro assays because of their high tissue specificity. Chemicals that are tumorigenic in both sexes of both species are logically more likely to be tumorigenic in a third species than are those that are tumorigenic in only one sex of the exposed species. Therefore, while positive genetic toxicity test results are not predictive of all carcinogens, a consistent positive response among the multiple endpoints in these assays is more likely to identify chemicals with the potential for trans‐sex/species carcinogenesis. Such trans‐sex/species carcinogens may have the most direct implication for human health effects.
Mutagenicity studies in salmonella: Residues of ozonated and/or chlorinated water fulvic acids
Authors: Kowbel David J. , Ramaswamy S. , Malaiyandi M. , Nestmann Earle R.
Published: February 2007

Aqueous residues of ozonated, chlorinated, and ozonated/chlorinated water fulvic acids (WFA) were tested for induction of His+ reversion in Salmonella typhimurium strain TA100 in fluctuation tests for mutagenicity. The data suggest that ozonation of natural organics present in sources of drinking water can prevent subsequent formation of by‐products of chlorination that are mutagenic in bacteria. Ozonation of the WFA at different pH and at varying dose levels produced residues that were not or were only weakly mutagenic. Chlorination of WFA or of previously ozonated WFA led to residues that were highly mutagenic. However, mutagen formation in the ozonated/chlorinated residues could be prevented, depending upon the pH of the WFA solutions during ozonation—mutagenicity decreased as pH increased. This decrease in mutagenicity is associated with previous observations of enhanced ozone decomposition into its highly reactive oxidant species at higher pH. Since ozonation seems to be more effective at alkaline pH, alkaline raw water sources seem to be the best candidates for water treatment that involves ozonation.
Structure‐activity relationships (SARs) among mutagens and carcinogens: A review
Authors: Frierson Manton R. , Klopman Gilles , Rosenkranz Herbert S.
Published: February 2007

This review is an introduction to methods for evaluating structure‐activity relationships (SARs), and, in particular, to those methods that have been applied to study mutagenicity and carcinogenicity. A brief history and some background material on the earliest attempts to correlate molecular structure and biological activity are included. Most of the discussion focuses on modern methods utilizing extrather‐modynamic and physical property variables such as the Hansch method and SIMCA, and approaches based on molecular connectivity such as the ADAPT, CASE, and Enslein methods. In general, the latter class is potentially the most useful in the study of the large and structurally diverse databases so often encountered in the study of mutagenicity and carcinogenicity. They also are not very sensitive to lab‐to‐lab variances in reported activities and outright misclassifications in activities of some compounds. This is chiefly because the statistical treatments used in these methods tend to dilute the importance of outliers. The methods using physicochemical and extrathermodynamic variables are especially important in relatively small, congeneric databases and can help fine‐tune the role of physicochemical properties in mechanistic hypotheses. All of the above methods have been used to look at mutagenicity and carcinogenicity and some of the results reported in the literature are reviewed here. As far as specific methods go, ADAPT, CASE, SIMCA and the Enslein approach all seem to have similar classification powers (in the range of 75–95%), depending very much on the database studied. The emphasis in this review is on showing that the use of these computer‐aided storage, retrieval and analysis techniques is a timely approach to predicting and even understanding the toxicity of environmental substances. However, each of the methods discussed is still under development, and their potential usefulness for predictive purposes is still being explored.
The use of metabolic inhibitors to compare the excision repair of pyrimidine dimers and nondimer dna damages in human skin fibroblasts exposed to 254‐NM and sunlamp‐produced >310‐NM ultraviolet radiation
Authors: Rosenstein Barry S. , Chao Chuck C.‐K. , Ducore Jonathan M.
Published: February 2007

Normal human skin fibroblasts were exposed to either 0–5 J/m2 of 254‐nm ultraviolet (UV) radiation or 0–50 kJ/m2 of the Mylar‐filtered UV (>310 nm) produced by a fluorescent sunlamp. These cells were then incubated for 0–20 min in medium containing 10 mM hydroxyurea (HU) and 0.1 mM 1‐β‐D‐arabinofuranosyl cytosine (ara C), and the yield of DNA strand breaks was measured by means of the alkaline elution technique. For cells irradiated with 254‐nm UV, which results primarily in the formation of cyclobutane pyrimidine dimers, a rapid increase in DNA strand breaks was detected following incubation with these metabolic inhibitors. In contrast, only a low level of strand breaks formed in cells incubated with HU and ara C after irradiation with approximately equitoxic fluences of sunlamp UV >310 nm, which mainly causes the induction of nondimer DNA lesions. Hence, these results are consistent with the conclusion that the pathways involved in the repair of nondimer DNA damages induced by UV wavelengths >310 nm differ from the repair of pyrimidine dimers.
Urine recovery experiments with quercetin and other mutagens using the ames test
Authors: Busch David B. , Hatcher James F. , Bryan George T.
Published: February 2007

Recovery from urine of the mutagenic activity of 2‐anthramine, cyclophosphamide, 7,12‐dimethylbenz[a]anthracene, 6‐chloro‐9‐((3‐(2‐chloroethylamino)‐propyl)amino)‐2‐methoxyacridine dihydrochloride (ICR‐191), mitomycin‐C, nitrofurantoin, and quercetin was studied with several of the Ames tester strains using acetone‐extracted XAD‐2 columns with yields ranging from 27% to 79%. Dose responses of the pure chemicals were also studied, and results showed TA 97 to be far more susceptible to quercetin mutagenesis than TA 1537. Reducing pour plate agar volume enhanced mutagenesis.
Formaldehyde‐induced acentric chromosome fragments and chromosome stickiness in Chortophaga neuroblasts
Authors: Dowd Mary Ann , Gaulden Mary Esther , Proctor Bertha L. , Seibert G. Burton
Published: February 2007

Embryos of the grasshopper Chortophaga viridifasciata were exposed in vitro to formaldehyde (FA), as formalin, at concentrations ranging from 10−8 M (0.0003 ppm) to 10−3 M (30 ppm) at 38°C. A low frequency of distinct acentric chromosome fragments (0.02–0.04/cell) was observed in the neuroblasts after 1 hr exposure to 7.5 × 10−4 or 10−3 M FA plus 3 hr recovery, but not at lower concentrations, even with 4 hr exposure. There was no obvious relation between distinct fragment frequency and concentration of FA. Neuroblasts with sticky chromosomes were observed at 10−4, 7.5 × 10−4, and 10−3 M FA, the percent of cells with slight, moderate, or severe stickiness varying with FA concentrations. Fragments were associated with the sticky chromosomes. The frequency of these sticky fragments at the two higher concentrations (0.15–0.30/cell) was greater than the frequency of distinct fragments. It is concluded that the distinct acentric fragments induced by FA result from breakage at a single sticky point (slight stickiness) between separating sister chromatids. The chromosome effects observed probably result from the action of daughter products that are formed by the interaction of FA with culture medium components, especially the fetal calf serum.
Potentiation of sister chromatid exchange by 3‐aminobenzamide is not modulated by topoisomerases or proteases
Authors: Morgan William F. , Doida Yukio , Fero Matthew L. , Xi‐Cang Guo , Shadley Jeff D.
Published: February 2007

Poly(ADP‐ribose) is synthesized in response to DNA strand breaks and covalently modifies numerous intracellular proteins. We have proposed that this modification regulates, i.e., inhibits, the activity of these enzymes, e.g., topoisomerases and proteases, which could otherwise cause additional DNA damage or alterations in chromatin structure. Inhibition of poly(ADP‐ribose) polymerase by 3‐amino‐benzamide (3AB) in cells exposed to DNA‐damaging agents would, according to this proposal, eliminate the regulatory role of ADP‐ribosylation. When Chinese hamster ovary cells are cultured with methyl methanesulfonate (MMS) and 3AB, a synergistic increase in sister chromatid exchange frequency is observed. We investigated the regulatory role of poly(ADP‐ribose) polymerase to see if topoisomerases or proteases are involved in this synergistic increase. Cells were exposed to MMS or the intercalating agent 4′‐(9‐acridinylamino)methanesulfon‐m‐anisidide (m‐AMSA), 3AB, and either the topoisomerase inhibitor novobiocin or the protease inhibitor antipain. Neither novobiocin nor antipain affected the synergistic response of MMS and 3AB or the additive response of m‐AMSA and 3AB. These results suggest that topoisomerases or proteases do not account for the effect of 3AB on sister chromatid exchange frequency after DNA damage.
Analysis of the interlaboratory and intralaboratory reproducibility of the enhancement of simian adenovirus SA7 transformation of syrian hamster embryo cells by model carcinogenic and noncarcinogenic compounds
Authors: Schechtman L. M. , Hatch G. G. , Anderson T. M. , Putman D. L. , Kouri R. E. , Cameron J. W. , Nims R. W. , Spalding J. W. , Tennant R. W. , Lubet R. A.
Published: February 2007

The intralaboratory and interlaboratory reproducibility of a DNA virus (SA7) transformation enhancement assay was investigated using nine carcinogenic and noncarcinogenic compounds representing a variety of chemical classes. By the use of standardized procedures designed to limit assay variables, replicate assay data were collected in two independent laboratories and analyzed for concurrence. The carcinogens, 7, 12‐dimethylbenz(a)anthracene, benzo(a)pyrene, and N‐methyl‐N′‐nitro‐N‐nitrosoguanidine yielded reproducible dose‐dependent cytoxicity and positive transformation effects (defined as statistically significant [p ≦ 0.05] enhancement of virus transformation at two or more consecutive dose levels) in all experiments in both laboratories. The carcinogens lead chromate, diethylnitrosamine, 4‐nitroquinoline‐N‐oxide, and 2‐acetylaminofluorene demonstrated enhancement of SA7 transformation at two or more dose levels in 40–50% of the assays. The noncarcinogenic structural analogs anthracene and pyrene consistently did not produce positive assay responses when tested at dose levels up to the limits of solubility. Good interlaboratory concurrence was demonstrated for these model compounds in the Syrian hamster embryo cell‐SA7 assay.
Influence of various parameters on benzo(a)pyrene enhancement of adenovirus SA7 transformation of syrian hamster embryo cells
Authors: Lubet R. A. , Nims R. W. , Kiss E. , Kouri R. E. , Putman D. L. , Schechtman L. M.
Published: February 2007

Several of the major variable factors in the Syrian hamster embryo/simian adeno‐virus SA7 (SHE/SA7) viral enhancement assay were identified and the effects of these parameters on assay sensitivity were assessed. The extent of dose‐dependent cytotoxicity and enhancement of SA7 transformation of primary SHE target cells by benzo(a)pyrene was examined through analysis of data obtained from 37 assays performed over a 2‐year period. The variables analyzed for contribution to assay sensitivity included (a) the number of SA7‐induced transformed SHE cell foci enumerated in ten replicate dishes in the negative control condition (background focus count) (range: 26–139); (b) the age of the SHE cell cultures at the time of exposure to benzo(a)pyrene (range: 72–144 hr postseeding); and (c) the source of the pregnant hamsters used to prepare the primary SHE cells (Wilmington colony vs Lakeview colony, Charles River Laboratories, Inc., Wilmington, MA). The benzo(a)pyrene‐induced cytotoxicity and enhancement of SA7 transformation responses were found to be independent of each of these variables, within the range of values tested.
Mutagenicity study on pyrazole, seven pyrazole derivatives, and two nitroimidazoles with the L‐arabinose resistance test of Salmonella typhimurium
Authors: Alejandre‐Durán Encarnación , Ruiz‐Rubio Manuel , Claramunt Rosa M. , Lápez Concepción , Pueyo Carmen
Published: February 2007

The mutagenicity of pyrazole and seven pyrazole derivatives (4‐nitropyrazole, 4‐bromopyrazole, 1‐methyl‐4‐nitropyrazole, 3,5‐dimethyl‐4‐nitropyrazole, 1‐methyl‐4‐bromopyrazole, 4,4′‐dinitro‐1, 1′‐methylene‐dipyrazole and 4,4′‐dibromo‐1, 1′‐methylene‐dipyrazole) has been investigated with the L‐arabinose forward mutation assay of Salmonella typhimurium. Two nitroimidazoles (1‐methyl‐5‐nitroimidazole and metronidazole) were included as reference drugs. The mutagenicity of each chemical was determined by both preincubation and liquid tests, in the presence or absence of S9 microsomal fraction. The mutagenic response was expressed as the absolute number of L‐arabinose resistant mutants growing in selective plates, supplemented with traces of D‐glucose. Strain BA13 with a wild‐type lipopolysaccharide barrier was used as a comparison to the deep rough derivative BA9. No mutagenic effect was detected with pyrazole and two of its derivatives, 1‐methyl‐4‐bromopyrazole and 4,4′‐dibromo‐1,1′‐methylene‐dipyrazole. The other five pyrazole derivatives were mutagenic to different degrees, although their mutagenic potencies were always considerably lower than those of the two nitroimidazoles. The results suggest that 4‐nitropyrazoles, as well as 4,4′‐dinitro‐1, 1′‐methylene‐dipyrazoles, should be investigated further as alternatives to, or even substitutes for, the currently used nitroimidazoles.
Bioassay of environmental nickel dusts in a particle feeding ciliate
Authors: Smith‐Sonneborn Joan , Leibovitz Brian , Donathan Robert , Fisher Gerald L.
Published: February 2007

The ciliated protozoan Paramecium was used to quantitate cytotoxic and genotoxic effects of nickel particles. The biological response of these eukaryotic cells to pure nickel powder and iron‐nickel powder was assayed and compared to the effect of the inorganic carcinogen nickel subsulfide. Cytotoxicity was determined by the percent survival of treated cells. Genotoxicity was indicated by significant increases in the fraction of nonviable offspring (presumed index of lethal mutations) found after self‐fertilization (autogamy) in parents from the nickel‐treated versus neutral control groups. The cells were exposed to the dusts and the biological effects determined. Only the nickel subsulfide consistently showed a significant increase in offspring lethality.
Environmental tobacco smoke: Comparative characterization by mutagenicity assays of sidestream and mainstream cigarette smoke
Authors: Löfroth Göran , Lazaridis Georgios
Published: February 2007

Mainstream cigarette smoke particles were collected by means of a smoking machine, and sidestream particles were collected from the room in which the smoking took place. The particles were extracted by sonication with acetone, and the extracts were solvent‐exchanged to dimethyl sulfoxide. The samples were tested for mutagenicity in the Ames Salmonella/microsome assay. The mainstream extract is preferentially mutagenic in the presence of S9, with about 30,000 revertants/cigarette in TA98, but has little or no activity in its absence. The sidestream extract is also mutagenic in the presence of S9 with TA98, and this activity is mainly due to basic compounds. Sidestream smoke is also significantly mutagenic in the absence of S9 in the strain TA100 as well as in TA97 and TA104. This “direct” activity is due to components that are labile. The response of sidestream particles is 10,000‐20,000 revertants/cigarette in TA98+S9 and TA100‐S9 when the collection is performed in a room where the particle concentration is modulated by deposition to surfaces. Sidestream particles collected on glass fiber filter and by electrostatic precipitation (ESP) with a commercial air cleaning device gave essentially the same mutagenic response, showing that ESP sampling may be an alternative to filter sampling for environmental tobacco smoke (ETS) in indoor environments. ESP sampling in children's rooms in smoking and nonsmoking homes showed that 5–10% of the tobacco smoke emitted in the smoking homes entered the child's room, demonstrating that diffusion of pollutants is faster than ventilation in modern buildings with low ventilation rates.
Influence of mouse liver stored vitamin A on the induction of mutations (Ames tests) and SCE of bone marrow cells by aflatoxin B1, benzo(a)pyrene, or cyclophosphamide
Authors: Qin S. , Huang C. C.
Published: February 2007

Male mice (C57BL/6J) at 2 weeks of age were divided into two groups and maintained on a vitamin A‐deficient or vitamin A‐ (retinyl acetate) supplemented diet. After 8 weeks, the average liver vitamin A concentration of mice fed on vitamin A‐deficient or ‐supplemented diet was 36 ± 7 μg/g vs 287 ± 22 m̈g/g, respectively. Uninduced liver S9 fractions were prepared from both groups of mice and used to activate (with cofactors) the precarcinogens aflatoxin B1 (AFB), cyclophosphamide (CPP), dimethylbenz(a)anthracene (DMBA), and benzo(a)pyrene (BP) in the Salmonella mutagenicity assay. S9 fraction prepared from both groups of mice failed to activate CPP to metabolites mutagenic in tester strains TA100 and TA1535 or to activate DMBA to metabolites mutagenic in TA100, but effectively activated AFB and BP to metabolites mutagenic in TA98. Comparison of activation activities of S9 prepared from liver of mice fed a high or low level of vitamin A was made with T98 treated with AFB or BP using three doses of S9 (50, 100, and 200 m̈l/plate). S9 fractions from mice with a high liver vitamin A level were consistently less potent than S9 fractions from mice with a low liver vitamin A level in activating AFB to its mutagenic metabolities. This effect was not observed in BP‐treated plates. Administration of AFB to groups of mice with a high liver vitamin A level induced significantly less SCE in bone marrow cells than did administration of AFB to mice with a low liver vitamin A level. This differential sensitivity was not observed when the two groups of mice were treated with either BP or CPP. The possible relationship between vitamin A levels in vivo and mutagenesis or carcinogenesis are discussed briefly.
Genotoxic effects in cultured mammalian cells produced by low pH treatment conditions and increased ion concentrations
Authors: Brusick David
Published: February 2007

A workshop describing the mutagenic effects of low pH and high osmotic levels on cultured mammalian cells was held at the 16th Annual Meeting of the Environmental Mutagen Society in Las Vegas, Nevada, 1985. Data presented in this summary report were generously provided by the five invited speakers.
Genotoxicity studies of methyl isocyanate in salmonella, drosophila, and cultured chinese hamster ovary cells
Authors: Mason J. M. , Zeiger E. , Haworth S. , Ivett J. , Valencia R.
Published: February 2007

The genotoxic effects of methyl isocyanate (MIC) were investigated using four short‐term tests: the Salmonella reversion assay (Ames test), the Drosophila sex‐linked recessive lethal assay, and the sister chromatid exchange (SCE) and chromosomal aberration assays in cultured Chinese hamster ovary (CHO) cells. No evidence was found for the induction of mutations in either Salmonella or Drosophila. MIC did, however, induce SCEs and chromosomal aberrations in CHO cells both in the presence and absence of Aroclor‐induced rat liver S‐9.
Methyl isocyanate: An evaluation of in vivo cytogenetic activity
Authors: Tice Raymond R. , Luke Carol A. , Shelby Michael D.
Published: February 2007

The ability of inhaled methyl isocyanate (MIC) to induce genotoxic and cytotoxic damage in vivo was evaluated by assessing the induction of chromosomal aberrations (CAs) and sister chromatid exchanges (SCEs) in bone marrow metaphase cells, the induction of micronuclei in polychromatic erythrocytes (MN‐PCEs), and the inhibition of bone marrow cellular proliferation and erythropoiesis. B6C3F1 mice were exposed to MIC by two exposure regiments: (a) in two experiments, male mice only were exposed to 3, 10, and 30 ppm for 2 hr(b) in four experiments, male and female mice were exposed to 1 and 3 ppm (in one experiment, to 6 ppm, also, 6 hr per day for 4 consecutive days. The various cytogenetic endpoints were analyzed in bone marrow and peripheral blood (4‐day exposure regimen only) samples taken from bromodeoxyuridine tablet‐implanted animals killed 11 to 22 hr after cessation of the exposure to MIC. Exposure to MIC for 2 hr induced a significant delay in cellular proliferation but did not induce a significant increase in CAs, SCEs (evaluated at 3 and 10 ppm, only) or in bone marrow MN‐PCEs. Also, this exposure regimen did not inhibit the rate of erythropoiesis. Following exposure to MIC for 4 days, a weak but significant increase in CAs and SCEs was observed in male (in one experiment) and in female (in two experiments) mice. The induction was especially apparent in the single experiment in which mice were exposed to 6 ppm MIC. At this concentration, a significant increase in MN‐PCEs in peripheral blood was observed in male but not female mice. Delay in bone marrow cell proliferation was observed in male mice beginning at 3 ppm and in female mice at 6 ppm. The 4‐day exposure regimen resulted also in a depressed rate of erythropoiesis, with male mice appearing to exhibit greater depression than female mice. The results demonstrate that exposure to MIC by inhalation results in bone marrow damage, indicating the systemic genotoxic/cytotoxic activity of MIC and/or reactive metabolites.
Induction of sperm abnormalities in mice by quercetin
Authors: Rastogi P. B. , Levin R. E.
Published: February 2007

Sperm abnormalities in mice induced by IP injection of quercetin at total cumulative doses of 16, 32, 80, and 160 mg/kg b. wt. administered fractionally on 5 successive days were quantitated by assessing the percent of sperm with abnormal heads or tails. Five weeks after the final injection a maximum of 14.8% abnormal sperm were observed with a total quercetin dose of 80 mg/kg b. wt. compared to the spontaneous control of 0.98%. Concomittantly, a 32.1% reduction in testicular weight occurred compared to control mice which was coincident with a reduction in sperm count of 28 %.
Re‐evaluation of 1,2‐dimethylhydrazine in the mouse bone marrow micronucleus assay: Observation of a positive response
Authors: Ashby J. , Mirkova E.
Published: February 2007

1,2‐Dimethylhydrazine showed dose‐related clastogenic activity in the mouse bone marrow micronucleus assay following intubation of an aqueous solution of the dihydrocholoride salt (10 and 50 mg/kg, 24 h sampling). These data counter earlier reports of the inactivity of this material in a rat and a mouse micronucleus assay, following its intraperitoneal injection. In the present study, 2,000 polychromatic erythrocytes were assessed per animal. However, had only 300 cells been assessed for micronucleated polychromatic erythrocytes, it is shown that a negative result could have been recorded. This may explain the earlier and negative results.
Mutagenicity and clastogenicity of acrylamide in L5178Y mouse lymphoma cells
Authors: Moore Martha M. , Amtower Amanda , Doerr Carolyn , Brock Karen H. , Dearfield Kerry L.
Published: February 2007

Acrylamide was tested without exogenous activation in L5178Y/TK+/− −3.7.2C cells for mutation at the thymidine kinase locus and for clastogenicity. Acrylamide gave a positive induced mutagenic response (approximately 70 mutants/106 survivors) when tested at 600–650 μg/ml. The highest dose tested (850 μg/ml) resulted in an induced mutant frequency of approximately 380 mutants/106 survivors (survival = 13%). Acrylamide induced almost exclusively small‐colony mutants, indicating that it might be acting by a clastogenic mechanism. As predicted, acrylamide was clastogenic, inducing both chromatid and chromosome breaks and rearrangements. A clearly positive clastogenic response was observed at both the 750 μg/ml and 850 μg/ml doses, which showed 16 and 64 aberrations per 100 cells, respectively (background = 3 aberrations per 100 cells). These studies indicate that the L5178Y/TK+/− mouse lymphoma assay can detect some chromosomal mutagens (clastogens) that show little activity in other single gene mutation assays, the CHO/HPRT and Salmonella.
Mutagenicity testing of 5‐(4‐nitrophenyl)‐2,4‐pentadien‐1‐al (Spy Dust) and its metabolites in vitro and in vivo
Authors: Zeiger Errol , Shelby Michael D. , Ivett James , McFee Alfred F.
Published: February 2007

5‐(4‐Nitrophenyl)‐2,4‐pentadien‐1‐al (NPPD; spy dust) was tested for mutagenicity in Salmonella and for its ability to induce chromosome aberrations and sister chromatid exchanges (SCE) in cultured Chinese hamster ovary (CHO) cells and mouse bone marrow. Two metabolites of NPPD produced by the rat, 4‐nitrobenzoic acid (NBA) and 4‐nitrohippuric acid (NHA), were also tested in Salmonella and CHO cells. NPPD was mutagenic in Salmonella, and induced low‐level increases in chromosome aberrations and SCE in bone marrow. It did not induce aberrations in CHO cells. NBA was positive in Salmonella and CHO cells, while NHA was negative. The mutagenicity of NPPD in Salmonella was partially, but not completely, eliminated in a strain lacking one of the bacterial nitroreductases.
Evaluation of pluronic® polyol F127 as a vehicle for petroleum hydrocarbons in the Salmonella/microsomal assay
Authors: Marino Dale J.
Published: February 2007

Complex hydrocarbon mixtures have proven difficult to evaluate in in vitro mutagenicity assays owing to their insolubility in aqueous environments. Pluronic® Polyol F127 (BASF Wyandotte, Parsippany, NJ), prepared as a 50% (w/w) solution in absolute ethanol, proved effective in emulsifying various petroleum hydrocarbon fractions. Its effectiveness in the Salmonella/microsomal assay was evaluated using model solutions each comprising a polycyclic aromatic hydrocarbon (PAH) dissolved in mineral oil. The PAHs used were benzo(a)pyrene, 3‐methylcholanthrene, and 7,12‐dimethylbenzanthracene. Model solutions were evaluated neat and as emulsions with the Pluronic® F127 solution or Tween 80. Similar levels of each PAH were prepared in dimethyl sulfoxide (DMSO) for comparison. Cytotoxicity and mutagenesis were evaluated in the preincubation technique using strain TA97. Little or no cytotoxicity or mutagenesis was evident for model solutions tested neat. However, emulsification of these PAH‐laden mixtures with the Pluronic® F127 solution yielded cytotoxic and mutagenic responses similar to, or greater than, those observed for PAHs delivered in DMSO. Model mixtures emulsified with Tween 80 were less active. Study results demonstrate that Pluronic® F127, prepared as a 50% (w/w) solution in absolute ethanol, is an effective vehicle for evaluating the mutagenic potential of complex hydrocarbon mixtures containing PAHs in the Salmonella/microsomal assay. Since PAHs are a class of insoluble hydrocarbons, the results also suggest the potential usefulness of the Pluronic® F127 solution to detect the mutagenicity of other insoluble hydrocarbons and hydrocarbon mixtures.
Plasmid pKM101 muc‐− and muc‐+ mediated anthracycline mutagenicity and cytotoxicity in Salmonella typhimurium
Authors: Thomas Harvey F. , Cole Janice A. , Freeman Brantley
Published: February 2007

Muc (mutagenesis: UV:chemical) genes of plasmid pKM101, along with the chromosomal gene recA are known to be important constituents for full expression of inducible error‐prone DNA repair in Salmonella typhimurium. This study investigates the affects of muc+ pKM101 and three of its derivatives bearing muc− point mutations (pGW1, pGW16, and pGW21) on spontaneous as well as anthracycline‐induced back mutation to histidine independence. Different base‐substitution and frameshift his− tester strains of S typhimurium were treated with the anthracyclines daunomycin, Adriamycin, carminomycin, and 4‐demethoxy‐doxorubicin. In many cases the muc− plasmids did mediate dose‐dependent anthracyline mutagenicity as measured by His+ reversion. In many of the his− strains, the presence of muc+ pKM101 or muc− plasmids also resulted in a concomitant elevation of spontaneous reversion patterns over those seen with the plasmid‐free parent strains. As a result, the sensitivity of most strains to anthracycline mutagenicity (based on His+ revertants/spontaneous background/μg anthracycline) was not enhanced by muc+ pKM101 or its muc− mutant derivatives. In contrast, the low incidence of spontaneous His+ reversion in strains of the hisD3052 series, along with the inherently greater sensitivities of these strains to anthracycline‐induced reversion, demonstrate most dramatically the mutagenesis‐enhancing effects of muc+ pKM101 and muc− plasmids. In these and other cases in which muc− plasmid effects on enhancement of strain sensitivity to mutagenesis or cytotoxicity are observed, the overall spectrum of enhancement is the following, in decreasing order: pKM101 > pGW16 > pGW1 > no plasmid = pGW21. Possible correlations are drawn between muc− plasmid effects on anthracycline‐induced mutagenesis and cytotoxicity.
Genotoxicity of three‐carbon compounds evaluated in the SCE test in vitro
Authors: von der Hude Wilhelm , Scheutwinkel Michael , Gramlich Ulrike , Fibler Brigitte , Basler Armin
Published: February 2007

Three‐carbon chemicals (chlorinated and nonchlorinated, saturated and unsaturated, hydroxy‐ and oxo‐hydrocarbons) were assayed for genotoxicity. The sister chromatid exchange test in vitro served as the test system. Without S9 mix, the nonchlorinated solvents 1‐propanol, 2‐propanol, and 2‐propanone (acetone) did not increase the SCE frequencies. All chlorinated 3‐C hydrocarbons, except 1,2,3‐trichloropropane, proved to be potent SCE inducers in V79 cells without S9 mix.

In the presence of S9 mix, the results obtained with the nonchlorinated solvents were also negative, whereas 1,2,3‐trichloropropane was transformed to SCE‐inducing metabolites. The addition of S9 mix resulted in an increased SCE rate for 2,3‐dichloropropanol, whereas genotoxicity of 2,3‐dichloropropene, 1,2‐di‐chloropropane, 1,3‐dichloropropene, and 1,3‐dichloropropanone was reduced. 1,3‐dichloropropanol, 1,3‐dichloropropene, and epichlorohydrin were substantially inactivated by S9 mix in the V79/SCE test.

It can be concluded that the reactivity of the saturated dichloro compounds in the SCE test depends on the degree of oxidation. There is no general difference between the reactivity of α,β‐dichloro and α,ω‐dichloro compounds.
Evaluation of aflatoxin B1 mutagenesis: Addition of glutathione and glutathione‐S‐transferase to the salmonella mutagenicity assay
Authors: Jorgensen Karen V. , Clayton J. Wesley , Price Ralph L.
Published: February 2007

The effects of glutathione (GSH) and the combination of GSH and glutathione‐S‐transferase (GST) on aflatoxin B1 (AFB1) mutagenesis in the Salmonella mutagenicity assay using Salmonella typhimurium strains TA98 and TA100 were tested. Ten concentrations of AFB1 (0‐1.0 μg/plate) were added to a liver microsomal homogenate (S9 mix) or to S9 mix containing GSH or S9 mix containing the combination of GSH+GST. One‐third of the samples were plated directly. Two‐thirds were incubated for 30 min at 37°C prior to plating, and of those, half included bacteria. All samples were plated according to Ames et al [1975]. The results show that the addition of GSH and GSH+GST affected AFB1 mutagenesis by forming the AFB1‐GSH conjugate and decreasing the availability of AFB1‐8,9‐epoxide. The effect of GST on GSH activity varied with the strain because of the different amounts of S9 mix used. The formation of the AFB1‐GSH conjugate was verified by using reverse‐phase high‐performance liquid chromatography for quantitation of AFB1 and detection of AFB1‐GSH.
Perspective in the environmental health sciences: A meeting report
Authors: Goldstein Linda L. , Rosenkranz Herbert S.
Published: February 2007

On June 26 and 27, 1986, the Charles A. Dana Foundation sponsored the Conference on the Environmental Health Sciences at Case Western Reserve University School of Medicine in Cleveland, Ohio. Hosted by Dr. Herbert S. Rosenkranz, Chairman of the Department of Environmental Health Sciences at Case Western Reserve University, the conference brought together, for the first time, sixteen academic leaders in the field of environmental health from around the country. In five sessions held over two days, the mission of the participants was to discuss the setting of policies and curricular standards for environmental health and to explore the possibility of forming a cohesive advisory group to serve at the national level, on behalf of the discipline.
A variance formula for gene mutation studies using agar plates
Authors: Javitz Harold S.
Published: February 2007

Previous attempts to derive a formula for the variance of the estimated mutant fraction have assumed (without providing justification) that variance components due to treatment, growth and dilution, and plating, are additive. We have derived a variance formula that does not depend on this assumption. Our formula also includes variability attributable to imprecise counting of cells to be plated in the non‐selective conditions.
Sister chromatid exchanges in the lymphocytes of control women, pregnant women, and women taking oral contraceptives: Effects of cell culture temperature
Authors: Ghosh Reita , Ghosh P. K.
Published: February 2007

The incidence of sister chromatid exchange (SCE) was investigated in the lymphocytes of control women, pregnant women, and women using oral contraceptives after culture at 37°C and 40°C. At 37°C, the mean frequency of SCE (MEAN ± S.E.) was found to be 7.91 ± 0.30 in pregnant women and 8.53 ± 0.29 in oral contraceptive users which were significantly higher than the SCE value of 5.56 ± 0.21 found in control women. Increase in growth temperature to 40°C elevated the SCE frequency to 11.86 ± 0.44 in pregnant women, 12.76 ± 0.46 in oral contraceptive users and 7.24 ± 0.26 in control women. These data indicate that there is a differential induction of SCEs following increased cell culture temperature in the lymphocytes of pregnant women and oral contraceptive users, compared with control women.
Heat shock and herpes virus: Enhanced reactivation without untargeted mutagenesis
Authors: Lytle C. David , Carney Paula G.
Published: February 2007

Enhanced reactivation of Ultraviolet‐irradiated virus has been reported to occur in heat‐shocked host cells. Since enhanced virus reactivation is often accompanied by untargeted mutagenesis, we investigated whether such mutagenesis would occur for herpes simplex virus (HSV) in CV‐1 monkey kidney cells subjected to heat shock. In addition to expressing enhanced reactivation, the treated cells were transiently more susceptible to infection by unirradiated HSV. No mutagenesis of unirradiated HSV was found whether infection occurred at the time of increased susceptibility to infection or during expression of enhanced viral reactivation.
Sister chromatid exchange induction near the baseline with low doses of the alkylating agent ccnu
Authors: Best Robert G. , McKenzie Wendell H.
Published: February 2007

Sister chromatid exchange (SCE) and cell‐cycle kinetics were examined at near‐baseline levels in human peripheral lymphocytes exposed to low doses of the potent SCE inducer 1‐(2‐chloroethyl)‐3‐cyclohexyl‐1‐nitrosourea (CCNU), in vitro. A preliminary SCE dose‐response curve was determined with a broad range of doses of CCNU using a single donor. SCE induction was approximately linear over the entire dose range from 5 to 200 μM CCNU. Cell cycle kinetics were retarded in a dose‐dependent manner. A second dose‐response curve from the same donor was constructed using several doses of CCNU between 0.5 and 10 μM to evaluate linearity and uniformity of SCE response near baseline levels. SCE induction was approximately linear between 1.0 and 10.0 μM CCNU. Finally, SCE and cell‐cycle kinetics were examined in 12 donors at doses of 1.0, 5.0, and 10.0 μM CCNU to evaluate the reproducibility of near‐baseline SCE induction over a range of subjects. Cell‐cycle kinetics were retarded at all three doses with a highly significant increase in SCE frequencies at 5.0 and 10.0 μM CCNU. These data suggest that increases in SCE less than twice background can be reliable indicators of genotoxic exposure.
Genotoxic effects of some systemic pesticides: In vivo chromosomal aberrations in bone marrow cells in rats
Authors: Adhikari N. , Graver I. S.
Published: February 2007

The genotoxic effects of five pesticides (benomyl, 2,4‐D, dimecron, monocrotophos, and vitavax) were evaluated in the rat bone marrow cytogenetic assay. The spectrum of aberrations observed included chromatid breaks, chromatid fragments, ring chromosomes, dicentric chromosomes, and chromosome fragments. It was observed that 2,4‐D, dimecron, and vitavax were clastogenic, but the results obtained with benomyl and monocrotophos were equivocal.
Weak and unexpected mutagenicity to salmonella of the rat hepatocarcinogen methapyrilene
Authors: Ashby J. , Callander R. D. , Paton D. , Zeiger Errol , Ratpan Flora
Published: February 2007

The rat liver carcinogen methapyrilene is shown to be a selective mutagen to strain TA1535 of Salmonella typhimurium when tested in the absence of S9 mix and using the standard plate‐incorporation assay protocol. The activity observed was weak but was reproducible for a range of samples on many occasions of test and was not due to impurities. These data contrast with six earlier reports of the inactivity of this chemical in the Salmonella mutation assay.
Synergistic killing of bacteria and phage by polystyrene and ultraviolet radiation
Authors: Hoerter J. , Eisenstark A.
Published: February 2007

Far‐untraviolet (254 nm) irradiation of bacteria and bacteriophage in polystyrene containers resulted in lower number of survivors, compared with irradiation in glass containers. This effect was not observed with near‐UV (300‐400 nm) irradiation.
Increased sister chromatid exchange associated with smoking and coffee consumption
Authors: Reidy John A. , Annest Joseph L. , Chen Andrew T. L. , Welty Thomas K.
Published: February 2007

Sister chromatid exchange (SCE) is a very sensitive cytogenetic assay for detecting exposure to chemical mutagens and carcinogens. One application of SCE is the monitoring of populations believed to be exposed to such agents. We have, however, relatively little knowledge about common lifestyle factors that may influence SCE and therefore complicate any study designed to examine the effects of exposure to genotoxins. In this study, we assessed the effect of cigarette smoking and coffee consumption on SCE. Smoking was associated with an increase of approximately 2 SCEs per cell and a decrease in cell proliferation. A positive linear relationship between SCE and coffee consumption was also observed. This effect was similar for smokers and nonsmokers. Additionally, the folk acid content of cell culture medium seemed to affect neither SCE nor cell proliferation.
Promutagen activation by freshly isolated and cryopreserved rat hepatocytes
Authors: Loretz Linda J. , Wilson Alan G. E. , Li Albert P.
Published: February 2007

The metabolic activation of promutagens by freshly isolated and cryopreserved rat hepatocytes was compared using the Chinese hamster ovary/hypoxanthine‐guanine phosphoribosyltransferase assay (CHO/HGPRT). Cryopreserved rat hepatocytes were equivalent to freshly isolated hepatocytes in their ability to metabolize dimethylbenz(a)anthracene (DMBA) and dimethylnitrosamine (DMN) to active mutagens. Similar dose‐response curves were observed using either freshly isolated or cryopreserved hepatocytes as activating systems after treatment with DMBA (0.1‐1 μg/ml) and DMN (0.075‐0.6 mg/ml). Our results suggest that cryopreserved hepatocytes are similar to freshly isolated hepatocytes as an experimental system for studies on promutagen activation.
Nitrilotriacetic acid (NTA) induces aneuploidy in drosophila and mouse germ‐line cells
Authors: Costa R. , Russo A. , Zordan M. , Pacchierotti F. , Tavella A. , Levis A. G.
Published: February 2007

The ability of nitrilotriacetic acid (NTA) to induce aneuploidy was studied in the germ line of both Drosophila and the mouse. The Free Inverted × Chromosomes (FIX) genetic system, adopting a brooding scheme, was used to detect induced aneuploidy in Drosophila, and a cytogenetic method based on chromosomal counting in secondary spermatocytes was used in the mouse.

In Drosophila a highly significant (P < 0.001) increase of aneuploidies was produced by NTA (5 × 10−2 M), which was greater than that produced by colchicine (7.5 × 10−6 M) and 5‐fluorodeoxyuridine (10−4 M), which were used as positive controls. Brooding effects were observed with NTA, which produced a maximum induction of chromosomal gain in brood I, suggesting a possible stage‐specific action during meiosis. The ability of NTA (275 mg/kg body weight) to induce meiotic aneuploidy (hyperhaploidy) also was confirmed in the mouse (P < 0.001), where all the aneuploidies detected were attributable to treatment of the metaphase I stage.
Genotoxicity evaluation of the tricyclic antidepressants amitriptyline and imipramine using human lymphocyte cultures
Authors: Saxena Renu , Ahuja Y. R.
Published: February 2007

Two tricyclic antidepressants, amitriptyline and imipramine, were evaluated for their in vitro cytogenetic effects in human lymphocyte cultures.

Peripheral blood cultures from three normal healthy donors were set up for 72 hr for each of the drugs. The drugs were added at the start (72‐hr exposure), 24 hr (48‐hr exposure), and 48 hr (24‐hr exposure) after initiation of the cultures. The concentrations evaluated at each exposure time were 50, 250, 1,000, and 10,000 ng/ml for amitriptyline and 25, 500, and 5,000 ng/ml for imipramine. The first two concentrations correspond to the plasma levels of the respective drugs after therapeutic doses. All treatments for a donor were given at the same time. Untreated cultures served as controls for the baseline frequency of the parameters assayed. The parameters assayed were chromosome aberrations, mitotic index, and sister chromatid exchanges (SCEs).

Amitriptyline was found to be nongenotoxic at plasma levels by all the parameters assayed. However, frequencies of chromosome aberrations and SCEs were significantly increased at concentrations 4 and 40 times the plasma level (1,000 and 10,000 ng/ml) although the actual increase was small. The mitotic index was not affected at any concentration.

Though imipramine showed a significant increase in chromosome damage at the upper plasma level and at concentrations higher than that, SCE frequency was significantly increased only at concentration higher than the plasma level (5,000 ng/ml), the actual increase being small for both these parameters. The mitotic index was not affected at any concentration.

These results suggest mat amitriptyline may be a slightly safer drug than imipramine from a genetic point of view.
A role for Connexin43 during neurodevelopment
Authors: Wiencken‐Barger Amy E. , Djukic Biljana , Casper Kristen B. , McCarthy Ken D.
Published: February 2007

Connexin43 (Cx43) is the predominant gap junction protein expressed in premitotic radial glial cells and mature astrocytes. It is thought to play a role in many aspects of brain development and physiology, including intercellular communication, the release of neuroactive substances, and neural and glial proliferation and migration. To investigate the role of Cx43 in brain physiology, we generated a conditional knockout (cKO) mouse expressing Cre recombinase driven by the human GFAP promoter and a floxed Cx43 gene. The removal of Cx43 from GFAP‐expressing cells affects the behavior of the mice and the development of several brain structures; however, the severity of the phenotype varies depending on the mouse background. One mouse subline, hereafter termed Shuffler, exhibits cellular disorganization of the cortex, hippocampus, and cerebellum, accompanied by ataxia and motor deficits. The Shuffler cerebellum is most affected and displays altered distribution and lamination of glia and neurons suggestive of cell migration defects. In all Shuffler mice by postnatal day two (P2), the hippocampus, cortex, and cerebellum are smaller. Disorganization of the ventricular and subventricular zone of the cortex is also evident. Given that these are sites of early progenitor cell proliferation, we suspect production and migration of neural progenitors may be altered. In conclusion, neurodevelopment of Shuffler/Cx43 cKO mice is abnormal, and the observed cellular phenotype may explain behavioral disturbances seen in these animals as well as in humans carrying Cx43 mutations. © 2007 Wiley‐Liss, Inc.