bluuebunny/crossref_metadata_2025 huggingface dataset:
Relationship between ossification and body weight of the CD‐1 mouse fetus exposed in utero to anticonvulsant drugs
Authors: Sucheston Martha E. , Hayes Thomas G. , Eluma Fabian O.
Published: February 2006

The anticonvulsant drugs carbamazepine, Na valproate, and phenytoin have been suspected as a cause of human congenital defects. The malformations produced may include cleft lip and/or palate, heart defects, skeletal defects, and low body weights. Since the toxic effects of these anticonvulsant drugs manifest themselves in terms of fetal growth retardation, evaluation of the state of ossification attained in the fetus is important. In the present study, pregnant CD‐1 mice received on gestational days 8–16 an oral dose of 375, 563, 938 mg/kg of carbamazepine; or 225, 338, 563 mg/kg of Na valproate; or 50, 75, 125 mg/kg of phenytoin. These groups were compared to two control groups. On day 17, the dams were killed by cervical dislocation and one‐third of the live fetuses were weighed and fixed for skeletal examination. Photographs were taken of the fore‐ and hindlimb skeletons. From these photographs, the length and width measurement of ossified regions of the humerus and femur were determined using a Zeiss Video‐Plan Morphometrics Computer. Of the three anticonvulsant drugs studied, the greatest correlation between reduced fetal weights and retarded ossification of the long bones was phenytoin at the 125 mg/kg dosage. Our results also showed that long bone ossification, when compared to the fetal weight, indicated that 59–66% of variability in weight is predictable by bone measurements.
Short‐term developmental toxicity testing: Considerations in the validation process
Authors: Kimmel G. L.
Published: February 2006

The development and validation of short‐term tests that assess developmental toxicity have received increased attention due to the large number of agents that need to be tested and the desirability to reduce the overall use of animals in research and safety evaluation. The Short‐Term In Vivo Reproductive Toxicity Test was developed to help meet this need and to provide a method for prioritizing agents for further testing. This report provides a brief overview of the risk assessment process and a focus on the aspects of test validation that should be considered when evaluating short‐term tests. It is intended to give the reader an appreciation for many of the considerations that must go into developing and validating a short‐term test, and to indicate areas within the risk assessment process where inclusion of such a test may be appropriate.
Further evaluation of an in vivo teratology screen
Authors: Kavlock Robert J. , Short Robert D. , Chernoff Neil
Published: February 2006

The in vivo teratology screening procedure described previously [Chernoff N, Kavlock RJ: J Toxical Environ Health 10:541–550, 1982] was further evaluated using a total of 46 chemicals in 50 different treatment regimens. Pregnant CD‐1 mice were generally treated by oral gavage on days 8–12 of gestation at a dose level predicted from a preliminary range finding study to induce a slight degree of maternal toxicity. The effects on early postnatal growth and viability were compared to results generated from standard mouse teratology bioassays as reported in the literature (there were nine regimens for which no valid comparisons could be made). The procedure correctly categorized 25 of the 30 treatment regimens which were considered developmentally toxic in the mouse, as well as nine of 11 which were considered to be nondevelopmentally toxic in the mouse. Thus, based upon the criteria used in the present study, the assay correctly classified 83% of the chemicals tested as to their effect in a standard mouse bioassay. The five nonconcurring negative findings were likely due to a combination of pharmacokinetic differences between the studies, as well as to the cessation of dosing on day 12, while critical events of organogenesis are still occurring. The assay achieves the requirements for a teratology screening system, but improved predictability would result from the addition of a lower dose level and extension of the dosing period to include later stages of organogenesis.
Currently used alternatives to the Chernoff‐Kavlock short‐term in vivo reproductive toxicity assay
Authors: Christian M. S. , Hoberman A. M. , Lochry E. A.
Published: February 2006

The Chernoff‐Kavlock assay was initially designed to identify developmental toxins within a relatively short time frame, using a mammalian system. Although it has been used for initial prioritization of multiple agents studied concurrently or for dosage‐range evaluations, it has not gained wide usage in a commercial setting. As proposed, use of the Chernoff‐Kavlock assay in an industrial setting is relatively inefficient, in terms of animal usage and data produced, when compared with available alternative study designs. Only a single dosage level was studied, and the data obtained from the assay do not provide sufficient information to meet the minimum requirements of safety evaluations submitted for regulatory review. For these reasons, other methods of prioritization are generally used. Alternate methods used for initial screening and prioritization at our laboratory are: (1) the hydra developmental toxicity assay; and (2) a dosage‐range study in rats. When the results of several pilot and definitive developmental toxicity assays performed in rats were compared, it appeared that the described pilot study served the dual functions of predicting and prioritizing the developmental hazard of the test agent and providing a dosage‐range study that identified the appropriate dosages to be tested in the definitive study.
Assessment of adult toxicity in developmental versus prechronic toxicology studies
Authors: Schwetz B. A. , Moorman M. P.
Published: February 2006

There is no general agreement at the present time regarding the role of maternal toxicity in the manifestation of teratogenic effects or developmental toxicity in animal studies designed to identify the teratogenic potential of chemicals. A survey of 90 teratology studies published in the recent literature revealed wide differences in the amount and nature of maternal toxicity data collected in teratology studies. In addition, the use of the maternal toxicity data in the process of interpreting developmental toxicity varied widely between authors. In comparison with teratology studies, there is more uniformity in the collection of toxicity data in prechronic toxicity studies. The guidance provided by various government agencies on the parameters to be observed in prechronic toxicity studies is much more explicit than that provided regarding the collection of maternal toxicity data in teratology studies. Because of the likely importance of maternal toxicity in the conduct and interpretation of teratology studies, it is recommended that more maternal toxicity data be collected in teratology studies and the parameters be standardized between laboratories so that data are comparable. Guidelines should be established for the use of maternal toxicity data in the interpretation of the results of teratology studies.
Approaches to defining the relationship of maternal and developmental toxicity
Authors: Schardein James L.
Published: February 2006

Maternal and developmental toxicities reported in the literature were examined in an attempt to define more clearly their relationship. Relationships were difficult to ascertain because maternal toxicity end points are not clearly defined, or even assessed, in every study. However, maternal toxicity accompanied by developmental toxicity is the most common outcome of in vivo testing. Approaches to define these associations have included assessment of acute maternal toxicity and teratogenicity; evaluation of maternal toxicity and its association to developmental toxicity in general, and to malformations, specifically; and examination of developmental variations, embryolethality, and altered growth. None has demonstrated an unequivocal relationship between specific maternal and developmental toxicities: Developmental disruption appears not to result unconditionally from maternal toxicity. Maternal “stress” appears to have some impact on development but resists further definition at this time. Variations in association may be due to the extent to which maternal homeostasis has been compromised. Several quantitative approaches to relating maternal toxicity and developmental toxicity in animal systems (ie, relative teratogenic index, adult/developmental toxicity ratio) may provide the most satisfactory means of evaluating developmental toxicity testing for assessment of hazard.
Comparison of maternal and fetal toxic dose responses in mammals
Authors: Rogers John M.
Published: February 2006

The role of maternal toxicity in adverse developmental outcome and the importance of maternal toxicity as a factor in developmental risk assessment have received increasing attention in the recent literature. This paper reviews these concepts and some of the experimental approaches that have been used to assess their importance. The often non‐parallel nature of maternal and fetal toxic dose response curves makes specific comparisons of maternal and fetal toxic doses quite difficult. The use of specific ratios of maternal to fetal toxic doses, such as the A/D ratio and the Relative Teratogenic Index is discussed, including some of the difficulties encountered in assigning such ratios, and a compilation of A/D ratios for compounds for which this ratio could be estimated in more than one mammalian species is included. Results of our studies on the fungicide, dinocap, for which the A/D ratio does not seem to be consistent across species, are briefly reviewed. Maternal toxicity is usually evaluated only on the basis of significant mortality, dose‐related weight loss, or obvious external observations. Some examples of other more specific indications of maternal toxicity that may be involved in teratogenesis are presented.
Developmental toxicity of dinocap in the mouse is not due to two isomers of the major active ingredients
Authors: Rogers John M. , Gray L. Earl , Carver Brenda D. , Kavlock Robert J.
Published: February 2006

Technical‐grade dinocap, a complex‐mixture fungicide, is teratogenic in the CD‐1 mouse, causing cleft palate and otolith defects. In this study we compared the developmental toxicities of 2,4‐dinitro‐6‐(1‐methylheptyl)phenyl crotonate and 2,6‐dinitro‐4‐(1‐methylheptyl)phenyl crotonate, model isomers of the major active ingredients of technical dinocap, to the known teratogenicity of the technical compound. Individual isomers, both isomers combined, or technical dinocap were administered to pregnant mice on days 7–16 of gestation. Some dams were killed at term and litters were removed, dead fetuses and resorptions were counted, and live fetuses were weighed and preserved in Bodian's fixative for examination for cleft palate. Other treated dams were allowed to give birth: postnatal viability and growth, development of swimming behavior, and otolith formation were evaluated. As in previous studies, technical‐grade dinocap caused cleft palate and weight deficits in fetuses at term and increased neonatal mortality and abnormal swimming behavior, torticollis, and deficient otolith formation in surviving pups. Neither of the purified isomers exhibited any developmental toxicity when administered under identical conditions. Thus, it is concluded that these isomers are not the active teratogenic component(s) in technical‐grade dinocap.
Late‐onset cerebellar degeneration in mice induced transplacentally by methylnitrosourea
Authors: Smith Sylvia B. , Brown Cathy B. , Wright Mary E. , Yielding K. Lemone
Published: February 2006

A late‐onset degenerative disease in the cerebellum was produced in the offspring of mice exposed to 1 mg/kg of the direct‐acting DNA alkylating agent methylnirosourea (MNU) on day 16 of gestation. This intrauterine exposure to MNU also provoked a progressive retinal degeneration that was described elsewhere. Mild ataxia in the MNU‐exposed animals was expressed by 20 weeks of age. Although animals appeared normal in the immediate post‐natal period, quantitative histological evaluation of cerebellar coronal sections indicated that MNU‐exposed animals had a significantly greater number of pyknotic Purkinje cells than age‐matched controls. The number of pyknotic Purkinje cells declined with age in the drug exposed animals; however, the percentage of pyknotic Purkinje cells to total number of Purkinje cells still was greater in MNU‐induced animals at 36 weeks than in controls, suggesting that a slow degenerative process was ongoing in the MNU‐exposed animals. Furthermore, the folia were grossly disrupted in 90% of the older MNU‐exposed animals (ages greater than 12 weeks), suggesting permanent cerebellar disruption macroscopically. Such intrauterine exposure to low doses of alkylating agents may be potentially useful in modeling degenerative neuronal diseases.
Variable patterns in anticonvulsant drug‐induced malformations in mice: Comparisons of phenytoin and phenobarbital
Authors: Finnell Richard H. , Shields Heidi E. , Chernoff Gerald F.
Published: February 2006

Anticonvulsant drugs are known to induce a varied pattern of malformation in both humans and in experimental rodent models. Often the clinical overlap between the pattern of defects induced by different anticonvulsant drugs is so striking that many clinicians question the role these compounds play relative to the existing maternal seizure disorder in the etiology of the observed malformations. In three inbred mouse strains exposed to phenytoin or phenobarbital in utero, the pattern of malformation differed markedly. From the types of anomalies observed, it is apparent that phenobarbital induced more malformations, while phenytoin produced a higher frequency of anomalies related to incomplete development. Thus, while there exists a certain degree of similarity between some of the minor features characteristic of each drug‐induced syndrome, there are distinct differences in pregnancy outcome in experimental animals exposed to these drugs. Given the fact that phenobarbital induces more malformations that can be traced to exposure during early organogenesis, it may be wise to consider a therapeutic strategy in which phenytoin is utilized only during organogenesis, and is then replaced with phenobarbital for the remainder of the pregnancy.
Regulatory decision making and the need for and the use of exposure data on pesticides determined to be teratogenic in test animals
Authors: Wang G. M.
Published: February 2006

Major efforts have been made in the development of a system that takes the reviewer step‐by‐step by the use of the developmental toxicity and exposure assessment data to a recommended risk assessment and risk management position. Thereupon a set of regulatory decisions can be arrived at that are backed up by sound scientific analysis. Examples of pesticides that have been found to be teratogenic in animal testing are provided to demonstrate the process of use of exposure data in making risk assessment and management decisions.
Quantitative risk assessment: Qualms and questions
Authors: Karstadt Myra
Published: February 2006

Within the past ten years, quantitative risk assessment has come to play a central role in the federal regulation of carcinogens. Federal agencies use quantitative risk both to set priorities and to establish exposure levels. The use of quantitative risk assessment is mandated by Executive Orders requiring performance of cost‐benefit analysis, although performance of assessments has often been ascribed to the Supreme Court decision in the 1980 “benzene” case.

Quantitative risk assessment is a deeply flawed methodology with results that give a false sense of certainty and objectivity. Also, the nature of quantitative risk assessment methodologies currently in use makes it difficult for members of the public to scrutinize and assess regulatory actions.

Quantitative risk assessment should not be used for regulation of carcinogens, especially not for establishing exposure levels. Instead, alternative methodologies should be utilized, which are appropriately reflective of uncertainty and which are more readily accessible to public scrutiny and participation.
Effects of the antioxidant butylated hydroxytoluene (BHT) on retinal degeneration induced transplacentally by a single low dosage of n‐methyl‐N‐nitrosourea (MNU)
Authors: Smith Sylvia B. , Cooke Carol Beck , Lemone Yielding K.
Published: February 2006

A 1 mg/kg dose of the DNA alkylating agent, N‐methyl‐N‐nitrosourea (MNU), when administered on day 16 of gestation provokes a progressive retinal degeneration in CD‐1 albino mice reared under standard fluorescent lighting conditions (12 hr light : 12 hr dark); this degeneration begins at about 4 weeks post‐natally and worsens with age. It is accelerated by constant fluorescent light exposure but is retarded greatly by constant darkness, suggesting the importance of secondary insults in the post‐natal period for development of the degenerative disease. To determine whether the secondary photochemical damage might be specifically blocked, MNU‐exposed and control animals in the present study were fed an antioxidant‐enriched diet of Purina mouse chow supplemented with 0.75% butylated hydroxytoluene (BHT). A second group of MNU‐exposed and control animals were fed a non‐BHT supplemented standard Purina mouse chow diet. Systematic measurements of the number of rows of photoreceptor cell nuclei, the thickness of the inner/outer segment layer, and the thickness of the whole retina were made, to quantify any degenerative changes in animals 2, 4, 6, and 8 weeks of age. By 8 weeks, retinas of BHT‐fed, MNU‐exposed animals were significantly thicker and had more rows of photoreceptor cell nuclei than regular‐diet, MNU‐exposed animals. Moreover, the retinas of BHT‐fed animals, both for MNU‐exposed and controls, demonstrated sporadic morphologic changes in the form of circular configurations composed of ganglion cells, arcades of nuclear and plexiform layers, and, in one control animal, a hyperplastic nodule. These experiments suggested that MNU‐induced retinal degeneration may be retarded by a BHT‐enriched diet; however, continuous high doses of this compound pre‐ and post‐natally may induce other retinal abnormalities.
Living in a chemical world: Actions and reactions to industrial carcinogens
Authors: Infante Peter F. , Pohl Gwen K.
Published: February 2006

The synthesis of chemical substances has resulted in technological benefits for society and has also caused increased chemical exposures and risks of related cancers. Although progress has been made in cancer treatment, there has been little improvement in real survival time for people who develop the major forms. Therefore, the need to prevent cancer is paramount. The greatest chemical exposures and risk of associated preventable cancers are found in the workplace, but generally environmental exposures receive greater public attention.

Chemical causes of cancer have been identified, and in some cases decades have passed before controls have been adequately instituted. Some companies show a high regard for worker health, while others may minimize or misrepresent the hazards to exposed workers. Medical personnel receive relatively little education in the recognition of industrially related diseases. Workers are often unaware of their risks or are trapped in jobs because of economic necessity, and government has been slow to regulate, in part because of lobbying efforts.

Experience has shown that when chemicals are regulated relatively soon after they are identified as hazardous, lives are saved, and the industry is economically healthy; however, when regulation is delayed, thousands of people die unnecessarily, and the cost to industry and society are phenomenal. Society has the opportunity to apply the knowledge learned from these experiences as we address current problems of chemical pollution.
Beneficial effects of ascorbic acid on preimplantation mouse embryos after exposure to cyclophosphamide in vivo
Authors: Vogei Richard , Spielmann Horst
Published: February 2006

To study mechanisms of embryotoxicity in early pregnancy, we have evaluated the genotoxic and embryolethal effects of ascorbic acid (AA) alone or in combination with cyclophosphamide (CPA). Female mice were exposed on day 3 of pregnancy. Embryotoxicity was investigated at term and genotoxicity shortly after treatment using the chromosomal aberration test and the sister chromatid exchange (SCE) assay as sensitive end points. Additionally, cytotoxic effects were determined by a proliferation test. AA was not found to be embryotoxic, cytotoxic, or genotoxic when given alone. In combination with 10 mg/kg CPA, however, which induced 50% aberrant metaphases, 100% increase SCE frequency, and a strong inhibition of cell proliferation, AA in a dose range of 25–1,600 mg/kg did not change SCE and proliferation, but reduced the rate of aberrant metaphases significantly. This anticlastogenic effect was clearly correlated to a beneficial effect on embryolethality at term when 200 mg/kg ascorbic acid was given in combination with 40 mg/kg CPA.

The results suggest that during early pregnancy AA is not genotoxic even at so‐called megadoses doses, but it seems to protect early embryos against damage induced by genotoxic agents like CPA.
Elevated expression of proto‐oncogenes accompany enhanced induction of heat‐shock genes after exposure of rat embryos in utero to ionizing irradiation
Authors: Higo Hiromi , Lee Juing Yi , Satow Yukio , Higo Ken‐Ichi
Published: February 2006

We have recently found that the effects of exposing rat embryos in utero to teratogens capable of producing cardiac anomalies were expressed later as enhanced induction of heat‐shock proteins (hsp70 family) when embryonic hearts were cultured in vitro. However, it remained to be determined whether heat‐shock proteins are induced in vivo after exposure to teratogens. The heat‐shock response in some mammalian systems is known to be accompanied by elevated expression of proto‐oncogenes. Using gene‐specific DNA probes, we examined the levels of the expression (transcription) of heat‐shock protein genes and two nuclear proto‐oncogenes, c‐fos and c‐myc, in the embryos removed from irradiated pregnant mother rats 4 or 5 days after the irradiation. We found that the levels of expression in vivo of the hsp70 and c‐myc genes in the irradiated embryos increased by approximately twofold as compared with those in the control. The expression in vivo of the c‐fos gene was not detected in either the irradiated or non‐irradiated embryos. After 0.5‐hr incubation in vitro of the embryos, however, the expression of the c‐fos gene in the irradiated embryos was highly enhanced whereas the control showed no changes. Although the exact functions of these gene products still remain obscure, the enhanced expression of hsp70 gene(s) and the nuclear proto‐oncogenes observed in the present study may reflect repair of intracellular damages and/or regeneration of tissue by compensatory cell proliferation, processes that may disturb the normal program of organogenesis.
Hydrogen peroxide mutagenicity towards Salmonella typhimurium
Authors: Kensese Sontin Mossanda , Smith Leland L.
Published: February 2006

In bioassays conducted under controlled, comparable conditions, weak direct mutagenicity responses were observed for hydrogen peroxide in the standard (Ames test) agar plate incorporation bioassay with Salmonella typhimurium strains TA97, TA98, TA102, and TA1537, in a 20 min preincubation test with strains TA97, TA98, TA100, TA102, TA1537, and TA1538, and in a liquid incubation modification using strain TA1537. These results conclusively demonstrate that hydrogen peroxide is a weak mutagen, especially in strains that are sensitive to oxidative damage under suitable bioassay conditions.
Micronucleus formation by benzene, cyclophosphamide, benzo(a)pyrene, and benzidine in male, female, pregnant female, and fetal mice
Authors: Harper Barbara L. , Ramanujam V. M. Sadagopa , Legator Marvin S.
Published: February 2006

Male, female, pregnant female, and fetal ICR mice were compared for their acute sensitivity to four single doses of model carcinogens, as measured by micronucleus formation in polychromatic erythrocytes 24 h after treatment in adult bone marrow and fetal liver at days 17–19 of gestation. Cyclophosphamide caused a dose‐responsive increase in micronuclei in all groups, without a consistent difference based on gender or pregnancy. At doses of 50 and 75 mg/kg given orally to the pregnant female, the fetuses were three to six times as sensitive as was the mother. Benzo(a)pyrene showed a similarly increased sensitivity of the fetus relative to the other groups, although it is a much weaker clastogen. Benzidine did not cause an increase in micronuclei in any group, although it was thought that the fetal liver might have been sensitive enough to detect it, relative to adult bone marrow. Benzene caused much less response in females than in males and almost no response in pregnant females and their fetuses, even though pregnant females metabolized at least half as much of the total dose as did the males (as measured by the presence of urinary metabolites of benzene). No single metabolite of benzene in the urine was consistently correlated with micronucleus formation in the bone marrow. Several factors must be interacting in different ways for different chemicals to influence their clastogenicity.
Rapid evaluation of topoisomerase inhibitors: Caffeine inhibition of topoisomerases in vivo
Authors: Shin Cha‐Gyun , Strayer John M. , Wani Maqsood A. , Snapka Robert M.
Published: February 2006

Caffeine was found to inhibit both type I and type II topoisomerases in vivo as judged by its effects on replicating simian virus 40 (SV40) chromosomes. The study was facilitated by the use of a rapid filter assay for the detection and characterization of topoisomerase inhibitors. The assay, which requires neither purified enzymes nor substrates, was able to identify both antagonists and poisons of type I and type II topoisomerases.
cytogenetic surveillance of workers exposed to genotoxic chemicals: Preliminary experiences from a prospective cancer study in a cytogenetic cohort
Authors: Sorsa Marja , Ojajärvi Anneli , Salomaa Sisko
Published: February 2006

Cytogenetic endpoints, conventionally chromosomal aberrations, and later sister chromatid exchanges and micronuclei have long been used to assess exposure of human populations to genotoxic agents. Although the adverse nature of somatic chromosome damage is recognized at the group level, no ill‐health manifestations have been causally related to cytogenetic damage at the individual level. In work‐related exposures, e.g., ethylene oxide, styrene, benzene, vinyl chloride, and alkylating anticancer agents have been shown to induce somatic chromosomal damage in several studies. For all of these, a carcinogenic risk to humans has also been documented.

The possible association of somatic chromosome damage and cancer will be elucidated in a Nordic prospective study. The objective is to find out the significance of a high or low score in any of the cytogenetic parametres to risk of cancer. In the Finnish part of the cohort of 806 individuals, 10 cases of cancer were observed during the first follow‐up period. Although the cohort is young and the numbers small, a slightly significant (P = 0.04) trend was observed for individuals with cancer and a score of chromosomal aberrations. No trend was observed for sister chromatid exchanges.

The application of cytogenetic surveillance is still not routine methodology, but it is useful and informative in carefully controlled study designs. Special efforts should be directed toward combining different disciplines, i.e., cytogenetics, adduct monitoring, and end‐effect epidemiology, in order to reach quantitativeness in risk assessment.
Cytotoxicity, anchorage‐independent growth, and DNA adduct formation in human neonatal fibroblasts by 1, 2, 3, 4‐tetrahydro‐7, 12‐dimethylbenz(a)‐anthracene (TH‐DMBA), its six aryl fluoro regioisomers, and an exo methylene tautomer
Authors: Kumari H. Lalitha , Milo George E. , Witiak Donald T.
Published: February 2006

1, 2, 3, 4‐Tetrahydro‐7, 12‐dimethylbenz(a)anthracene (TH‐DMBA), its six possible fluoro‐substituted regioisomers, and the C‐7 exo methylene tautomer of the 11F derivative have been investigated for their cytotoxicity and for their ability to induce anchorage‐independent growth and to form adducts in human neonatal foreskin fibroblasts. All compounds tested exhibited a low level of cytotoxicity, determined as percent cloning efficiency, up to a final concentration of 30 μ/ml. Except for 5F‐TH‐DMBA and the C‐7 exo methylene tautomer, all compounds induced anchorage‐independent growth of neonatal foreskin fibroblasts in soft agar at all concentrations tested (1, 3, 10, and 30 μ/ml). The C‐7 exo methylene tautomer induced anchorage‐independent growth only at a concentration of 10 μ/ml. Among the compounds tested the 6F derivative was the most effective compound at 1 μ/ml. The D‐ring fluoro isomers induced anchorage‐independent growth at a frequency comparable to TH‐DMBA itself, with the 11F derivative being the least effective of the four D‐ring regioisomers. All compounds except 5F‐TH‐DMBA formed detectable adducts with cellular DNA as determined by 32P postlabeling procedures, when the cells were treated at 1 μ/ml. Two adducts were detected in cells treated with TH‐DMBA and four adducts were detected in DNA obtained from cells treated with 6F‐TH‐DMBA. The level of bonding for the D‐ring fluoro isomers was quantitatively less and sometimes qualitatively different than that for TH‐DMBA. For the D‐ring compounds, the ability to induce anchorage‐independent growth frequency correlated with the total quantity of adduct formed. The C‐7 exo methylene tautomer formed a single adduct and the level of bonding was less than one adduct per 109 nucleotides. Analysis of these results led to the proposal that the planar anthracene ring structure (rings B, C, and D) of TH‐DMBA and possibly oxidative metabolism at benzylic carbon 4 of the A‐ring are important to DNA bonding and initation of induction of anchorage‐independent growth.
Simultaneous induction of mutagenic and cancerogenic effects in T × HT mice with transplacental ethylnitrosourea treatment
Authors: Schmahl Wolfgang , Neuhäuser‐Klaus Angelika , Leierseder‐Bauer Maria , Luz Arne
Published: February 2006

The relationship between mutagenesis and carcinogenesis was investigated in T × HT crossbred mice using diaplacental application of ethylnitrosourea (ENU) at different stages of embryonal development. Mutagenesis was detected by induction of coat color spots, and the carcinogenic response was investigated in a long‐term follow‐up study of the F1‐generation. The animals were particularly sensitive to induction of tumors at the central nervous system (CNS)‐skull/vertebra interface (30% and 20% in ENU‐treated male and female offspring, respectively, compared with < 1% in controls). There was a correlation between the appearance of these tumors and the presence of color spots. This correlation was low but statistically significant in female offspring. Three other types of tumors showed a correlation with the presence of coat color spots. Liver tumors were significantly increased in color spot‐positive females but unchanged in males. Lung tumors were reduced in color spot‐positive males and appeared earlier in color spot‐positive females. There was a lower incidence of lymphoma/leukemia in all spot‐positive mice. The reduction in tumor incidence beyond the spontaneous rate in spot‐positive animals might be caused by a high cytolethal response to ENU in the relevant organs and tissues.
Mutagenic and teratogenic effects of cyclophosphamide on the chick embryo: Chromosomal aberrations and cell proliferation in affected and unaffected tissues
Authors: Novotná Božena , Jelínek Richard
Published: February 2006

Chromosomal aberrations and cell proliferation were analyzed in the chick embryo blood, limb bud, and facial tissues 12 and 24 hours after cyclophosphamide (CP) administration on day 3. The cytogenic findings were compared with teratogenic effects evaluated on incubation day 8. Low dose (0.3 μg) resulting in heart defects exclusively, increased the frequency of aberrant cells with simultaneous depression of cell proliferation in blood only. High dose of CP (6 μg), besides the heart defects, also induced facial clefts and limb malformations, and strong clastogenic effects associated with mitotic inhibition were observed in all tissues investigated. The results support the idea that the consequences of mutagenic action of cyclophosphamide—cell cycle delay and excessive death of cells with unstable aberrations—result in abnormal morphogenesis.
Dangerous properties of petroleum‐refining products: Carcinogenicity of motor fuels (Gasoline)
Authors: Mehlman Myron A.
Published: February 2006

Gasoline contains large numbers of dangerous and cancer‐causing chemicals such as benzene, butadiene, toluene, ethylbenzene, xylene, trimethyl pentane, methyltert‐butylether (MTBE) and many others. For the U.S. alone approximately 140 billion gallons of gasoline were consumed in 1989. An increase in only ten cents per gallon in price of gasoline generates 14 billion dollars in extra profit per year for oil industry cartel.

Laboratory animals exposed to gasoline developed cancers in different tissues and organs. A number of epidemiological studies in humans provide evidence of increased cancer risk of leukemia, kidney, liver, brain, lymphosarcoma, lymphatic tissue pancreas and other tissues and organs.
Sensitivity of cultured lymphocytes from patients with nevoid basal cell carcinoma syndrome to ultraviolet light and phytohemagglutinin stimulation
Authors: Ferraro P. , Celotti L. , Furlan D. , Pattarello I. , Peserico A.
Published: February 2006

DNA repair and replication after in vitro UV irradiation were determined in cultured peripheral blood lymphocytes from 6 patients with nevoid basal cell carcinoma syndrome (NBCCS) and from a group of control donors. DNA repair synthesis (UDS) was measured in unstimulated lymphocytes by incubation with 3H‐TdR in the presence of hydroxyurea for 3 and 6 h after UV irradiation (6‐‐48 J/m2). DNA replication was measured in PHA‐stimulated lymphocytes, UV‐irradiated or mock‐irradiated, by incubation with 3H‐TdR for 24 h. The effect of the mitogen was followed during 5 days after stimulation by determining the incorporation of 3H‐TdR, the increase of cell number, and the mitotic index. NBCCS and control lymphocytes showed equal sensitivity to UV light in terms of UDS and reduced response to PHA. On the contrary, the mitotic index and the number of cells in stimulated cultures were significantly lower in the affected subjects. These data suggest an altered progression along the cell cycle, which could be characteristic of stimulated NBCCS lymphocytes.
Evaluation of the genotoxicity of gentian violet in bacterial and mammalian cell systems
Authors: Aidoo Anane , Gao Ning , Neft Robin E. , Schol Henry M. , Hass Bruce S. , Minor Toni Y. , Heflich Robert H.
Published: February 2006

Previous studies indicate that gentian violet (GV), a triphenylmethane dye used in agriculture and human medicine, is a clastogen in vitro and a carcinogen in chronically exposed mice and rats. Data on its genotoxic activity, however, have been incomplete and partly contradictory. Mutagenesis and DNA damage experiments were conducted to re‐evaluate the genotoxic potential of GV in both bacterial and mammalian cell systems. GV was mutagenic in Salmonella typhimurium tester strains TA97 and TA104, but there was little mutagenic activity detected in strains TA98 and TA100. A rat liver homogenate fraction (S9) tended to increase mutagenicity. The major microsomal metabolites of GV, pentamethylpararosaniline and N,N,N′,N′‐tetramethylpararosaniline were less mutagenic in TA97 and TA104, while N,N,N′,N"‐tetramethylpararosaniline was a weak mutagen in Salmonella. GV was not mutagenic in Chinese hamster ovary (CHO) cell strain CHO‐K1‐BH4, and was a questionable mutagen in CHO‐AS52 cells. While GV produced DNA damage as measured by sedimentation of nucleoids derived from B6C3F1 mouse lymphocytes treated in vitro, no damage was found in lymphocytes isolated from mice dosed with GV. GV was also a weak producer of gene amplification in an SV40‐transformed Chinese hamster cell line. The results indicate that GV is a point mutagen in bacteria; however, since similar exposure conditions produced weak mutagenic activity in mammalian cells, GV may be carcinogenic by virtue of its clastogenic activity.
Interaction of drugs with extranuclear genetic elements and its consequences
Authors: Ebringer Libor
Published: February 2006

Bacterial ancestry of mitochondria and plastids is now generally accepted. Both organelles contain their own DNA and transcription‐translation apparatus of a prokaryotic type. Due to this fact these systems carry bacteria‐like properties. Thus organellar DNA and ribosomes are essentially different from nuclear DNA and cytoplasmic ribosomes in physical as well as in functional respects. Due to the bacterial character of both types of organelles they are susceptible to various antibacterial chemicals. Inhibitors of bacterial protein synthesis inhibit mitochondrial (plastidial) biogenesis. Therefore the cellular content of mitochondria (plastids)‐made proteins decreases during cytoplasmic turnover or cell division in the presence of these drugs. Such drug activity consequently leads to a reduced capacity for oxidative phosphorylation or photosynthesis. Organellar genomes are less stable and more sensitive to mutagenesis as compared to nuclear genome. It means also that genotoxic agents induce various disorders of mitochondrial (plastidial) functions. Impairments in the respiratory chain are associated with structural as well as functional abnormalities of mitochondria. These are clinically expressed mostly in tissues with a high demand for ATP: brain, heart, skeletal muscle, and retina. On the other hand, some antibacterial inhibitors of mitochondrial biogenesis (e.g., tetracyclines) inhibit selectively tumor cell proliferation. Therefore they may be considered for use in anticancer therapy.

The article summarizes the response of mitochondria and plastids in various organisms to drugs and environmental xenobiotics. Various model organisms suitable for detection of xenobiotic effect on mitochondria (plastids) are presented as well as the possible consequences of such interaction.
Detection of 6‐thioguanine‐resistant spleen lymphocytes in different mouse strains by autoradiography
Authors: Hüttner Edith , Braun Rolf , Wielgosz Szesny M. , Schöneich Jörg
Published: February 2006

The variant frequencies for 6‐thioguanine‐resistant spleen cells in different mouse strains have been estimated by autoradiography for animals without chemical treatment and in cases of in vivo mutagen dosage with ethylnitrosourea and cyclophosphamide, respectively. In untreated mice, the following variant frequencies have been found: C57B1/6J, 2.84 × 10−5;NMRI, 3.04 × 10−5;DBA/2J, 5.91 × 10−5. The selective concentration of 6‐thioguanine was 100 μM for strains NMRI and DBA, while in the case of C57B1 with this concentration, no variant cells could be counted and a selective concentration of 50 μM was chosen. Treatment with 70, 140, and 210 mg/kg ethylnitrosourea resulted in increased variant frequencies in cells isolated 8 or 15 days later. On the other hand, doses of 20, 60, and 120 mg/kg cyclophosphamide did not result in a clear dose‐response relationship of variant frequency in cells isolated 1, 8, 15, 22, and 29 days after treatment. These data are discussed with respect to findings in human populations exposed occupationally to cyclophosphamide.
Rules of molecular geometry for predicting carcinogenic activity of unsubstituted polynuclear aromatic hydrocarbons
Authors: Flesher James W. , Myers Steven R.
Published: February 2006

The rules of molecular geometry for predicting carcinogenic activity of polynuclear aromatic hydrocarbons (PAH) have been applied to a series of 50 unsubstituted PAH, and predicted carcinogenic activity is in good agreement with the results of testing for complete carcinogenic activity in mice and/or rats. The rules were developed from a knowledge of the center or centers of highest chemical or biochemical reactivity and are consistent with a unified hypothesis which states that the first step in the metabolic activation of unsubstituted PAH is the biochemical introduction of a methyl group. This bioalkylation reaction 1) takes place between certain PAH and S‐adenosyl‐L‐methionine and is catalyzed by cytosolic methyltransferase, 2) offers a means of probing for centers of reactivity in PAH, 3) provides a biochemical link between unsubstituted preprocarcinogens of aromatic type ArX and alkyl‐substituted procarcinogens of aromatic type ArCH2X (where X = H), and 4) makes it possible to include compounds of both aromatic types, in a consistent theory of aromatic hydrocarbon activation which incorporates alkyl substitution. The present study reveals that there are structural determinants of carcinogenicity.
Analysis of rat lymphocyte activation of benzo[a]pyrene, 2‐acetylaminofluorene, and several of their metabolites to mutagenic and DNA‐damaging species in vitro
Authors: Gao Ning , Aidoo Anane , Heflich Robert H.
Published: February 2006

Rat lymphocytes are a potentially useful and convenient cell system for monitoring the genotoxic effects of chemicals in vivo, but little is known about the ability of these cells to metabolize promutagens to genotoxic species. In this study, Fischer 344 rat lymphocytes were treated in vitro with benzo[a]pyrene (BaP), 2‐acetylaminofluorene (2‐AAF), and several of their metabolites, and DNA damage was measured using nucleoid sedimentation analysis. Of the BaP derivatives, BaP 4,5‐oxide and BaP 7,8‐diol‐9,10‐epoxide decreased lymphocyte nucleoid sedimentation, whereas BaP and BaP 7,8‐dihydrodiol had little effect. Among the 2‐AAF derivatives, N‐acetoxy‐2‐AAF, N‐hydroxy‐2‐AAF, and N‐hydroxy‐2‐aminofluorene damaged rat lymphocyte nucleoids, whereas 2‐AAF, 2‐aminofluorene, and fluorene produced little detectable damage. The decrease in nucleoid sedimentation produced by N‐hydroxy‐2‐AAF was not inhibited by paraoxon, salicylamide, or 2‐aminofluorene, whereas paraoxon inhibited damage produced by N‐acetoxy‐2‐AAF. In co‐culture assays, rat lymphocytes increased the mutagenicity of N‐hydroxy‐2‐AAF in Salmonella typhimurium strain TA98, but mediated little or no mutagenic response with BaP, BaP 7,8‐dihydrodiol, and 2‐AAF. Also, human lymphocytes, but not rat lymphocytes, mediated a positive mutagenic response with BaP 7,8‐dihydrodiol in Chinese hamster ovary UV5 cells. Although rat lymphocytes may metabolize certain proximal genotoxic chemicals to DNA‐damaging species (e.g., N‐hydroxy‐2‐AAF), these data suggest that in vivo lymphocyte DNA damage is more likely to result from lymphocytes encountering reactive chemical derivatives produced by other cells. It is also clear that differences exist between the ability of human and rat lymphocytes to activate promutagens, and this may impact on the use of the rat model to predict the genotoxicity of chemicals in humans.
In vitro reduction of peroxidation in UVC‐irradiated cell cultures by concurrent exposure with Bowman‐Birk protease inhibitor
Authors: Baturay Nesrine Z. , Roque Henry
Published: February 2006

Bowman‐Birk protease inhibitor (BBI), extracted from soybean flour, has been shown to protect cells from damage after UVC irradiation. BBI also reduces the number of transformed foci following exposure to the same doses of UVC irradiation. Proteins have been reported to act as antioxidants by their sheer intracellular bulk. In this study we assessed BBI's ability to attenuate UVC‐irradiation‐induced peroxidation.

Balb c/3T3 cells, clone A31, subcloned by this laboratory were used. Treatment groups consisted of cells suspended in Hanks' Balanced Salt Solution with and without 100 μg/ml of a crude soybean extract containing the Bowman‐Birk protease inhibitor. Cells were exposed to irradiation in suspension by using GE (G8T5) germicidal lamps which deliver predominantly 254 nm light at 0.385 J/m2/s as measured at the cell surface. Aliquots of UVC‐exposed cells were homogenized in HBSS in the presence and absence of 100 μg/ml BBI with a motor‐driven Teflon pestle and assayed for peroxidation.

The results showed significant (P < 0.0001) protection afforded by BBI against peroxidation at 1.16 (65%), 1.93 (60%, and 2.70 (48%) J/m2 of UVC irradiation. Transformation was reduced to 0.33 foci per dish at all levels of radiation exposure.
An appreciation of the maximum tolerated dose: An inadequately precise decision point in designing a carcinogenesis bioassay?
Authors: Clayson David B. , Iverson Frank , Mueller Rudolf
Published: February 2006

Cancers arise in specific tissues. One difficulty with the present definitions of the Maximum Tolerated Dose (MTD), as they pertain to the rodent cancer bioassay, is that they base MTD on relatively crude parameters associated with the well‐being of the entire animal rather than with the lack of specific tissue toxicity. Additional factors that could be included in the MTD definition, or could be separately determined, are addressed. Many of these factors refer to toxic behavior in one or a few tissues and, if used in setting the MTD, may mask more relevant events occurring at higher dose levels in other tissues. Reducing the MTD to a level that fails to take into account pesticide or drug‐related toxicity may lead to the loss of relevant information in the bioassay. It is concluded, therefore, that there are two possible approaches to a more appropriate use of the MTD. The highest dose of the test agent (MTD) may be chosen (i) to lie below the thresholds of carcinogenicity‐related non‐genotoxic toxicity or (ii) the present high level MTD may continue to be used and tumors that arise may be classified as being irrelvant to humans at some or all exposure levels. The latter approach is to be preferred. It has the potential to avoid missing high level effects of the test agent that may be relevant to the human population. © 1992 Wiley‐Liss, Inc.
Diphenylhydantoin is not genotoxic in a battery of short‐term cytogenetic assays
Authors: Kindig D. , Garriott M. L. , Parton J. W. , Brunny J. D. , Beyers J. E.
Published: February 2006

5,5‐Diphenylhydantoin (DPH) is an antiepileptic drug associated with an increase in malformations in infants born to women taking DPH during pregnancy. Positive and negative results have been reported by various investigators for in vivo and in vitro chromosome aberration (CAB) assays, in vivo and in vitro sister chromatid exchange (SCE) assays, and in vivo micronucleus tests (MNT). In this laboratory, DPH was tested in an in vitro CAB assay using Chinese hamster ovary cells with and wihout an S‐9 activation system, an in vivo SCE assay in female CD‐1 mice, an in vivo MNT, using both male and female CD‐1 mice, and a transplacental micronucleus test. The results from this comprehensive battery of cytogenetic tests were uniformly negative and support a conclusion that the known teratogen, DPH, is not clastogenic. © 1992 Wiley‐Liss, Inc.