bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of exposing rat embryos in utero to physical or chemical teratogens are expressed later as enhanced induction of heat‐shock proteins when embryonic hearts are cultured in vitro
Authors: Higo Hiromi , Higo Ken‐Ichi , Lee Juing Yi , Hori Hiroshi , Satow Yukio
Published: February 2006

In order to get more insight into the effects of teratogens on developing embryos, we investigated the protein synthesis patterns of the target organs isolated from teratogen‐treated embryos. Rat embryos were either irradiated in utero with either 252Cf fission neutrons or 60Co gamma rays on day 8 of gestation or treated in utero with a bis(dichloroacetyl)diamine (a chemical teratogen) on days 9 and 10. Hearts were removed from the embryos on day 12 and were incubated in vitro at 37°C in the presence of [35S]methionine for up to 8 hr. The newly synthesized labeled proteins were then analyzed qualitatively by two‐dimensional polyacryl‐amide gel electrophoresis. Enhanced and prolonged induction of a family of heat‐shock (stress) proteins with a molecular weight of about 70,000 (SP70s) was observed as compared with those of controls. Among the teratogen‐treated hearts, those with gross malformations already detectable at this early stage showed especially higher inductions of SP70s than did the others. The abnormal expression of SP70s observed in the present study appears to be a reflection of persisting cellular (tissue) damage inflicted by the teratogens, and the extent of the induction may be indicative of the degree and/or type of the damage. Such persisting defects in surviving cells, manifested by abnormal induction of SP70s in the present study, might be related to malformation of embryonic hearts.
The antioxidant butylated hydroxytoluene can retard cerebellar degeneration induced transplacentally by a single low dosage of N‐methyl‐n‐nitrosourea
Authors: Smith Sylvia B. , Cooke Carol Beck , Lemone Yielding K.
Published: February 2006

Late‐onset cerebellar degeneration can be induced transplacentally in mice by a single low‐dose (1 mg/kg) injection of the direct‐acting DNA alkylating agent N‐methyl‐N‐nitrosourea (MNU) on day 16 of pregnancy. The offspring develop a mild ataxia that manifests by 12–16 weeks of age postnatally when the animals are challenged with a motor coordination task. Morphological evidence of degeneration includes pyknosis of Purkinje cells and abnormal foliation patterns. Additionally, these animals demonstrate a progressive retinopathy characterized by thinning of the nuclear and plexiform layers of the retina. Efforts to retard the cerebellar degeneration were undertaken in the present study. MNU‐exposed and control animals were fed a standard mouse chow diet supplemented with 0.75% butylated hydroxytoluene (BHT), an antioxidant. This supplementation commenced 24 h following exposure to the teratogen and continued throughout the life of the offspring. A second group of MNU‐exposed and control mice were fed a non‐BHT‐supplemented, standard Purina mouse chow diet. Quantitative histological evaluation of cerebellar coronal sections indicated that by 4 weeks of age BHT‐fed, MNU‐exposed mice had significantly fewer pyknotic Purkinje cells than non‐BHT‐fed, MNU‐exposed animals. This was true for the vermal, paravermal, and lateral areas of the cerebellum. The findings suggest the usefulness of teratogenic models of degenerative diseases for the testing of potential intervention strategies.
Genotoxic effects of estrogens in epithelial cells from the neonatal mouse uterine cervix: Modifications by metabolic modifiers
Authors: Hillbertz‐Nilsson Kerstin , Forsberg John‐Gunnar
Published: February 2006

Epithelium from the neonatal mouse uterine cervix and uppermost part of vagina was cultured in vitro. The culture medium was supplemented with 17β‐estradiol (E2; 10–6–10–5 M) or diethylstilbestrol (DES; 10–8–10–5 M) alone or in combination with different metabolic modifiers (α‐naphthoflavone, β‐naphthoflavone, phenobarbital, metyrapone, indomethacin) with postulated activating or inhibitory effects on DES metabolizing enzymes (cytochrome P‐448‐ and P‐450‐dependent microsomal monooxidases, prostaglandin cyclooxygenase). E2 at 10–5 M and DES at 10–6 and 10–5 M concentrations increased the incidence of cells with a high number of sister chromatid exchanges (high‐frequency chromatid exchange cells, HFCEC). Indomethacin partially depressed DES‐induced HFCEC, whereas the incidence was increased by α‐naphthoflavone, which may be a result of stimulation of the fetal type of P‐448‐dependent enzyme activity or of DES increasing the metabolic activation of α‐naphthoflavone. Phenobarbital and β‐naphthoflavone did not affect the incidence of DES‐induced HFCEC. Metyrapone alone induced the highest incidence of HFCEC observed in this study, and this effect was inactivated by phenobarbital and/or DES. The mechanisms behind these results are discussed. This study shows that E2 and DES have a genotoxic effect (sister chromatid exchanges) in vitro in epithelial cells from the same target organ as in which epithelial aberrations occur after in vivo estrogen treatment in the neonatal period. The difference in incidence of tetraploid cells between stroma and epithelium is stressed (< 5% vs. 16–47% depending on experimental group).
Comparison of DNA alkylation, fragmentation, and repair in maternal and fetal tissues of pregnant rats treated with a single dose of ethyl methanesulfonate, ethyl‐N‐nitrosourea, N‐nitrosodiethylamine, and methyl‐N‐nitrosourea
Authors: Robbiano Luigi , Parodi Angela , Venturelli Simonetta , Brambilla Giovanni
Published: February 2006

The occurrence and persistence of DNA damage, as detected by the alkaline elution technique, have been studied in some tissues of both fetal and adult Sprague‐Dawley rats (18th day of gestation) after administration of a single equimolar dose (0.5 mmol/kg) of ethyl methanesulfonate (EMS), N‐ethyl‐N‐nitrosourea (ENU), N‐nitrosodiethylamine (NDEA), and N‐methyl‐N‐nitrosourea (MNU). EMS, ENU, and MNU, injected intravenously, produced a statistically significant increase of DNA elution rate, which is considered indicative of DNA fragmentation, in both maternal and fetal liver, kidney, and brain. NDEA, introduced by gastric gavage, induced DNA breaks in both liver and kidney of dams, but only in the liver of fetuses. The frequency of DNA lesions was found to vary with the four alkylating agents and in the three organs tested, to exhibit a different time course, and usually to be higher in maternal than in fetal tissues. Results provided by the concomitant determination of DNA binding levels demonstrated a satisfactory correlation with the amounts of DNA fragmentation. In contrast, the values of both these parameters did not show any positive correlation with the different susceptibility of the three organs to tumor induction. In conclusion, these findings suggest that when a compound is not available in radiolabeled form, measurement of DNA fragmentation may represent a useful alternative to the determination of DNA binding level in order to obtain information on the distribution of its reactive species in maternal and fetal tissues.
Teratogenicity and toxicity of ethylene glycol monomethyl ether (2‐methoxyethanol) in Drosophila melanogaster: Involvement of alcohol dehydrogenase activity
Authors: Eisses Karel T. H.
Published: February 2006

Ethylene glycol monomethyl ether (EGME) or 2‐methoxyethanol (2‐ME) is a toxic compound that has teratogenic effects in developing organisms similar to its oxidation product methoxyacetic acid (MAA). Alcohol dehydrogenase (ADH) is supposed to be involved with the oxidation of EGME into MAA. It has been shown here that Drosophila ADH dehydrogenates EGME in an in vitro test system. Three strains of Drosophila melanogaster, genetically different from each other with regard to ADH activities (qualitatively and quantitatively), were used to test the in vivo involvement of ADH with the oxidation of EGME into MAA. The toxicity of EGME in egg‐to‐adult and third instar larval‐to‐adult development is concentration and strain dependent. Higher detoxification occurs in the strain with increased ADH activity. EGME is much more toxic than its oxidation product MAA at the level of adult eclosion. Teratogenic effects were observed in the ADH‐negative strain in spite of lacking ADH activity. EGME is apparently a teratogenic compound by itself.
Chromosomal aberrations as a contributing factor for tumor promotion in the mouse skin
Authors: Kaina Bernd
Published: February 2006

Tumor promotion in mouse skin can be dissected in two stages: stage I (conversion) and stage II. Whereas for stage II clonal expansion of transformed cells is believed to play a major role, the mechanism(s) underlying conversion is still a matter of debate. Because conversion can be achieved upon treatment with phorbol ester tumor promoters prior to initiation, it is unlikely to represent simply proliferative stimulation of initiated cells (due to epigenetic changes induced). Since tumor promoters exert clastogenic activities and, on the other hand, the clastogen methyl methanesulfonate proved to be convertogenic, the possibility arises that chromosomal changes are involved in conversion. Based on this hypothesis, several findings concerning the action of tumor promoters and the process of tumor promotion in the mouse skin system are discussed and interpreted: the frequency, reversibility, and transient nature of conversion, dependence of tumor promotion on DNA synthesis, induction of DNA breaks by tumor promoters, and the protecting effect of scavengers of free radicals. A model is presented suggesting tumor formation in mouse skin (and other systems) to proceed in discrete, genetically determined steps. Initiation is considered to be due to the induction of point mutations in a dominant‐acting oncogene that becomes thereupon activated, whereas the decisive event in the conversion stage of tumor promotion is the induction of numerical and/or structural chromosomal changes with the consequence of loss or inactivation of gene(s) involved in suppression of the tumor phenotype.
Common Biochemical Pathway of Dysmorphogenesis in Murine Embryos: Use of the Glucocorticoid Pathway by Phenytoin
Authors: Kay Elizabeth D. , Goldman Allen S. , Daniel Jon C.
Published: February 2006

Phenytoin (5,5‐diphenylhydantoin), a common anticonvulsant drug, is known to produce anomalies in the craniofacial region of animals and humans. Furthermore, recent evidence suggests that phenytoin disrupts craniofacial and neural tube morphogenesis by inhibiting the arachidonic acid cascade, a pathogenesis already implicated for glucocorticoids and hyperglycemia in the palate. This study tested the hypothesis that phenytoin interferes with the arachidonic acid cascade via the same biochemical pathway demonstrated for glucocorticoids. The proposed pathway was tested at two levels. First, indomethacin, an inhibitor of the enzyme cyclooxygenase, was used in culture to block the correction of phenytoin‐induced defects by arachidonic acid. Second, cortexolone, an anti‐glucocorticoid that binds at the glucocorticoid receptor binding site, was tested for its ability to prevent phenytoin‐induced teratogenicity. Eighty‐four percent of the embryos cultured in phenytoin and 93% of those cultured in phenytoin plus arachidonic acid and indomethacin had neural tube and/or craniofacial deformities. In contrast, only 14% of the embryos cultured in phenytoin plus cortexolone were affected. Indomethacin itself produced anomalies in 83% of the exposed embryos. These data are consistent with the hypothesis that the teratogenic action of phenytoin in murine embryo cultures occurs via the glucocorticoid anti‐inflammatory pathway. Thus, the glucocorticoid receptor appears to be responsible for mediating phenytoin‐induced teratogenicity.
Species differences in carcinogenicity: The role of metabolism in human risk evaluation
Authors: Green Trevor
Published: February 2006

The assessment of human risk from exposure to chemicals is frequently confounded by differences in response to those chemicals in laboratory animals. In many cases, the basis of the species differences is differences in metabolism and pharmacokinetics. Since metabolic and pharmacokinetic data are readily accessible in both laboratory animals and humans, risk assessments can be significantly improved if these data are incorporated into the process. Several chlorinated solvents, perchloroethylene, trichloroethylene, and methylene chloride, have been investigated, and the species differences in carcinogenicity have been shown to be the result of pharmacokinetic differences between the species. The rates of metabolism of these chemicals have been measured in rats and mice in vivo, and the species differences observed have been reproduced in vitro using tissue fractions and hepatocytes. Identical experiments have been carried out using human tissues. The results of these studies together with species‐specific physiological parameters have been used in a mathematical model to predict human cancer risk over a wide range of exposures. This approach provides both an explanation for the species differences in response to these chemicals and also a more rational approach to human risk assessment.
Genetic activity profiles in the testing and evaluation of chemical mixtures
Authors: Waters Michael D. , Claxton Larry D. , Frank Stack H. , Brady Ann L. , Graedel Thomas E.
Published: February 2006

Some knowledge of the potential genetic activity of a complex environmental mixture may be gained from an assessment of the genetic activity of its component chemicals. The expanded Genetic Activity Profile (GAP) data base provides a computer‐generated graphic representation of genetic bioassay data as a function of dose of the substance tested. In addition, the Atmospheric Chemical Compound (ACC) data base contains information on chemical structures, properties, detection methods, and sources of chemicals found in ambient air. Using the combined data bases, the quantity of an individual chemical present within a mixture or fraction of a mixture may be related to the quantity (lowest effective dose, LED) of the chemical, by itself, required to demonstrate a positive response in one or more genetic bioassays.
Potentiation of ethyl methanesulfonate‐induced germ cell mutagenesis and depression of glutathione in male reproductive tissues by 1,2‐dibromoethane
Authors: Teaf Christopher M. , Bishop Jack B. , Harbison Raymond D.
Published: February 2006

EDB significantly depressed GSH in caput and cauda epididymis, but not in testis, 2 hours following injection. This depression was dose‐related. EDB enhanced EMS‐induced dominant lethal mutations at mating weeks 2 and 3 (of 6). At mating week 2 the fetal death rate was increased two‐fold, while at week 3, the fetal death rate had increased to nearly three‐fold greater than the EMS‐only controls. Enhancement of fetal death rate was confined to postimplantation loss. As with EMS alone, the EDB potentiation of EMS‐induced mutations was limited to postmeiotic stages of spermatogenesis. EDB also enhanced alkylation of rat spermatozoa by labeled EMS. Depression of GSH in reproductive tissues is correlated with a potentiation of dominant lethal mutations, as well as an increase in the binding of EMS to sperm heads.
Estrogen effects on chromosome number and sister chromatid exchanges in uterine epithelial cells and kidney cells from neonatal mice
Authors: Forsberg John‐Gunnar
Published: February 2006

Female mice of the NMRI strain were treated with a high dose (5 μg) or a low dose (10−2 μg) of the synthetic estrogen diethylstilbestrol (DES) or estradiol‐17β (E2) 48 and 24 hr, 24 hr, or 12 hr before death on day 3 after birth. Cultures of epithelium from the uterine cervix and uterine horns as well as of kidney cells were set up, and, after a culture period of 3 days, cells were studied for chromosome number and sister chromatid exchanges. Cervical cells from control females had a distinct peak of tetraploid cells, which was depressed after DES treatment but less so after E2 treatment. The tetraploid peak was lower in uterine horn epithelium and was less influenced by DES. No indications were obtained for an estrogen‐induced aneuploid cell population in the uterine horn epithelium or cervical epithelium. The incidence of cells with a high number of sister chromatid exchanges (HFCECs) was only about one‐fifth in uterine horn epithelium from control females compared with cervical epithelium from the same females. In the cervical epithelium, DES at both dose levels and E2 at the high dose induced an increased and similar incidence of HFCECs (from about 12.5% in controls to about 35%), which was not related to estrogen exposure time. In uterine horn epithelium, the incidence of HFCECs increased from 2.5% in controls to 8.5% after DES treatment. It is postulated that there is a more pronounced genomic plasticity in the cervical epithelium compared with the uterine horn epithelium. The estrogens had no effect on chromosome number or HFCECs in kidney cells.
Effects of QA 208‐199 and its metabolite 209‐668 on embryonic development in vitro after microinjection into the exocoelomic space or into the amniotic cavity of cultured rat conceptuses
Authors: Bechter R. , Terlouw G. D. C. , Lee Q. P. , Juchau M. R.
Published: February 2006

The objective of this study was to determine the direct embryotoxic effects in vitro of N‐hydroxy‐N‐methyl‐7‐propoxy‐2‐naphthalene‐ethanamine (QA 208‐199, QAB) and of one of its metabolites, 7‐propoxy‐naphthalene‐2‐ylacetic acid (209–668, QAA), after circumventing the bioconverting conceptal membranes. The compounds were, therefore, microinjected either into the exocoelomic space or into the amniotic cavity of rat conceptuses of 10 d at prenatal age at doses of up to 84.9 ng (QAA) and 180 ng (QAB) per conceptus respectively. The conceptuses were subsequently cultured for 28 h after which their development was assessed. QAB produced marginal effects on embryonic differentiation only after microinjection of the compound into the amniotic cavity. Dysmorphogenic effects, however, occurred in a dose‐dependent fashion after either exocoelomic or intraamniotic microinjections of the compound. The frequencies and types of anomalies were similar after either exposure route and consisted predominantly of anomalies associated with axial rotation. QAA also impaired embryonic differentiation at only the high dose level of 84.9 ng per embryo and after intraamniotic injections only. Dysmorphogenic effects were observed in all experimental groups, although the differences were not statistically significant when compared with the concomitant controls. An increased proportion of anomalies observed were in the cephalic region as compared to the defects produced by QAB. These data suggest that QAA most probably is not the QAB metabolite responsible for the embryotoxic action of QAB in vitro. Furthermore, the results tend to confirm the suggested involvement of the visceral yolk sac membrane in mediating QAB embryotoxicity.
Induction of micronuclei in rat bone marrow by four model compounds
Authors: Shi Xiao‐Chun , Krishna Gopala , Ong Tong‐Man
Published: February 2006

Studies have been performed to determine the dose and sampling time responses of micronuclei after Sprague‐Dawley rats were treated wth triethylenemelamine, mitomycin C. dimethylbenzanthracene, and vincristine by a single intraperitoneal injection. Three doses were tested for each compound. Animals were sacrificed 24, 48, and 72 h after chemical treatment. Slides prepared from the bone marrow were stained with May‐Gruenwald and Giemsa stains. The number of micronucleated polychromatic erythrocytes among 2,000 polychromatic erythrocytes (PCEs) and the ratio of PCEs to normochromatic erythrocytes were determined for each animal. The results show that all four compounds cause micronucleus formation in rat bone marrow. The peak response sampling time, either 24 or 48 h posttreatment, is dependent on the chemical as well as the dose. In all cases, however, an increase in the micronucleated PCEs was detected 24 h after chemical treatment. These results seem to indicate that two sampling times, 24 and 48 h, may be adequate for the micronucleus assay using rat bone marrow cells. © 1992 Wiley‐Liss, Inc.
Evaluation in a battery of in vivo assays of four in vitro genotoxins proved to be noncarcinogens in rodents
Authors: Allavena Alessandra , Martelli Antonietta , Robbiano Luigi , Brambilla Giovanni
Published: February 2006

2‐Chlorethanol, 8‐hydroxyquinoline, 2,6‐toluenediamine, and eugenol, previously found to behave as genotoxins in in vitro systems and as noncarcinogens in rodents, were evaluated for their ability to induce genotoxic effects in vivo. Rats were given by gavage a single or two successive doses equal to one‐half the corresponding LD50, killed at different times after treatment, and examined for the following end points: the frequency of both micronucleated polychromatic erythrocytes in the bone marrow and micronucleated hepatocytes (after partial hepatectomy); the in vivo—in vitro induction of DNA fragmentation, as measured by the alkaline elution technique, and of unscheduled DNA synthesis, as measured by autoradiography, in hepatocyte primary cultures. The two latter end points were also evaluated after in vitro exposure of hepatocytes to log‐spaced subtoxic concentrations. 2‐Chloroethanol, 8‐hydroxyquinoline, and eugenol never produced effects indicative of genotoxic activity. The same happened with 2,6‐toluenediamine, with the exception of a significant increase over controls in the amounts of DNA damage and repair displayed by hepatocyte cultures obtained from rats given two 1/2 LD50 separated by a 24 h interval. Our results, which, apart the above mentioned exception, are in concordance with the rodent carcinogenicity results, contribute to underline the role of in vivo short‐term tests for the detection of potential genotoxic carcinogens. © 1992 Wiley‐Liss, Inc.
Further in vitro and in vivo mutagenicity assays with thiram and ziram fungicides: Bacterial reversion assays and mouse micronucleus test
Authors: Crebelli R. , Zijno A. , Conti L. , Crochi B. , Leopardi P. , Marcon F. , Renzi L. , Carere A.
Published: February 2006

The fungicides thiram and ziram have been assayed in a battery of nine bacterial strains of different genetic specificity. The results obtained suggest the induction of excisable DNA lesion(s), and indicate similar mutability of strains with AT or GC base pairs at target sites. This mutagenic profile is clearly distinct from that of oxidative mutagens, and it does not support the proposed role of oxidative stress in the mechanism of dithiocarbamates mutagenicity in bacteria.

Furthermore, the bone marrow micronucleus test has been carried out in B6C3F1 mice with intraperitoneal administration of high grade thiram (12.5–50 mg/kg) and ziram samples (2.5–10 mg/kg in males, and 5–20 mg/kg in females). Thiram produced a significant increase of micronucleated PCEs in male mice sampled 48 h after treatment with 25, 37.5, and 50 mg/kg. No significant increase was detected in treated females. Ziram, tested in a lower range of doses because of its higher toxicity, resulted negative in both sexes. Both the acute toxicity and the ratio polychromatic/normochromatic erythrocytes indicated some sex specificity in the toxic effects induced by these dithiocarbamates in the B6C3F1 mouse. © 1992 Wiley‐Liss, Inc.
A novel model to assess developmental toxicity of dihaloalkanes in humans: Bioactivation of 1,2‐dibromoethane by the isozymes of human fetal liver glutathione S‐transferase
Authors: Mitra Ashoke , Hilbelink Don R. , Dwornik Julian J. , Kulkarni Arun
Published: February 2006

Glutathione S‐transferase (GST) isozymes from human fetal liver (16–18 weeks gestation) were purified by affinity chromatography followed by ion‐exchange high performance liquid chromatography (HPLC). The purified isozymes were used to investigate toxicity of 1,2‐dibromoethane(EDB) in an in vitro model of rat embryos in culture as passive targets. At least five isozymes of GST were found in the human fetal liver. Two anionic forms [pI values 5.5 (P‐2) and 4.5 (P‐3)] and one basic form [pI value 8.7 (P‐6)] were clearly separated. The presence of two near‐neutral forms was also identified. All the isozymes of the human fetal liver GSTs tested metabolized EDB (specific activities were 2.1, 7.0, and 2.0 μmol of GSH consumed/min/mg protein for P‐2, P‐3, and P‐6 isozymes, respectively). Covalent binding of EDB to DNA and protein was 144% and 212% higher, respectively, with the P‐3 anionic isozyme when compared to the P‐6 basic isozyme of GST. No covalent binding to either protein or DNA was observed with the P‐2 isozyme.

EDB bioactivation by the GST isozyme P‐3 (15 units; 1 unit = 1 nmol of GSH consumed/min) resulted in toxicity to cultured rat embryos. Significant reductions of crown rump length, yolk sac diameter, and the composite score of morphological parameters (Brown and Fabro method) were observed. The central nervous system, optic and olfactory systems, and the hind limb were most significantly affected. The results of this investigation suggest that EDB may be classified as a suspected developmental toxicant in humans. © 1992 Wiley‐Liss, Inc.
N‐nitrosoheptamethyleneimine‐induced pulmonary and esophageal carcinogenesis and effects of concomitant treatment with bleomycin in rats
Authors: Yoshida Yasunori , Hasegawa Ryohei , Takaba Katsumi , Imaida Katsumi , Ito Nobuyuki
Published: February 2006

The combination effects of bleomycin with N‐nitrosoheptamethyleneimine (NHMI) or dihydroxy‐di‐N‐propylnitrosamine (DHPN) on pulmonary carcinogenesis were investigated. Male F344 rats were given NHMI (20 or 40 ppm) or DHPN (200 ppm) in the drinking water and intraperitoneally injected with bleomycin (1 mg/kg) once a week for 18 weeks and then killed at week 24. Many rats treated with NHMI died before the termination of the experiment due to toxicity or development of advanced esophageal carcinomas, considered to be the main cause of death. Detailed histological examination performed on rats killed at week 24 revealed no statistically significant effects of bleomycin on NHMI or DHPN induction of neoplastic lesions in the lung or esophagus, although pulmonary carcinomas were only found in two rats treated with NHMI plus bleomycin.

Under the present experimental conditions, NHMI exerted stronger carcinogenic activity in the esophagus than in the lung, and no obvious modifying effects of simultaneously administered bleomycin were evident on NHMI‐ or DHPN‐induced pulmonary carcinogenesis. © 1992 Wiley‐Liss, Inc.
Transplacental genotoxicity of triethylenemelamine, benzene, and vinblastine in mice
Authors: Xing S. G. , Shi X. , Wu Z.‐L. , Chen J.‐K. , Wallace W. , Whong W.‐Z. , Ong T.
Published: February 2006

Transplacental cytogenetic effects of triethylenemelamine (TEM), benzene, and vinblastine on maternal mice and their fetuses have been investigated using micronucleus and sister chromatid exchange (SCE) as genetic endpoints. CD‐1 mice were treated on day 14 and 15 of gestation with TEM (0.125, 0.25, and 0.5 mg/kg), benzene (439, 878, and 1,318 mg/kg), and vinblastine (0.5, 1, and 2 mg/kg) by intraperitoneal injection at 24 hr intervals, and sacrificed 40 hr after the first injection. Erythrocytic precursor cells in maternal bone marrow and fetal livers (2–4) from each pregnant mouse were used for the micronucleus and/or the SCE analyses. Significant dose‐related increases in both micronuclei and SCE were found in maternal bone marrow and fetal liver following TEM treatment. Benzene at the highest dose (1,318 mg/kg) also caused a significant increase in micronuclei and SCE in both maternal bone marrow and fetal liver cells. The embryonic genotoxic effect of TEM was much higher than that of benzene for both genetic endpoints, and the frequency of micronuclei induced by benzene was higher in fetal liver than in maternal bone marrow cells. Vinblastine, a spindle poison, induced micronuclei but not SCE. Micronuclei induction by vinblastine was 7 fold greater in maternal bone marrow than in fetal liver cells. All three chemicals were cytotoxic in maternal bone marrow cells, but not in fetal liver cells except for TEM, which showed a weak cytotoxicity in fetal liver cells in the micronucleus assay. These results indicate that TEM, benzene, and vinblastine are transplacental genotoxicants in mice. © 1992 Wiley‐Liss, Inc.
Changes in secondary structure of DNA of rat embryos following treatment with diethylnitrosamine and methylazoxymethanol acetate in vivo
Authors: Catalano Alberto , Huang Peter H. T.
Published: February 2006

Diethylnitrosamine (DEN) and methylazoxymethanol acetate (MAM) are not transplacental carcinogenic but embryotoxic to Wistar rats when administered by i.p. injection on day 12 of gestation. MAM, a weak teratogen to rats during this period, induced a dose dependent increase in the number of resorptions to 15% and 40% of the litters following doses of 15 and 25 mg/kg bw, respectively. Rats similarly treated with 70, 150, and 180 mg DEN/kg bw resulted in increases in total DNA mass of day 13 embryos by 31%, 45% and 52%, respectively, compared to the saline treated controls. Twenty percent reduction in total DNA amount was detected following 25 mg MAM/kg bw.

Benzoylated DEAE‐cellulose (BD‐cellulose) chromatography fractionates DNA on the basis of secondary structure by stepwise elution of double‐stranded DNA with 1.0M NaCl solution (SE‐DNA) followed by elution of DNA containing single‐stranded regions with caffeine solution (CE‐DNA). Day 13 embryonic DNA was monitored by in vivo labelling with [methyl‐3H]‐thymidine (3H‐TdR) on days 6 and 7 of gestation. Significant increases in percentages of caffeine‐eluted DNA (%CE‐DNA) compared to control values were detected 24 h after treatment of day 12 embryos with 70, 150, and 180 mg DEN/kg bw. Such increases were not observed after MAM.

Incorporation of [methyl‐14C]‐thymidine (14C‐TdR) into eymbryonic DNA demonstrated the effects of treatment with these compounds on DNA synthesis in vivo. When compared to saline controls, DEN induced significant increases in 14C‐TdR incorporation into embryo DNA, 1 h prior to analysis, but the increases were not proportional to the doses administered. Similar analysis of MAM treated samples showed no significant changes to %CE‐DNA values. The relative %CE‐DNA is expressed as the ratio of the percentage of caffeine‐eluted 14C‐labelled DNA to %CE‐DNA (i.e., %CE‐14C‐DNA:%CE‐3H‐DNA). In the majority of control embryos the 14C‐specific activity of CE‐DNA was higher than the 14C‐specific activity of SE‐DNA. No significant change to relative %CE‐DNA values of embryos to those of the controls was observed 24 h after treatment of day 12 gestation rats with single doses of DEN and MAM.

The results of this study support the hypothesis that initiation mechanisms of teratogenesis and transplacental carcinogenesis are different. The pertinence of %CE‐DNA and realative %CE‐DNA values to teratogenesis and transplacental carcinogenesis is also discussed. © 1992 Wiley‐Liss, Inc.
Valproic acid induced abnormal development of the central nervous system of three species of amphibians: Implications for neural tube defects and alternative experimental systems
Authors: Oberemm A. , Kirschbaum F.
Published: February 2006

Embryos of Ambystoma mexicanum, Xenopus laevis, and Hyperolius viridiflavus taeniatus were exposed to various concentrations of valproic acid (VPA: 0.1, 1, 5, 10 mM) from blastula stage (S) 9 on up to advanced gastrulation of control embryos (S 11 1/2–12). At 10 and 5 mM VPA early development was affected in all species tested. However, the most pronounced effects occurred in Ambystoma: the neural folds appeared delayed and showed a flattened and wavy shape; the neural tube was not formed and embryos successively died. In Xenopus and Hyperolius (10, 5 mM VPA) the beginning of gastrulation was delayed up to neurulation of control embryos. In Xenopus many of the embryos completed neurulation, whereas some embryos exposed to 10 mM VPA showed neural tube defects (NTDs) of different type and degree (open neural tube at different regions of the dorsum). In Hyperolius neural folds arose around the blastoporus and fused later on (earlier in embryos treated with 5 mM VPA), but the shape of these embryos was abnormal and the development was not continued (pronounced effect at 10 mM VPA). Comparing the three species, Xenopus proved to be the least sensitive species (at 5 mM VPA 14.2% NTDs of total malformations compared to 100% in the other species). The most sensitive species, Ambystoma, developed head‐oedema at 1 mM VPA, whereas the anurans were not affected. Our results suggest a similar mechanism of VPA‐induced NTDs in mammals and amphibians. © 1992 Wiley‐Liss, Inc.
Effect of acyl derivatives of 4,4,4‐trifluoro‐1‐phenyl‐1,3‐butanedione on 2‐acetylaminofluorene‐induced glutathione S‐transferase positive foci in the rat liver
Authors: Hasegawa Ryohei , Wang Ching Y. , Kim Dae‐Joong , Lee Mei‐Sie , Yoshida Yasunori , Ito Nobuyuki
Published: February 2006

Acyl derivatives of 4,4,4‐trifluoro‐1‐phenyl‐1,3‐butanedione (TFPB), 1‐benzoyl‐2‐tri‐fluoromethyl‐2‐acetoxyethene (BTAE), and 1‐benzoyl‐2‐trifluoromethyl‐2‐(4‐methyl‐thio)benzoyloxyethene (BTME), were synthesized and investigated for inhibition of tRNA binding by N‐acethoxy‐2‐acetylaminofluorene (N‐AcO‐AAF), and induction of glutathione S‐transferase placental form (GST‐P) positive foci in the rat liver by 2‐acetylaminofluorene (2‐AAF). Male F344 rats were given BTAE or BTME intraperitoneally and 2‐AAF by intragastric intubation. Two weeks following the treatment, the rats were maintained on the diet containing 0.05% phenobalbital for an additional 6 weeks and then killed. Development of GST‐P positive foci was not affected by concomitant treatment with BTAE or BTME. These two compounds inhibited the in vitro binding of N‐AcO‐AAF to tRNA. Thus, although these diacylmethane derivatives had the in vitro inhibitory activity, they did not inhibit tumor‐initiating activity of 2‐AAF in the rat liver. © 1992 Wiley‐Liss, Inc.
Stage‐specific effects of teratogens on sea urchin embryogenesis
Authors: Graillet C. , Pagano G. , Girard J.‐P.
Published: February 2006

The effect of direct (chlorambucil and allopurinol) and indirect (cyclophosphamide) teratogens on the fertilization and early development of sea urchin embryos has been investigated. Fertilization was affected by none of the drugs tested. Continuous exposure of embryos to chlorambucil (10−6 to 3 × 10−4 M) starting after fertilization delayed the first cleavage and hatching. Developmental defects in chlorambucil‐treated embryos consisted mainly of blastula and gastrula‐arrested embryos and in a limited number (25%) of plutei with malformed gut or skeleton. Post‐hatching exposure to chlorambucil led to malformed plutei only. Early (pre‐hatching) exposure to allopurinol (10−6 to 10−3 M) did not affect cleavage but induced developmental defects in a ratio comparable to chlorambucil. Post‐hatching exposure to allopurinol failed to affect the embryogenesis. The indirect teratogen cyclophosphamide (10−6 to 3 × 10−5 M) had no effect on the early embryogenesis. Results were discussed in view of using sea urchin embryos to detect and analyze the early mechanisms of teratogenic action. ©1993 Wiley‐Liss, Inc.
Mammary gland morphology and responsiveness to regulatory molecules following prenatal exposure to diethylstilbestrol
Authors: Vassilacopoulou Dido , Boylan Elizabeth S.
Published: February 2006

Female ACI rats were exposed to diethylstilbestrol (DES) in utero to evaluate the effects on the peri‐pubertal mammary gland with respect to 1) mammary gland morphology, 2) sensitivity to natural and synthetic estrogens, and 3) sensitivity to endogenous epidermal growth factor (EGF). Pregnant rats were injected with vehicle (sesame oil) or DES (total dose, 8.0 μg) on days 15 and 18 of gestation. DES‐exposed and control offspring were ovariectomized at 34 days of age and sacrificed at day 53 to ascertain the morphology of the mammary glands in peri‐pubertal rats. Elvax pellets containing 5 or 11 ng 17 β‐estradiol (E2) or DES were implanted subcutaneously adjacent to the third mammary gland pair. Furthermore, additional groups of rats were subjected to bilateral sialoadenectomy at the day of ovariectomy to remove the major source of endogenous EGF. A significant proportion of mammary glands of DES‐exposed animals exhibited atypical mammary gland morphology, with approximately 25% displaying hypo‐differentiation, and about 5% with aberrant hyper‐proliferation. From the pellet implantation experiments, the DES‐exposed glands were found to be refractory to stimulation by 5 and 11 ng DES; however, there was no significant difference in the degree of local stimulation elicited by either dose of E2. Sialoadenectomy at d34 had no apparent effect on mammary gland morphology in either the DES‐exposed or vehicle‐exposed groups. These data support the premise that the mammary gland of the peri‐pubertal ACI rat is morphologically and physiologically aberrant as a function of transplacental exposure to DES, with a significant percentage hypo‐differentiated and refractory to subsequent hormonal stimulation. ©1993 Wiley‐Liss, Inc.
Lack of promotion of N‐butyl‐N‐(4‐hydroxybutyl)nitrosamine‐initiated urinary bladder carcinogenesis in mice by rat cancer promoters
Authors: Tamano Seiko , Asakawa Emiko , Boomyaphiphat Pleumjit , Masui Tsuneo , Fukushima Shoji
Published: February 2006

The effects of dietary exposure to sodium L‐ascorbate (Na‐AsA), butylated hydroxyanisole (BHA), and diphenyl on the development of urinary bladder tumors in a mouse two‐stage carcinogenesis model were examined. Male B6C3F1 mice received 0.05% N‐butyl‐N‐(4‐hydroxybutyl) nitrosamine (BBN) in the drinking water for 4 weeks and were then treated with 5% Na‐AsA, 1% BHA, or 1% diphenyl for 32 weeks. None of these chemicals enhanced the development of either preneoplastic or neoplastic lesions in the urinary bladder. Furthermore, DNA synthesis levels of urinary bladder epithelium in mice treated with each substance alone for 8 weeks were not elevated significantly, although Na‐AsA was associated with a significant increase in the urinary pH value and Na+ concentration. The results indicate that Na‐AsA, BHA, and diphenyl do not exert an enhancing influence on mouse bladder carcinogenesis, in clear contrast to the case in the rat. ©1993 Wiley‐Liss, Inc.
Modification of hepato‐ and renal carcinogenesis by catechol and its isomers in rats pretreated with N‐ethyl‐N‐hydroxyethylnitrosamine
Authors: Okazaki Shuzo , Hoshiya Toru , Takahashi Satoru , Futakuchi Mitsuru , Saito Koichi , Hirose Masao
Published: February 2006

Modifying effects of catechol, resorcinol, and hydroquinone on second stage hepato‐ and renal carcinogenesis was investigated in rats pretreated with N‐ethyl‐N‐hydroxyethyl‐nitrosamine (EHEN).

Groups of twenty 6‐week‐old Wistar/Crj male rats were treated with 0.1% EHEN in the drinking water for 3 weeks. Starting 1 week after the termination of EHEN treatment, they were given a diet containing 0.8% catechol, 0.8% resorcinol, or 0.8% hydroquinone or basal diet for 36 weeks. Further groups of 15 rats were each treated with the same doses of phenolic compounds or basal diet alone without EHEN pretreatment. All surviving animals were killed at the end of week 40 when histopathological assessment revealed significant reduction of the numbers per rat of hepatocellular adenomas and hepatocellular carcinomas by resorcinol, whereas hydroquinone significantly enhanced the numbers per rat of renal microadenomas and renal cell tumors. On the other hand, the number of α2u‐globulin positive tubules in the animals treated with hydroquinone was significantly lower than controls, without any alteration in bromodeoxyuridine (BrdU) incorporation. Lipid peroxidation, as evaluated by thiobarbituric acid reactive substance (TBARS), was at control levels in the kidneys of rats treated with hydroquinone throughout the experiment.

The results showed that the known renal carcinogen hydroquinone potently enhances the second stage of EHEN‐induced renal carcinogenesis, while its isomer resorcinol inhibited hepatocarcinogenesis. α2u‐Globulin and lipid peroxidation may not play roles in hydroquinone‐associated promotion of renal carcinogenesis. ©1993 Wiley‐Liss, Inc.
Modifying influence of prior treatment with toxic agents on induction of preneoplastic and neoplastic lesions in a medium‐term multi‐organ carcinogenesis bioassay
Authors: Hagiwara A. , Tiwawech D. , Imaida K. , Tamano S. , Shibata M.‐A. , Fukushima S. , Ito N.
Published: February 2006

The modifying potential of prior administration of toxic agents was investigated in our multi‐organ carcinogenesis model using male F344/DuCrj rats with the aim of assessing the link between tissue damage and initiation. Animals were administered one of four toxic agents for 8 wk, and then treated with N‐diethylnitrosamine (DEN, 100 mg/kg body weight (b.w.), intraperitoneally (i.p.), single injection), N‐methylnitrosourea(MNU, 20 mg/kg b.w., i.p., four times during wk 9 and 10), and dihydroxy‐di‐N‐propylnitrosamine (DHPN, 0.1% in drinking water, during wk 11 and 12) for multi‐organ carcinogenesis. All surviving rats were killed at the end of wk 36, and the major organs carefully examined for preneoplastic and neoplastic lesion development. Immunohistochemical demonstration of glutathione S‐transferase placental form (GST‐P) positive foci was also performed to facilitate quantitative assessment of liver lesion development. D‐galactosamine (300 mg/kg b.w., i.p., once a week), a hepatotoxin, significantly inhibited the induction of GST‐P positive foci, while 4,4′‐diaminodiphenylmethane (DDPM, 0.1 % in diet), a bile duct proliferator which is itself a hepatocarcinogen, possessed enhancing activity. DDPM, also a goitrogen, clearly inhibited the development of follicular cell tumors in the thyroid. Uracil (3.0% in diet), which is an inducer of papillomatosis in the urinary bladder, did not exert any enhancing potential on bladder carcinogenesis. Bleomycin (2 mg/kg b.w., i.p., twice a week), which is an alveolar epithelium injuring agent, also did not modify the induction of alveolar epithelium proliferative lesions. These results indicate that prior organ injury by toxic agents does not always act to enhance sensitivity to carcinogenesis. ©1993 Wiley‐Liss, Inc.
Clastogenic effects of the dithiocarbamate fungicides thiram and ziram in Chinese hamster cell lines cultured in vitro
Authors: Mosesso P. , Turchi G. , Cinelli S. , Di Chiara D. , Fiore M. , Palitti F.
Published: February 2006

We report here the results obtained using the dithiocarbamate fungicides thiram and ziram to investigate the induction of chromosomal aberrations (CAs) in Chinese hamster ovary (CHO) cells both in the absence and presence of S9 metabolism, and in a Chinese hamster epithelial liver (CHEL) cells which retain metabolic competence to activate different classes of promutagens/procarcinogens. Both thiram and ziram proved to be strong chromosome breaking agents in the CHEL cells and CHO cells in the presence of S9 metabolism. These findings suggest that thiram and ziram require metabolic conversion to become genetically active, and corroborate the evidence that CHEL cells are suitable to activate and detect a broad spectrum of chemical procarcinogens including these two pesticides. © 1994 Wiley‐Liss, Inc.
Effect of phenacetin pretreatment on renal pelvic carcinogenesis by N‐butyl‐N‐(4‐hydroxybutyl)nitrosamine in NON/Shi mice of both sexes
Authors: Murai Takashi , Mori Satoru , Hosono Motoko , Takeuchi Yasuyoshi , Oohara Tadao , Makino Susumu , Takeda Reiji , Fukushima Shoji
Published: February 2006

Influences of phenacetin (PH) pretreatment on renal pelvic carcinogenesis induced by N‐butyl‐N‐(4‐hydroxybutyl)nitrosamine (BBN) were examined in NON/Shi mice of both sexes. Histopathological examination revealed that PH pretreatment enhanced not only the induction of urinary tract carcinoma but also distant metastasis of renal pelvic carcinoma by BBN in male mice. The high incidence of urinary tract carcinoma by PH pretreatment might be due to hydronephrosis and epithelial proliferative lesions enhanced by PH, since a single treatment of PH induced hydronephrosis in all mice and simple hyperplasia in 70–80% of mice used. © 1994 Wiley‐Liss, Inc.
Profile of procarbazine‐induced embryotoxicity in an embryo hepatocyte Co‐culture system and after in utero glutathione depletion
Authors: Ebron‐McCoy Marian T. , Nichols Harriette P. , Andrews James E. , Kavlock Robert J.
Published: February 2006

Procarbazine (PCZ) is an antineoplastic agent useful in the treatment of Hodgkin's disease, brain tumors, and chronic leukemia. PCZ is dysmorphogenic to developing embryos exposed in vivo or cultured in the serum of PCZ‐treated rats. However, embryos directly cultured with PCZ (up to 400 μg/ml) or PCZ plus S‐9 liver fractions are unaffected. Since intact liver cells provide several advantages over hepatic subcellular fractions for in vitro bioactivation, we exposed rat embryos to PCZ in an embryo/hepatocyte co‐culture system. Gestation day (GD) 9.5 rat embryos exposed to 0, 200, 300, or 400 μg PCZ/ml in the presence of untreated or phenobarbital induced male rat hepatocytes failed to display toxicity. However, in a companion study GD 9.5 rat embryos cultured in the serum from PCZ‐treated rats exhibited developmental deficiencies. Studies have shown that the formation of toxic metabolites can result from glutathione (GSH) conjugation of toxicants in the liver. Therefore, in a second set of experiments, rat embryos were cultured in serum from rats pretreated with two GSH depleters (phorone and buthionine sulfoximine) and subsequently dosed with PCZ. Effects on development were enhanced when embryos were cultured in the serum from PCZ‐treated/GSH depleted rats. These data indicate that PCZ requires in vivo activation to be dysmorphogenic and further suggest that the metabolite(s) responsible for procarbazine embryotoxicity are formed readily under conditions of low GSH levels. This argues against a glutathione conjugate as the ultimate toxicant. © 1995 Wiley‐Liss, Inc.

This article is a US Government work and, as such, is in the public domain in the United States of America.
In vitro transforming effect of the fungicides metalaxyl and zineb
Authors: Perocco Paolo , Colacci Annamaria , Bonora Bruna , Grilli Sandro
Published: February 2006

The cytotoxic effects and the transforming properties of two fungicides, metalaxyl and zineb, whose mutagenic or carcinogenic activity has not been clarified yet, were analyzed in the in vitro BALB/c 3T3 cell transformation test both in the presence and in the absence of an exogenous metabolizing system. Zineb was completely detoxified when the exogenous metabolizing system was added to the target cells to increase their inherent metabolic capacity. Metalaxyl induced cell transformation at any assayed dosage, i.e., 500, 250, and 50 μg/ml, in the presence of bioactivation, and at the highest dosage (500 μg/ml) in the absence of bioactivation. The transforming effect was detectable only in the level‐II transformation cultures and it was likely linked to the induction of additional cell proliferation which allowed obtaining the transformation amplification in these experimental conditions. © 1995 Wiley‐Liss, Inc.