Showing papers in "Teratogenesis Carcinogenesis and Mutagenesis in 1999"
TL;DR: A wide range of experimental designs are used in investigations using the Comet assay, but it is important to recognize that the sample rather than the cell is the experimental unit, and interpretation of the results should be related to identifying changes of biological importance rather than relying solely on the P values of specific statistical tests.
Abstract: A wide range of experimental designs are used in investigations using the Comet assay. The statistical issues associated with this assay are however not particularly unusual or difficult. It is important however to recognize that the sample rather than the cell is the experimental unit. Statistical analyses based upon measures from the individual cells can lead to serious misinterpretation of results. Interpretation of the results of the assay should be related to identifying changes of biological importance rather than relying solely on the P values of specific statistical tests. Teratogenesis Carcinog. Mutagen. 19:109–119, 1999. © 1999 Wiley-Liss, Inc.
119 citations
TL;DR: Food sources that yield chlorophyll derivatives may play a significant role in cancer prevention, and antimutagenic effect towards 3-methylcholanthrene (3-MC) is exhibited, suggesting that the porphyrin nucleus may complex directly with the mutagen.
Abstract: Much attention in recent years has been given to the antigenotoxicity of chlorophyll. Chlorophyll, however, is known to be converted into pheophytin, pyropheophytin, and pheophorbide in processed vegetable food and following ingestion by humans. Studies were conducted on the antimutagenic and tumoricidal potencies of these compounds. All the chlorophyll derivatives tested exhibit identical antimutagenic effect towards 3-methylcholanthrene (3-MC), suggesting that the porphyrin nucleus may complex directly with the mutagen. It does not exclude, however, another mechanism of activity involving inactivation the enzymatic transformation of 3-MC. In contrast, the action of N'-nitro-N'-nitrosoguanidine (MNNG) depends upon structural differences between the chlorophyll derivatives. It is significantly lower when the phytol-containing pheophytin and pyropheophytin are tested as to that of the phytol-lacking pheophorbide. The higher concentrations of the chlorophyll derivatives were required to reduce the mutagenicity of MNNG than needed for 3-MC. The cytotoxicity of chlorophyll derivatives against tumor cells also was evaluated. The cellular uptake and inhibition of myeloma cell multiplicity were found to be greater for pheophorbide than for pheophytin. Calculated on the amount of cell associated chlorophyll derivative, however, pheophytin was more cytostatic/cytotoxic than pheophorbide. The results presented in this report indicate that food sources that yield chlorophyll derivatives may play a significant role in cancer prevention. Teratogenesis Carcinog. Mutagen. 19:313-322, 1999.
90 citations
TL;DR: The antimutagenic effect of ethanolic extract of propolis (EEP) and honeybee venom, both collected in the State of São Paulo, Brazil, was assessed by the Salmonella/microsome assay upon direct- and indirect-acting mutagens.
Abstract: The antimutagenic effect of ethanolic extract of propolis (EEP) and honeybee (Apis mellifera) venom, both collected in the State of Sao Paulo, Brazil, was assessed by the Salmonella/microsome assay upon direct- and indirect-acting mutagens. EEP had inhibitory effect (in an ascending order) on the mutagenicity power of daunomycin (TA102), benzo(a)pyrene (TA100), and aflatoxin B(1)(TA98) and the venom acted against the mutagenicity of 4-nitro-o-phenylenediamine (TA98) and daunomycin (TA102). Teratogenesis Carcinog. Mutagen. 19:403-413, 1999.
75 citations
TL;DR: It is shown that these compounds could produce DNA damage in human blood cells in the Comet assay and also, that rat liver microsomes could abolish the effect of DEHP, and in the intact animal, no response may be observed.
Abstract: Phthalate esters are among the most extensively used industrial chemicals and are widely distributed in the environment. Di-(2-ethylhexyl)phthalate (DEHP) and its hydrolysis product mono-(2-ethylhexyl)phthalate (MEHP) have been examined for genotoxic activity on previous occasions. Only MEHP was found to cause chromosome damage in CHO cells but was without effect in the sister chromatid exchange and hypoxanthine guanine phosphoribosyl assay. DEHP was found to be a weak direct acting mutagen in Salmonella typhimurium strain TA100, the mutagenic activity of which could be abolished by rat liver microsomes (S9 mix). The clastogenicity and weak mutagenicity suggest a possible contributory role for these compounds in the observed carcinogenicity of the phthalates, which have been thought predominantly to be linked to cancer pathology through proliferation of hepatic peroxisomes. The present study showed that these compounds could produce DNA damage in human blood cells in the Comet assay and also, that rat liver microsomes could abolish the effect of DEHP. Thus in the intact animal, no response may be observed.
60 citations
TL;DR: The results clearly indicate the exacerbated effect of turmeric and curcumin on radiation-induced clastogenicity, suggesting that these antioxidants are also potentiating agents depending on the experimental conditions.
Abstract: The effect of turmeric and curcumin, two natural antioxidants, on the frequencies of chromosome aberrations induced in Chinese hamster ovary (CHO) cells by gamma-radiation was investigated. Cells were treated with three concentrations of each drug, turmeric (100, 250, and 500 microg/ml) and curcumin (2.5, 5, and 10 microg/ml), and then irradiated (2.5 Gy) during different phases of the cell cycle. Turmeric was not clastogenic by itself, whereas curcumin at 10 microg/ml enhanced the chromosomal damage frequency. Neither of the two antioxidants showed protective effect against the clastogenicity of gamma-radiation. Instead, an obvious increase in the frequencies of chromosome aberrations was observed when turmeric at 500 microg/ml was associated with gamma-radiation during G2/S phase, and curcumin at 10 microg/ml plus gamma-radiation during S and G2/S phases of the cell cycle. The results clearly indicate the exacerbated effect of turmeric and curcumin on radiation-induced clastogenicity, suggesting that these antioxidants are also potentiating agents depending on the experimental conditions.
56 citations
TL;DR: It is clearly demonstrates that curcumin does not inhibit DNA damaging action of hexavalent chromium in human lymphocytes and GM cells, and the total effect of chromium andCurcumin is additive.
Abstract: Human population can be considered as a subject of combined exposure to chemicals. Hexavalent chromium is a well-known mutagen and carcinogen. Curcumin, a popular spice and pigment, is reported to have antineoplastic properties. The single cell gel electrophoresis (Comet assay) is a sensitive technique that allows detecting double- and single-strand DNA breaks caused by a broad spectrum of mutagens. In the present work the ability of curcumin to reduce DNA damage induced by chromium in human lymphocytes and gastric mucosa (GM) cells was investigated by using the comet assay. Chromium at 500 microM evoked DNA damage measured as significant (P < 0.001), about a two-fold increase in comet tail moment of both lymphocytes and GM cells. Curcumin at 10, 25, and 50 microM also damaged DNA of both types of cells in a dose-dependent manner: the increase in the tail moment reached about twenty times of the control value (P < 0.001). The combined action of chromium at 500 microM and curcumin at 50 microM resulted in the significant (P < 0.001) increase in the comet tail moment of both types of cells. In each case, treated cells were able to recover within 60 min. Our study clearly demonstrates that curcumin does not inhibit DNA damaging action of hexavalent chromium in human lymphocytes and GM cells. Moreover, curcumin itself can damage DNA of these cells and the total effect of chromium and curcumin is additive. Further studies are needed to establish the role of interaction of curcumin with DNA in carcinogenesis.
50 citations
TL;DR: Speciation of tissue total arsenic into iAs, methylarsonic acid (MMA), and dimethylarsinic acid (DMA) permits us to link tissue iAs and HO enzyme induction and there was a sublinear relationship between administered arsenite and biological effect in rats.
Abstract: Rat heme oxygenase (HO) activity was used as a specific (among forms of arsenic) and sensitive biomarker of effect for orally administered sodium arsenite in rats. Time course studies showed that HO was induced in rat liver from 2 to 48 h in both rat liver and kidney. Hepatic and renal inorganic arsenic (iAs) concentrations were high at times preceding a high degree of HO induction. At times following pronounced HO induction, tissue dimethylarsinic acid concentrations were high. Dose-response studies of arsenite showed substantial HO induction in liver at doses of 30 micromol/kg and higher and in the kidney at doses of 100 micromol/kg and higher. Doses of 10 (in liver) and of 30 micromol/kg (in kidney) sodium arsenite given by gavage did not significantly induce rat HO activity. Speciation of tissue total arsenic into iAs, methylarsonic acid (MMA), and dimethylarsinic acid (DMA) permits us to link tissue iAs and HO enzyme induction. There was a linear relationship between tissue inorganic arsenic (iAs) concentration and tissue HO in individual rats (r(2) = 0.780 in liver and r(2) = 0.797 in kidney). Nonlinear relationships were observed between administered arsenite dose and either liver or kidney iAs concentration. Overall, there was a sublinear relationship between administered arsenite and biological effect in rats. Teratogenesis Carcinog. Mutagen. 19:385-402, 1999. Published 1999 Wiley-Liss, Inc.
45 citations
TL;DR: The analysis of DNA migration and the distribution of DNA damage showed that there are differences in sensitivity between organs and tissues to this compound, and the sensitivity of SCGE assay allows the detection of long term DNA damage and the possibility to compare it in various tissues and target organs.
Abstract: Vanadium pentoxide (V 2 O 5 ) was tested for its ability to induce genotoxic damage in six different organs (liver, kidney, lung, spleen, heart, and bone marrow) of mice by using the alkaline Single Cell Gel Electrophoresis (SCGE) assay. Animals were sacrificed 24 h after i.p. administration of the vanadium pentoxide of 23.0, 11.5, or 5.75 μg/g (corresponding to the LD 50 , 1/2 LD 50 and 1/4 LD 50 , respectively). In all tissues and organs evaluated (except for bone marrow), V 2 O 5 increased the number of cells with damage. Our results showed that i.p. injection of V 2 O 5 induced DNA damage in different organs and tissues, and that this kind of damage can be observed even 24 h after treatment. The analysis of DNA migration and the distribution of DNA damage showed that there are differences in sensitivity between organs and tissues to this compound. In addition the sensitivity of SCGE assay allows the detection of long term DNA damage and the possibility to compare it in various tissues and target organs.
42 citations
TL;DR: Investigation of the effects of VC on human peripheral blood lymphocytes in vitro and its anticlastogenic effect on chromosomal aberrations induced by doxorubicin found the efficiency of VC in protecting against chromosome damage was dependent on the doses used.
Abstract: Some chemotherapeutic approaches have proposed the use of antioxidants such as vitamin C (VC) to minimize the cytotoxicity and damage induced in normal tissue by antitumor agents that produce free radicals. Nevertheless, VC can also be cytotoxic, genotoxic, and harmful when combined with antitumor agents in human cells in vitro. The present study was undertaken to investigate the effects of VC (100, 200, 500, and 1,000 μg/ml) on human peripheral blood lymphocytes in vitro and its anticlastogenic effect on chromosomal aberrations induced by doxorubicin (DXR). VC did not show a clastogenic effect by itself, except at 1,000 μg/ml. At the concentration of 100 or 200 μg/ml of VC, administered in pre-, post-, or simultaneous treatment, there was a significant reduction in both chromosome aberrations and number of abnormal metaphases induced by DXR. At the doses of 500 or 1,000 μg/ml, VC did not present the same protective effect and was cytotoxic. Under the present experimental conditions, the efficiency of VC in protecting against chromosome damage was dependent on the doses used. Teratogenesis Carcinog. Mutagen. 19:53–59, 1999. © 1999 Wiley-Liss, Inc.
41 citations
TL;DR: A statistically significant increase in the frequency of chromosomal damage was observed when the middle and the highest concentrations of curcumin were associated with DXR during the G1/S, S, and S/G2 phases of the cell cycle.
Abstract: Curcumin, a natural phenolic compound, is gaining importance as a free radical scavenger. This study was undertaken to investigate the modulatory effects of curcumin on the chromosomal damage induced by the antitumoral doxorubicin (DXR), a known free radical generator, in Chinese hamster ovary cells in culture. Cells were treated with three concentrations of curcumin (2.5, 5, or 10 microg/ml) and then treated with DXR (1.0 microg/ml) during different phases of the cell cycle. The results show that curcumin induces chromosomal damage in CHO at the highest concentration when compared to the untreated control. Neither treatment with curcumin shows a reduction in the clastogenicity of DXR. Instead, a statistically significant increase in the frequency of chromosomal damage was observed when the middle and the highest concentrations of curcumin were associated with DXR during the G1/S, S, and S/G2 phases of the cell cycle. The results clearly indicate the potentiating effect of curcumin on DXR-induced chromosomal damage.
36 citations
TL;DR: A possible role for GM-CSF in modulating teratogen-induced effects, a process in which maternal immune responses such as splenocytes proliferation and cytokine production might be involved is suggested.
Abstract: It was already shown that stimulation of the maternal immune system by allogeneic or xenogeneic leukocytes is capable of affecting embryonic responses to teratogenic insults and various cytokines, including granulocyte macrophage-colony stimulating factor (GM-CSF), were implicated as mediators of this effect. Therefore, in the present study we tried to assess the ability of GM-CSF to modulate teratogenic activity, along with possible changes in systemic as well as local maternal immune responses, that might be involved in the process. Thus, the percentage of cyclophosphamide (CP)-treated embryos exhibiting limb malformations was shown to decrease significantly following GM-CSF administration. This effect was found to be comparable to that demonstrated by intrauterine leukocytes administration. GM-CSF treatment resulted in a significant enhancement in maternal splenocytes Con-A-induced proliferation, as well as Interleukin-2 (IL-2) and IL-3 production. Examination of leukocyte cell surface antigens expressed by splenocytes revealed no statistically-significant changes in the level of the T lymphoid antigens Thy-1, CD5, CD4, CD8 and CD3, the macrophage antigen Mac-1, and the adhesion molecules LFA-1alpha, LFA-1beta and L-Selectin, following GM-CSF immunostimulation. In parallel, immunohistochemical analysis of the uteroplacental unit revealed Mac-1 and to a lesser extent LFA-1beta-positive cells localized to the myometrium and the placenta in both the control and the GM-CSF-treated groups, while no cells expressing Thy-1, CD3, CD4, CD8, or LFA-1alpha could be demonstrated. Our results suggest a possible role for GM-CSF in modulating teratogen-induced effects, a process in which maternal immune responses such as splenocytes proliferation and cytokine production might be involved.
TL;DR: T-DCTN is not genotoxic nor cytotoxic to mouse bone marrow cells, submitted to acute intraperitoneal treatment in vivo, by micronucleus (MN) and chromosomal aberration (CA) tests.
Abstract: The genotoxic action of three doses of trans-dehydrocrotonin (t-DCTN), an active ingredient obtained from the bark extracts of an Amazon native plant, Croton cajucara, were examined in Swiss mouse bone marrow cells in vivo, submitted to acute intraperitoneal treatment, by micronucleus (MN) and chromosomal aberration (CA) tests. The statistical tests (Anova and Tukey) made to compare the results obtained in each of the three doses of t-DCTN with the negative-control group showed that the frequencies of MN and mitotic index were equal to the negative-control and that the frequencies of CA were lower than that observed in the negative-control. Therefore, based on our results it can be said that t-DCTN is not genotoxic nor cytotoxic to mouse bone marrow cells, submitted to acute intraperitoneal treatment in vivo. Teratogenesis Carcinog. Mutagen. 19:377-384, 1999.
TL;DR: Results of this study suggest that the differential sensitivity of tested organ systems to CP may be associated with differences in cellular events following CP-initiated cell death, and that the input of p53 in determining the response of these organ Systems to CP-induced teratogenic insult may be different.
Abstract: This study was aimed at characterizing the temporal patterns of cell responses and p53 protein expression in the limbs, head, and liver of embryos responding to cyclophosphamide (CP)-induced teratogenic insult. ICR murine embryos were examined 24, 48, or 72 h after injection of 40 mg/kg CP on day 12 of pregnancy. The cellular events and temporal pattern of p53 protein expression were determined by FACS analysis and by TUNEL (apoptosis) in the head, limbs, and liver of the embryos. All tested organs showed apoptosis and a significantly decreased proportion of live cells after 24 h. Subsequent events were organ-dependent. In the liver, there were no dysmorphic events at any time and excessive cell death had been almost compensated for by 48 h. Compensation was preceded by G1 arrest and accompanied by an increased level of p53 protein in surviving cells. Excessive cell death in the head and the limbs resulted in structural anomalies. In the head, there was an increased level of p53 protein and G1 arrest after 24 h and the number of live cells at 48 h was equal to that seen in earlier samples, despite apoptosis. In the limbs, however, only isolated viable cells were seen by 48 h, but there was no increased level of p53 protein or G1 arrest. Results of this study suggest that the differential sensitivity of tested organ systems to CP may be associated with differences in cellular events following CP-initiated cell death. They also suggest that the input of p53 in determining the response of these organ systems to CP-induced teratogenic insult may be different. Teratogenesis Carcinog. Mutagen. 19:353–367, 1999. © 1999 Wiley-Liss, Inc.
TL;DR: The phenoxyacetates 2,4-D and 4-CPA were evaluated using the Drosophila melanogaster wing spot test, which assesses for somatic mutation and recombination events, and induced a weak but significant increase in the frequency of two of the categories of recorded spots.
Abstract: The phenoxyacetates 2,4-D and 4-CPA were evaluated for genotoxicity using the Drosophila melanogaster wing spot test, which assesses for somatic mutation and recombination events. Third-instar larvae trans-heterozygous for two recessive mutations affecting the expression of wing trichomes, multiple wing hairs (mwh), and flare (flr) were treated by chronic feeding with different concentrations of the two chemicals. Feeding lasted until pupation of the surviving larvae and the genotoxic effects induced were evaluated in adults for the appearance of wing-blade cell clones with the mwh, flr, or mwh-flr phenotypes. Exposure to 2,4-D, at the highest concentration evaluated (10 mM), induced a weak but significant increase in the frequency of two of the categories of recorded spots: large single and total spots; in contrast, the 4-CPA treatments failed to induce any significant increase in the frequency of evaluated spots. When the heterozygous larvae for mwh and the multiple inverted TM3 balancer chromosome were treated with the chemicals, no increases were detected, either after the 2,4-D nor the 4-CPA treatments.
TL;DR: It was considered that weak developmental toxicity of indium by oral administration was due to low exposure concentrations in the embryo, and that embryotoxic concentrations were comparable to the serum concentration at a developmentally toxic dose by intravenous administration in an in vivo experiment.
Abstract: Developmental toxicity of indium was examined using rat embryo culture with reference to toxicokinetics. Rat embryos at day 9.5 of pregnancy were cultured for 48 h under various exposure conditions to indium trichloride. Indium was embryotoxic to cultured rat embryos at concentrations ranging from 25 to 50 microM for 24 h exposure according to the embryonic age, and the exposure concentration was more critical than the exposure time. The embryotoxic concentrations were comparable to the serum concentration at a developmentally toxic dose by intravenous administration in an in vivo experiment. It was considered from these results that the developmental toxicity of indium is a direct effect on the embryo or yolk sac and that weak developmental toxicity of indium by oral administration was due to low exposure concentrations in the embryo.
TL;DR: KDA induced DNA damage in the three cell types, confirming that KDA is genotoxic in a range of mammalian cells.
Abstract: In previous studies. N-(N'-acetyl-L-propyl)-N-nitrosoglycine (APNG) has been shown to be a potent mutagen in a variety of genotoxicity assays and a carcinogen in a limited cancer study. APNG decomposes to a carboxymethyldiazonium ion, which can also be generated from potassium diazoacetate (KDA). KDA is particularly interesting because it is a stable nitrosated derivative of glycine, one of the most common dietary amino acids. KDA has been shown to produce more O 6 carboxymethyl-and O 6 methyl-adducts than APNG, so it was anticipated that it might also be a potent genotoxic agent. Thus in the present study KDA has been investigated in the single cell gel electrophoresis (Comet) assay, which primarily measures DNA strand hreakage. Since KDA has been shown to be formed in the gut, the genotoxic effects of KDA were investigated in vitro in human adenocarcinoma colon Caco-2 cells, and in rat primary colon cells and compared to responses from human peripheral lymphocytes. KDA induced DNA damage in the three cell types, confirming that KDA is genotoxic in a range of mammalian cells.
TL;DR: DNA damage as measured in the Comet assay in peripheral blood lymphocytes is not a sensitive genotoxic marker of aflatoxin exposure in this population and there were no associations between cytogenetic damage, GSTM1 genotype, age, sex, lifestyle factors, and Comet response at the individual level.
Abstract: The single cell gel electrophoresis assay (Comet assay) was used to measure DNA damage in peripheral lymphocytes from a group of individuals from The Gambia in order to determine whether such damage could be associated with increased exposure to aflatoxin in this population. Responses obtained were correlated to responses previously obtained [1] in a cross-sectional study in the same individuals of various cytogenetic alterations [chromosomal aberrations, micronuclei (crest positive and negative staining), and sister chromatid exchanges], and aflatoxin-albumin adducts. Analysis of variance methods were used to assess the effects of smoking, GSTM1 genotype, sex, age, and smoking status. A comparison was also made between The Gambian individuals and a group of healthy, non-smoking volunteers in the United Kingdom where aflatoxin exposure would be expected to be low. From the earlier study [1], it was determined that the levels of the sister chromatid exchanges and micronuclei were higher in The Gambian group than in a European group where aflatoxin exposure was lower, but that there were no correlations between the adduct levels and the cytogenetic abnormalities at the individual level. In the present study, DNA damage as measured in the Comet assay was not significantly higher than in the healthy United Kingdom volunteers. In addition, there were no associations between cytogenetic damage, GSTM1 genotype, age, sex, lifestyle factors (smoking and aflatoxin exposure), and Comet response at the individual level. Comet response was higher in females than males in The Gambia if one outlier was excluded from analysis and not taking into account other sources of variability. It would appear that DNA damage as measured in the Comet assay in peripheral blood lymphocytes is not a sensitive genotoxic marker of aflatoxin exposure in this population. Teratogenesis Carcinog. Mutagen. 19:147–155, 1999. © 1999 Wiley-Liss, Inc.
TL;DR: The evidence gathered in this in vitro study suggests that phenothiazines can undergo peroxidative N-demethylation via lipoxygenase pathway, and an enzymatic nature of the reaction is suggested.
Abstract: Several phenothiazine derivatives have been shown to cause reproductive toxicity. The biochemical mechanisms responsible for these effects are not fully understood at present. In this study, we investigated hydrogen peroxide-dependent oxidation of six phenothiazines by purified lipoxygenase from soybean (SLO) and human term placenta (HTPLO). Chlorpromazine was employed as the prototype phenothiazine drug. Chlorpromazine was easily demethylated releasing formaldehyde when incubated at pH 7.0 and 6.5 with SLO or HTPLO, respectively, in the presence of hydrogen peroxide. The reaction was linear with respect to time, exhibited dependence on the amount of enzyme, and the concentration of chlorpromazine and hydrogen peroxide. Under the optimal assay conditions, the estimated Vmax values for chlorpromazine N-demethylation were 139 and 7.2 nmoles/min/mg of SLO and HTPLO, respectively. Collectively, the results suggest an enzymatic nature of the reaction. In the presence of gossypol and NDGA, the classical inhibitors of different lipoxygenases, the formaldehyde production was significantly decreased, as expected. Similar to SLO, the generation of chlorpromazine cation radical, an initial oxidation product with an absorption maximum at 525 nm, was also observed with HTPLO. The radical generation was detectable only under acidic conditions (pH 3.5-4.5). The formaldehyde production was also decreased by BHT and BHA, suggesting a radical nature of the SLO-mediated chlorpromazine N-demethylation. Reduced glutathione, ascorbate, and dithiothreitol suppressed the rate of SLO-dependent formaldehyde generation, presumably due to the reduction of the cation radical back to chlorpromazine in a concentration-dependent manner. Besides chlorpromazine, SLO also oxidized promazine, triflupromazine, trifluperazine, trimeprazine, and perphenazine, albeit at different rates, in the presence of hydrogen peroxide. The evidence gathered in this in vitro study suggests that phenothiazines can undergo peroxidative N-demethylation via lipoxygenase pathway. The role of this biochemical mechanism in the in vivo developmental toxicity of phenothiazines remains to be established.
TL;DR: It was suggested that the formation of selenodiglutathione and the oxidative stress were involved in the embryotoxicity of seenite and selenate, respectively.
Abstract: Effects of depletion of reduced glutathione (GSH) on selenium (Se) embryotoxicity in cultured rat embryos were examined. Rat embryos at day 9.5 of gestation were cultured for 48 h in the presence of Se as either sodium selenite at 10 and 20 microM or sodium selenate at 30 and 100 microM. Embryonic GSH was depleted by the addition of 0.1 mM of L-buthionine-[S,R]-sulfoximine (BSO) without embryotoxicity, i.e., significant growth retardation and malformation of the embryos. Selenite at 10 microM or selenate at 100 microM significantly increased the incidence of malformation of the embryos. The incidence of selenite-induced malformation of the embryos at 20 microM was significantly decreased with BSO. On the contrary, the incidence of selenate-induced malformation at 30 microM was significantly increased with BSO. It was noted that the major malformed regions of the embryos by the embryotoxic concentration of BSO alone were the same to those affected by selenite or selenate. It was considered from these results that embryonic GSH was involved in the embryotoxicity of selenite and selenate. The embryotoxicity of selenate may not be mediated through the reduction to selenite. It was suggested that the formation of selenodiglutathione and the oxidative stress were involved in the embryotoxicity of selenite and selenate, respectively.
TL;DR: Commercial products of agronomic crop plants may become a reliable and inexpensive source of phytonutrients, such as antimutagenic food supplements, and commercial soybean products and by-products may be a source of chemoprotective food additives.
Abstract: Commercial products of agronomic crop plants may become a reliable and inexpensive source of phytonutrients, such as antimutagenic food supplements. We previously demonstrated that PCC, an ethanol extract of a commercial soybean processing by-product, was able to repress induced genomic DNA damage, whole cell clastogenicity, and point mutation in mammalian cells. In this paper we separated PCC into a series of chemically defined fractions and determined their ability to repress induced mutagenic damage in Chinese hamster lung cells, Chinese hamster ovary cells and human lymphocytes. Fraction PCC70 (PCC 70% methanol eluate) contained the flavonoids from PCC and daidzin and genistin repressed 2-acetoxyacetylaminofluorene (2AAAF)-induced DNA damage measured with single cell gel electrophoresis. Genistein, however, enhanced the genotoxic impact of 2AAAF. Fraction PCC100 (PCC 100% methanol eluate) had the greatest level of antigenotoxic activity against 2AAAF in CHO cells and repressed the genotoxic capacity of the dietary carcinogen 2-amino-3-methylimidazo-(4,5-f)quinoline (IQ) in human lymphocytes. These data indicate that commercial soybean products and by-products may be a source of chemoprotective food additives.
TL;DR: Data indicate significant differences in the developmental responses to d-AZA of the mouse and the rat, which may reflect interspecies Differences in the temporal expression of genes involved in morphogenesis and/or the methylation patterns of such genes.
Abstract: 5-aza-2'-deoxycytidine (d-AZA) causes temporally related defects in the developing mouse. Treatment of 1.0 mg/kg on gestation day (GD) 8 results in axial skeletal defects; on GD9, cleft palate and vertebral defects; on GD10, hindlimb phocomelia; and on GD11, digital defects. An unusual aspect of d-AZA teratogenicity in mice is that the phocomelia appears to be specific to the hindlimb, and the forelimb is not similarly affected regardless of treatment day. The current study was initiated to evaluate the embryonic response of another species, the rat, to this unique teratogen. Pregnant Sprague Dawley (CD) rats were treated with d-AZA or vehicle control. The compound was administered i.p. on GD9, 10, 11, or 12 to parallel developmental staging of the mouse. The highest dose (1.0 mg/kg) elicited effects indicating increased sensitivity to the compound in the rat as compared to the mouse. GD9 treatment was characterized by massive resorptions; GD10, by a predominance of axial skeletal defects and cleft palate; GD11, by a predominance of forelimb phocomelia and missing ribs; and GD12 by hindlimb phocomelia and forelimb digit defects. These data indicate significant differences in the developmental responses to d-AZA of the mouse and the rat. This may reflect interspecies differences in the temporal expression of genes involved in morphogenesis and/or the methylation patterns of such genes. Molecular data generated in the mouse will be compared to that of the rat to further characterize the developmental dynamics responsible for the interspecies differences. Teratogenesis Carcinog. Mutagen. 19:329-338, 1999.
TL;DR: The present results indicate that ENU and MNU each has a unique effect on the spermatogenic cells, likely due to the different mechanism of action of ENU at the level of DNA and also to the physiological differences between different germ-cell stages.
Abstract: An autoradiographic procedure was used to measure unscheduled DNA synthesis (UDS, DNA repair synthesis) in spermatogonial and postspermatogonial cell stages of mice after treatment with two doses of N-ethyl-N-nitrosourea (ENU) and N-methyl-N-nitrosourea (MNU). Significant levels of UDS were measured in type A spermatogonia, meiotic spermatocytes, round spermatids, and early elongating spermatids but not in mature spermatids. The extent of UDS varied according to the germ cell stage and the dose. At equimolar concentrations, MNU was more efficient than ENU in eliciting a UDS response in all germ cells. After ENU treatment, type A spermatogonia showed the highest UDS response, while round and elongating spermatids showed the lowest. After MNU treatment, pachytene spermatocytes exhibited the highest UDS response while type A spermatogonia showed the lowest. The high UDS response of type A spermatogonia to ENU parallels the well-known high mutational sensitivity of spermatogonia to this chemical. Similarly, the high UDS response observed in meiotic spermatocytes and early spermatid stages after MNU treatment correlates with the high mutational sensitivity of postspermatogonial stages to MNU. Thus, the present results, like the specific locus mutation studies, indicate that ENU and MNU each has a unique effect on the spermatogenic cells. This effect is likely due to the different mechanism of action of ENU and MNU at the level of DNA and also to the physiological differences between different germ-cell stages. Teratogenesis Carcinog. Mutagen. 19:339-351, 1999. Published 1999 Wiley-Liss, Inc.
TL;DR: It was concluded that anatomical defects (skeletal malformation or variation) in rabbits were induced by intravenous etoposide treatment during early organogenesis and that they occurred in the presence of maternal toxicity.
Abstract: The developmental effect of the topoisomerase inhibitor, etoposide, was investigated in pregnant rabbits given intravenous doses during early organogenesis. Does received 0, 0.25, 0.5, 1, or 2 mg/kg/day on days 7 through 9 of gestation. Fetal parameters were evaluated on day 28 of gestation. Live fetuses were examined for gross, visceral, and skeletal malformations and variations. In addition, telencephalon in embryos 8 h following the final treatment was examined histologically. No change in general condition was observed in any does, but a significant decrease in body weight gain during the pregnancy and enlargement of the liver resulting from marked fatty change were observed in does treated with etoposide at 2 mg/kg/day. Etoposide had neither lethal nor growth retarded effects on embryos/fetuses. However, axial skeletal malformation and extra ribs had a low incidence but were significant in the group treated with etoposide at 2 mg/kg/day, whereas no significant increases in external malformations in term fetuses nor in pyknotic cells in the ventricular zone of telencephalon in embryos were noticed in any etoposide-treated groups. It was concluded that anatomical defects (skeletal malformation or variation) in rabbits were induced by intravenous etoposide treatment during early organogenesis and that they occurred in the presence of maternal toxicity.
TL;DR: There was an increased incidence and earlier onset of liver tumours induced in F1 male offspring from F0 males treated with 1.75 g/kg bwt, but not in the female offspring, and no predisposition of F1 animals to tumours although observation times were shorter.
Abstract: In order to investigate the alleged potential of paternally administered urethane to cause foetal abnormalities and heritable tumours, male CD-1 mice were treated with urethane, either acutely by intraperitoneal injection at doses of 1.25 and 1.75 g/kg bodyweight (bwt) or sub-chronically in the drinking water at 1.25 for 10 weeks, and 3.75 mg/ml for 9 weeks or vehicle for the control groups. They were mated to untreated females 1 week later. Uterine contents of half the pregnant females were examined just before full term, while the remaining females were allowed to deliver their litters. The resulting F1 offspring were observed for approximately 18 months and 12 months for acute and sub-chronic exposures respectively and subjected to necropsy examination. Some of the mice treated acutely with 1.75 g/kg bwt exhibited partial infertility but none of those treated with 1.25 g/kg bwt had an adverse effect on their reproductive ability. There was no genetic effect of acute urethane treatment on male germ cells as indicated by dominant lethality. After birth, there was an increase (P < 0.05) in post-implantation deaths possibly due to perinatal mortality. There was an increased incidence and earlier onset of liver tumours induced in F1 male offspring from F0 males treated with 1.75 g/kg bwt, (20.7% vs. 10.1%, P = 0.026) but not in the female offspring. F1 males from both treatment groups had mean bodyweights significantly higher than controls (P < 0.01). Some males from each dose group of the acute study were examined using the restriction site mutation assay involving analysis of exon sequences. No mutations were identified in testes, liver or spleen of DNA isolated from the urethane-treated animals. No reproductive or genetic effects were seen with sub-chronic treatment at either 1.25 or 3.75 mg/ml urethane nor was there any predisposition of F1 animals to tumours although observation times were shorter.
TL;DR: The time course for the accumulation of AFB1-DNA adducts in rat liver slices incubated with AFB1 at 0.5 microM has been investigated and the relationship between adduct formation and AFB1 concentration over a wide concentration range in liver slices has been determined.
Abstract: An immunoslot blotting (ISB) method for quantitating aflatoxin B1-DNA adduct levels has been developed and used to examine the relationship between dose and hepatic aflatoxin B1-DNA adduct levels in rats fed aflatoxin B1 (AFB1) in the diet at dose levels of between 0.5 and 10 micrograms/kg/day. The method has also been used to examine the dose-response relationship for adduct formation in precision-cut rat liver slices incubated with AFB1 at concentrations between 0.01 and 2 microM. For the feeding studies, groups of male Fisher F344 rats were given AFB1 in the diet for periods of 1 to 10 weeks and hepatic DNA adduct levels determined using ISB. The time for adduct levels to reach steady-state conditions was determined in animals given approximately 10 micrograms of AFB1/kg/day and steady-state levels at lower concentrations measured. The time course for the accumulation of AFB1-DNA adducts in rat liver slices incubated with AFB1 at 0.5 microM has been investigated and the relationship between adduct formation and AFB1 concentration over a wide concentration range in liver slices has been determined.
TL;DR: The interaction between these two compounds with regard to teratogenicity in NMRI mice was investigated and the influence of HU on the 6-MPr-induced DNA modification was determined by measuring the incorporation of 6-thioguanine into the DNA of day-11 embryos.
Abstract: Hydroxyurea (HU) and 6-mercaptopurine riboside (6-MPr) are used as cytostatic chemotherapeutics. Their teratogenic potential in experimental animals has been well known for several decades. Generally, it is assumed that the toxicity of both agents is due to an interference with enzymes of DNA synthesis. In the case of 6-MPr, it was speculated that the teratogenicity in rodents might be paralleled by or even correlated to an incorporation of 6-thioguanine into the DNA of the embryos. In this study, the interaction between these two compounds with regard to teratogenicity in NMRI mice was investigated. Dose-response data of 6-MPr (s.c.-treatment) were published earlier. First, a dose-response study with HU alone (i.p.-treatment) was performed. From these data, the doses for the combination study were derived: HU 250 mg/kg (NOAEL dose) and 6-MPr 16 mg/kg (strongly teratogenic dose). In all experimental groups the substances were administered to the dams once on day 11 of gestation. Combination effects were investigated applying various dosing regimens. In group I treatment was simultaneous, in group II HU was administered 2 h before 6-MPr, in group III 2 h after 6-MPr. The differences in the overall frequency of gross structural abnormalities were moderate. However, when analysing the effects in more detail (single abnormalities), group III exhibited great differences: 1) 6-MPr co-treatment increased the frequency of HU effects (skull defects) and 2) HU co-treatment decreased the frequency of 6-MPr effects (limb defects) when compared to the findings of the dose-response studies. In addition, the influence of HU on the 6-MPr-induced DNA modification was determined by measuring the incorporation of 6-thioguanine into the DNA of day-11 embryos. As expected, the HU co-treatment corresponding to the group III dosing regimen of the teratogenicity experiment decreased the incorporation rate by ca. 40%. This was in parallel to the decrease in the frequency of 6-MPr effects in the teratogenicity III group. This finding may be considered as a further indication that in the case of 6-MPr, DNA modification is accompanying teratogenicity.
TL;DR: PHT monotherapy appears to have genotoxic effect as expressed by the induction of increased SCE rates in treated epileptics, while disease does not play any role in inducing genetic damage as shown by no difference in SCE frequencies between control subjects and PHT-untreated epileptic patients.
Abstract: Phenytoin (PHT) is a widely prescribed antiepileptic drug. Its potential to interact with genetic material was investigated in a set of 30 epileptic patients (age 10-30 years) prior to and following the administration of PHT over a period of 9 months (grouped in a multiple of 3 months) and 40 control subjects in relation to age, sex, duration of drug therapy, and plasma concentration of PHT, using the sister chromatid exchange (SCE) frequency assay. Plasma levels of the phenytoin were measured by biochemical assay in epileptic patients before and after the PHT therapy. The peripheral blood lymphocytes were cultured and harvested at 72 h. The frequency of SCE was significantly higher (P < 0.001) in both age groups (10-20 and 21-30 years) for PHT-treated epileptics compared to PHT-untreated and control subjects. However, there were no considerable variations in SCE finding between the control and PHT-untreated patients. Between the two age groups, a significantly higher SCE frequency was observed in PHT-treated patients (P < 0.01) in the older age group (21-30 years). Mean SCE frequency did not differ between the male and female in the controls, PHT-untreated, or treated epileptics. Correlation between the plasma concentration of PHT and the incidence of SCE among 30 patients was insignificant. PHT monotherapy appears to have genotoxic effect as expressed by the induction of increased SCE rates in treated epileptics, while disease does not play any role in inducing genetic damage as shown by no difference in SCE frequencies between control subjects and PHT-untreated epileptic patients.
TL;DR: Results, from the quantitative analysis of small GST-P+ foci as end point marker lesions, indicate that the liver tumor modifying potential of test chemicals can be evaluated in rats by using an initiation/promotion protocol for urinary bladder carcinogenesis.
Abstract: Male F344 and Wistar Shionogi (WS) rats were treated with 0.05% N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) for 20 weeks and then killed at week 36 (experiment 1). Although reduction of body weight increase was found, no effects on liver weights were noted. Formalin-fixed and paraffin-embedded liver tissues from rats killed terminally were cut and stained for glutathione S-transferase placental form (GST-P) immunohistochemically. Marked elevation of quantitative values of small GST-P positive (GST-P + ) foci were apparent in both strains of rat administered BBN. In experiment 2, both sexes of F344 rats were given 0.05% BBN in the drinking water for 4 weeks and then fed diet containing 0 or 5.0% sodium L-ascorbate (SA) for 32 weeks. No body and liver weight changes were evident in any group. Quantitative values for small GST-P + foci were increased in both sexes of rats exposed to BBN but were not modified by additional SA treatment. Thus, it was confirmed that the selective bladder carcinogen BBN also acts as a liver carcinogen. These results, from the quantitative analysis of small GST-P + foci as end point marker lesions, indicate that the liver tumor modifying potential of test chemicals can be evaluated in rats by using an initiation/promotion protocol for urinary bladder carcinogenesis.
TL;DR: The induction of micronuclei in polychromatic erythrocytes of bone marrow of Pzh:SWISS mice after combined treatment with X-rays and cyclophosphamide (CP) or X-ray and mitomycin C (MMC) were investigated and mutagenic effects were found.
Abstract: The induction of micronuclei in polychromatic erythrocytes of bone marrow of Pzh:SWISS mice after combined treatment with X-rays and cyclophosphamide (CP) or X-rays and mitomycin C (MMC) were investigated. Combinations of high (1.00 Gy + 100 mg/kg bw CP and 1. 00 Gy + 5.25 mg/kg bw MMC) and low (0.25 Gy + 25 mg/kg bw CP and 0. 25 Gy + 1.75 mg/kg bw MMC) doses were used. Both chemicals enhanced the mutagenic effects caused by irradiation. After combined treatment with high doses of X-rays + CP and X-rays + MMC at different sample times increases in frequency of micronuclei were observed. Mutagenic effects were found also after treatment with two low doses, when irradiation alone produced no effects. The effects of combined treatments are generally similar to the additive effect of the single treatments.
TL;DR: Both aromatic amines are genotoxic, taking into account the results obtained by the other researchers who reported mutagenic activity of 2,4-dimethylaniline in a Salmonella typhimurium assay and DNA repair test using Chinese hamster hepatocytes.
Abstract: The alkaline single cell gel electrophoresis (SCGE) or "comet" assay under alkaline conditions was used to measure DNA damage in the liver cells of B6C3F1 male mice exposed to 2,4-dimethylaniline and 2, 4,6-trimethylaniline. Cells embedded in agarose were lysed, subjected briefly to an electric field, stained with a fluorescent DNA-binding stain, and viewed using a fluorescence microscope. The effect of 2,4-dimethylaniline and 2,4,6-trimethylaniline was studied after a single intraperitoneal injections at doses equal to 100, 200 mg/kg and 150, 300 mg/kg b.w., respectively. It was found that 2, 4-dimethylaniline and 2,4,6-trimethylaniline were able to damage DNA in the liver cells of the mice. As has been published elsewhere, the DNA damaging effect of the studied compounds were also observed in bone marrow cells of the mice. In conclusion, taking into account the results mentioned above and the results obtained by other researchers who reported mutagenic activity of 2,4-dimethylaniline and 2,4,6-trimethylaniline in Salmonella typhimurium assay and in a DNA repair test using Chinese hamster hepatocytes, it can be stated that both aromatic amines are genotoxic. Teratogenesis Carcinog. Mutagen. 19:323-327, 1999.