Publications

Sugasawa K., Shimizu Y., Iwai S., Hanaoka F (Cellular Physiology Laboratory, RIKEN, The Institute of Physical and Chemical Research, 2-1 Hirosewa,Wako, Saitma 351-0198, Japan): A molecular mechanism for DNA damage recognition by the xeroderma pigmentosum group C protein complex. DNA Repair, 1(1), 2002, 95-107. [35 Ref]

The XPC-HR23B complex is involved in DNA damage recognition and the initiation of global genomic nucleotide excision repair (GG-NER). Our previous study demonstrate that XPC-HR23B recognizes and binds DNA containing a helix distortion, regardless of the presence or absence of damaged bases. Here, we describe an extended analysis of the DNA binding specificity of XPC-HR23B using various defined DNA substrates. Although XPC-HR23B showed significantly higher affinity for single-stranded DNA than double-stranded DNA, specific secondary structures of DNA, involving a single and double strand junction, were strongly preferred by the complex. This indicates that the presence of bases, which cannot form normal Waston-Crick base pairs in double stranded DNA, is a critical factor in determining the specificity of XPC-HR23B binding. A DNase I footprint analysis, using a looped DNA substrate, revealed that a single XPC-HR23B complex protected a distorted site in an asymmetrical manner, consistent with the preferred secondary structure. The specific binding of XPC-HR23B is undoubtedly an important molecular process, based on which NER machinery detects a wide variety of lesions that vary in terms of chemical structure during DNA repair.

Nouspikel T., Hanawalt PC (Department of Biological Science, Stanford University, Stanford, CA 94305-5020, USA): DNA repair in terminally differentiated cells. DNA Repair, 1(1), 2002, 59-75. [110 Ref]

Terminally differentiated cells do not replicate their genomic DNA, and could therefore dispense with the task of removing DNA damage from the non-essential bulk of their genome, as long as they are able to maintain the integrity of the genes that must be expressed. There is increasing experimental evidence that this is indeed the case, at least for some repair pathways such as nucleotide excision repair (NER). In this review, we examine a number of terminally differentiated cell systems in which it has been demonstrated that DNA repair is attenuated at the global genome level, but maintained in expressed genes. How these cells manage to repair transcribed genes in not yet fully elucidated, but there are indications that the transcription coupled repair (TCR) pathway could maintain integrity of the transcribed strand (TS) in the active genes. We have observed in neurons that the non-transcribed strand (NTS) of active genes is also well repaired, a phenomenon that we have named differentiation associated repair (DAR). It is conceivable that DAR is necessary to maintain the integrity of the template strand that is needed by TCR to complete the repair of lesions in the TS of essential expressed genes with high fidelity.

Barzilai A., Rotman G., Shiloh Y (Department of Neurobiochemistry, George S, Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv 69978, Israel): ATM deficiency and oxidative stress: a new dimension of defective response to DNA damage. DNA Repair, 1(1), 2002, 3-25. [203 Ref]

ATM is one of the sentries at the gate of genome stability. This multifunctional protein kinase orchestrates the intricate array of cellular responses to DNA double strand breaks. Absence or inactivation of ATM leads to the pleiotropic genetic disorder ataxia-telangiectasia (A-T), whose hallmarks are neuronal degeneration, immunodeficiency, genomic instability, premature aging and cancer predisposition. Several features of the complex clinical and cellular phenotype of A-T are reminiscent of other syndromes involving neurodegeneration, premature aging or genomic instability. A common denominator of many of these conditions is the perturbation of the cellular balance of reactive oxygen species, which leads to constant oxidative stress. Of these disorders, ATM deficiency is one of the most extensively studied with regard to the genome instability-oxidative stress connection. This connection may provide new insights into the phenotypes associated with genetic deficiencies of DNA damage responses, and point to new strategies to alleviate some of the their clinical symptoms.

Amer SM., Fahmy MA., Aly FAE., Farghaly AA (Department of Genetics and Cytology, National Research Center, Dokki, Cairo, Egypt): Cytogenetic studies on the effect of feeding mice with stored wheat grains treated with malathion. Mut Res, 513(1-2), 2002, 1-10. [32 Ref]

The cytogenetic effect of malathion residues in wheat grains stored for different periods of time (4, 12, 24 weeks) was evaluated in Swiss mice. The studies included: (1) chromosomal aberrations analysis in bone-marrow and spermatocyte cells: (2) chromosomal aberrations and sister chromatid exchange (SCE) analysis in spleen cell culture from mice fed with stored wheat grains. The tested doses were 8.36 (applied dose), 25.08 and 41.80 mg malathion/kg wheat grains. The results demonstrated that the cytogenetic effect induced in different mouse tissues by malathion residues was dose-dependent and increased with increasing of both feeding and storage periods. Feeding mice with wheat grains stored for 4 weeks had a non-significant effect with respect to the induction of chromosomal aberrations or SCEs. Significant chromosome damage and increase of SCEs were observed in mice fed with wheat grains stored for 12 weeks. The maximum effect was recorded in mice fed for 12 weeks with the grains treated with the highest tested dose and stored for 24 weeks. However, mitomycin C i.p. injected in mice at 1 mg/kg body weight (b.w.) (positive control) induced higher effect. The percentage of chromosome aberrations reached 13.60 +- 0.98, 13.60 +- 0.77 and 11.73 +- 0.98 (P
Cavallo D., Marinaccio A., Perniconi B., Tomao P., Pecoriello V., Moccaldi R., Iavicoli S (Department of Occupational Medicine, ISPESL-National Institute for Occupational Safety and Prevention Via Fontana Candida, 1,00040 Monteporzio Catone, Rome, Italy): Chromosomal aberrations in long-haul air crew members. Mut Res, 513(1-2), 2002, 11-15. [17 Ref]

The increasing use of air travel suggests the need for risk assessment and cytogenetic analysis of flight personnel, to check for the risk of developing cancer. Taking into consideration occupational risk and possible confounding factors, we used traditional cytogenetic, the micronucleus test and fluorescent in situ hybridization (FISH) analysis to study 48 male crew members working on long-haul flights and a control group of 48 ground staff. Compared to controls, we detected a significant increase in the relative risk of gaps and breaks (adjusted odds ratio (OR adj)-7.8; 95 percent confidence interval (CI) -2.4-24.9) and of translocations (ORadj-5.1:95percent CI 1.5-17.3) in crew members. With a non-significant difference in the other chromosomal aberrations. The possibility of a correlation between translocations and cancer risk highlights the need for preventive measures for aircraft personnel.

Syng-sai C., Baul TSB., Chatterjee A (Genetics Laboratory, Department of Zoology, North- Eastern Hill University, Shillong 793022, India): Antiproliferative and cytotoxic effect of a novel organotin compound on mammalian cells both in vitro and in vivo. Mut Res, 513(1-2), 2002, 49-59. [45 Ref]

Organotin compounds are organometallic compounds showing various toxicological properties. Several organotin compounds also showed an antineoplastic effect. However, their relative mutagenic potential is not well established. In this study Et2SNCl2 L [L = N (2-pyridylmethylene)4-toluidine] OTC) has been subjected to investigation for its cytotoxic effect in mouse bone marrow cells (BMCs) and human peripheral blood lymphocyte cells (HPBLs). The Sn-N bond in OTC is 2.46 A which is greater than 2.39 A and therefore, a better formation of tin-DNA complex can be expected. The present data indicate that OTC induced significant delay in cell kinetics and sister chromatid exchanges (SCEs) in both BMCs and HPBLs, whereas, induction of chromosome aberrations was found only in HPBLs. The presence of buthionine sulfoximine (BSO) modulated cellular sensitivity towards OTC in both cell systems. It may be inferred that the OTC could bind on DNA more easily owing to its structural advantage and this may explain the induction of DNA damage and the delay in cell proliferation. Since the cytotoxic effect of OTC is more in glutathione depleted cells, the concentrations of OTC may be reduced to get an antitumour effect in GSH-depleted cells and thus minimizes its toxic side effect.

Idaomar M., Hamss RE., Bakkail F., Mezzoug N., Zhiri A., Baudoux D., Serrano AM., Liemans V., Moraga AA (Departmento de Biologie,Faculte des Sciences, Unite de Biologie Cellulaaire et Moleculaire (BCM), Universite Abdelmalek Essaadi, BP 2121, 93002 Tetouan, Morocco): Genotoxicity and antigenotoxicity of some essential oils evaluated by wing spot test of drosophila melanogaster. Mut Res, 513(1-2), 2002, 61-68. [40 Ref]

Essential oils extracted from the medicinal plants; Helichrysum italicum, Ledum groenlandicum and Ravensara aromatica, together with their mixture were tested for their genotoxic and antigenotoxic activities against urethane, a well-known promutagen. We have adopted the somatic mutations and recombination test (SMART) in the wings of Drosophila melanogaster. Three days old larvae, trans-heterozygous for two genetic markers mwh and flr, were treated by essential oil and/or urethane. A negative control corresponding to solvent was also used. Our results do not show any significant effect of the oils tested but they reduce the mutation ratio resulting from urethane. The mixture of the three oils at equal volume seems to be the most effective. The antimutagenetic effect of these oils could be explained by the interaction of their constituents with cytochrome P-450 activation system leading to a reduction of the formation of the active metabolite. The effect could also be attributed to certain molecules that are involved in these oils.

Miloshev G., Mihaylov I., Anachkova B (Institute of Molecular Biology, Bulgarian Academy of Sciences, 1113 Sofia, Bulgaria): Application of the single cell gel electrophoresis on yeast cells. Mut Res, 513(1-2), 2002, 69-74. [15 Ref]

In the present paper, we have applied the single cell gel electrophoresis (SCGE) assay on yeast cells treating Saccharomyces cerevisiae cells with hydrogen peroxide and methyl methanesulfonate (MMS), two DNA damaging agents. In order to overcome the problem with the yeast cell wall that prevented DNA to be extended by the electric field, we disintegrated the cell wall after embedding the cells in agarose. A characteristic picture of comets with residual nuclei and tails was observed and the length of the comet tails was dependent on the concentration of the damaging agents. Yeast cells developed comets at concentrations at least 10 times lower than the concentrations at which comets begin to appear in mammalian cells after treatement with the two genotoxic agents. The higher sensitivity of the yeast comet assay and the fact that S. cerevisiae is one of the most thoroughly studied and easy to work with eudarotic model system suggest that the proposed method could be an useful tool for investigation of the DNA damaging activity of potential genotoxins. Amanuma K., Tone S., Saito H., Shigeoka T., Aoki Y (Division of Environmental Health Science, National Institute for Environmental Studies, Onogawa, Tsukuba 305-0053, Japan): Mutational spectra of benzo[a]pyrene and MeIQx in rpsL transgenic zebrafish embryos. Mut Res, 513(1-2), 2002, 83-92. [36 Ref]

To evaluate the rpsL transgenic zebrafish (Brachydanio rerio) mutation assay. we treated the embryos with benzo[a]pyrene {B[a]P} (10ug/ml) or 2-amino 3,8-dimethylimidazol[4,5-flquinoxaline (MeIQx) (300 ug/ml) for 16h and determined the mutation spectra. These treatments were previously reported to induce mutant frequencies that were 4.3 and 2.4 times the control value, respectively. In the B[a]P-treated group, half of the mutations were single base substitutions, 74 percent of which occurred at G:C base pairs. Among G:C base pair substitutions, G:C to T:A and G:C to C:G transversions were predominant, suggesting that B[a]P induced mutations in zebrafish embryos by mechanisms previously described in mammalians tissues. In the MeIQx treated group, about 60 percent of the mutations were deletions. Some specific mutations were found, but the compound primarily amplified the background mutation level; improvement in the conditions of treatment may be required for elucidating MeIQx mutagenesis in this system. This study showed that transgenic zebrafish may be a useful tool for detecting mutagens in aquatic environments and for elucidating mutagenic mechanisms. Kumar TR., Doreswamy K., Shrilatha B., Muralidhara (Department of Biochemistry and Nutrition,Central Food Technological Research Institute, Mysore 570013, Karanataka, India): Oxidative stress associated DNA damage in testis of mice: induction of abnormal sperms and effects on fertility. Mut Res, 513(1-2), 2002, 103-111. [43 Ref] Our previous work has shown that prooxidant treatment has the propensity to induce male-mediated dominant lethal (DL) type mutations in mice. The present investigation is aimed to understand the effect of oxidative stress (OS) on DNA damage in testis, epididymal sperms and its propensity to induce sperm head abnormalities as well as its implications on male fertility in mice. Initially, employing two organic hydroperoxides, (t-butyl hydroperoxide, t-bHP and cumene hydroperoxide, cHP)as model prooxidants, induction of oxidative stress was ascertained following single/multiple sublethal doses. Further, the multiple exposure model was utilized to characterize effects on testicular weights, histoarchitecture, caudal sperm counts, lipid peroxidation, DNA damage and frequency of abnormal sperms. Single sublethal doses (1/20, 1/10 and 1/5 LD50) of t-bHP and cHP administered (i.p.) to adult mice resulted in only a marginal increase (20 percent at the highest dosage) in testicular MDA levels. However, multiple doses (1/10 and 1/5 LD50 per day for 5 days) induced marked OS in testis and epididymal sperms as evidenced by a marked increase in lipid peroxidation at 24 h after the last dose. This was associated with significant increase in the DNA damage (FADU assay) in the testicular tissue. While caudal sperm counts determined at all sampling weeks showed no treatment related alterations, analysis for head abnormalities revealed nearly 2-3 fold increase in the percent abnormal sperms among the hydroperoxide treated mice during the first 3 weeks. Furthermore, mating of prooxidant treated males sequentially for a period of 5 weeks with untreated females resulted in a significant reduction in average pup number per litter during the first 3 weeks. These results suggest that oxidative stress in testicular milieu is associated with DNA damage and produces higher frequency of abnormal sperms with significant effect on male fertility.

Win W., Cao Z., Peng X., Trush MA., Li Y (Department of Pharmaceutical Sciences, College of Pharmacy and Allied Health Professions, St. Albert Hall, St. John's University, 8000 Utopia Parkway. Jamaica, NY 11439, USA): Different effects of genistein and resveratrol on oxidative DNA damage in vitro. Mut Res, 513(1-2), 2002, 113-120. [32 Ref]

Previous studies have demonstrated that phenolic compounds, including genistein (4',5,7-trihydroxysoflavone) and resveratrol (3,4',5-trihydroxystilbene),are able to protect against carcinogenesis in animal models. This study was undertaken to examine the ability of genistein and resveratrol to inhibit reactive oxygen species (ROS)-meditated strand breaks in oX-174 plasmid DNA, H2O2/Cu(II) and hydroquinone/Cu(II) were used to cause oxidative DNA strand breaks in the plasmid DNA. We demonstrated that the presence of genistein at micromolar concentrations resulted in a marked inhibition of DNA strand breaks induced by either H2O2/Cu(II) or hydroquinone/Cu(II). Genistein neither affected the Cu(II)/Cu(I) redox cycle nor reacted with H2O2 suggest that genistein may directly scavenge the ROS that participate in the induction of DNA strand breaks. In contrast to the inhibitory effects of genistein, the presence of resveratrol at similar concentrations led to increased DNA strand breaks induced by H2O2/Cu(II). Further studies showed that in the presence of Cu(II), resveratrol, but not genistein was able to cause DNA strand breaks. Moreover, both Cut(II)Cut(I) redox cycle and H2O2 were shown to be critically involved in resveratrol/copper-mediated DNA strand breaks. The above results indicate that despite their similar in vivo anticarcinogenic effects, genistein and resveratrol appear to exert different effects on oxidative DNA damage in vitro.

Kreja L., seidel HJ (Institute of Occupational, Social and Environmental Medicine, University of Ulm, Frauensteige 10, D-89075 Ulm, Germany): Evaluation of the genotoxic potential of some microbial volatile organic compounds (MVOC) with the comet assay, the micronucleus assay and the HPRT gene mutation assay. Mut Res, 513(1-2), 2002, 143-150. [29 Ref]

Microbial volatile organic compounds (MVOC), metabolites of fungi detected in indoor moulds and in working places in compost facilities are considered as a potential health hazard. Their toxicological relevance, however, is largely unknown and data are rare. The aim of this study was to evaluate in vitro the genotoxic, clastogenic and mutagenic potential of same typical MVOC. For the study of DNA damage human lung carcinoma epithelial A549 cells. V79 Chines hamster fibroblasts and human peripheral blood cells were exposed and subjected to the alkaline comet assay (single cell test). Taking the Chinese hamster V79 cell line as a target clastogenic effects were studied by the micronucleus test and mutagenic effects by the hypoxanthine-guanine phosphoribosyl transferase gene mutation test (HPRT test). The cytogenic effects of MVOC were assessed by a clonogenic assay using the A549 cell line. The alkylating agent methyl methanesulfonate (MMS) was taken as a positive control. The results indicate that MVOC induced DNA damage is only seen in conditions in which also cytotoxic effects are observed. Clastogenic and mutagenic effects could not be detected.

Purschke M., Jacobi H., Witte I (Carlvon Ossietzky Universitat Oldenburg, FB Biology and ICBM, Postfach 2503, D-26111 Oldenburg, Germany): Differences in genotoxicity of H2O2 and tetrachlorohydroquinone in human fibroblasts. Mut Res, 513(1-2), 2002, 159-167. [26 Ref]

During autoxidation of the pentachlorophenol (PCP0 metabolite tetrachlorohydroquinone (TCHQ) the semiquinone is formed as well as reactive oxygen species (ROS). It was examined if OH or the semiquinone are the cause of TCHQ induced genotoxicity by direct comparison of TCHQ and H2O2 induced DNA damage in human cells. All endpoints tested (DNA damage, DNA repair, and mutagenicity) revealed a greater genotoxicity potential for TCHQ than for H2O2. In the comet assay, TCHQ induced DNA damage at lower concentrations than H2O2. The damaging rate by TCHQ (tail moment (tm)/concentration) was 10-fold greater than by H2O2 DNA repair was lower for TCHQ than for H2O0 treatment. This was shown by measuring DNA repair in the unscheduled DNA synthesis (UDS) assay and the persistence of the DNA damage in the comet assay. In contrast to H2O0, TCHQ in non-toxic concentrations was mutagenic in the hypoxanthine-guanine phosphoribosyltranferase (HPRT) locus of V79 cells. Finally, there were also differences observed in cytotoxicity (3-(4,5-dimethylthiazol-2-y) 2,5-diphenyl tetrazolium bromide (MTT) assay) of TCHQ and H2O2, Whereas the TCHQ cytotoxicity was enhanced during a 21 h recovery phase, the H2O2 cytotoxicity did not change. The results demonstrated that the pronounced genotoxic properties of TCHQ in human cells were not caused by OH radical but more likely by the tetrachlorosemiquinone (TCSQ) radical.

Dreeier J., Breitmaier EB., Gocke E., Apfel CM., Page MGP (Department of Biological Technologies, F. Hoffmann-La Roche Ltd. Pharma Division, CH-4002 Basel, Switzerland): Direct influence of S9 liver homogenate on fluorescence signals: impact on practical applications in a bacterial genotoxicity assay. Mut Res, 513(1-2), 2002, 169-182. [20 Ref]

Assays based on the bacterial SOS-response offer the possibility of automatization of genotoxicity testing for screening of large compound libraries. While existing assays use colorimetric detection or luminescence read-out, we describe here the use of a fluorescence-based system to achieve high sensitivity of detection required for assay miniaturization. Three commonly used fluorophores -fluorescein, DDAO and resorufin-are evaluated. Experimental evidence is given that S9 liver homogenate contains a heat-labile, reversible fluorophore-binding activity and therefore, significantly reduces fluorescence intensities. We have worked out simple solutions to overcome the S9 related interference in order to be able to establish a robust bacterial genotoxicity assay.

Noda Y., Suzuki T., Kohara A., Hasegawa A., Yotsuyanagi T., Hayashi M., Sofuni T., Yamanaka K., Okada S (Department of Biochemical Toxicology, Nihon University College of Pharmacy, 7-7-1 Narashinodai, Funabashi-shi 274-8555, Japan): In vivo genotoxicity evaluation of dimethylarsinic acid in Muta TM mouse. Mut Res, 513(1-2), 2002, 205-212. [23 Ref]

Dimethylarsinic acid (DMA) induces DNA damage in the lung by formation of various peroxyl radical speceis. The present study was conducted to evaluate whether arsenite or its metabolite, DMA, could initiate carcinogenesis via mutagenic DNA lesions in vivo that can be submitted to oxidative damage. A transgenic mouse model, Muta TM Mouse, was used in this study and mutations in the LacZ transgene and in the endogenous cII gene were assessed. When DMA was intraperitoneally injected into Muta TM Mice at a dose of 10.6 mg/kg per day for 5 consecutive days, it caused only a weak increase in the mutant frequency (MF) of the lacZ gene in the lung, which was at most 1.3 fold higher than in the untreated control animals. DMA did not appreciably raise the MF in the bladder or bone marrow. Further analysis of the cll gene in the lung, the organ in which DMA induced the DNA damage , revealed only a marginal increase in the MF. Following DMA administration, no change in the cII mutation spectra was observed, except for a slight increase in the G:C to T:A transversion. Administration of arsenic trioxide (arsenite) at a dose of 7.6 mg/kg per day did not result in any increase in the MF of the lacZ gene in the lung, kidney, bone marrow, or bladder. Micronucleus formation was also evaluated in peripheral blood reticulocytes (RETs). The assay for micronuclei gave marginally positive results with arsenite, but not with DMA. These results suggest that the mutagenicity of DMA and arsenite might be too low to be detected in the Muta TM Mouse.

Wiseman BS., Werb Z (Department of Anatomy, University of California, 513 Parmassus Avenue, San Francisco, CA 94143, USA): Stromal effects on mammary gland development and breast cancer. Science, 296(5570), 2002, 1045-1049. [43 Ref][Rev]

Breast cancer manifests itself in the mammary epithelium, yet there is a growing recognition that mammary stromal cells also play an important role in tumorigenesis. During its developmental cycle, the mammary gland displays many of the properties associated with breast cancer, and many of the stromal factors necessary for mammary development also promote or protect against breast cancer. Here we review our present knowledge of the specific factors and cell types that contribute to epithelial stromal crosstalk during mammary development. To find cures for diseases like breast cancer that rely on epithelial-stromal crosstalk, we must understand how these different cell types communicate with each other.

Taylor JP., Hardy J., Fischbeck KH (Neurogenetics Branch, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892, USA): Toxic proteins in neurodegenerative disease. Science, 296(5575), 2002, 1991-1995. [75 Ref][Rev]

A broad range of neurodegenerative disorders is characterized by neuronal damage that may be caused by toxic, aggregation prone proteins. As genes are identified for these disorders and cell culture and animal models are developed, it has become clear that a major effect of mutations in these genes is the abnormal processing and accumulation of misfolded protein in neuronal inclusions and plaques. Increased understanding of the cellular mechanisms for disposal of abnormal proteins and of the effects of toxic protein accumulation on neuronal survival may allow the development of rational, effective treatment for these disorders.

Harstad EB., Klaassen CD (Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas 66160-7140, USA): Gadolinium chloride pretreatment prevents cadmium chloride induced liver damage in both wild-types and MT-Null Mice. Toxicol Appl Pharmacol, 180(3), 2002, 178-185. [28 Ref]

The heavy metal cadmium (Cd) causes hepatotoxicity upon acute administration. Kupffer cells, the resident macrophages of the liver, have been suggested to play a role in Cd-induced hepatotoxicity. Gadolinium chloride (GdCl3) may prevent Cd-induced hepatotoxicity by suppressing Kupffer cells. However, GdCl3 also induces the Cd-binding protein, metallothionein (MT). Therefore, this study was conducted to determine whether GdCl3 prevents Cd-induced hepatotoxicity via the induction of MT. Hepatic MT and Kupffer cell counts were analyzed 24 h after wild type (WT) mice were administered saline or 10, 30 or 60 mg GdCl3/kg. GdCl3 induced MT in a dose-dependent manner without affecting nonprotein sulfhydryl content. All examined doses of GdCl3 were effective at eliminating Kupffer cells from the liver. To examine the hepatoprotective effects of GDCl3 WT and MT null mice were pretreated with saline or 10, 30, or 60 mg GdCl3 24 h prior to a hepatotoxic dose of Cd (2.5 mg Cd/kg). Blood and livers were removed 16 h later and analyzed for hepatotoxicity as well as MT, Cd, and Kupffer cell content. Hepatotoxicity was alleviated in both WT and MT-null mice that were pretreated with 30 or 60 mg GdCl3/kg , indicating that MT induction is not required for the hepatoprotective effects of GdCl3. Hepatic Cd content was not decreased by GdCl3 , demonstrating that GdCl3 does not negatively affect Cd distribution to the liver. Kupffer cells were depleted at all three doses of GdCl3, whereas hepatoprotection was only observed at doses of 30 and 60 mg GdCl3/kg. This does not rule out Kupffer cells in the mechanism of Cd-induced hepatotoxicity, but it does suggest that GdCl3 exerts hepatoprotective effects on the liver aside from depleting Kupffer cells. In summary, these data substantially rule out MT induction and decrease the importance of Kupffer cells as mechanisms of GdCl3-induced protection from Cd-induced hepatotoxicity.

Copple BL., Wooley B., Banes A., Ganey PE., Roth RA (Department of Pharmacology and Toxicology Michigan State University, East Lansing, Michigan 48824, USA): Anticoagulants prevent monocrotaline-induced hepatic parenchymal cell injury but not endothelial cell injury in the rat. Toxicol Appl Pharmacol, 180(3), 2002, 186-196. [41 Ref]

Monocrotaline (MCT) is a pyrrolizidine alkaloid plant toxin that produces hepatotoxicity in humans and animals. Human exposure to MCT occurs through consumption of contaminated grains and herbal medicines. Administration of MCT to rats stimulates activation of the coagulation system and fibrin deposition in the liver. Fibrin deposition occurs simultaneously with endothelial cell damage and prior to hepatic parenchymal cell injury. Accordingly, the hypothesis that activation of the coagulation system is required for MCT induced liver injury was tested. Treatment of rats with either heparin or warfarin significantly reduced MCT induced activation of the coagulation system and the increase in alanine aminotransferase activity in the plasma, a biomarker of hepatic parenchymal cell injury. Histopathological examination of liver sections revealed that heparin decreased parenchymal cell necrosis but did not affect central venular endothelial cell damage, congestion and dilation of the sinusoids, or hemorrhage in the liver. Morphometric analysis revealed that 28 percent of the areas of livers from MCT-treated rats contained regions of coagulative necrosis, whereas less than 5 percent of the area of livers from rats treated with MCT and hepatin contained these regions. By contrast, neither heparin nor warfarin prevented MCT-induced damage to endothelial cells in the liver as estimated by increased plasma hyaluronic acid concentration. These results suggest that activation of the coagulation system is required for MCT induced parenchymal cell injury but not endothelial cell injury in the liver.

Liu BH., Yu FY., Chan MH., Yang YL (Department of Life Science, Chung Shan Medical University, No. 110, Sec.1, Chien-Kuo N. Road, Taichung, Taiwan, 402, Taiwan): The effects of mycotoxins, fumonisin B1 and aflatoxin B1, on primary swine alveolar macrophages. Toxicol Appl Pharmacol, 180(3), 2002, 197-204. [61 Ref]

Mycotoxins were fungal metabolites that were widely present in feed and food; some of them were known to associate with human and animal disease. In the present study, the effects of fumonisin B1 (FmB1) and aflatoxin B1 (AFB1) on swine alveolar macrophages (AM) were examined by exposing primary cultures of swine AM to various concentrations of mycotoxins. Incubation of AM with 5 ug/ml of FmB1 for 72 h led to a reduction in the number of viable cells to 65 percent of the control levels. In the presence of 1.5 ug/ml of AFB1 the viability of AM falls to less than 41 percent of controls after 24 h exposure. FmB1, but not AFB1, induced the apoptosis of swine with evidence of DNA laddering and nuclear fragmentation However, both FmB1 and AFB1 exposure induced the expression of apoptosis-related heat shock protein 72 (HSP72) in AM. Swine AM treated with 50 ng/ml of FmB1 and 100 ng/ml of AFB1 for 24h led to a reduction in phagocytic ability to approximately 55 and 36 percent of the control levels, respectively. Incubation of AM with FmB1 (2 and 10 ug/ml) for 24 h dramatically decreased the mRNA levels of interleukin-1B (IL-1B) and tumor necrosis factor-a TNF-a). However, AFB1 treatment did not affect the expression of IL-1B and TNF-a mRNA. The results suggest that both FmB1 and AFB1 are immunotoxic to swine AM but that they exert their toxic effects via different biochemical mechanisms.

Mobley JA., Brueggemeier RW (Division of Medicinal Chemistry and Pharmacognosy College of Pharmacy, The Ohio State University Columbus, Ohio 43210, USA): Increasing the DNA damage threshold in breast cancer cells. Toxicol Appl Pharmacol, 180(3), 2002, 219-226. [45 Ref]

The biochemical role of estrogens in the development of estrogen-dependent breast cancer remains to be elucidated, and the involvement of estrogens in tumor initiation remains controversial. Reports regarding estrogen-mediated DNA damage include the induction of 8-oxo-2' deoxyguanosine (8-oxo-dG) in vitro and in vivo, indicating a role for oxidative stress in tumor initiation and/or progression. However, DNA isolation, cellular DNA repair, and high antioxidant status have made the measurement of 8-oxo-dG in vivo and in cell culture somewhat challenging. In this regard, a potentiation in DNA damage can be achieved by depleting cellular stores of glutathione. We chose to deplete glutathione in the estrogen receptor (ER)-positive MCF-7 breast cancer cell line with a y-glutamylcysteine transpeptidase enzyme inhibitor buthionine sulphoximine (BSO) for the purpose of studying estrogen-induced DNA damage. Treatment of GSH-depleted MCF-7 cells with 10 uM 2-OH-E2 or 4-OH-E2 for 30 min resulted in a statistically significant increase in 8-oxo-dG/10 5 dG of 127 and 160 percent, respectively. A potentiation in catechol estrogen-induced DNA damage was observed with the addition of copper (II) chloride for both 2-OH-E2 and 4-OH-E2 by 165 and 200 percent, respectively In addition, 100 nM and 1.0 uM estradiol increased DNA damage in a dose response-like fashion by 145 and 189 percent, respectively. The depletion of GSH by BSO may prove to be an advantageous technique for the study of DNA damage in cells otherwise resistant to oxidative stress and/or alkylating agents and has proven useful in the study of estrogen-induced oxidative DNA damage in a highly reproducible and sensitive manner.

Igisu H., Matsuoka M (Institute of Industrial Ecological Sciences, University of Occupational and Environmental Health, 1-1, Yahata-Nishi-Ku, Kitakyushu 807-8555, Japan): Acrylamide encephalopathy. J Occup Hlth, 44(2), 2002, 63-68. [21 Ref][Rev]

Although the toxic effects of acrylamide on the peripheral nervous system have been well established, less attention has been paid to its effects on the brain. Nevertheless, (1) ultrahigh field magnetic resonance imaging revealed enlarged cerebral ventricles and cisterns in rats treated with acrylamide 50 mg/kg/d for 8 d, (2) creatine kinase (CK) activities were suppressed in the brain from such rats and mice, (3) CK activities in the brains of mice intoxicated with acrylamine changed in parallel with neurological dysfunction, (4) in human cases of acrylamide intoxication due to occupational exposure, symptoms suggesting brain involvement were seen in a few, and (5) cases of acrylamide intoxication due to contaminated well water showed unquestionable signs of brain dysfunction including mental confusion with hallucinations. These indicate that acrylamine can cause encephalopathy in animals as well as in humans, and that the spectrum of impairment of the nervous system can vary greatly depending on the mode of exposure.

Ito T., Yoshitome K., Horike T., Kira S (Faculty of Education, Okayama University, 3-1-1 Tsushima-naka, Okayama City, 700-8530, Japan): Distribution of inhaled m-xylene in rat brain and its effect on GABA a receptor binding. J Occup Hlth, 44(2), 2002, 69-75. [22 Ref]

Organic solvents generally depress the central nervous system (CNS), similarly to volatile anesthetics. The precise mechanism of their action on the CNS, however, is not fully understood, and remains to be clarified. This study is focused on how inhaled m-xylene distributes in the brain, and whether region specific change in GABA a receptor binding takes place due to the exposure. To conduct this study , we first developed a simple exposure system suitable for inhalation experiments with small animals. Using this system, six-week-old male Sprague-Dawley rats were exposed to m-xylene vapor (2000 ppm) 4 h/d, for 5 consecutive days. At the end of the exposure, m-xylene levels in four different regions of the brain were measured by head-space gas chromatography. Also 14 um-thick frozen sections of the brain were made, and [35S] t-butylbicyclophosphorothionate (TBPS) binding autoradiography was performed. Uneven distribution of m-xylene in the brain was observed. The concentration in the cerebellum (976+-93.4 ug/g tissue) was the highest, while that in the cerebral cortex (467 +-43.6 ug/g tissue) was the lowest. [35S] TBPS binding was significantly greater in the molecular layer of the cerebellum (control: 12.6+-0.64, m-xylene 16.0+-1.34 fmol/mg tissue). These results suggest that m-xylene is distributed unevenly in the rat brain, and acute exposure to m-xylene at a high concentration alters [35S]TBPS binding, which may reflect changes in GABA a receptor characteristics.

Shin YC., Yi GY., Kim TK., Sohn NS., Park J., Chung HK., Yoo C., Lee JH., Kim Y* (Department of Occupational and Environmental Medicine, Ulsan University Hospital, 290-3 Cheonha-Dong, Dong-Ku, Ulsan 682-060, Korea): Evaluation of exposure to 2,2-dichloro-1,1,1-trifluoroethane (HCFC-123) in air-conditioner manufacturing workers and their health effects in South Korea. J Occup Hlth, 44(2), 2002, 108-111. [11 Ref]

Hepatic dysfunction in workers exposed to HCFC-123 has recently been reported, but information on the level of exposure to the material and the related hepatotoxicity in workers handling the material is very limited. This study evaluated the exposure status in some Korean workers handling the chemical and whether they have liver dysfunction. We investigated all four factories manufacturing large-sized industrial air-conditioners operating in South Korea in 1998. The range and geometric mean of the estimated 8-hour TWA concentrations at two factories were 2.2 ppm (0.89-4.2 ppm) and 32.5 ppm (4.9-113.9 ppm), respectively. It was estimated that the HCFC-123 concentrations at the other two factories were lower than those at the former two. Liver function was within the normal range in all the exposed workers. The present study did not show any health effect of HCFC-123 on workers. Worker exposure in this study was intermittent and lower than in studies which reported cases of liver dysfunction, so that the level and pattern of exposure to HCFC-123 would be very important as a cause of liver dysfunction. Our study showed that liver dysfunction did not occur at an exposure concentration less than 32.5 ppm as an 8-hour TWA of HCFC-123.

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