P XIII.99 Comparative evaluation of the estrogenic and toxicological potential of environmental compounds with estrogenic activity

S116

S-X1I1: Identification and evaluation of environmental mutagens, ecogenotoxicology

Ip XIII.9SI

Strong genotoxlc activity of fraction PM 10 and especially of fraction PM 1.5 of airborne particulates on human trachen-bronchlal epithelial cells (lIne BEAS-

lB) In vitro Claudia Hornberg, Lolita Maciuleviciute, N.H. Seemayer, Eva Kainka. Med· ical Institute 0/ Enuironmental Hygiene 0/ the University. Our/ittstrape 53. D-40223 Dusseldorf; Germany Airborne particulates containing more than SOO mostly organic substances pose a health risk to humans by inhalation. Recent epidemiological studies indicate an association between fine particulate pollution and excess mortality. Airborne particulates were collected in the city of DUsseldorf with a low volume Dichotomous sampler (Graseby-Anderson) obtaining the coarse fraction PM 10 (10 um) and fine fraction PM 2.S (2.S 11m). Glass fiber filters were extracted with dichloromethane and chemical substances transferred quantitatively to dimethyl sulfoxide. As a sensitive bioassay for detection of genotoxic activity of extracts induction of "sister chromatic exchanges" was evaluated dose-dependently. As target cells we utilized the human bronchial epithelial cells of hne BEAS-2B. The fraction PM 10 and especially the fraction PM 2.5 exerted strong genotoxic activity disclosed by induction of sister chromatid exchanges corresponding to extremely low volumes of collection from 0.24 cbm of air and less. We must consider a permanent body burden and health risk to humans by inhalation of fraction PM 10 and especially offraction PM 2.S. Keyword(s): airborne particulates; PM 10; PM 2.S; human bronchial epithehal cells; sister chromatid exchanges

Ip XIII.961

Genotoxlclty evaluation of Ihe 9C trlmethylbenzenes (TMB)

Ewa Janik-Spiechowicz, Kalina WyszyJIska, Eli:bieta Dziubaltowska. 10M.

Institut Nofer; P.O. Box 199. 90-950 LOdt. Poland We have studied genotoxic activity oftbree positional isomers: hemimellitene (1,2,3 • TMB; CAS No. S26-73-8), pseudocumene (1,2,4 - TMB; CAS No. 9S-63-6) and mesitylene (I ,3,S - TMB; CAS No. 108-67-8). They an: solvents widely used as components of industrial and commercial dyes and varnishes. The following were analysed: reverse mutation in S. typhimurium tester strains and incidence of micronuclei and sister chromatid exchange in bone marrow of Imp:Balb/c mice. The direct mutagenic activities showed 1,2,3 - TMB in bacterial cells. It was higher for the frameshift-mutated strains (TA97a, TA98) than for the base-pair substitution-mutated strains (TAIOO, TAI02). The compounds did not cause micronuclei induction in bone marrow of mice. Significant responses in SCE induction, compared with the level of those changes in the controls were observed after intrapentoneal administration of all three compounds: 1,2,3 - TMB 730, 1470 and 2200 mg/kg; 1,2,4 - TMB 900, 1800 and 2700 mg/kg and 1,3,S - TMB 1800 and 2700 mg/kg. These data gave a limited evidence for genetic activity of 1,2,3 -TMB and no evidence for genetic activity 1,2,4 - TMB and 1,3,S - TMB. Keyword(s): 9C Trimethylbenzenes; Genotoxicity

Ip XIII.971

Analysis of micronuclei In erythrocytes of rals treated with TCDD In a liver tumor promotion study

Jorma Mliki-Paakkanen, Raimo Pohjanvirta, Jouni Tuomisto, Mikko Unkila, Matti Viluksela, Jouko Tuomisto. National Public Health Institute. Kuopio.

FIN.70701. Finland The frequencies of micronucleated erythrocytes (MNEs) were analysed in 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) -treated HanlWistar (Kuopio; HIW) and Long-Evans (TurkulAB; L·E) rats. The strains differ by a factor of > 1000 in their sensitivities to the acute toxicity of TCDD, the former being resistant Initiated (partially hepatectomized and treated with nitrosodiethylamine) and non-initiated (neither hepatectomized nor treated with

NDEA) animals were treated s.c. with TCDD once a week for 20 weeks. The total doses were 0, 1.7, 17, or 170 J.lg/kg for HIW rats and 0, 0.17, I. 7 or 17 J.lg/kg for L-E rats. The frequencies of polychromatic erythrocytes with micronuclei (MNPCEs) were slightly increased in the bone marrow of initiated and non-initiated HIW rats treated with 170 fLglkg of TCDD. Micronucleated erythrocytes were more frequent, compared with controls, also in peripheral blood of initiated HIW rats dosed with 170 J.lg/kg of TCDD. A very slight, nonsigmficant increase in micronucleated immature erythrocytes was observed in the bone marrow of initiated L-E rats of the 17 ug/kg dose group, and in peripheral blood of non-initiated L-E rats treated with 17 J.lg/kg of TCDD. The highest TCDD dose increased the frequency of immature red blood cells in both strains. In L-E rats, the increase was obvious only in peripheral blood. Keyword(s): TCDD; Promotion study; Erythrocyte micronuclei

Ip XIII.981

DNA adduct detection In the Uver of the dragonnet Call1onymus lyra and the flounder Plalchlhys flesus: A potential blolndlcator of pollutant

K. Boillot l , T. Burgeor', J.P. Arnould", A. Pfohl-Leszkowiez", I Ecole Nationale superieure agronomique: Lab. Toxicologie et Securite Alimentaire, 145 avenue de Muret. F·31076 Toulouse. France; 2 Laboratoirede Toxicolo: gie, FacuJte de Pharmacie F-80D037 Amiens, France; J Lab. Ecotoxicologie marine. IFREMER. F-44D37 Nantes, France Enzyme linked immunosorhent assay (ELISA) and 32 P-postlabelling were used to investigate DNA adduct formation in two sentinel species in field study. The dragonnet Callionymus lyra and the flounder platchthys flesus were trawled respectively in the Seme bay and estuary. Chemicals compounds analyses in this area showed a great contamination with PCB and PAH. Differents DNA·adduct patterns were observed in the two different species collected in the five different areas located in the bay and the estuary. In the bay, total DNA adduct level is higher after butanol extraction (2 to 30 fold) compared to nuclease PI enrichment and could reach ISO adducts per 10 9 nucleotides. Cross reactivity with Benzo diol epoxide DNA adduct shows contamination of the bay station by benzo(a)pyrene. Very few adducts are detected in the station near the mouth of Seine. In the estuary, other pollutants than PAH have induced DNA-adducts. All these differences indicate that the pollutants an: not similar in the estuary and in the mouth of the river. These results suggest that detection of DNA adduct could be applied as a bioindicator of exposition to pollutant in field study. Keyword(s): DNA adduct; bioindicator; pollution

Ip XIII.991

Comparallve evaluation of the estrogenic and tOXIcological potential of environmental compounds with eslrogenlc acllvlty

Manfred Schlager", Franz Krassnig!f, Peter Maria Eckl l . /Instituteo/Genet_ ics and General Biology. University 0/ Salzburg, Hellbrunnerstr: 34. A-5020 Salzburg, Austria; 2 FederalE1Wironment Agency, Innsbrucke»Bundesstra,Pe 47. A-5020 Salzburg, Austria Reports of abnormal sexual development of fish species, the findings indicat_ ing a reduction of male fertility during the last decades together with results implicating the possible relation to an increased breast and prostate cancer rate, have focused common interest on compounds exhibiting estrogenic properties. The interest increased significantly when synergistic activation of estrogen receptors with combinations of such environmental chemicals was reported. This synergistic interaction of different compounds may have profound environmental implications. To assess the potency of such combinations for estrogenic activity we use a technique based on rainbow trout hepatocytes incubated in a medium free of growth factors or estrogenic chemicals for the production of vitellogenin (VTG) as a marker for the estrogenic potency. VTG levels were detected by an Elisa-technique, Furthermore the cyto- and genotoxic as well as mitogenic potential of the compounds with

S-XIII: Identification and evaluation of environmental mutagens, ecogenotoxicology estrogenic activity is determined in primary rat hepatocytes. We are currently evaluating the toxicological potential of selected chemicals (4-nonylphenol, a-endosulfan, chlordane, DDT, atrazine, dieldrin and tamox ifen). Prehminary results indicate a trend to dose-dependent increases of DNA synthesis and cell div ision. Moreover we show that the highest levels of VTG production were induced by DDT. In contrast to previous results no additive effect of combinations such as a-endosulfan + chlordane, dieldrin + chlordane, atrazine and dieldrin, a-endosulfan + atrazine, which mimic estrogenic effects, have been observed.

S 117

IChEl and gives reason to suggest that these children have an increased cancer risk. Using FISH technique with whole-chromosome specific DNa probes the stable ChA 's were studied in various gro ups of exposed people . Their incidence was increased in these people as compared to the control ones of the same ages . The results obtained are in agreement with the hterature data available and show an obvious late cytogenetic effects of Chernobyl accident. They seem to be useful for identification of groups with increased health risk and for retrospective biodosimetry. Keyword(s) : Chromosomal damages; Chernobyl

Keyword(s): estrogenic activity; vitellogenin synthesis; rat and fish hepatocytes

Ip XIII.1021 Ip XIII.tOOI

DNA damage and repair capacity In blood cells of people exposed to lead In battery factories

Evaluation of mutagenic potential of air pollutJon edslln!: during wood conversion operations

Ka lma Wyszynska, Ewa Janik-Spiechowicz, Wieslawa Chwialkowslca-Liro.

10M. LOdi. Poland Helena Groot de Restrepo", Diana Sicard', Maria Mercedes Torres', Camilo Uribe 2 • I Laboratorio de Genetica Humana, Uniuersidad de los Andes.

Colombia; JClinica de Toxicologia, Santa Fe de Bogota. Colombia In order to investigate the presence of genotoxic effects in blood cell samples, a human population exposed to lead in battery industries was studied. Single strand breaks and the interference with DNA repair processes, after an X-ray exposure of 300 cGy, were analized using the Comet Assay. Cells proliferation kinetics (Replication Index, RI) and Sister Chromatid Exchanges (SCE) were also studied in a total of 56 people. The workers were classified in four groups according to the blood levels of lead (2186 ug/dl) . Thirteen people not occupationally exposed to this metal were selected as the group of the lowest level., No significant differences were found in the DNA damage previous to the irradiation. A significant difference in DNA damage (199 .8 +1- 82.0 um and 322 .7 +1- 45.7 um) was found between the cells of the people with lowest and the highest blood lead levels respectively after the irradiation. Within the various groups, no differences were found in DNA migration after 120 minutes of repair. The lymphocyte cultures were harvested at 72 hours , no significant differences were found for the replication index. No differences were found in the frequency of sister chromatid exchanges. The results suggest that the capacity of DNA repair in cells of people exposed to lead is not altered. Nevertheless, in combination with x-rays an interaction with lead was observed in the increase of DNA single strand breaks measured immediately after the irradiation.

Ip XIII.10ll

Late eytegenetle effects of Chernobyl accident In humans

I. Vorobtsova. Central Reseaah Institute of Roentgenology and Radiology. St. Petersburg. 189646, Russia Several cohorts of people suffered from Chernoby! accident were studied cytogenetically: "liquidators" Igroup U, children of liquidators IChU, children evacuated from contaminated areas IChEl, as well two control groups:adults IACI and children IChC/. The following cytogenetic endpoints were estirnated:unstable and stable chromosome aberrations ICW in lymphocytes, micronuclei IMNI in lymphocytes and erytroeytes, in vitro chromosomal radisensitivity of lymphocytes Idose -1 .5 Gyl . The frequency of unstable ChA was found to be higher in I., ChI. and CbE as compared to the groups AC and Che correspondingly - mainly due to increased number of single fragments . It gives reason to suggest that spontaneous mutagenesis is elevated in previously exposed people. In I. and ChE the incidence of dicentrics and rings was also increased. In group I. the ir frequency was the higher the earlier after clean -up working cytogenetic study was carried out The frequencies of MN in lymphocytes and erythrocytes were higher ih I. and ChI. as compared to corresponding controls. The interindividual variability of the number of MN as well as of ChA was increased in I., ChI., ChE due to persons with high frequency of these events. In vitro chromosomal radiosensitivity of lymphocyres of ChI. and ChE exceeded that in group of ChC. Tbis finding provides evidence on genetic instability of children irradiated prezigotically IChU or postnataly

The aim of our stud ies was to assess the mutagenic hazards during manufacturing of wood products. The studies were carried out in selected work-Sites in a furniture factory and a woodworking plant. We tested mutagenicity of airborne particulate matters and substances extracted from worker's urine samples . Mutagenic activity was evaluated by the Salmonella plate incorporation assay with TA98 tester strain, without and with S9 mix, The obtained results have shown that the processes such as milling, sanding, and turning significantly increase the number of mutagenic agents in working environment. The level of inhalation exposure to mutagens in above listed workplaces was much higher (40-205 rev/m") than that observed in urban aerosol (4-15 rev/m l ). Observed high activity of mutagenic agents in the air was confirmed by results that come from urine samples of furniture factory workers . Keyword(s) : Wood conversion; Mutagenicity; Ames test

Ip XII1.1031

The Inlluence of anthrax vaccine on mutagenesls

Arm~n Nersesyan'P, Leven Mkrtchiarr" , J State University. Yerevan. Armenia; • Cancer Research Centre. Yerevan. Armenia The use of anthrax spores as biological weapon was recently discussed (the NYAS, USA). Anthrax vaccine (AV) produced in Armenia is widely used for prevention of anthrax infection in animals. The aim of this work was to study the inlluence of AVon spontaneous and induced mutagenesis, because it is known that some vaccines (B. abortus, BCG) have clastogenic activity. The experiments were performed with male rats and mice . We have shown that I.D50 of AV is about 5 million of spores both for rats and mice . Rodents were injected subcutaneously with AV at doses equal to 115 and 1/10 of I.DSO and were killed I, 3, 5, 7 and 10 days after injection. In the second experiment 15 days after vaccination with AV (1110 of LD50) rodents received cyclophosphamide or adriblastin at doses induced about 25% of cells with chromosomal aberrations (CA). Positive control animals received only clastogens, Bone marrow cells were studied for CA 24 h after the last injection. The results of the experiments have shown that AV did not increase the level of cells with CA in bone marrow cells of both species of rodents. In bone marrow of rats and mice immunized with AV the number of cells with CA induced by both mutagens significantly decreased compared with controls . Hence, AV is not c1astogenic for rats and mice, and at the same time immunization of rodents with AV can protect them against the clastogenic effect of some chemical agents . Keyword(s): Anthrax vaccine; Rodents; Mutagenesis