Tetrachlorodibenzo-p-dioxin Research Articles

Polychlorinated dibenzo-p-dioxin and dibenzofuran concentrations in the serum samples of workers at continuously burning municipal waste incinerators in Japan

OBJECTIVESTo find whether concentrations of polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs) in serum increased in workers at municipal incinerators that burn continuously.METHODS30 Workers employed at three municipal waste incineration...

Evaluation of a Reporter Gene System Biomarker for Detecting Contamination in Tropical Marine Sediments.

A major challenge in conducting field assessments of potential ecological impacts is optimizing the number of samples and the costs. This is especially important in light of the growing concern over the presence of persistent organic contaminants, such as PCBs, dioxins, furans, and PAHs in sediments. A reporter gene system (RGS) assay that measures induction of the CYP1A1 gene and transcription of P450 enzyme systems is often used to assess potential toxicity of these compounds in environmental samples (1, 2, 3). RGS has gained acceptance as an inexpensive, rapid method for screening environmental samples for contaminants (4, 5). The RGS approach has been validated in the laboratory with pure compounds, known chemical mixtures, and from field-collected sediments by comparing RGS system response and chemical concentrations (1, 2, 3, 6). Our study differed from other validation studies in two respects: we used field-collected sediments over a wide range of contaminant concentrations and evaluated RGS response to sediment samples containing a mixture of dioxins, furans, and PAHs. With few exceptions, the previous validation studies using field-collected samples used fairly small sample sizes and generally evaluated one chemical group (3, 6). Few studies have compared large numbers of samples containing both PAHs and 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD) over the range reported here. The purpose of this study was to determine if there is a high correlation between RGS response and chemistry results from the same samples. If this proves to be the case, the assay could be used to screen large areas at a relatively low cost. Samples exhibiting high responses could be targeted for further characterization using more precise, but costly, GC/MS methods. Matched sediment samples (n = 31) were collected off the northwestern shore of Johnston Island, adjacent to potential sources of PAHs and 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD) (Site 1), and at sites farther removed from the potential contaminant sources (Sites 3, 4). To ensure blind analysis of the samples, P450 RGS assays were conducted by MEC Analytical Systems (Carlsbad, California) and Columbia Analytical Services (Vista, California) while chemical analyses were completed by the Toxic Contaminant Research Laboratory at Wright State University (Dayton, Ohio). The RGS assay used in this study was developed by Anderson et al. (1) and detailed methods have been described (1, 7). Induction in the assay is dependent upon the aryl hydrocarbon receptor (AhR) activation pathway. AhR ligands, including planar PCBs, PAHs, and TCDD, bind to AhR, activating it and resulting in its translocation to the nucleus of the cell. In the nucleus, the activated

Blood lipid concentrations of dioxins and dibenzofurans causing chloracne.

Chloracne is caused by exposure to certain halogenated polycyclic hydrocarbons such as polychlorinated dibenzodioxins (PCDDs) and dibenzofurans (PCDFs). In chronic exposure it is not known what level of intoxication, represented by the level in blood lipids, is sufficient to cause chloracne. Blood levels of the congeners of PCDD/Fs were determined in four groups of humans. One group had clinically visible chloracne due to exposure in a hexachlorobenzene workshop of a large chemical factory. A second group was exposed in the same workshop, but had no skin changes. There were two control groups: one non-exposed group of maintenance workers from the same chemical factory, and one group of healthy individuals living elsewhere. Blood levels were converted to toxicity equivalents of tetrachlorodibenzo-p-dioxin (TCDD). In the chloracne group blood levels in toxicity equivalents (TEQs) ranged from 1168 to 22,308 pg/g blood lipid. In the exposed without chloracne this ranged from 424 to 662 pg/g. It is concluded that the level to develop chloracne is between 650 and 1200 pg/g TEQ. The contribution of TCDD was rather small, and the main causative congeners were the hexachlorinated dibenzodioxins and dibenzofurans (HxCDD/Fs); lipid-based blood levels in absolute amounts that may cause chloracne are in the range of 2-3.5 ng/g HxCDD, and 2-5 ng/g HxCDF.

Role of HSP90 in mediating cross-talk between the estrogen receptor and the Ah receptor signal transduction pathways

Tetrachlorodibenzo- p-dioxin (TCDD)-mediated gene transactivation via the Ah receptor (AhR) has been shown to be dependent upon estrogen receptor (ER) expression in human breast cancer cells. We have investigated the 90-kDa heat shock protein (HSP90) as a mediator of cross-talk between the AhR and the ER signal transduction pathways. The effect of HSP90 overexpression on receptor activity was determined by transient transfection assays using a HSP90 expression vector. Ligand-inducible gene expression was inhibited when the HSP90 expression vector was cotransfected with a TCDD-responsive reporter plasmid. However, overexpression of HSP90 did not block induction of an estrogen-responsive reporter plasmid. To determine whether ER facilitates AhR signaling through its ability to squelch HSP90, two vectors expressing protein products that bind HSP90 were transfected into MDA-MB-231 cells. Introduction of (i) He11, an ER deletion mutant that does not bind DNA, and (ii) the ligand-binding domain of human AhR, both led to increased basal and TCDD-inducible CYP1A1 expression. Finally, the subcellular distribution of HSP90 was investigated in human breast cancer cell lines. These studies showed HSP90 to be primarily cytoplasmic in ER-positive cell lines, whereas in matched ER-negative cell lines HSP90 was distributed equally between the cytoplasm and nucleus. Taken together, these results demonstrate that HSP90 can regulate AhR activity in vivo, and that Ah-responsiveness is dependent upon cellular ER content through a mechanism that involves HSP90.

Reproductive Endocrine Disruption by Environmental Xenobiotics that Modulate the Estrogen-Signaling Pathway, Particularly Tetrachlorodibenzo-p-dioxin (TCDD)

Environmental pollutants are known to exert endocrine-disrupting effects on several hormonal axes of animals, including reproduction and development. Many of these xenobiotics modulate the estrogen-receptor signaling pathway(s) in agonistic or antagonistic ways. Some of the compounds are themselves estrogenic (so-called xenoestrogens, environmental estrogens, or ecoestrogens), and are classified as synthetic estrogens, phyto- or fungal estrogens, alkylphenol ethoxylates; certain non-coplanar polyclorinated biphenyls (PCBs), etc. Other molecules are anti-androgenic, e.g., p, p'-dichloro-diphenyl-dichloroethylene (DDE); while still others disrupt estrogen function in a manner that is dose, species and tissue specific, e.g., certain co-planar PCBs and dioxin-like molecules (e.g., tetrachlorodibenzo-p-dioxin [TCDD]). Exposure to some compounds has been correlated with skewing of sex ratios in aquatic species, and feminization and demasculinization of male animals; declines in human sperm counts; and overall diminution in fertility of birds, fish, and mammals. This review will cover these various xenobiotics and evaluate them for their estrogen-modulating effects; and then further concentrate on TCDD specifically. Dioxins are produced as by-products of herbicide overuse, from paper bleaching, plastics manufacture, and waste incineration. TCDD has been correlated with altered fecundity and endometriosis in monkeys, and with certain cancers in experimental animals and humans. TCDD is also correlated with reproductive deficits in many laboratory species. In summary, we believe that some of the reproductive deficits from endocrine-disrupting xenobiotics may be attributable to the modulation of the estrogen-signaling pathway, in positive and negative manners, depending on dose, species, and tissue specificity.

Potential alterations in gene expression associated with carcinogen exposure in Mya arenaria

Gonadal cancers in soft-shell clams (Mya arenaria) have been found at high prevalences (20-40%) in populations in eastern Maine. The aetiology of these tumours is unknown. We hypothesized that gene expression would be altered in gonadal tumours and that examination of gene expression patterns would provide some information as to the mechanism of tumour development. To investigate this hypothesis, we initiated a broad search for differentially expressed genes using differential display polymerase chain reaction (dd-PCR) to compare RNA from tumour and normal gonadal tissue. We identified two classes of genes whose expression may be altered in the gonadal tumours: genes involved in biosynthetic processes and genes with possible roles in signal transduction. We also investigated the hypothesis that environmental contaminants, such as tetrachlorodibenzo-p-dioxin (TCDD), may play a role in the development of these tumours. To investigate this hypothesis, we performed a short-term exposure of M. arenaria to [3H]TCDD. Tissues were sampled up to 2 weeks after a 24-h exposure to 10 pptr or 2000 pptr of [3H]TCDD in the water. Using dd-PCR, we identified potential alterations in expression of genes associated with cell proliferation: heparan sulphate proteoglycan, E3 ubiquitinating enzyme and p68 RNAhelicase/initiation factor eIEF-4A. There were no observable histopathological alterations in gonadal or gill tissue from exposed animals. These results suggest possible early changes in gene expression indicative of environmental exposures.

Toxicity of hercides 2,4-D and MCPA for Rats and Rabbits

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National Annual Dioxin Emissions Estimate for Hazardous Waste Incinerators

ABSTRACT Reducing emissions of polychlorinated dibenzo-p-di-oxins and dibenzofurans, commonly known as di-oxins, is a high priority for environmental regulatory bodies throughout much of the world. In the United States, Section 112 (c)(6) of the Clean Air Act (CAA) requires the Environmental Protectio n Agency (EPA) to identify and control emissions from sources that are responsible for at least 90% of the overall emissions of seven targeted hazardous air pollutants, including dioxins.1 On April 19, 1996, the EPA proposed Maximum Achievable Control Technology (MACT) Standards for Hazardous Waste Combustors2 (HWCs). In that preamble, the EPA estimated annual dioxin emissions from the nation's hazardous waste incinerators (HWIs) to be 79 grams expressed as 2,3,7,8 tetrachloro dibenzo-p-dioxins (TCDD) international toxic equivalents (ITEQs). However, early EPA dioxin emission estimates from medical waste incinerators and cement kilns were significantly overestimated;3-5 so, the following independent national dioxin emissions estimate for HWIs was prepared. This estimate corrects the errors in the EPA's HWI emissions database, uses an updated inventory of HWIs in the United States, and applies statistical imputation techniques that take maximum advantage of the limited dioxin emissions data for HWIs.

Protein kinase C activity is required for aryl hydrocarbon receptor pathway-mediated signal transduction.

The role of protein kinase C (PKC) in the human aryl hydrocarbon receptor (hAhR) signal transduction pathway was examined in cell lines stably transfected with pGUDLUC6.1, in which luc+ is solely controlled by four dioxin-responsive elements (DREs). These cell lines, P5A11 and HG40/6, were derived from HeLa and HepG2 cells respectively. Simultaneous treatment of these cells with 2,3,7,8, -tetrachlorodibenzo-p-dioxin (TCDD) and phorbol-12-myristate-13-acetate (PMA) enhanced trans-activation of the reporter construct several-fold relative to cells treated with TCDD alone. PKC inhibitors block the PMA effect and hAhR-mediated signal transduction, demonstrating these processes require PKC activity. Examination of other independently generated, HeLa-derived cell lines stably transfected with pGUDLUC6.1 demonstrates the PMA effect in P5A11 cells is not a clonal artifact. Transient transfections indicate the PMA effect is not due to a luciferase message/gene product stabilization mechanism or stimulation of the basal transcription machinery. Examination of cytosolic preparations demonstrates PKC stimulation or inhibition does not alter hAhR and hAhR nuclear translocator protein levels or TCDD-induced down-regulation of hAhR levels. Similarly, examination of nuclear extracts indicated PKC stimulation or inhibition does not alter nuclear AhR levels or hAhR/hAhR nuclear translocator protein heterodimer DRE-binding activity as assessed by electrophoretic mobility shift assay. These results demonstrate a PKC-mediated event is required for the hAhR to form a functional transcriptional complex that leads to trans-activation and that the DRE is the minimal DNA element required for PMA to enhance AhR-mediated trans-activation.

Levels and membrane localization of the c-K-ras p21 protein in lungs of mice of different genetic strains and effects of 2,3,7,8- tetrachlorodibenzo-p-dioxin (TCDD) and Aroclor 1254

Mutational activation of the K-ras oncogene often occurs in human and mouse lung adenocarcinomas. Since K-ras p21 functions in trans-membrane signaling, we have investigated whether the amount of this protein in lung cell membranes is a variable that could influence lung tumorigenesis, either due to genetic differences or in response to tumor promoters. The six mouse strains assessed showed little difference in the total lung K-ras p21 after immunoprecipitation and immunoblotting. However, amount of ras p21 in the membrane fraction showed significant differences, with C57BL/6 and BALB/c having 3-5-fold more than NIH Swiss, AKR and DBA mice. Interestingly, a congenic AKR strain having the Ahr(b-1) Ah receptor allele from C57BL/6 mice (designated AKR.B6Ah) had high lung membrane K-ras p21 similar to that of C57BL/6. To test for possible changes related to lung tumor promotion, mice were treated with a promotional dose of TCDD (5 nmol/kg). After 48 h C57BL/6 lungs showed an increase in p21 in both total and membrane fractions. BALB/c, DBA and Swiss mice showed an increase only in membranes. There was no change in the AKR and AKR.B6Ah. Aroclor 1254 (250 mg/kg) caused an increase in membrane/cytosol ratio in Swiss mice. Thus the membrane:cytosol K-ras p21 ratio may be influenced by the Ahr phenotype, and TCDD and PCBs can induce p21 or increase its membrane level in certain strains, but these properties are not fully dependent on Ahr receptor type. In confirmation of the relevance of these findings for the tumor target cell type, the immortalized alveolar type 2 E10 cell line presented K-ras p21 in membrane, and this was increased 4-fold by treatment with 10 nM TCDD.

Polychlorinated naphthalenes in urban air

Air samples were collected at an urban site (Chicago, IL) and a semi-urban location (Downsview, Ontario) in February and March 1995 to measure atmospheric concentrations and particle/gas partitioning of polychlorinated naphthalenes (PCN). PCNs were quantified by gas chromatography — negative-ion mass spectrometry (GC-NIMS) against Halowax 1014, a commercial mixture of PCN compounds. GC-flame ionization responses were used to assign mass percent contributions to congeners in Halowax 1014. The reliability of this method was checked using pure compounds and was within ± 20% of the true value for 5 out of 6 congeners tested. Mean atmospheric concentrations (ΣPCN) at the urban and semi-urban site were 68 pgm −3 ( n = 14) and 17 pgm −3 ( n = 2), respectively. Several bioaccumulating PCNs which have “dioxin-like” toxicity similar to coplanar PCBs were quantified. The tetrachlorodibenzodioxin (TCDD) toxicity contribution of one of these PCNs in Chicago air was estimated to outweigh the contributions of some of the coplanar PCBs. The fraction of PCNs on particles exhibited a stepwise increase with homolog group. A need for relevant physicochemical data for this group of compounds was identified.

Toxicity and bioaccumulation of 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin in long‐term tests with the freshwater benthic invertebrates Chironomus tentans and Lumbriculus variegatus

AbstractTwo species of freshwater benthic invertebrates, Chironomus tentans and Lumbriculus variegatus, were exposed to three dietary concentrations of 2,3,7,8‐ tetrachlorodibenzo‐p‐dioxin (TCDD), and toxicity and bioaccumulation were determined. No toxic effects were observed in full life cycle tests with either species at tissue residue concentrations up to 9,533 ng TCDD/g lipid. The observed lack of sensitivity of the two species to TCDD was consistent with a presumed absence of the aryl hydrocarbon receptor in aquatic invertebrates. Predictions of lipid‐normalized tissue concentrations were made based on lipid‐normalized TCDD concentrations in the food and were within 15% of targeted concentrations in both species. Depuration studies indicated that TCDD elimination followed first‐order kinetics, with elimination rate constants of 0.0014 to 0.0022 h−1 for L. variegatus and 0.0070 to 0.0099 h−1 for C. tentans. Half‐lives ranged from 315 to 495 h in L. variegatus and from 70 to 99 h in C. tentans. The ability of invertebrates to accumulate relatively high concentrations of TCDD in the absence of toxic effects may be relevant to the transfer of contaminants through aquatic food webs to potentially sensitive vertebrate species.

Individual-based modeling of PCBs effects on young-of-the-year largemouth bass in southeastern USA reservoira

Effects of polychlorinated biphenyls (PCBs) on young-of-the-year (YOY) largemouth bass ( Micropterus salmoides) are simulated using an individual-based population model. The model simulates the daily development, growth, and survival of largemouth bass from eggs to the end of their first growing season (October 15) in a single, well-mixed box. The model was configured for southeastern USA reservoirs, mostly using data from two Tennessee River impoundments. PCBs exposure levels are expressed as tetrachlorodibenzo- p-dioxin (TCDD) concentrations in largemouth bass tissue. PCBs effects of increased mortality and reduced growth (via decreased feeding and increased metabolic rate) are imposed on modelled individuals dependent on specified exposure concentrations. Monte Carlo following fie model predictions of largemouth bass were analyzed; number density (number/ha), biomass density (kg/ha), mean length (cm), and mean condition factor ( K), all on October 15, and survivorship (fraction of eggs surviving to October 15). PCBs concentrations between 0 and 20 ppm were stimulated. Predicted bass number density and biomass density increased with increasing spawner density show a leveling off with increasing spawner density, implying density-dependence (due to bass consumption reducing prey densities) was operating. PCBs effects were apparent but small relative to natural variation in model predictions. Number density, biomass density, mean condition factor K, and survivorship all decreased, and mean length slightly increased, with increaseing PCNs levels. Predicted PCBs effects for a chronically exposed population (fewer spawners) were less at low exposure levels and greater at high exposure levels than those for an initially-healhy population. Field-based comparisons of YOY densities have a low power for distinguishing PCBs effects from natural interannual variability. While the direct effects of PCBs were relatively small. PCBs exposure may increase the risk of populations to natural and other anthropogenic stresses. Some caution should be used when short-term predictions are used to assess long-term consequences of contaminant exposure. Accurate prediction of PCBs effects require realistic representation of YOY largemouth bass growth rates and better method for estimating exposure in nature.

Accumulation, depuration and hepatic mixed-function oxidase enzyme induction in juvenile rainbow trout and lake whitefish exposed to dietary 2,3,7,8-tetrachlorodibenzo -p-dioxin

Juvenile rainbow trout ( Oncorhynchus mykiss) and lake whitefish ( Coregonus clupeaformis) were exposed to three concentrations (40, 190, 400 pg g −1) of dietary 2,3,7,8-[ 3 H] tetrachlorodibenzo-p-dioxin (TCDD) to compare bioaccumulation and hepatic monooxygenase enzyme (MO) induction. Fish were exposed for 30 days followed by a 180 day depuration phase. Differences in the accumulation and depuration of TCDD were found between rainbow trout and lake whitefish, despite similar body size and lipid content. Assimilation efficiencies of TCDD were greater in lake whitefish (66–76%) than rainbow trout (43–58%), but TCDD half lives were shorter in lake whitefish (32–39 days) than in rainbow trout (73–83 days). Biomagnification factors (BMF) ranged from 1.6 to 1.8 in rainbow trout and from 0.8 to 0.9 in lake whitefish, confirming the known potential for biomagnification of TCDD in aquatic food webs. Reverse phase HPLC showed that a majority of the radioactivity in the rainbow trout bile was TCDD, with minor amounts present as a hydroxylated TCDD and as a glucuronide conjugate. MO enzyme induction, measured by ethoxyresorufin- O-deethylase (EROD), was observed in the rainbow trout after 10 days of exposure to 400 pg g −1 TCDD, and in the lake whitefish after 5 days of exposure to 380 pg g −1 TCDD. The whole fish threshold concentration for EROD induction by TCDD ranged between 15 and 45 pg g −1 (wet weight) for both species. EROD activity returned to control levels 120 and 80 days after the cessation of the treatments in the rainbow trout and lake whitefish, respectively. Growth rates were significantly reduced in trout and whitefish at whole fish concentrations (wet weight) of 150 ± 4.6 and 85 ± 8.3 pg g −1, respectively. Histological effects of the TCDD were found in the spleen and liver of the rainbow trout which had whole fish concentrations (wet weight) of 150 ± 4.6 pg mg −1and 72 ± 8.0 pg mg −1 TCDD, respectively.

Inhibition of increases of transcription factor mRNAs during differentiation of primary rat adipocytes by in vivo 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) treatment

Understanding the differentiation pathway of adipocytes is an important first step for controlling human and animal fat deposition. Although many studies have been done on adipogenesis, most have utilized established cell lines rather than isolated primary cells. We have studied primary preadipoce differentiation to determine whether the cell lines reflect the situation in vivo. In this study, mRNA of several transcription factors and adipocyte-related enzymes, isolated from cultured differentiating primary rat inguinal and epididymal cells, followed the same pattern of change during differentiation as seen in differentiating 3T3-L1 cells. As the cells differentiated, mRNA for C/EBPα, PPARγ2, aP2 and lipoprotein lipase (LPL) increased, C/EBPβ decreased and CHOP remained at a low level. Previously we have shown that in vivo treatment with TCDD (2,3,7,8- tetrachlorodibenzo- p-dioxin) inhibits in vitro adipogenesis and the increase of mRNAs for glycerol-3-phosphate dehydrogenase and LPL (Tox. Lett. 84:55, 1996). TCDD treatment in vivo also inhibited the increase of mRNA for the PPARγ2, aP2 and C/EBPα during differentiation of the isolated preadipocytes. C/EBPβ and CHOP mRNAs were unaffected. Due to the similarity of changes of the transcription factor mRNAs for primary and 3T3-L1 cells during differentiation and after TCDD treatment, 3T3-L1 cells appear to provide a good model for more clearly defining the route of adipogenesis and TCDD inhibition.

Biomonitoring environmental contaminants near a municipal solid-waste combustor

Tetrachlorodibenzo-p-dioxin (TCDD), tetrachlorodibenzofuran (TCDF) and selected metal concentrations were measured in eggs and nestlings of anhingas (Anhinga anhinga) and white ibises (Eudocimus albus) collected from a colony next to a municipal solid-waste (MSW) combustor and ash landfill. Most of the measured residues, including TCDD, TCDF, arsenic, beryllium, cadmium and nickel, remained at pre-operational levels during the first five years of facility operation. Selenium (in anhingas) and mercury (in both anhingas and ibises) occurred at their lowest concentrations in samples collected during the fifth year of facility operation (Year-5). Alternatively, concentrations of lead in ibis nestlings were highest in Year-1 and Year-5 compared to Year-0. The MSW combustor could neither be ruled out nor confirmed as the source of this lead.

Inhibitors of Preadipocyte Differentiation Induce COUP-TF Binding to a PPAR/RXR Binding Sequence

Inhibition of preadipocyte differentiation by 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD) or retinoic acid (RA) identified another transcription factor which appears to be important for preadipocyte differentiation. Within 15 min of treating 3T3-L1 cells with TCDD, the aryl hydrocarbon receptor (AhR) is present within the cell nucleus, and increased binding of COUP-TF to an oligomer of the PPARγ2/RXR binding sequence (ARE7) occurs. Following 2 days of RA treatment, increased binding of COUP-TF to the ARE7 oligomer also occurs. In untreated preadipocytes, COUP-TF mRNA increased at confluence and then decreased after induction. TCDD treatment did not alter COUP-TF mRNA changes. Dephosphorylating the nuclear extracts from TCDD and RA-treated cells eliminated binding of COUP-TF to ARE7. This is the first indication that COUP-TF may play a role in preadipocyte differentiation and that COUP-TF binding to DNA is correlated with TCDD and RA-induced phosphorylation.

Cause specific mortality and cancer incidence among employees exposed to 2,3,7,8-TCDD after a 1953 reactor accident.

To evaluate the long term health consequences of past occupational exposure to 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD). Cancer incidence and cause specific mortality were examined up to and including 1992 in a group of 243 men with external comparisons and internal dose-response analyses. Model based estimates of TCDD dose (expressed in micrograms/kg body weight) were developed for all cohort members with an approach that incorporated detailed accounts of each employee's work activities, analyses of TCDD in blood lipid of 138 employees, and internally derived estimates of elimination rates of TCDD. The estimated dose of TCDD for 135 men was > or = 0.1 microgram/kg body weight and for 69 men > or = 1 microgram/kg body weight. Increased cancer risk ratios were found with higher doses of TCDD and longer interval since first exposure for all sites combined and digestive and respiratory cancers in particular. Within the high dose group (> or = 1 microgram/kg body weight), total cancer mortality was increased > or = 20 years after first exposure (13 cases, standardised mortality ratio (SMR) 1.97, 95% confidence interval (95% CI) 1.05-3.36) as was respiratory cancer (six cases, SMR 3.06; 95% CI 1.12-6.66). Among current cigarette smokers, 12 cancer deaths occurred in the high dose group (SMR 3.42, 95% CI 1.77-5.97) compared with seven deaths at lower doses of TCDD (SMR 1.29, 95% CI 0.52-2.66). Regression analyses based on the Cox's proportional hazards model provided further evidence of a relation between cumulative dose of TCDD and occurrence of both overall and digestive cancer. No evidence of an effect of TCDD on overall mortality or deaths due to circulatory disease was found and no cases of non-Hodgkin's lymphoma or soft tissue sarcoma have been found to date. Our findings are consistent with a carcinogenic effect induced by TCDD at doses > or = 1 microgram/kg body weight. With such a small cohort, the risk estimates are not very stable and could be affected by selection and confounding.

Latent effects of pesticides and toxic substances on sexual differentiation of rodents.

In humans and rodents, exposure to hormonally active chemicals during sex differentiation can produce morphological pseudohermaphrodism (Schardein, 1993; Gray, 1992). For example, hormonally active drugs like DES (estrogenic), Danazol (androgenic), and progestins cause urogenital malformations in the reproductive tracts of humans and rodents. The current discussion will present new information on the effects of toxic chemicals and pesticides that act on reproductive development via novel mechanisms, including germ cell toxicity, antiandrogenicity, and Ah-receptor binding. Information will be presented that describes how exposure during critical stages of life to synthetic chemicals present in our environment, such as benzidine-based dyes, antiandrogenic fungicides, 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD), and PCB congener 169, result in abnormal rodent sex differentiation. In rodents, perinatal exposure to fetal germ cell toxicants reduced the reproductive potential of female, and permanently reduced sperm production in male progeny. Phenotypic sex differentiation, however, was unaffected by these germ cell toxicants. In contrast, antiandrogenic drugs and fungicides induced profound alterations in phenotypic sex differentiation. Effects such as hypospadias, ectopic testes, vaginal pouches, agenesis of the ventral prostate, and nipple retention in male rats were observed commonly. Although these antiandrogens induced no permanent effects in female progeny, another class of chemicals, the Ah-receptor mediated toxicants, did reduce fertility in both male and female rat offspring. Cauda epididymal sperm numbers were reduced permanently in TCDD-exposed male rat and hamster progeny, and female progeny displayed malformations of the external genitalia. Other toxicants produced dramatic alterations of sex differentiation (uterus unicornis, agenesis of the vas and epididymis, and undescended testes), via mechanisms that have not been characterized yet. Since these adult/pubertal alterations resulted from gestational and/or neonatal exposures, future studies should include a comprehensive assessment of reproductive function after perinatal exposure because the developing animal is extremely sensitive to toxicants during sex differentiation, and many of the effects are difficult to detect until late in life.

In vitro modulation of 17-β-estradiol-induced vitellogenin synthesis: Effects of cytochrome P4501A1 inducing compounds on rainbow trout ( Oncorhynchus mykiss) liver cells

Cytochrome P4501A1 (CYP1A1) induction, particularly in liver, is a useful marker of exposure of fish to polycyclic- and halogenated-aromatic hydrocarbons. However, the relationship between toxicity and CYP1A1 induction in fish is uncertain. Some compounds that induce CYP1A1 are antiestrogenic in mammalian bioassay, and this effect is linked to aryl hydrocarbon (Ah) receptor and/or increased catabolism of 17-β-estradiol. Liver of fish synthesizes estrogen-inducible egg yolk precursor protein vitellogenin (Vg) which is critical for oocyte maturation and ovarian development. To determine if CYP1A1-associated endocrine modulation could occur in fish liver, primary cultures of rainbow trout liver cells were co-administered 17-β-estradiol and CYP1A1 inducing compounds or mixtures. Protein synthesis and enzyme activity of cells were optimal when cultured in a modified HEPES buffered Medium 199 at 15 °C. Vg and albumin (Alb), estimated by ELISA measurement of concentration in the media 48 h after treatment, formed the basis for the test. Equivalent viability (mitochondrial dehydrogenase activity) and secretory functional capacity (Alb synthesis) were estimated and correlated with other results. In descending order, 2,3,4,7,8-pentachlorodibenzofuran (10 −12 to 10 −8 M) > 2,3,7,8- tetrachlorodibenzo-p- dioxin ( TCDD) ≊ 2,3,7,8- tetrachlorodibenzofuran (10 −11 to 10 −8 M) > β- naphthoflavone (10 −7 to 10 −6 M) inhibited Vg synthesis in 17-β-estradiol treated liver cells. Potency of inhibition directly related to strength as an inducer of CYP1A1 protein. At 10 −8 M, PCB congeners 77, 126, 156 did not inhibit Vg synthesis and induced no to moderate levels of CYP1A1 protein or EROD activity. At 10 −8 M, congener 114, a weak EROD inducer, potentiated Vg synthesis relative to cells treated with 17-β-estradiol alone. The results of this study increase our understanding of the consequences of hepatic CYP1A1 induction, forewarn of reproductive impairment of sexually maturing fishes exposed to CYP1A1 inducing compounds and argue for further, more detailed in vivo investigation.