Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on guinea pig heart muscle

Cardiac effects of TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) were examined using atrial muscle isolated from young or mature guinea pigs fed ad libitum, from young animals 10 or 20 days after a single ip injection of 1 microgram TCDD/kg body wt, and from control guinea pigs pair-fed to the TCDD-treated animals. Basal contraction force of left atrial muscle obtained from TCDD-treated and pair-fed control guinea pigs 10 days after treatment was significantly increased compared to atrial muscle obtained from ad libitum control animals. Atrial muscle from TCDD-treated animals 20 days after treatment had a significantly lower basal force of contraction while that from pair-fed animals was significantly higher relative to atrial muscle from ad libitum control animals. These results suggest that the TCDD-induced change in basal atrial contraction force is in part due to undernutrition and that prolonged exposure to TCDD inhibits this response observed in pair-fed animals. Furthermore, prolonged TCDD treatment shifted the dose-response curve for the positive inotropic effect of isoproterenol to the right with no statistically significant change in the maximal inotropic effect obtained with high concentrations of isoproterenol. The TCDD-induced changes in force of contraction and the dose-response curve for the positive inotropic effect of isoproterenol were similar in young TCDD-treated and mature control guinea pigs. TCDD did not alter the basal contraction frequency or the dose-response curve for the positive chronotropic effect of isoproterenol. There was no effect on cardiac lipoprotein lipase activity at 1 microgram/kg, but in animals administered 4 micrograms TCDD/kg activity was reduced. It was concluded that TCDD adversely affects atrial muscle of guinea pig heart.     

Intermediary metabolism of the mature rat following 2,3,7,8-tetrachlorodibenzo-p-dioxin treatment

Changes in body weight, feed intake, hepatic cellularity, and intermediary metabolism were assessed in the mature male (450 g) rat following 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) administration. All animals were schedule-fed (8-hr feeding period/24 hr) and treated with a single oral dose of either TCDD (75 micrograms/kg) or vehicle. Blood and tissues were sampled 16 to 18 hr following the end of the feeding period on 2, 4, 6, and 8 days post-treatment. Mature rats treated with TCDD exhibited a slight but progressive reduction in both body weight and feed intake throughout the 8-day experimental period. An increase in liver mass that was apparent at 2 days and plateaued by 4 days after TCDD treatment was associated with a decrease in the concentration of DNA per gram of wet liver. However, the total liver content of DNA in TCDD-treated rats remained similar to pair-fed animals. Thus, TCDD treatment produced liver enlargement in the mature rat that was the result of hepatocellular hypertrophy and not an increase in cell number. Hepatic glycogen content in TCDD-treated rats was threefold higher than their pair-fed counterparts at 2 to 6 days post-treatment, and this augmentation would account, in part, for the hypertrophy of the liver cell found after administration of TCDD. Plasma glucose and lactate concentrations were similar in TCDD-treated and pair-fed rats, suggesting that the Cori cycle remained unaltered following TCDD administration. Likewise, heart and gastrocnemius glycogen concentrations were similar in all experimental groups. Urinary excretion of urea, ammonia, and creatinine was comparable in TCDD-treated rats and their pair-fed counterparts, indicative of a nitrogen balance that was not disturbed by TCDD. Plasma glutamine concentrations in TCDD-treated rats tended to be reduced and were significantly lower at Day 6 post-treatment when compared to those of pair-fed counterparts, suggestive that amino acid release from muscle was not enhanced in TCDD-treated rats. Likewise, plasma concentrations of branched-chain amino acids, which are metabolized to a large extent in muscle, tended to be lower on Day 6 following TCDD treatment. Yet at Day 6 post-treatment, the circulating concentrations of amino acids that are metabolized by the liver were elevated in TCDD-treated animals. TCDD administration also resulted in an increase in total hepatic protein concentration which was evident at 4 days and increased progressively at 6 and 8 days post-treatment. Liver content of phospholipids also increased gradually following administration of TCDD.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on the hepatic metabolism of testosterone in the rat

A dose of 20 micrograms/kg of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was given to adult male rats intragastrically and the metabolism of [4-14C]-testosterone by liver homogenate was examined 1 week later. The overall catabolism of testosterone was significantly reduced by the TCDD treatment. Dihydroxysteroids (intermediate reduced metabolites) were increased, while the production of polar non-conjugated metabolites (end products) was diminished.     

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on [3H]thymidine incorporation into rat liver deoxyribonucleic acid

The effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on [3H]thymidine incorporation into hepatic DNA was studied in rats. In non-hepatectomized male and female animals, incorporation measured at the peak of the first round of liver DNA synthesis after TCDD treatment (10 micrograms/kg) was similar to that of control animals. In contrast, the first round of [3H]thymidine incorporation after a 1/3 hepatectomy was enhanced 3-fold in TCDD-treated rats. The enhanced response to 1/3 hepatectomy was produced by doses of TCDD ranging from 1 to 30 micrograms/kg with an apparent ED50 of 5 micrograms/kg. Enhanced incorporation was observed when the 1/3 hepatectomy was performed 5-10 days after an ED50 dose and it returned to the control level after 20 days. This enhanced response was not preceded by changes in food consumption or hepatic activities of ornithine decarboxylase (ODC), tyrosine aminotransferase (TAT) or gamma glutamyl transpeptidase (GGT) when compared to respective control values. Also, the enhanced incorporation was not necessarily due to removal of 1/3 of the liver because it was also seen in TCDD-treated rats that were laparotomized. The mechanism of enhancement in laparotomized animals does not appear to involve a diminished response of the liver to the inhibitory effects of adrenal hormones on liver DNA synthesis. This was suggested by the finding that an adrenalectomy prior to the laparotomy did not block the enhanced incorporation of [3H]thymidine into hepatic DNA. The mechanism by which TCDD enhances the first round of liver DNA synthesis after a 1/3 hepatectomy or laparotomy remains to be determined.     

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on preimplantation mouse embryos

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a potent environmental contaminant that can exert developmental toxicity. To investigate the stage-specific effects of TCDD on preimplantation embryos, we exposed mouse embryos to TCDD at different stages (1-, 2-, and 8-cell) and collected them at different stages of development (the 1- or 2-, 8-cell, and blastocyst stage, respectively). Semiquantitative RT-PCR revealed increased constitutive gene expression of the arylhydrocarbon receptor (AhR) and AhR nuclear translocator (Arnt) at the 1-cell stage, decreased expression at the 2- to 8-cell stage, and increased expression again at the blastocyst stage, and addition of TCDD to media did not affect their mRNA levels. Interestingly, no cytochrome P4501A1 (CYP1A1) mRNA was detected in embryos at the 1-, 2-, and 8-cell stages after exposure to 10 nM TCDD for 12 or 24 h, whereas CYP1A1 mRNA was significantly increased at the blastocyst stage in response to TCDD, and its induction was found to be concentration-dependent on TCDD exposure from 0.01 to 10 nM for 24 h. In addition, no significant differences in development rate of preimplantation embryos, cell number of blastocyst embryos, or apoptotic indices, such as TUNEL-positive cell number or Bax/Bcl-2 expression ratios were observed at the blastocyst stage between TCDD-exposed groups and non-exposed group. These results suggest that the sensitivity to TCDD differs with the embryonic stage, which may reflect an ability of embryos to adapt to environmental stressors, such as dioxins.     

Activity of thyroid hormone-inducible enzymes following treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) caused a depletion of serum thyroxine, but paradoxically did not change L-3,5,3'-triiodothyronine (T3) levels in serum of rats. The activities of the thyroid-regulated enzymes alpha-glycerol phosphate dehydrogenase (GPD) and malic enzyme (ME) were determined in livers of normal and thyroidectomized (THX) rats treated with 0.1 to 100 nmol TCDD/kg body weight. Mitochondrial GPD activity did not change significantly as a function of TCDD dose in either normal or THX rats. ME activity was induced by TCDD in a dose-dependent fashion, but only in non-THX animals. The absence of ME induction in THX rats treated with TCDD indicates that TCDD is not intrinsically thyromimetic. The dependence of ME induction on thyroid hormones is much like the thyroid-hormone-dependent, multihormonal induction of ME by insulin and glucocorticoids. However, TCDD had no additive or synergistic effects on induction of ME activity in THX rats fed T3. A 30% decrease in steady-state plasma T3 levels of T3-fed animals treated with TCDD relative to T3-fed controls suggested that T3 catabolism was more rapid in TCDD-treated rats than controls. Thus a thyroid-hormone-dependent, multihormonal interaction is suggested as the basis for induction of ME by TCDD, but a strictly T3-dependent process has not been ruled out.     

Effects of in utero and lactational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on rat follicular steroidogenesis

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a widespread environmental pollutant and causes adverse effects on female reproduction when administered to rats. Our aims were to study effects of gestational and lactational exposure to TCDD on ovarian steroidogenesis and steroidogenic enzyme expression of offspring on postnatal day (PND) 14 in the rat and sensitivity of enzymatically isolated ovarian follicles to TCDD in vitro. Synthetic estrogen diethylstilbestrol (DES) was used as a treatment control. Serum progesterone (P4) level in offspring increased significantly on PND 14 in the TCDD (1 microg/kg)-exposed group while body weight, FSH and E2 levels were not changed. In ovarian follicles of offspring on PND 14 in the TCDD-exposed groups, protein expression of cytochrome P-450 aromatase, cytochrome P-450 cholesterol side-chain cleavage, steroidogenic acute regulatory protein, 3beta-hydroxy-steroid-dehydrogenase/Delta(5)-Delta(4) isomerase type 1, or P4 receptor was not affected. TCDD decreased E2 and P4 production in ex vivo follicle culture. DES at a dose level of 0.1mg/kg was dystocic while a dose 0.02 mg/kg increased ovarian ex vivo E2 and testosterone production without affecting P450arom activity indicating stimulation of early steps of steroidogenic pathway. Data suggests that TCDD has multiple targets in ovarian steroidogenesis, but the inhibitory action represented as decreased follicular steroid hormone production ex vivo is not apparent at the ovarian protein expression. Furthermore, TCDD had no direct effect on immature rat ovarian steroidogenesis in vitro suggesting that the follicle culture method is not a sensitive method to study the mechanisms of TCDD action.     

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on the in vivo and in vitro dechlorination of pentachlorophenol

The metabolism of pentachlorophenol has been studied in the rat after pretreatments with phenobarbital, 3-methyl-cholanthrene or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In addition to the previously identified metabolite, tetrachloro-p-hydroquinone, trichloro-p-hydroquinone has been identified in urine as a metabolite. The formation of the latter represents a type of dechlorination different from that of the formation of tetrachlorohydroquinone. The inducing agents, 3-methylcholanthrene and TCDD have similar effects on the dechlorination and increase the formation of tetrachloro-p-hydroquinone more pronounced than does phenobarbital. In contrast to phenobarbital they also increase the formation of trichloro-p-hydroquinone and the total elimination of pentachlorophenol and its metabolites. The in vivo findings are supported by in vitro studies with microsomes from rats pretreated with phenobarbital or TCDD. Use of the inhibitor -diethylaminoethyl-diphenyl propylacetate (SKF 525-A) in vitro showed a more pronounced inhibition on microsomes from phenobarbital-treated rats than on microsomes from untreated or TCDD-treated rats.Gas chromatography-mass spectrometry have been used for the identification and quantification of pentachlorophenol and its metabolites.

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Zusammenfassung Der Metabolismus von Pentachlorphenol nach Vorbehandlung der Versuchstiere (Ratten) mit phenobarbital, 3-Methylcholantren oder 2,3,7,8-Tetrachlordibenzo-p-Dioxin (TCDD) ist untersucht worden. Zu dem schon früher nachgewiesenen Metaboliten Tetrachlor-p-Hydrochinon wurde nun auch Trichlor-p-Hydrochinon als Harnmetabolit festgestellt. Die Bildung des letzteren stellt eine andere Art von Dechlorierung dar als diejenige die bei der Entstehung von Tetrachlor-p-Hydrochinon vorliegt. 3-Methylcholantren und TCDD haben ähnlichen Einfluß auf die Dechlorierung und steigern die Bildung von Tetrachlor-p-Hydrochinon mehr ausgeprägt als es bei phenobarbital der Fall ist. Im Gegensatz zu phenobarbital steigern sie auch die Bildung von Tri-chlor-p-Hydrochinon sowie die totale Eliminierung von Pentachlorphenol und von Metaboliten. Die in vivo-Befunde werden von in vitro-Studien mit Mikrosomen von mit phenobarbital oder TCDD vorbehandelten Ratten gestützt. Anwendung des Inhibitors -Diethylaminoethyl-Diphenyl-Propylacetat (SKF 525-A) zeigte in vitro eine ausgeprägtere Inhibition der Mikrosomen von mit phenobarbital behandelten Ratten als der Mikrosomen von unbehandelten oder TCDD-behandelten Ratten. Nachweis und Bestimmung von Pentachlorphenol und seinen Metaboliten wurden gaschromatographisch-massenspektrometrisch durchgeführt.

     

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on behavior of monkeys in peer groups.

Adult female rhesus monkeys were fed diets containing 0, 5, or 25 ppt 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) for approximately 4 years. They were bred to unexposed males during TCDD exposure (Experiment 1) and again after TCDD exposure ended (Experiment 2). Offspring from both experiments were weaned at 4 months and socialized for 1.5 h/day in groups of four monkeys each beginning at approximately 8 months of age. Each social group contained both control and TCDD-exposed monkeys. In Experiment 2, the offspring were later placed in new social groups containing only monkeys from the same TCDD exposure condition. The TCDD-exposed offspring born concurrent with maternal TCDD exposure (Experiment 1) initiated more rough-tumble play, retreated less during play bouts, and were less often displaced from preferred positions in the playroom. They also engaged in more self-directed behaviors. The behavior of offspring born after maternal TCDD exposure ended (Experiment 2) was not altered when they were socialized with control monkeys. However, some behavioral changes did emerge when they were placed in social groups containing only TCDD-exposed monkeys.     

Response of the antral mucosa of the rat stomach to 2,3,7,8-tetrachlorodibenzo-p-dioxin.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) produces a striking hypergastrinemia in rats that is thought to mediate the antiatrophy effect of TCDD on the oxyntic gland mucosa of the stomach. However, effects of TCDD on the antral mucosa, which is the origin of most physiologically released gastrin and is not a target for the trophic action of gastrin, has yet to be thoroughly investigated. Also gastrin release from gastrin-containing cells (i.e., G-cells) in the antral mucosa is inhibited by the paracrine secretion of somatostatin from D-cells in the antrum. Our purpose was to determine if the antral mucosa is affected by the trophic influence of TCDD and if alterations in antral mucosa levels of gastrin or somatostatin cause the hypergastrinemia. TCDD (100 micrograms/kg, Day 14 post-treatment) had a trophic effect on the antral mucosa. This was demonstrated histologically and by significant increases in antral wet weight and antral mucosa height. In contrast, pair-fed control rats that lost the same amount of body weight developed antral mucosa atrophy. With respect to serum and antral levels of gastrointestinal hormones, TCDD produced a 7- to 10-fold increase in serum gastrin concentrations that was not detected until Day 14 post-treatment. In contrast, serum gastrin concentrations in pair-fed control rats were comparable to those of control rats. The number of G-cells in the antral mucosa was not affected by either TCDD treatment or paired-feed restriction. These findings demonstrate that hypergastrinemia in TCDD-treated rats is not caused by reduced feed intake or antral G-cell hyperplasia. A major finding was that antral mucosa levels of both gastrin and somatostatin were decreased significantly in TCDD-treated rats. However, the temporal development and dose-dependence of these TCDD effects on antral hormone levels were quite different than those for hypergastrinemia. TCDD-induced decreases in antral levels of gastrin and somatostatin were detected 1 week earlier than hypergastrinemia. Also, the ED50 of TCDD on Day 14 post-treatment for the decrease in antral mucosa content and concentration of gastrin (29 and 22 micrograms/kg, respectively) and somatostatin (24 and 19 micrograms/kg, respectively) was less than that for hypergastrinemia (46 micrograms/kg). These time- and dose-dependent differences demonstrate that hypergastrinemia in TCDD-treated rats is not a consequence of reduced antral levels of gastrin or somatostatin. We conclude that the antral mucosa, an epithelial tissue not responsive to the proliferative effect of gastrin, is nevertheless a target for the trophic influence and gastrointestinal hormone-altering effects of TCDD.     

Biochemical and functional effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on the heart of female rats

Biochemical, functional and morphologic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on the hearts of female rats were examined. Six days after the treatment of rats with TCDD, the blood pressures and resting heart rates were significantly less than in control animals. Treated animals were also less responsive to the effects of the beta-1 agonist, (-)isoproterenol. No histopathologic changes were observed in the heart although extensive centrilobular necrosis occurred in the liver after TCDD administration. Serum levels of thyroxine were 66% less than in control animals. Marked lipid peroxidation was produced in the liver with small but significant increases occurring in the heart. TCDD administration had no effect on catalase activity in the heart, but produced a 20% decrease in superoxide dismutase activity relative to control animals. The effects of TCDD on cardiac function do not appear to be due to a direct action of the xenobiotic on the heart but possibly to a down-regulation of beta-receptors in the heart as a result of the hypothyroid state.     

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin, kepone, and polybrominated biphenyls on transport systems in isolated rat hepatocytes.

Adult male rats recived intraperitoneal injections of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, 10 μg/kg), polybrominated biphenyls (PBBs, 200 mg/kg), Kepone (5 mg/kg/day for 5 days), or vehicle, and isolated hepatocytes were prepared 10, 3, or 1 day after injection of TCDD, PBBs, or Kepone, respectively. All treatments significantly increased the hepatic microsomal content of cytochrome P-450. The initial velocity of uptake (V₀; measured within the first 2 min), steady-state concentration (SS, measured over 1 hr), and an indirect estimate of an efflux rate constant, k_cf, were determined for ouabain and procaine amide ethobromide (PAEB). TCDD decreased V₀ and SS of ouabain but had no effect on k_cf for ouabain or any parameter of PAEB transport. PBBs tended to increase k_cf and significantly decreased SS of ouabain. The k_cf for PAEB was significantly increased, but V₀ and SS were not changed in PBB-treated rats. Kepone had no effect on V₀ of either substrate but significantly decreased SS and increased k_cf for ouabain. These data demonstrate that the ouabain transport system is selectively inhibited by TCDD and that PBBs may nonspecifically increase the rate of efflux of actively transported substrates from cell to medium.     

The in vivo biotransformation of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the rat

This study presents evidence for the in vivo biotransformation of TCDD in the rat. Three male rats with indwelling bile loop cannulas were given repeated daily po doses of 15 μg [¹⁴C]TCDD/kg body weight. After either two, four, or six doses, the total output of bile from one rat was collected for 24 hr following the last dose. Biliary ¹⁴C activity was excreted at a rate similar to the excretion of ¹⁴C activity in the feces of normal (noncannulated) rats given po doses of [¹⁴C]TCDD. Therefore it is not likely that enterohepatic recycling plays a significant role in the retention of ¹⁴C activity following a dose of [¹⁴C]TCDD. High-pressure liquid chromatography of the bile from these rats showed the presence of at least eight radioactive peaks and very little, if any, unchanged TCDD. These metabolites were all more polar than TCDD, and the chromatographic profile was altered following incubation of the bile with β-glucuronidase. These data, in conjunction with previous studies, indicate that the metabolic transformation of TCDD in the liver may be the rate-limiting step in the elimination of TCDD from the body.     

Effects of short-term exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin on microRNA expression in zebrafish embryos

Although many drugs and environmental chemicals are teratogenic, the mechanisms by which most toxicants disrupt embryonic development are not well understood. MicroRNAs, single-stranded RNA molecules of ~ 22 nt that regulate protein expression by inhibiting mRNA translation and promoting mRNA sequestration or degradation, are important regulators of a variety of cellular processes including embryonic development and cellular differentiation. Recent studies have demonstrated that exposure to xenobiotics can alter microRNA expression and contribute to the mechanisms by which environmental chemicals disrupt embryonic development. In this study we tested the hypothesis that developmental exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a well-known teratogen, alters microRNA expression during zebrafish development. We exposed zebrafish embryos to DMSO (0.1%) or TCDD (5 nM) for 1 h at 30 hours post fertilization (hpf) and measured microRNA expression using several methods at 36 and 60 hpf. TCDD caused strong induction of CYP1A at 36 hpf (62-fold) and 60 hpf (135-fold) as determined by real-time RT-PCR, verifying the effectiveness of the exposure. MicroRNA expression profiles were determined using microarrays (Agilent and Exiqon), next-generation sequencing (SOLiD), and real-time RT-PCR. The two microarray platforms yielded results that were similar but not identical; both showed significant changes in expression of miR-451, 23a, 23b, 24 and 27e at 60 hpf. Multiple analyses were performed on the SOLiD sequences yielding a total of 16 microRNAs as differentially expressed by TCDD in zebrafish embryos. However, miR-27e was the only microRNA to be identified as differentially expressed by all three methods (both microarrays, SOLiD sequencing, and real-time RT-PCR). These results suggest that TCDD exposure causes modest changes in expression of microRNAs, including some (miR-451, 23a, 23b, 24 and 27e) that are critical for hematopoiesis and cardiovascular development.     

Comparative activities of 2,3,7,8-tetrachlorodibenzo-p-dioxin and progesterone as antiestrogens in the female rat uterus

The comparative antiestrogenic effects of progesterone and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on cytosolic and nuclear estrogen (ERc and ERn, respectively) and progesterone (PRc and PRn, respectively) receptor levels were determined in female Long Evans rats. Estradiol treatment typically increases ER and PR levels in target cells or tissues and these proteins have been proposed as markers of estrogen action. 2,3,7,8-TCDD causes a dose-dependent decrease in uterine ERc, ERn, PRc, and PRn levels which persist up to 7 days. Progesterone treatment caused significant decreases in uterine ERc, ERn, and PRn levels; however, after 7 days, the effects of the hormone on receptor levels were diminished. The effects of 2,3,7,8-TCDD and progesterone on hepatic ER and PR levels were comparable to those observed in the uterus. Treatment of the rats with estradiol (5 micrograms/kg), estradiol (5 micrograms/kg) plus progesterone (1 mg/animal), or 2,3,7,8-TCDD (80 micrograms/kg) showed that both progesterone and 2,3,7,8-TCDD significantly antagonized the estradiol-mediated increases in uterine (and hepatic) ERc, ERn, PRc, and PRn levels and for 2,3,7,8-TCDD the decreased receptor levels persisted for up to 7 days. In vitro studies with freshly isolated uterine strips demonstrated that both 2,3,7,8-TCDD and progesterone antagonized the estradiol-mediated increases in ER and PR levels. Previous studies suggest that the antiestrogenic activity of progesterone is due, in part, to the induction of proteins which are involved in decreasing ERn levels in target cells. Moreover in the uterine strip assay procedure, it was previously shown and reproduced in this study that the decrease in uterine ERn by progesterone was inhibited by both protein and RNA synthesis inhibitors over a 4-hr incubation period. In contrast, the 2,3,7,8-TCDD-mediated decrease in uterine ERn was inhibited only by actinomycin D and not by cycloheximide or puromycin. These in vitro studies thus confirm that both progesterone and 2,3,7,8-TCDD exhibit comparable antiestrogenic effects in vivo and in vitro; however, the results suggest that these effects are expressed through different mechanisms.     

Gestational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin induces developmental defects in the rat vagina.

At puberty, female rats exposed in utero to 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD) exhibit a persistent thread of mesenchymal tissue surrounded by keratinized epithelium that partially occludes the vaginal opening. Our objective was to determine the earliest time during fetal development that morphological signs of this vaginal canal malformation could be detected and to obtain greater insight into mechanisms involved in this effect. Pregnant rats were administered a single dose of vehicle (control) or TCDD (1.0 microg/kg, po) on gestation day (GD) 15 and were sacrificed on GD 18, 19, 20, and 21 for histological evaluation of female. Gestational exposure to TCDD affected vaginal morphogenesis as early as GD 19, 4 days after exposure of pregnant dams. In exposed fetuses, the thickness of mesenchymal tissue between the caudal Mullerian ducts was increased, which resulted in a failure of the Mullerian ducts to fuse, a process normally completed prior to parturition. In addition, TCDD exposure appeared to inhibit the regression of Wolffian ducts. Thus, TCDD interferes with vaginal development by impairing regression of the Wolffian ducts, by increasing the size of interductal mesenchyme, and by preventing fusion of the Mullerian ducts. Taken together, these effects appear to cause the persistent vaginal thread defect observed in rats following in utero and lactational TCDD exposure.     

Arrest of rat molar tooth development by lactational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin

The interference with tooth development by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was studied in dioxin-resistant Han/Wistar rats. Lactating dams were given a single dose of 50 or 1000 microg TCDD/kg body wt 1 day after delivery and the pup heads were analyzed radiographically or histologically at postnatal days 9 and/or 22. Of 19 animals studied histologically, 10 lacked one or more third molars, which were at the bud stage at the start of the experiment. A higher proportion of pups exposed to the higher dose (9/13) lacked third molars than those exposed to the lower dose (1/6) (27/52 and 2/24 teeth missing, respectively). Missing upper third molars (19/38) were more frequent than were lower (10/38). The development of the third molars present was retarded. The root tips of the more advanced first and second molars were prematurely closed and root formation was arrested, but eruption was not affected. Dentinogenesis of the continuously erupting lower incisor teeth was preeruptively arrested because of pulpal cell death. All the teeth of the control rat pups developed normally. In contrast to the control pups, none of the 11 experimental pups examined radiographically (6 exposed to the higher dose and 5 to the lower) showed mineralization of their third molar cusps. The results show that the effects of TCDD on rat tooth development depend on not only the dose but also the tooth type and developmental stage. Inasmuch as early tooth development is under the control of inductive interactions between the epithelium and the mesenchyme, the interference by TCDD with tooth morphogenesis with the consequent arrest of development is likely to involve epithelial-mesenchymal signaling.     

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on indocyanine green blood clearance in rhesus monkeys.

To assess effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on liver function adult male rhesus monkeys were treated with a single oral dose of acetone/corn oil (control) or 5, 25, or 75 micrograms/kg TCDD. Each monkey was used as its own control and indocyanine green (ICG) blood clearance and the following serum enzymes: glutamic pyruvate transaminase (SGPT), sorbitol dehydrogenase (SDH) and gamma glutamyl transpeptidase (gamma GTP), were measured at regular intervals for 4 weeks before and 17 weeks after treatment. In control monkeys ICG blood clearance and serum enzymes were similar before and after treatment. However, in the monkey that received 5 micrograms/kg TCDD there was a mild increase in ICG blood clearance followed by a slight decrease. The magnitude of this biphasic change was greater in monkeys that received 25 and 75 micrograms/kg TCDD and the decrease in clearance was invariably associated with a 1--2-week period before the monkeys died. SDH and SGPT activities were elevated at some time during the course of intoxication in all TCDD-treated monkeys but gamma GTP activity was not altered. The monkey treated with 5 micrograms/kg TCDD survived but monkeys treated with 25 and 75 micrograms/kg died 4--6 weeks after treatment. Light microscopy of the livers of TCDD-treated monkeys that died revealed fatty infiltration with minimal hepatocellular necrosis.     

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on hepatic and uterine estrogen receptor levels in rats

Administration of a single dose of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, 20 or 80 micrograms/kg) resulted in significantly decreased hepatic and uterine estrogen receptor (ER) levels in 25-day-old female Long-Evans rats. By contrast, estradiol (5 and 15 micrograms/kg) administration increased hepatic and uterine ER levels, while a combination of 2,3,7,8-TCDD plus estradiol resulted in uterine and hepatic ER levels which were similar or lower than those observed after treatment with only 2,3,7,8-TCDD. In addition, 2,3,7,8-TCDD significantly decreased the effects of estradiol on uterine wet weight increase. Competitive binding studies showed that estradiol did not bind to the aryl hydrocarbon (Ah) receptor and that 2,3,7,8-TCDD did not bind to the ER. The effects of structure on the activity of polychlorinated dibenzo-p-dioxin congeners on their activity to down-regulate hepatic and uterine ER levels were determined by using 2,3,7,8-TCDD, 1,2,3,7,8-pentachlorodibenzo-p-dioxin (PeCDD), 1,3,7,8-TCDD, and 1,2,4,7,8-PeCDD. Both 2,3,7,8-TCDD and 1,2,3,7,8-PeCDD exhibit high affinities for the Ah receptor and at dose levels of 80 micrograms/kg the hepatic ER levels were decreased 42 and 41%, respectively, and uterine ER levels were decreased 53 and 49%, respectively. 1,3,7,8-TCDD and 1,2,4,7,8-PeCDD bind less avidly to the Ah receptor and at dose levels of 400 micrograms/kg these compounds decreased hepatic ER levels 36 and 40%, respectively, and uterine ER levels 21 and 24%, respectively. These results support a role for the Ah receptor in the down-regulation of uterine and hepatic ER levels in the female rat by 2,3,7,8-TCDD and this effect may be associated with the decrease in spontaneous mammary and uterine tumors observed in female rats treated with 2,3,7,8-TCDD.