Mechanism of thiazopyr-induced effects on thyroid hormone homeostasis in male Sprague-Dawley rats.

Chronic administration of thiazopyr in the diet at dose levels of 1000 and 3000 ppm, but not 100 ppm, has demonstrated an increase in thyroid follicular cell tumors in male Sprague-Dawley rats. In the studies reported here we have evaluated the mechanism of thiazopyr-induced thyroid tumors by studying the effect of thiazopyr on a number of endpoints that indicate hypothalamic-pituitary-thyroid homeostasis. At a dose level of 3000 ppm, thiazopyr caused a marked depression in circulating levels of T4 as soon as 7 days after commencement of treatment. Concurrent with this decrease in T4 was an increase in TSH levels, an increase in thyroid and liver weights, a three- to sixfold increase in hepatic T4-uridine diphosphate glucuronosyl transferase (UDPGT) activity, and increases in thyroid follicular cell hypertrophy and hyperplasia. Dose-related changes associated with thiazopyr treatment were significant increases in liver weight, thyroid weight, and hepatic T4-UDPGT activity at high doses. Increased levels of serum TSH, T3, and rT3, decreased levels of T4, and an increased incidence of thyroid follicular cell hypertrophy and hyperplasia were observed 56 days after the initiation of 3000 ppm thiazopyr. All the changes, except thyroid weight, were partially or completely reversible upon removal of thiazopyr from the diet. Increased thyroid T4 elimination, primarily via increased hepatic conjugation by T4-UDPGT, resulting in decreased serum T4, appeared to be responsible for the increased TSH levels. The sustained increase in TSH by thiazopyr appears responsible for the stimulation of the thyroid follicular cells resulting in follicular cell hypertrophy, hyperplasia, and ultimately neoplasia. In summary, evidence is presented for a hormonally mediated, threshold-dependent process for the development of thyroid follicular cell tumors from high-dose thiazopyr administration in male rats. This mechanism is not considered to be relevant to humans, since the thyroid of humans is much less sensitive to this pathogenic phenomenon than rodents.

Mode of action of thyroid tumor formation in the male Long-Evans rat administered high doses of alachlor.

Chronic administration of alachlor in the diet at a level of 126 mg/kg/day has previously been shown to cause an increase in benign thyroid follicular cell tumors in male Long-Evans rats. Studies were conducted to elucidate the mechanism of the alachlor-induced thyroid tumors in the male rat by evaluating changes in parameters that are collectively associated with a hormonally mediated mode of action for thyroid neoplasia. Male Long-Evans rats were administered 126 mg alachlor/kg body wt/day via the diet for up to 120 days. One group of animals was removed from alachlor-treated diet after 60 days and received untreated diet for an additional 60 days. Liver and thyroid weights and serum levels of triiodothyronine (T3), thyroxine (T4), and thyroid-stimulating hormone (TSH), as well as hepatic uridine diphosphate glucuronosyl transferase (UDPGT) activity, were determined at 7, 14, 28, 60, and 120 days of treatment. Liver and thyroid weights, hepatic UDPGT activity, and circulating levels of TSH were significantly increased in animals administered alachlor. These increases were seen as early as 7 days after alachlor administration. Circulating levels of T4 in alachlor-treated animals were significantly decreased compared with controls at 7 days, but had returned to control levels by 60 days. T3 levels were elevated at all time points except at 28 days. The changes in TSH and T3 levels, hepatic UDPGT activity, and liver weights were all reversible on elimination of alachlor from the diet. Thyroid weights did not completely return to control levels after removal of alachlor from the diet, although some recovery was evident. The results of this study clearly suggest that alachlor-induced thyroid neoplasia, observed in previous chronic bioassays with alachlor, was associated with increases in circulating TSH levels. Increased metabolism of T4 via hepatic enzymatic conjugation (i.e., T4-UDPGT) appeared to be responsible for the increased TSH levels. These effects were shown to be reversible on cessation of exposure to alachlor. In summary, evidence is presented for a hormonally mediated process for the development of thyroid follicular cell tumors.

Biochemical toxicology and disposition of Therminol 66 heat transfer fluid after inhalation or after dietary administration to male Sprague-Dawley rats.

The objectives of this study were to determine the disposition of Therminol 66 in rats and to determine the effects of this heat-transfer fluid on liver and kidney microsomal drug-metabolizing enzymes. Therminol 66 was administered to male Sprague-Dawley rats at various doses as either a single oral administration at 0, 100, or 300 mg/kg, or as a single 6-h inhalation exposure at 0 or 350 mg/m3. Animals were killed 48 h after gavage or after termination of inhalation exposure. Additional groups of animals were exposed to Therminol 66 via the diet at 0, 100, 500, or 5000 ppm for 14 d, or via repeated inhalation exposure at 0, 25, 250, or 1200 mg/m3 for 6 h/d for 14 d. These exposure scenarios represent approximately equivalent doses of Therminol 66 by the different routes of administration. No change in body weight was observed after acute oral or inhalation exposure, and little change in body weight was observed in animals administered Therminol 66 via the diet except at the highest dose. There was no change in kidney weight, and liver weights were increased only at the higher doses of Therminol 66. The body weight gain of animals exposed to Therminol 66 via inhalation decreased in a dose-dependent manner over the 2-wk exposure period. Results from the disposition study indicated that Therminol 66 did not appear to accumulate in the tissues examined and did not appear to be extensively absorbed after a single oral dose of 300 mg/kg. The whole-body elimination half-life was approximately 14 h and occurred primarily via the feces. There was no significant induction of hepatic aryl hydrocarbon hydroxylase (AHH) activity after single oral or inhalation exposures to Therminol 66. Ethoxycoumarin O-deethylase (ECOD) was significantly induced only in animals exposed to 350 mg/m3 via inhalation. Repeated dietary and inhalation exposures resulted in AHH and ECOD induction only at the highest doses, and the kidney appeared to be less sensitive than the liver. Animals exposed via inhalation demonstrated a greater hepatic inductive effect than did animals exposed via the diet, which may be due to absorption differences.(ABSTRACT TRUNCATED AT 400 WORDS)

Metabolism of glyphosate in Sprague-Dawley rats: tissue distribution, identification, and quantitation of glyphosate-derived materials following a single oral dose.

Five groups of male Sprague-Dawley rats were orally administered a mixture of [14C]- and [12C]-glyphosate (N-phosphonomethylglycine) at a dose level of 10 mg/kg body weight. The majority of radioactivity 2 hr after administration was associated with the gastrointestinal contents and small intestinal tissue. Approximately 35-40% of the administered dose was absorbed from the gastrointestinal tract, and urine and feces were equally important routes of elimination. The total body burden 7 days after administration was approximately 1% of the administered dose and was primarily associated with the bone. Total recovery for this study ranged from 95 to 102% of the administered dose. Metabolic profiles of tissues containing greater than 1% of the administered dose at various times after administration indicated that nearly 100% of the body burden of radioactivity was present as unmetabolized parent glyphosate. A minor component constituting less than 0.1% of the administered dose (less than 0.4 ppm) was observed in colon tissue from animals 2 hr after the administration of glyphosate and was also present in the GI contents of one animal 28 hr after administration of the radiolabel. The retention time for this metabolite was similar, but not identical, to the retention time for AMPA (aminomethylphosphonic acid), the major bacterial metabolite of glyphosate found in soil. Tissue extraction efficiency was always greater than 90% and stability assays indicated no significant effect of storage on either parent glyphosate or AMPA. The results from this study indicate that virtually no toxic metabolites of glyphosate were produced since there was little evidence of metabolism and essentially 100% of the body burden was parent compound with no significant persistence of material.

Age-related changes in dermal absorption of 2,3,7, 8-tetrachlorodibenzo-p-dioxin and 2,3,4,7,8-pentachlorodibenzofuran.

Changes in the structure and function of aged skin may alter percutaneous absorption of environmental compounds such as the halogenated aromatic hydrocarbons. TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) and 2,3,4,7,8-penta-chlorodibenzofuran (4PeCDF) were previously found to be poorly absorbed in 3 months male Fischer 344 rats within 3 days after dermal administration. In order to examine age-related changes in dermal absorption and changes in potential for systemic exposure, the absorption, distribution, and elimination of TCDD and 4PeCDF were examined in male Fischer 344 rats of various ages (TCDD: 10, 36, and 96 week; 4PeCDF: 10, 36, 64, 96, and 120 week). Each compound was applied at a dose of 0.1 mumol/kg and/or 0.04 mumol in 60 microliters acetone to a preclipped region of the back and covered with a stainless-steel perforated cap. Rats were housed in individual metabolism cages for 3 days. Dermal absorption of both compounds was decreased in older age groups compared to 10 week rats. The major tissue depots for both compounds were liver, adipose, skin, and muscle and in comparable age groups, the liver:fat ratio was greater in 4PeCDF-treated animals. Age-related changes in the distribution of the administered dose and the absorbed dose varied with the compound as well as the depot. Elimination of TCDD and 4PeCDF was limited at all ages. Results indicate that percutaneous absorption of these compounds is decreased in older animals, suggesting that systemic bioavailability may be decreased in older organisms following dermal exposure to TCDD or 4PeCDF.

Lack of in vivo DNA binding of mercaptobenzothiazole to selected tissues of the rat.

In this study, the in vivo binding of 14C-labelled 2-mercaptobenzothiazole (MBT) to DNA was investigated. Male and female Fischer 344 rats were gavaged with 375 mg MBT/kg body weight and killed 8 hours later. DNA was extracted from the liver, adrenal glands, pituitary gland, pancreas, and bone marrow and the amount of radioactivity associated with the DNA was determined. Results from this study indicate that MBT does not significantly bind to DNA from any of the tissues examined. CBI values for liver for the 3 methods of purification were -1-3 which are on the low end of the covalent binding index. The CBI values for the other tissues were always less than 1. Other chemicals with similar CBI values include estrone and diethylstilbesterol. Strong hepatocarcinogens such as dimethylnitrosamine and aflatoxin have CBI values ranging from 6000 to greater than 20000.

The biochemical toxicity of perfluorodecanoic acid in the mouse is different from that of 2,3,7,8-tetrachlorodibenzo-p-dioxin.

Perfluorodecanoic acid (PFDA) is an industrial surfactant that has been reported to produce signs of toxicity in rats similar to those due to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In order to characterize the biochemical toxicity of PFDA in the mouse and to determine whether PFDA toxicity is mediated by the Ah locus, congenic female C57BL/6J mice differing only at the Ah locus (normal homozygous responsive Ahb/b, heterozygous responsive Ahb/d, and homozygous nonresponsive Ahd/d) were administered a single oral dose of PFDA. The wild type (Ahb/b) mice were killed 2, 7, 14, or 30 days after administration of 0, 40, 80, 100, 120, or 160 mg PFDA/kg. Mice from the other two congenic strains were killed 30 days after dosing with 0, 40, 80, or 160 mg/kg. PFDA produced a 2.5-fold increase in absolute liver weight, a 5- to 15-fold increase in hepatic fatty acyl Co-A oxidase activity, and a 70% decrease in hepatic ethoxyresorufin O-deethylase (EROD) activity. These effects were dose and time dependent. Total hepatic lipids were increased at an early time point and at the lowest dose. At later time periods and/or higher doses, the lipid concentration was decreased approximately 20% from that of controls. Hepatic protein concentrations were depressed approximately 25% from control levels 30 days after treatment. There was little difference in any of these parameters between responsive (Ahb/b, Ahb/d) and nonresponsive (Ahd/d) mice. These results suggest that the Ah allele has little effect in regulating the toxicity of PFDA in the mouse and that the biochemical response to PFDA in the mouse is markedly different from that of TCDD. Furthermore, the biochemical response to PFDA in the mouse is different from that reported in the rat.

Differential effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on adipose tissue lipoprotein lipase activity in the guinea pig, rat, hamster, rabbit, and mink.

1. Adipose tissue lipoprotein lipase (LPL) activity of several different animal species was determined after i.p. administration of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). 2. TCDD caused a significant reduction in LPL activity and an increase in serum triglyceride concentration in guinea pigs, rabbits, and hamsters but not rats. 3. TCDD increased adipose tissue LPL activity of mink and lowered their serum triglyceride concentration. 4. Results of this study indicate that profound differences occur in lipid metabolism between various species in response to TCDD and these changes do not appear to be related to generalized toxicity such as wasting.

Comparative dermal absorption of 2,3,7,8-tetrachlorodibenzo-p-dioxin and three polychlorinated dibenzofurans.

Polychlorinated dibenzodioxins (PCDDs) and dibenzofurans (PCDFs) are toxic environmental contaminants which have the potential to accumulate in human tissues. In order to examine the potential for systemic exposure following dermal exposure, the absorption, distribution, and elimination of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 2,3,7,8-tetrachlorodibenzofuran (TCDF), 1,2,3,7,8-pentachlorodibenzofuran (1PeCDF), and 2,3,4,7,8-pentachlorodibenzofuran (4PeCDF) were evaluated in male F344 rats. TCDD (0.00015, 0.001, 0.01, 0.1, 0.5, and 1.0 mumol/kg) and the three PCDFs (0.1, 0.5, and 1.0 mumol/kg) were applied to a preclipped region on the back of the rat and covered with a perforated cap. The rats were held in individual metabolism cages for 3 days. In animals administered 0.1 mumol/kg, the absorption of TCDF was greater than that of 4PeCDF, 1PeCDF, and TCDD. Relative absorption (percentage of administered dose) declined with increasing dose while the absolute absorption (microgram/kg) increased nonlinearly with dose. Absorption of TCDF at 0.1 mumol/kg was 48% of the administered dose which was significantly greater than that of the other compounds. At this dose, absorption of 4PeCDF was greater than that of TCDD. Absorption at the higher doses was similar for all four compounds. Maximum relative absorption of TCDD (approximately 40% of the administered dose) was obtained at 0.001 and 0.00015 mumol/kg. Major tissue depots for these four chemicals included liver, adipose, skin, and muscle tissue; however, the liver:fat ratio for 4PeCDF was approximately fourfold higher than that for the other three compounds. When normalized to 100% of dose absorbed, the distribution of 4PeCDF-derived radioactivity in liver and adipose tissue was similar to that previously observed after oral and iv administration. In animals administered 0.1 mumol TCDF or 1PeCDF/kg, 56 and 32% of the respective absorbed dose was excreted as polar metabolites within 3 days. Very little of the absorbed dose of either TCDD (approximately 10%) or 4PeCDF (approximately 2%) was eliminated. Results indicate that the dermal absorption of these compounds is incomplete and that systemic toxicity following acute dermal exposure to levels found in the environment is unlikely.

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on serum insulin and glucose levels in the rabbit.

Serum insulin and glucose were measured in young male rabbits after a single intraperitoneal dose of 1 or 50 micrograms/kg of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Serum insulin levels in the high dosed rabbits were significantly decreased between 15 min and 8 h post treatment, equivalent after 24 h, significantly elevated 48 h post treatment, and they were not different at 10 days post-treatment when compared to weight matched and pair-fed controls. At the low dose, rabbits showed no differences in serum insulin from controls. In the high dose group, serum glucose levels were generally not different between treated and control animals, though there was a transient hyperglycemia 1 h after treatment, and both treated groups became hypoglycemic after ten days. The results indicate that TCDD altered serum insulin levels which were not coupled to changes in serum glucose.

Disposition of 1,2,3,7,8-pentachlorodibenzofuran in the rat.

1,2,3,7,8-Pentachlorodibenzofuran (1PeCDF) is one of several toxic polychlorinated dibenzofurans (PCDFs) which are ubiquitous environmental contaminants. Related in structure and toxicity to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), PCDFs have been detected in municipal and industrial effluents, PCB mixtures, and in a variety of antiseptics and preservative solutions. The objective of this study was to evaluate the distribution and elimination of 1PeCDF in the rat and to compare these parameters with that of 2,3,4,7,8-pentachlorodibenzofuran (4PeCDF) and 2,3,7,8-tetrachlorodibenzofuran (TCDF). After iv administration of 0.1 mumol [3H]1PeCDF/kg, 1PeCDF was rapidly cleared from the blood and distributed to the liver, muscle, skin, and adipose tissue in a manner similar to that for other dibenzofurans. The initial pool sizes of 1PeCDF-derived radioactivity in the liver, muscle, skin, and adipose tissue were 43,35,10, and 7% of the administered dose, respectively. In all cases, loss of radioactivity from these tissues could be described by exponential decay and the initial half-lives for these tissues were 1.36, 0.03, 13, and 1 day, respectively. After redistribution from the muscle, skin, and adipose tissues to the liver, 1PeCDF was metabolized to a polar metabolite(s) and excreted from the body via the bile into the feces. No parent compound was detected in the bile and fecal excretion was the major route of elimination. Most of the radioactivity in the urine was excreted within the first day, after which less than 0.5% of the dose/day was detected. More than half of the administered dose was excreted in the urine and feces within 2 days. The whole-body half-life of related compounds is 4PeCDF much greater than 1PeCDF greater than or equal to TCDF. Therefore, persistence appears to be inversely related to the metabolism of these compounds and metabolism is inhibited by chlorine-substituted carbon atoms adjacent to the oxygen atom in the dibenzofuran ring.

The acute toxicity of 2,3,4,7,8-pentachlorodibenzofuran (4PeCDF) in the male Fischer rat.

Polychlorinated dibenzofurans are ubiquitous environmental pollutants which have great potential for human exposure. To characterize the toxicity of 2,3,4,7,8-pentachlorodibenzofuran (4PeCDF), male F344 rats were administered a single oral dose of 0, 100, 250, 500, 1000, or 2000 micrograms 4PeCDF/kg. A progressive and dose-dependent loss of body weight was evident by 3 days after treatment. Signs of toxicity included piloerection, hair loss, hypoactivity, morbidity, and death. Death occurred as soon as 14 days after treatment and continued throughout the 35-day observation period. The LD50/35 was estimated to be 916 micrograms/kg with a 95% confidence interval of 565-1484 micrograms/kg. Dose-dependent increases were observed in serum cholesterol, triglyceride, and bile acid concentrations and in sorbitol dehydrogenase and aspartate aminotransferase activities. The hematocrit, hemoglobin, mean corpuscular volume, and mean corpuscular hemoglobin concentrations were depressed in a dose-dependent fashion. Hepatic ethoxyresorufin-O-deethylase (EROD) activity was increased in all treatment groups approximately 25 times above that of control animals. Lymphoid depletion in the thymus and spleen was observed in the three highest doses and thymic atrophy was present at all dose levels. Absolute liver weight and the liver:body weight ratio were significantly increased above controls. Hepatotoxicity was dose-dependent and was characterized by lipid accumulation resulting in hepatocytomegaly. Epithelial hyperplasia and focal ulcerations of the forestomach was observed in animals administered 500 micrograms 4PeCDF/kg. Spontaneous cardiomyopathy was exacerbated by treatment with 2000 micrograms/kg. Since 4PeCDF and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) produce a similar spectrum of toxic effects, the biochemical mechanism(s) of toxicity for these chemicals may be similar.

Reduction of adipose tissue lipoprotein lipase activity as a result of in vivo administration of 2,3,7,8-tetrachlorodibenzo-p-dioxin to the guinea pig.

Within 1 hr of intraperitoneal administration of 1 microgram 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)/kg, lipoprotein lipase (LPL) activity was reduced 38% from initial levels in the adipose tissue of the guinea pig. Maximal depression was observed after 2 days and persisted throughout the 10-day observation period. Oral administration of glucose restored LPL activity in TCDD-treated animals after 1 day but only partially after 2 and 5 days, and had no effect after 10 days of exposure. Although initial (2-day) serum insulin levels were depressed, the inability of glucose to restore LPL activity after prolonged exposure was not due to malabsorption of glucose nor to changes in serum thyroxine or insulin concentration. TCDD also inhibited the lipolytic pathway in the adipocyte, but had no effect on hormone sensitive lipase (HSL). Since HSL and LPL are reciprocally regulated, it was concluded that TCDD acts on the adipocyte to uncouple HSL-LPL reciprocity as well as to reduce LPL production.

Toxicity and disposition of 2,3,4,7,8-pentachlorodibenzofuran (4PeCDF) in the rhesus monkey (Macaca mulatta).

The toxicity and disposition of 2,3,4,7,8-pentachlorodibenzofuran (4PeCDF), a ubiquitous and acutely toxic environmental contaminant, was examined in three adult male Rhesus monkeys administered a single iv dose of 34 micrograms (0.1 mumol)/kg. Within 20 min, 4PeCDF was eliminated from the blood and was distributed to the liver, skin, adipose, and muscle tissues. Excretion occurred primarily via the feces with a minimum whole body half-life approximately 38 days. Within 7-14 days after administration, the packed cell volume and serum triglyceride and bile acid concentrations were significantly increased while serum cholesterol, protein, and albumin concentrations were decreased relative to pretreatment levels. Thyroid hormone levels were also altered with an increase in TSH and a decrease in T3 and T4 concentrations. After 28 days, two monkeys began exhibiting alopecia, hyperkeratinization of the toe and finger nails, facial chloracne-like lesions, and loss of body weight. They subsequently died 40 and 48 days after treatment. Similar symptoms of toxicity were observed in the third animal 58 days after 4PeCDF administration, but this animal appeared to fully recover and was administered 4PeCDF orally and [3H]1,2,3,7,8-pentachloro-dibenzofuran (1PeCDF) dermally 238 days after the initial iv dose. In this animal, approximately 2% of an oral dose of [14C]-4PeCDF was absorbed from the stomach and small intestine in 6 hr and was distributed mainly to the muscle and skin and less than 99% of a dermal dose of 1PeCDF remained at the site of application. Pathological findings in the monkeys that died indicated hyperplastic and metaplastic changes in the gastric mucosa, the Meibomian glands of the eyelid, and the ceruminous glands of the ear. Regression of these lesions was present in the surviving animal. Therefore, 4PeCDF produces dioxin-like toxicity in the monkey similar to that reported for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and in the same dose range.

2,3,7,8-Tetrachlorodibenzo-p-dioxin causes an increase in protein kinases associated with epidermal growth factor receptor in the hepatic plasma membrane.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), administered to male rats at a single intraperitoneal (IP) injection dose of 25 micrograms/kg causes down-regulation of epidermal growth factor (EGF) receptor in the plasma membrane of rat liver which starts after two days and continues throughout the experimental period (20 days). Using monoclonal antibody to EGF receptor, it was determined that TCDD-caused EFG receptor down-regulation in the rat liver was accompanied by increased protein kinase activity. Such an increase in the protein kinase activity involves, at least in part, an activation of protein tyrosine kinase. Examination of serum samples from control and treated rats revealed no detectable difference in the level of EGF itself or EGF receptor-reacting substances (eg, hormones and other growth factors). In vivo TCDD caused early eye opening and tooth eruption and poor body weight gain and hair growth in mouse neonates similar to those observed with exogenously administered EGF. The results indicate that such EGF receptor-mediated effect of TCDD has some toxocilogical significance in vivo. Although TCDD causes significant reduction in [125I]-EGF binding in the hepatic plasma membrane in susceptible strains of mice, it has only modest effects in tolerant strains. The results are consistent with the idea that the action of TCDD on the EGF receptor is mediated through the cytosolic/nuclear TCDD receptor, which is known to be regulated by the Ah locus.

Rabbit serum hypertriglyceridemia after administration of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD).

TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) caused a dose-dependent decrease of adipose tissue lipoprotein lipase (LPL) activity and caused a concomitant increase in serum triglyceride concentration in the rabbit 10 d after single ip administration of either 1 or 50 micrograms/kg. Hepatic low-density lipoprotein (LDL) binding was markedly depressed and serum cholesterol concentrations were modestly increased relative to pair-fed control animals. Serum glucose concentrations were significantly lower in the rabbit administered TCDD compared to ad libitum or pair-fed control animals, although little change was observed in serum insulin concentration. Electron microscopic examination of aortic arches 20 d after a single ip administration of 50 micrograms TCDD/kg revealed ruffling, denudation, and sloughing off of the cell surface and the appearance of macrophage-like structures in the intima and media of the endothelial cells. These alterations resemble preatherosclerotic lesions typical in animals with hyperlipidemia. It is proposed that TCDD causes hyperlipidemia in the rabbit through suppression of LPL activity and LDL receptor binding.

Disposition and excretion of 2,3,4,7,8-pentachlorodibenzofuran in the rat.

The disposition of 2,3,4,7,8-pentachlorodibenzofuran (PeCDF), a highly toxic environmental contaminant which accumulates in human tissues, was examined in the male Fischer rat after iv and oral exposure. Greater than 70% of an oral dose of 0.1, 0.5, or 1.0 mumol PeCDF/kg body wt was absorbed by the gastrointestinal system. After either oral or iv administration of 0.1 mumol/kg, the dibenzofuran was rapidly removed from the blood and accumulated in the liver and adipose tissue and to a lesser extent in the skin and muscle. Three days after administration, 70% of the iv dose of PeCDF was found in the liver, 7% in the fat, 1% in the skin, and 0.5% in the muscle. Route of exposure had little effect on tissue distribution. TLC analyses indicated that greater than 99% of the [14C]-PeCDF-derived radioactivity which had accumulated in the liver and adipose tissue was unmetabolized PeCDF which was eliminated very slowly (t1/2 = 193 and 69 days, respectively). The whole body half-life calculated from the daily fecal excretion rate was approximately 64 days. Excretion occurred primarily via the feces. No radioactivity was detected in expired air and less than 0.02% was detected in the urine. TLC analysis of fecal extracts indicated greater than 90% of the [14C]PeCDF-derived radioactivity in the feces was polar metabolites of the parent compound. Pretreatment with 500 micrograms PeCDF/kg body wt caused biliary excretion to nearly double. Treatment of bile with beta-glucuronidase or arylsulfatase had little effect on the chromatographic profile. Therefore, PeCDF was readily absorbed from the gastrointestinal tract, concentrated primarily in the liver, and was slowly eliminated from the body as polar metabolites. The long half-life and high body burden of PeCDF suggest that the toxicity of this chemical may be enhanced due to bioaccumulation upon chronic low-level exposure.

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.

TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) causes increases in protein kinases particularly protein kinase C in the hepatic plasma membrane of the rat and the guinea pig.

To study the cause of TCDD-evoked changes in the functions of plasma membrane constituents TCDD's effects on protein kinase activities in the liver of rats and guinea pigs were investigated. TCDD was found to cause a sharp increase in both c-AMP independent and dependent protein kinase activities in plasma membrane preparations from rat liver within 48 hours from the time of administration. Such effects reached maxima around day 20, and were quite noticeable even 40 days after a single administration of TCDD. As a result of SDS-polyacrylamide gel-electrophoresis (SDS-PAGE) analysis several substrate proteins for these increased protein kinases were observed. Among them are 170 K - 150 K bands, representing EGF receptor protein. TCDD was found to particularly stimulate protein kinase C which is known to influence many enzyme and receptor functions through protein phosphorylation. The possible significance of such an action of TCDD is discussed.

Effects of in vivo-administered 2,3,7,8-tetrachlorodibenzo-p-dioxin on receptor binding of epidermal growth factor in the hepatic plasma membrane of rat, guinea pig, mouse, and hamster.

The effect of in vivo-administered 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on epidermal growth factor (EGF) receptor activity of the rat hepatic plasma membrane was studied. TCDD causes a significant reduction in EGF binding at an early stage of toxicity (day 2) and at very low doses (1 microgram/kg, single i.p., rat). This reduction appears to be due to a decline in the number of receptors. There is a good correlation between levels of decline in EGF binding and loss of body weight among TCDD-treated rats. The reduction in EGF binding occurs at a relatively low dose in the guinea pig (a very sensitive species) and at high doses in the hamster (a tolerant species). Among three mice strains, TCDD (115 micrograms/kg, single i.p.) caused 98% reduction in EGF binding in the sensitive strains (C57BL/6J and CBA/J) but only a 50% reduction in the tolerant strain (AKR/J). To relate the above biochemical changes to in vivo effects, TCDD was postnatally administered (through mother's milk) to mouse neonates. The most prominent toxic manifestations were early eye opening and incisor eruption, loss in body weight gain, and retardation of hair growth. All of these symptoms have been ascribed to EGF effects. TCDD was also found to stimulate phosphorylation of the EGF receptor in the rat hepatic plasma membrane. This phosphorylation effect was observed at day 1 and persisted until the end of the test (day 10). It has long been recognized that agents causing reduction in number of EGF receptors (e.g., phorbol esters) elicit in vivo cellular responses that are similar to those caused by exposure to excess doses of growth factors. Accordingly, a hypothesis has been proposed to ascribe some of the EGF-like effects of TCDD, such as fatty infiltration of the liver and hyperplastic proliferation of gastric epithelia and epidermal cells to its action on the EGF receptor.