Hepatic Aryl Hydrocarbon Hydroxylase and Cytochrome P450 Induction following the Transpulmonary Absorption of TCDD from Intratracheally Instilled Particles

Hepatic Aryl Hydrocarbon Hydroxylase and Cytochrome P450 Inductionfollowing the Transpulmonary Absorption of TCDD from IntratracheallyInstilled Particles. NESSEL, C. S., AMORUSO, M. A., UMBREIT,T. H., AND GALLO, M. A. (1990). Fundam. Appl. Toxicol. 15, 500-509.Inhalation of particles contaminated with 2,3,7,8-tetrachlorodibenzo-p-dioxin(TCDD) will be an increasingly important route of human exposurein light of the increased utilization of municipal waste incinerationand the resultant emission of contaminated materials into theenvironment. The potential for pulmonary absorption of the compoundfrom respirable particles was assessed in the present studyfollowing the intratracheal instillation of TCDD (1) as a contaminantof gallium oxide particles and (2) in a corn oil vehicle. Groupsof five female Sprague-Dawley rats received 0, 0.005, 0.055,0.55, or 5.5 µg/kg TCDD in a single instillation and wereeuthanized 4 days later. Absorption was characterized by enzymeinduction [aryl hydrocarbon hydroxylase (AHH) activity and totalcytochrome P450] and histopathological examination of the liver.Induction of hepatic enzymes was dose-dependent with both treatmentregimes. Up to an 18-fold increase in AHH and an 80% increasein cytochrome P450 were observed in treated animals. Inductionwas slightly higher when animals received TCDD in com oil thanwhen animals received TCDD-contaminated particles and was relativelycomparable to induction following oral exposure. Similar resultswere obtained when animals were treated with particles contaminatedup to 4 weeks prior to instillation. Characteristics of TCDD-inducedhepatotoxicity, including enlarged hepatocytes and fatty infiltration,were apparent in treated rats, but were not present in vehicle-instilledanimals. These results indicate that systemic effects occurfollowing pulmonary exposure to TCDD and that inhalation maybe an important route Of exposure for TCDD.     

Aryl Hydrocarbon Hydroxylase Induction in Adult Rat Hepatocytes in Primary Culture by Several Chlorinated Aromatic Hydrocarbons Including 2,3,7,8-Tetrachlorodibenzo-p-dioxin

Aryl Hydrocarbon Hydroxylase Induction in Adult Rat Hepatocytesin Primary Culture by Several Chlorinated Aromatic HydrocarbonsIncluding 2,3,7,8-Tetrachlorodibenzo-p-dioxin. JANSING, R. L.,AND SHAIN, W. (1985). Fundam. Appl. Toxicol. 5, 713–720.The induction of aryl hydrocarbon hydroxylase (AHH) by Aroclor1254, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and other polychlorinatedorganic compounds was examined in primary cultures of adultrat hepatocytes isolated by a collagenase perfusion technique.Following exposure of fresh hepatocyte cultures to 1 µg/mlAroclor 1254, AHH induction was undetected for 48 hr and thenincreased dramatically up to 96 hr. Cultures maintained in controlmedium for either 24, 48, or 72 hr prior to a 24-hr exposureto Aroclor 1254 displayed significant inducible AHH which wassustained to 96 hr. AHH induction was extremely sensitive totwo planar polyaromatic hydrocarbons, 2,3,7,8-TCDD and 2,3,7,8-tetrachlorodibenzofuran,and the PCB congener 3,4,3',4'-tetrachlorobiphenyl, but insensitiveto 2,6-dichlorodibenzofuran, 2,5,2',5'-tetrachlorobiphenyl,2,4,5,2',4',5'-hexachlorobiphenyl, and hexachlorobenzene. Theinduction of AHH activity in primary cultures of adult rat hepatocytesmay represent a useful bioassay for screening extracts of foodstuffs,biological fluids, or environmental samples for dioxin-likeactivity     

2,3,7,8-Tetrachlorodibenzo-p-dioxin Pretreatment of Female Mice Altered Tissue Distribution but Not Hepatic Metabolism of a Subsequent Dose

2,3,7,8-Tetrachlorodibenzo-p-dioxin Pretreatment of Female MiceAltered Tissue Distribution but Not Hepatic Metabolism of aSubsequent Dose. CURTIS, L. R., JCERKVLIET, N. I., BAECHER-STEPPAN,L., AND CARPENTER, H. M. (1990). Fundam. Appl. Toxicol 14, 523–531.Lipid partitioning of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)inadequately explains its tissue distribution since higher concentrationsoccur in liver than fat except at high doses. This study providesIn vivo evidence that an inducible, saturable system plays apredominant role in disposition of [¹⁴C]TCDD in female miceat doses between 5 and 20 Mg/kg. Female C57BL/6J mice were gavagedwith 0, 5, or 15 µg, TCDD/kg, received a subsequent gavageof 5 or 20 11% [¹⁴C]TCDD after 6 days, and were killed 1 daylater. In mice pretreated with 5 and 15MgTCDD/ kg and subsequentlydosed with 20 µg [¹⁴C]TCDD/kg, liver weight and [¹⁴C]TCDDconcentration increased. Total liver [¹⁴C]TCDD burden increasedabout 50% in both pretreatment groups. Concentrations of [¹⁴C]TCDDin kidney, fat, heart, lung, gastrointestinal tract, but notplasma or splenic lymphocytes, decreased in a reciprocal manner.Alterations in absorption, concentrations of polar metabolitesof [¹⁴C]TCDD in liver, and hepatic lipid content failed to explainthese results. About 97% of hepatic ¹⁴C was hexane extractable.HPLC of this extract indicated [¹⁴C]TCDD was the only significantnonpolar form of radiolabel in liver. In mice pretreated with511% TCDD/kg and subsequently dosed with 5 µg [¹⁴C]TCDD/kg,a more marked pretreatment disposition response was observed.These results are consistent with a predominant role for aninducible, high affinity, low capacity system in whole animalpharmacokinetics of TCDD.     

Opposite Effects of 2,2',4,4',5,5'-Hexachlorobiphenyl and 2,3,7,8-Tetrachlorodibenzo-p-dioxin on the Antibody Response to Sheep Erythrocytes in Mice

The effect that cotreatment with 2,2',4,4',5,5'-hexachlorobiphe-nyl(PCB153) and 2,3,7,8-terrachlorodibenzo-p-dioxin (TCDD) hason the antibody plaque-forming cell (PFC) response to sheepred blood cells (SRBCs) was determined in female B6C3F1 mice.Groups of eight mice per group were given a single oral doseof PCB153 alone (0, 3.58, 35.8, or 358 mg/kg), TCDD alone (0,0.1, 1, or 10 µg/kg), and all possible combinations ofthese doses in corn oil 7 days prior to immunization with SRBCs.Separate groups of mice were given phenobarbital (PB) parenterallyby intraperito-neal injection at a dosage of 160 mg/kg/day for3 days. Four days after intravenous immunization, body, spleen,thymus, and liver weights and the PFC response to SRBCs weredetermined. Exposure to TCDD alone resulted in a dose-relatedsuppression of the PFC response, with significant suppressionat 1 and 10 µg/kg. In contrast, exposure to PCB153 aloneresulted in the enhancement of the PFC response at 358 mg/kg.Combined exposure to 358 mg/ kg PCB153 and TCDD resulted inno change (PCB153 + 0.1 µg/ kg TCDD) or suppression (PCB153+ 1 or 10 µg/kg TCDD) of the PFC response relative toPCB153 alone; however, the PFC response was enhanced (PCB153+ 0.1 µ/kg TCDD), unaffected (PCB153 + 1 µ/kg TCDD),or suppressed (PCB153 + 10 µ/kg TCDD) relative to cornoil controls. PB did not affect the PFC response to SRBCs, despitea 13-fold induction of hepatic pentoxyresorufin O-dealkylase(PROD) activity. These results suggest that PCB153 enhancementof the PFC response is not related to PROD induction and thatit acts as a functional antagonist rather than an aryl hydrocarbonreceptor or dispositional antagonist. By enhancing the PFC responseto SRBCs, PCB153 raises the "setpoint" response level. Consequently,cotreatment with an immunosuppressive dose of TCDD fails tosuppress the PFC response relative to corn oil controls, whileclearly suppressing it relative to the appropriate control,PCB153 alone.     

Bioavailability of Soil-Bound TCDD: Oral Bioavailability in the Rat

Bioavailability of Soil-Bound TCDD: Oral Bioavailability inthe Rat. SHU, H., PAUSTENBACH, D., MURRAY, F. J., MARPLE, L.,BRUNCK, B., DEI ROSSI, D., TEITELBAUM, P. (1988). Fundam ApplToxicol. 10, 648–654. The implications to the public healthof trace amounts of 2,3,7,8-TCDD in the environment are underevaluation by regulatory agencies in the United States and WesternEurope. One major consideration in such evaluations is the contributionto human exposure via ingestion of TCDD-contaminated soil. An80% figure is under consideration by some regulators for estimatedhuman exposure. A contractor for one agency has, in fact, useda value of 1007% bioavailability for estimating human bioavailability.Several studies have investigated the oral bioavailability ofTCDD from contaminated soil in animals. Most have reported estimatesof 25–50%, although one has reported <0.5 and 85%,depending on the source of the contaminated soil. This paperreports an oral bioavailability of approximately 43% in therat dosed with three environmentally contaminated soil samplesfrom Times Beach, Missouri. This figure did not change significantlyover a 500-fold dose range of 2 to 1450 ng TCDD/kg of body weightfor soil contaminated with approximately 2, 30, or 600 ppb ofTCDD. The relevance of animal oral bioavailability data forthe human remains to be evaluated. However, since regulatoryagencies use animal data for extrapolating to humans, the 43%or 25–50% figure would be more accurate than the 80 or100% estimates.     

Corticosterone Modulates Acute Toxicity of 2,3,7,8-Tetrachlorodibenzo-p-1 dioxin (TCDD) in Male Sprague--Dawley Rats

Corticosterone Modulates Acute Toxicity of 2,3,7,8-Tetrachlorodibenzo-p-dioxin(TCDD) in Male Sprague-Dawley Rats.GORSKI, J. R., ROZMAN, T.,GREIM, H., AND ROXMAN, K. (1988). Fundam Appl. Toxicol 11, 494-502.Bilateral adrenalectomy or adrenal demedullation was performedon male Sprague-Dawley rats by established surgical techniques.Subsequently, the dose-response (mortality and mean time todeath) to TCDD was determined in adrenalecto-mized (10, 20,40 µg/kg TCDD ip in 95:5 corn oil: acetone) or demedullated(15, 30, 60 µg/kg TCDD) rats. Adrenalectomy drasticallyincreased mortality and greatly shortened mean time to deathafter dosing with TCDD. More importantly, adrenalectomized TCDD-treatedrats died 3 of hypoglycemic shock without losing much body weightConversely, adrenal demedullation had no effect on mortalityor mean time to death caused by TCDD when compared to non-demedullatedTCDD-treated controls. Thus, it was concluded that the factors)modulating the acute toxicity of TCDD resides in the adrenalcortex and not in the medulla. Administration of corticosterone(25 ngjµl in drinking water) to adrenalectomized ratsreturned the toxicity of TCDD to levels seen in nonadrenalectomizedrats suggesting that this hormone is another key 3 factor (inaddition to the thyroid hormones) in the modulation of the acutetoxicity of TCDD. Corticosterone supplementation (25, 50, or100 µg/) to nonadrenalectomized rats, or to thy- roidectomized-adrenalectomizedrats (25 µg/ml), resulted in no additional beneficialeffect indicating that a factoids) other than thyroid hormonesand corticosterone is also involved in the 14 acute toxicityof TCDD     

Bioavailability of Soil-Bound TCDD: Dermal Bioavailability in the Rat

Bioavailability of Soil-Bound TCDD: Dermal Bioavailability inthe Rat. SHU, H., TEITELBAUM, T., WEBB, A. S., MARPLE, L., BRUNCK,B. DEI ROSSI, D., MURRAY, J., AND PAUSTENBACH, D. (1988). Fundam.Appl. Toxicol. 10, 335-343. 2,3,7,8-Tetrachlorodibenzo-p-dioxin(TCDD), an unwanted by-product formed during the manufactureof hexachlorophene and phenoxyherbicides, has been found asan environmental contaminant in many U.S. and Western Europeansites. This study examines in the rat the degree of dermal absorptionof TCDD bound to soil. Such information would assist regulatoryagencies in evaluating the degree of exposure of humans whocome in contact with TCDD-contaminated soil. Several parameterswhich may influence dermal absorption were studied, includingTCDD dose, duration of contact, presence of crankcase oil asa co-contaminant, and environmentally contaminated vs laboratory-preparedsoil. The dermal penetration of TCDD following 4 hr of contactwith skin was approximately 60% of that following 24 hr of contact(P 0.05). Following 24 hr of contact with the skin, the degreeof dermal uptake of TCDD contaminated soil was approximately1% of the administered dose. Under the conditions of the presentstudy, the degree of uptake does not appear to be influencedto any significant extent by the concentration of TCDD on soil,the presence of crankcase oil as co-contaminants, or by environmentallyvs laboratory-contaminated soil. Although a number of parametersexamined in this study did not significantly influence the degreeof dermal absorption of TCDD in the rat following 24 hr of contactwith the contaminated soil, the unqualified use of the 1% valueto estimate human exposure would overestimate human exposure,since there is general agreement among researchers that ratskin tends to be more permeable than human skin to highly lipid-solublecompounds such as TCDD.     

Relationship of Alterations in Energy Metabolism to Hypophagia in Rats Treated with 2,3,7,8-Tetrachlorodibenzo-p-dioxin

Relationship of Alterations in Energy Metabolism to Hypophagiain Rats Treated with 2,3,7,8-Tetrachlorodibenzo-p-dioxin. POTTER,C. L., MENAHAN, L. A., AND PETERSON, R. E. (1986). Fundam. Appl.Toxicol. 6, 89–97. Efficiency of energy utilization wasevaluated temporally in 2,3,7,8-tetrachlorodibenzo-p-dioxin(TCDD, 50 µg/kg)-treated male Sprague—Dawley rats(275–300 g), their pair-fed counterparts, and a groupwith ad libitum access to ground feed. TCDD treated rats exhibiteda progressive reduction in feed intake and body weight. Theweight loss of vehicle-treated rats, pair-fed to the TCDD-treatedgroup, was comparable to that found in rats receiving TCDD.Following treatment, rats administered TCDD were as efficientin absorbing feed energy from the gut as control rats. Thiswas evidenced by similar relative digestible energy values inTCDD-treated rats, their pair-fed partners, and a group withad libitum access to feed. Equivalent decreases in oxygen consumptionand carbon dioxide production in TCDD-treated rats and theirpair-fed counterparts, relative to rats with ad libitum accessto feed, suggested that the decrease in both of these parametersin TCDD-treated rats was secondary to hypophagia and/or weightloss. Decline of respiratory quotient (RQ) to almost 0.7 inboth TCDD-treated rats and their pair-fed counterparts is indicativeof fat combustion. By Day 17 post-treatment, RQ increased significantlyin the TCDD-treated and pair-fed groups possibly due to a limitationin the availability of lipid stores. Also, TCDD-treated ratsand their pair-fed partners diminished their water intake toa similar extent without reducing urine output. Likewise, urinaryexcretion of both energy and urea was decreased to the sameextent in rats treated with TCDD as it was in their pair-fedcounterparts. However, TCDD-treated rats tended to excrete moreurinary ammonia than their pair-fed partners on Days 10 and16 post-treatment. Thus, TCDD treatment does result in a reductionof caloric intake in the rat, yet efficiency of energy utilizationis preserved.     

Characterization of the Peak Period of Sensitivity for the Induction of Hydronephrosis in C57BL/6N Mice following Exposure to 2,3,7,8-Tetrachlorodibenzo-p-dioxin

Characterization of the Peak Period of Sensitivity for the Inductionof Hydronephrosis in C57BL/6N Mice following Exposure to 2,3,7,8-Tetrachlorodibenzo-p-dioxin.COUTURE, L. A., HARRIS, M. W., AND BIRNBAUM, L. S. (1990). Fundam.Appl. Toxicol. 15, 142–150. 2,3,7,8-Tetrachlorodibenzo-p-dioxin(TCDD) is an extremely potent teratogen in mice. Hydronephrosisand cleft palate are the most sensitive measures of teratogenicityin mice following exposure to TCDD and other structurally relatedpolyhalogenated aromatic hydrocarbons. Despite a relativelylong half-life, investigators have identified a critical windowfor the induction of cleft palate in C57BL/6N mice. To characterizethe critical period for renal teratogenesis, pregnant C57BL/6Nmice were treated once by gavage with 0–24 µg TCDD/kgbody wt on Gestation Day (GD) 6, 8, 10, 12, or 14. All damswere killed on GD 18, and the fetuses were examined for thepresence of hydronephrosis and cleft palate. Maternal liver-to-bodyweight ratios were significantly elevated above controls onall days, while maternal weight gain was unaffected. Fetal mortalitywas increased relative to controls only at 24 µg TCDD/kgon GD 6. There was no significant difference in fetal body weightsbetween control and TCDD-treated fetuses. The incidence of cleftpalate increased in a dose-related fashion from GD 6 to GD 12,and identification of GD 12 as the critical window for inductionof clefting of the hard palate was confirmed. Hydronephrosiswas observed at all dose levels, regardless of exposure day,and the incidence was close to 100% at 3 µg TCDD/kg andhigher doses on GD 12 and earlier. At all doses on GD 14, boththe incidence and severity of hydronephrosis were decreasedrelative to all other days. There was a dose-related increasein the severity of the renal lesion on each day, but betweenGD 6 and 12 severity was constant Thus, while palatal sensitivityto TCDD increased with gestational age between GD 6 and 12,there was no difference among these days in development of hydronephrosis.The data suggest, however, that on GD 14 the urinary tract maybe less sensitive to TCDD.     

Interactive Effects between 2,3,7,8-Tetrachlorodibenzo-p-dioxin and 2,2',4,4',5,5'-Hexachlorobiphenyl in Female B6C3F1 Mice: Tissue Distribution and Tissue-Specific Enzyme Induction

The distribution of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)and 2,2',4,4',5,5'-hexachlorobiphenyl (PCB 153) was studiedin female B6C3F1 mice. Single doses of TCDD alone (0, 0.1, 1,or 10 µg [³H]TCDD/kg), PCB 153 alone (0, 3.58, 35.8, or358 mg [¹⁴C]PCB 153/kg), and all possible combinations of thesedoses were administered in corn oil, po. At 7 days after dosing,11 different tissues were analyzed for radioactivity. When TCDDwas administered alone, TCDD-derived radioactivity distributedto all tissues in a dose-dependent manner, increasing with dosein the liver, while decreasing (as a percentage of the administereddose) in all other tissues. When PCB 153 was administered alone,the PCB 153 concentration was dose-dependently (percentage ofdose) decreased in liver, skin, lung, adrenals, kidney, andblood; no dosimetric effects were observed in the other organs.Coadminis-tration of low doses of both TCDD and PCB 153 resultedin little or no effect on the distribution of either compound.Interactive effects occurred in the pharmacokinetic behaviorof both compounds only at higher doses. For example, the amountof TCDD in the liver was increased by 358 mg PCB 153/kg. Inmost other organs administration of PCB 153 resulted in a dose-dependentdecrease in the TCDD content Coadministration of PCB 153 with10 µg TCDD/kg increased PCB 153 in the liver, but notin other tissues. These results clearly demonstrate that interactiveeffects on pharmacokinetic behavior occur only at high doses.     

The Effect to Pretreatment on the Biliary Excretion of 2,3,7,8-Tetrachlorodibenzo-p-dioxin, 2,3,7,8-Tetrachlorodibenzofuran, and 3,3',4,4'-Tetrachlorobiphenyl in the Rat

The Effect of Pretreatment on the Biliary Excretion of 2,3,7,8-Tetrachlorodibenzo-p-dioxin,2,3,7,8-Tetrachlorodibenzofuran, and 3,3',4,4'-Tetrachlorobiphenylin the rat. MCKINLEY, M. K., KEDDERIS, L. B., and BIRNBAUM,L. S. (1993). Fundam. Appl. Toxicol. 21, 425–432. The laterally halogenated chemicals 2,3,7,8-tetrachlorodibenzofuran(TCDF) and 3,3',4,4'-tetrachlorobiphenyl (TCB) exhibit the samespectrum of toxic effects as 2,3,7,8-tetrachlorodibenzo-p-dioxin(TCDD), the prototype and most toxic member of the halogenatedaromatic hydrocarbon family. Metabolism of all three compoundsappears to be the rate-limiting step for excretion, which isprimarily via the bile into the feces. Therefore, the biliaryelimination of TCDF, TCDD, and TCB was examined as an indirectmeasure of metabolism. Male F344 rats were anesthetized withpentobarbital, the bile duct was cannulated, and 0.1 µmol[³H]TCDD, [¹⁴C]TCDF, or [¹⁴C]TCB/kg body wt was administerediv. Bile was collected for 0–8 hr while the animals werekept under anesthesia. To determine if TCDF was able to induceits own metabolism in vivo, a single dose of 1.0 µmolTCDF/kg was administered to rats by oral gavage 3 days priorto iv injection of 0.1 or 0.3 µmol [¹⁴C]TCDF/kg. Biliaryexcretion and hepatic concentrations of [¹⁴C]TCDF were significantlyincreased in the pretreated animals. These results suggest anautoinduction of TCDF metabolism. Essentially all biliary [¹⁴C]TCDFradioactivity was attributable to metabolites. High-pressureliquid chromatography profiles of biliary radioactivity from0 to 4 hr were qualitatively different between naive and pretreatedrats. To determine if pretreatment with TCDD altered the metabolismof TCDF and vice versa, a single dose of 1.0 µmol TCDF/kgor 0.1 µmol TCDD/kg was administered by oral gavage 3days prior to iv injection of 0.1 µmol [³H]TCDD or [¹⁴C]TCDF/kg,respectively. TCDD pretreatment increased the metabolism andhepatic concentrations of [¹⁴C]TCDF in pretreated rats, whilebiliary elimination and hepatic concentrations of [³H]TCDD wereunaffected by pretreatment with TCDF. In a fourth experiment,the ability of TCDD to alter the metabolism of TCB, a laterallysubstituted polychlorinated biphenyl (PCB), was examined. Pretreatmentwith TCDD increased the metabolism of [¹⁴C]TCB by approximatelytwofold, but no differences in hepatic concentrations were seendue to the rapid elimination of TCB. Rats pretreated with Aroclor1254, a commercial mixture of PCBs, demonstrated significantlyincreased metabolism of [¹⁴C]TCB compared to the naive group.Therefore, under these experimental conditions, induction ofTCDF metabolism occurred in the rat upon pretreatment with eitherTCDD or TCDF at doses which also elicited enhanced hepatic uptake.TCDD and Aroclor 1254 induced the metabolism of TCB. These findingssuggest that repeated exposure to a chemical or to a mixtureof these halogenated aromatic hydrocarbons can result in morerapid metabolism and elimination from the body than followinga single, acute exposure.     

Differential Toxicity of 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) in C57BL/6J Mice Congenic at the Ah Locus

Differential Toxicity of 2,3,7,8-Tetrachlorodibenzo-p-dioxin(TCDD) in C57BL/6J Mice Congenic at the Ah Locus. BIRNBAUM,L. S., MCDONALD, M. M., BLAIR, P.C., CLARK, A. M., AND HARRIS,M. W. (1990). Fundam. Appl. Toxicol 15, 186–200. The acutetoxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was examinedin male C57BL/6J mice differing only at the Ah locus. Wild typemice (Ah^b/b; "b/b") were treated once with 0, 50, 100, 200,300, and 400 eg TCDD/kg po while congenic mice (Ah^d/d; "d/d")received a single dose of 0, 400, 800, 1600,2400, and 3200µgTCDD/kg. Mice were checked daily, weighed twice a week, andthose that survived, killed 35 days post-treatment. The LD5Ovalues were 159 and 3351 µg/kg for b/b and d/d mice, respectively.Mean time to death was 22 days and was independent of dose andgenotype. Decrease in body weight gain was noted in both strains5 days after treatment and occurred at doses 100 µ/kgin b/b mice and 1600 µg/kg in d/d mice. Dose-related increasesin liver weight (both absolute and relative to body weight)and decreases in thymus, spleen, testes, and epididymal fatpad weights were observed at 8–24–fold higher dosesin d/d than in b/b mice. A dose-related increase in segmentedneutrophils was observed in both strains. Serum chemistry valuesindicated that 8–24x greater doses of TCDD were neededto elevate sorbitol dehydrogenase, alanine aminotransferase,and 5'-nucleotidase and to decrease total and esterifled cholesterolin d/d than in b/b mice. Few effects were seen on total bileacids, serum triglycerides, glucose, or nonesterifled cholesterol.In the liver, hepatocellular cytomegaly, fatty change, and bileduct hyperplasia occurred in both strains in a dose-relatedmanner, as did thymic and splenic atrophy. Necrosis of germinalepithelium in the testes and edema in the stomach submu cosaoccurred at acutely toxic doses. These lesions also occurredat doses 8–24x greater in did than in b/b mice. Thus,the spectrum of toxicity is independent of the allele at theAh locus, but the relative dose needed to bring about variousacute responses is approximately 8–24x greater in congenicmice homozygous for the "d" allele than for the wild type animalscarrying two copies of the "b" gene.     

Comparison of the T Cell-Independent Antibody Response of Mice and Rats Exposed to 2,3,7,8-Tetrachlorodibenzo-p-dioxin

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is an environmentalcontaminant that produces adverse effects on the immune systemof experimental animals. In this study, the effect that TCDDhas on the antibody plaque-forming cell (PFC) response to theT cell-independent (TI) antigen trinitrophenyl-lipopolysaccharide(TNP-LPS) was compared in adult female B6C3F1 mice and F344rats. Mice or rats were given a single intraperitoneal injectionof TCDD at doses ranging from 1 to 30 µg/kg, 7 days priorto immunization with TNP-LPS by intravenous injection. Threedays later body, spleen, thymus, and liver weights were measuredand the PFC response to TNP-LPS was determined. Thymus weightswere decreased at 10 and 30 µg TCDD/kg, whereas spleenweights were decreased and liver weights increased in mice dosedat 3,10, and 30 µg/kg. Mice dosed at 10 and 30 µgTCDD/kg had suppressed PFC responses and serum hemagglutinationliters. In rats, thymus weights were decreased and liver weightsincreased at 3, 10, and 30 µg TCDD/kg; however, the PFCresponse and serum hemagglutination titers to TNP-LPS were suppressedonly at 30 µg/kg TCDD. TCDD did not affect splenic lymphocytesubsets evaluated by flow cytometry. These results indicatethat TCDD suppresses the TI antibody response to TNP-LPS inboth B6C3F1 mice and F344 rats, with mice more sensitive tosuppression by TCDD than rats.     

Endometriosis in Rhesus Monkeys (Macaca mulatta) Following Chronic Exposure to 2,3,7,8-Tetrachlorodibenzo-p-dioxin

Endometriosis in Rhesus Monkeys (Macaca mulatta) Following ChronicExposure to 2,3,7,8-Tetrachlorodibenzo-p-dioxin. RIER, S. E.,MARTIN, D. C., BOWMAN, R. E., DMOWSKI, W. P., AND BECKER, J.L. (1993). Fundam. Appl. Toxicol. 21, 433–441. The incidence of the reproductive disease endometriosis wasdetermined in a colony of rhesus monkeys chronically exposedto 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD or dioxin) fora period of 4 years. Ten years after termination of dioxin treatment,the presence of endometriosis was documented by surgical laparoscopyand the severity of disease was assessed. The incidence of endometriosiswas directly correlated with dioxin exposure and the severityof disease was dependent upon the dose administered (p <0.001). Three of 7 animals exposed to 5 ppt dioxin (43%) and5 of 7 animals exposed to 25 ppt dioxin (71%) had moderate tosevere endometriosis. In contrast, the frequency of diseasein the control group was 33%, similar to an overall prevalenceof 30% in 304 rhesus monkeys housed at The Harlow Primate Centerwith no dioxin exposure. This 15-year study indicates that latentfemale reproductive abnormalities may be associated with dioxinexposure in the rhesus. Therefore, the effects of this toxinmay be more diverse than previously recognized.     

Hypophagia-Induced Weight Loss in Mice, Rats, and Guinea Pigs Treated with 2,3,7,8-Tetrachlorodibenzo-p-dioxin

Hypophagia-lnduced Weight Loss in Mice, Rats, and Guinea PigsTreated with 2,3,7,8-Tetrachlorodibenzo-p-dioxin. KELLING, C.K, CHRISTIAN, B. J., INHORN, S. L., AND PETERSON, R. E. (1985).Fundam. Appl. Toxicol. 5, 700–712. C57BL/6 mice treatedwith 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; 360 µ/kg)displayed a significant reduction in feed intake and body weightuntil just before death, when they developed ascites and subcutaneousedema. This caused body weight of the mice that died to suddenlyincrease during the terminal stage of toxicity. TCDD-treatedmice that survived did not develop ascites or edema, and maintaineda body weight that was slightly less than that of pair-fed mice.Cumulative lethality in TCDD-treated mice (69%) was greaterthan that of pair-fed controls (14%). In guinea pigs treatedwith TCDD (2 µ/kg) both the time course and magnitudeof hypophagja were closely associated with weight loss. Pair-fedguinea pigs did not lose quite as much weight as TCDD-treatedanimals because their total body water content was higher. Waterintake in pair-fed guinea pigs was greater than that of TCDD-treatedanimals. The time course and magnitude of lethality tended tobe similar in TCDD-treated guinea pigs (81%) and pair-fed controls(64%). In Fischer F-344 rats treated with TCDD (100 µg/kg)body weight loss was associated with a reduction in both feedand water intake. The time course and magnitude of weight lossin TCDD-treated and pair-fed rats was essentially identical.Lethality was higher in TCDD-treated rats (95%) than pair-fedcontrol animals (48%). Taken together, these findings suggestthat hypophagia is responsible for the loss of adipose and leantissue in mice, guinea pigs, and rats treated with a LD70–95dose of TCDD. Under these dosage conditions, weight loss contributesmore to the lethality of guinea pigs than to that of FischerF-344 rats or C57BL/6 mice     

Use of Model-Based Compartmental Analysis to Study Effects of 2,3,7,8-Tetrachlorodibenzo-p-dioxin on Vitamin A Kinetics in Rats

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a highly toxic,widespread environmental contaminant that has dramatic adverseeffects on the metabolism of vitamin A. We used model-basedcompartmental analysis to investigate sites and quantitativeimpacts of TCDD on vitamin A kinetics in rats given an oralloading dose of TCDD in oil (3.5 µg/kg) followed by weeklymaintenance doses (0.7 µg/kg) or oil only. [³H]retinylin its plasma transport complex (experiment 1) or lymph containingchylomicrons labeled mainly with [³H]retinyl esters (experiment2) were administered iv, and tracer kinetics in plasma, liver,carcass, urine, and feces were measured for up to 42 days. TCDDtreatment caused significant reductions in liver vitamin A levelsand significant changes in tracer kinetics and tracer excretion.A four-compartment model was used to fit tracer data for experiment1; for experiment 2, compartments were added to describe themetabolism of newly absorbed vitamin A. The compartmental modelspredict that TCDD caused a slight delay in plasma clearance(via an increased recycling to plasma), and in liver processing,of chylomicronderived vitamin A. Models for both experimentspredict that TCDD exposure did not affect the fractional uptakeof plasma retinol from the rapidly turning-over extravascularpool, but it doubled the fractional transfer of recycled retholfrom slowly turning-over pools of vitamin A to plasma The residencetime for vitamin A was reduced by 70% in TCDD-treated rats,transfer into urine and feces was tripled, and vitamin A utilizationrates were significantly increased. Since our results do notindicate that retino1 esterification is inhibited, we hypothesizethat some of the significant effects of TCDD on vitamin A metabolismresult from increased catabolism and mobilization of vitaminA from slowly turning-over pools (especially the liver).     

Target Tissue Morphology and Serum Biochemistry following 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) Exposure in a TCDD-Susceptible and a TCDD-Resistant Rat Strain

Target Tissue Morphology and Serum Biochemistry following 2,3,7,8-Tetrachlorodibenzo-p-dioxin(TCDD) Exposure in a TCDD-Susceptible and a TCDD-Resistant RatStrain. POHJANVIRTA, R., KULJU, T., MORSELT, A. F. W., TUOMINEN,R., JUVONEN, R., ROZMAN, K.,MÄNNISTÖ, P., COLLAN,Y., SAINIO, E.-L., AND TUOMISTO, J. (1989). Fundam. Appl. Toxicol.12, 698–712. The mode of action of the highly toxic environmentalcontaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is unknown.It was recently discovered that two strains of rat, Long-Evans(L-E) and Han/Wistar (H/W), differ widely in susceptibilityto TCDD. Employing this strain divergence as a probe, the presentstudy set out to assess the role of various biochemical andmorphological effects in TCDD lethality. In the main experiment,the rats were treated once ip with 0, 5, 50, or (H/W) 500 µg/kgTCDD and killed 1 to 16 days postexposure. Several target organswere evaluated by light microscopy and a number of serum lipidand carbohydrate parameters as well as a few major regulatoryhormones were analyzed. The results demonstrated that most alterationscaused by TCDD were essentially similar in both strains. TCDDreduced circulating thyroxine to a slightly greater extent andmore permanently in the sensitive L-E strain. Moreover, a highlysignificant interaction on thyroid-stimulating hormone was foundamong strain, dose. and time. Serum concentrations of corticosteroneand free fatty acids were increased only in the L-E rats given50 µg/kg TCDD, i.e., at an apparent LDl00 dose level forthis strain. Yet, the most striking interstrain difference wasseen in the liver which was distinctly affected after Day 4in L-E rats given 50 µg/kg TCDD but only marginally affectedin rats from any H/W group. The lesion, while showing no necroticcell changes, was suggestive of plasma membrane damage, possiblyreflecting the production of free radicals. The relation ofthe findings to possible mechanisms of TCDD action is discussed.     

Genetically mediated induction of aryl hydrocarbon hydroxylase activity in mice by polychlorinated dibenzofuran isomers and 2,3,7,8-tetrachlorodibenzo-p-dioxin

Hepatic aryl hydrocarbon hydroxylase (AHH)-inducing potency of toxic polychlorinated aromatic hydrocarbons such as polychlorinated dibenzofurans (PCDFs), 3,4,5,3,4,5-hexachlorobiphenyl (HCB) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was studied in four inbred strains of mice with different phenotypes of Ah locus, i.e., AHH-responsive strains: C57BL/6N and AKR/Ms Qdj, and AHH-nonresponsive strains: DBA/2Cr Slc and Qdj; DDD. Eight individual PCDF isomers or TCDD were administered IP in doses of 30 g/kg; HCB was given in a dose of 120 g/kg. In AHH-nonresponsive strains of mice, only TCDD significantly induced hepatic AHH activity, while in AHH-responsive strains, 2,3,7,8-tetrachlorodibenzofuran(2,3,7,8-TCDF), 1,2,3,7,8-pentachlorodibenzofuran(1,2,3,7,8-PCDF) 2, 3, 4, 7, 8-pentachlorodibenzofuran (2, 3,4, 7, 8-PCDF), and TCDD significantly enhanced the enzyme activity, and the induced AHH activities with the three PCDF isomers were about 30–65% of those of TCDD. These results indicate that AHH responsiveness in mice segregates with the induction of AHH activity by PCDF isomers and may also segregate with the toxic potency of the isomers; i.e., toxic potencies of 2,3,7,8-TCDF, 1,2,3,7,8-PCDF, and 2,3,4,7,8-PCDF in AHH-responsive strains of mice may be much greater than those in AHH-nonresponsive strains of mice. Taking into account both the potent AHH inducibility and the high bioaccumulation of 2,3,7,8-TCDF, 1,2,3,7,8-PCDF, and 2,3,4,7,8-PCDF, these three PCDF isomers should be given greater attention with regard to environmental contamination.A part of this work was presented at the 41st annual meeting of the Japanese Society of Public Health, October 27–29, 1982, Fukuoka, Japan.     

Tissue Distribution and Toxicokinetics of 2,3,7,8-Tetrachlorodibenzo-p-dioxin in Rats after Intravenous Injection

Tissue Distribution and Toxicokinetics of 2,3,7,8-Tetrachlorodibenzo-p-dioxinin Rats after Intravenous Injection. WEBER, L. W. D., ERNST,S. W., STAHL, B. U., AND ROZMAN, K. (1993). Fundam. Appl. Toxicol.21, 523–534. Male Sprague-Dawley rats (240–290 g) received intravenouslya nonlethal (9.25 µg/kg) or a lethal (72.7 µg/kg)dose of ¹⁴C-labeled 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)administered as an emulsion. Animals were euthanized between5 min and 16 days (lethal dose) or 32 days (nonlethal dose)after treatment. Tissue distribution was considered completeafter 24 hr, as by this time radioactivity levels in white adiposetissue had reached a maximum. The highest levels of radioactivitywere found in liver (5% of dose/g tissue), followed by whitefat (1% of dose/g tissue); serum was lowest at 0.01% of dose/mlserum. Relatively high levels of radioactivity were also detectedin most known target organs of TCDD toxicity, e.g., brown fat,adrenals, and thyroid. The pattern of organ distribution ofTCDD was essentially the same after the lethal and the nonlethaldose, but did not follow a simple lipophilicity relationship,as levels in liver were higher than those in white fat, andthose in brain were extremely low. A pool of TCDD in liposomesinitially trapped in lung and spleen was redistributed within24 hr mainly to liver and adipose tissue. Affinity of TCDD tostorage fat seemed to play a more important role as a drivingforce for redistribution than did induction of cytochrome P4501A2. The terminal slope of elimination of TCDD from tissuesindicated a half-life of 16 days after the nonlethal dose. Afterthe lethal dose radioactivity declined in all tissues for 2to 8 days and then increased again, reflecting shrinking tissuevolumes as well as remobilization of TCDD caused by the processof body mass wasting. Distribution data for 17 tissues and serumwere subjected to regression analysis and resulted in up totwo uptake phases and up to three elimination phases for a giventissue. After the nonlethal dose TCDD was mainly excreted viafeces; combined urinary and fecal excretions occurred with abiological half-life of 16.3 ± 3.0 days. Much longerhalf-lives were detected in white fat and skin. After the lethaldose, the fecal excretion of TCDD-derived radioactivity decreasedafter 8 days, and urinary excretion increased starting 12 daysafter dosing. Radioactivity in liver and white fat and the extractableportion in feces was mainly unchanged TCDD, as determined bythin-layer chromatography. Radioactivity in urine indicatedthe presence of a metabolite(s) of TCDD only.     

Role of the Ah Locus in Suppression of Cytotoxic T Lymphocyte Activity by Halogenated Aromatic Hydrocarbons (PCBs and TCDD): Structure-Activity Relationships and Effects in C57BI/6 Mice Congenic at the Ah Locus

Role of the Ah Locus in Suppression of Cytotoxic T LymphocyteActivity by Halogenated Aromatic Hydrocarbons (PCBs and TCDD):Structure-Activity Relationships and Effects in C57B1/6 MiceCongenic at the Ah Locus. KERKVLIET, N. I., BAECHER-STEPPAN,L., SMITH, B. B., YOUNGBERG, J. A., HENDERSON, M. C, AND BUHLER,D. R. (1990). Fundam. Appl. Toxicol. 14, 532–541. Previousstudies have shown that the generation of cytotoxic T lymphocytes(CTL) following allogeneic tumor challenge is suppressed inAh-responsive C57B1/6 mice treated with a single oral dose ofthe toxic, Ah receptor-binding 3,4,5,3',4',5'-hexachlorobiphenyl(HxCB). The present studies have examined the specific roleof the Ah receptor in this immuno-toxic response by utilizingHxCB isomers of known, varied affinity for the Ah receptor aswell as by comparing effects of high-affinity Ah receptor ligands(3,4,5,3',4',5'-HxCB and 2,3,7,8-tetrachlorodibenzo-p-dioxin[TCDD]) on the CTL response of mice that differ only at theAh locus, that is, Ah-responsive (Ah1^*) and Ah-nonresponsive(Ah^dd) congenic C57B1/6 mice. Correlative changes in thymicweight, serum corticosterone (CS) levels, and spleen cellularitywere also measured. The potency of HxCB congeners (3,4,5,3',4',5'-;2,3,4,5,3',4'-; 2,4,5,2',4',5'-) and 2,3,7,8-TCDD to suppressthe CTL response, to reduce spleen cellularity, to cause thymicatrophy, and to elevate serum CS levels was directly correlatedwith the binding affinity of the congener for the Ah receptor.Furthermore, these parameters of immunotoxicity in Ah^*1 C57B1/6mice were significantly more resistant to alterations inducedby either 3,4,5,3',4',5'-HxCB or 2,3,7,8-TCDD as compared toAh1^* C57B1/6 mice. These results strongly support an Ah receptor-dependentimmunotoxic mechanism in suppression of the CTL response followingacute exposure to halogenated aromatic hydrocarbons.