Effect of the Peroxisome Proliferator, Ammonium Perfluorooctanoate (C8), on Hepatic Aromatase Activity in Adult Male CrI:CD BR (CD) Rats

The incidence of Leydig cell adenomas increases in CD rats fedfor 2 years with the hepatic peroxisome proliferator, ammoniumperfluorooctanoate (C8). Treatment with C8 increased the serumconcentration of estradiol in 2-week gavage studies, and feedingstudies at various time points up to 2 years, and was also accompaniedby increases in liver weight and hepatic ß-oxidationactivity. Since peroxisome proliferators induce both hepaticß-oxidation and specific cytochrome P450 enzymes,C8 may also induce aromatase (cytochrome P450-19A1), the cytochromeP450 mono oxygenase which converts androgens to estrogens. Thishypothesis was investigated in the present study. Adult maleCD rats were dosed daily by gavage for 14 days with 0, 0.2,2, 20, or 40 mg C8/kg body wt. An additional group, the pair-fedcontrol, was fed at a rate matched to the daily consumptionby the 40 mg C8/kg group. Treatment with C8 produced a dose-dependentdecrease in body weight, and increases in absolute and relativeliver weights, and in the protein yield of hepatic microsomes.These C8-induced changes were associated with a 2-fold increasein the serum concentration of estradiol and up to a 16-foldincrease in total hepatic aromatase activity. A significantlinear correlation was established between serum estradiol andtotal hepatic aromatase activity. The absolute weights and thearomatase activity of the testes were not affected by C8. Hepaticperoxisomal ß-oxidation activity and the microsomalconcentration of total cytochrome P450 were also increased byC8. A comparison of estimated EC50 values suggested that theseparameters may be less sensitive to induction by C8 than hepaticaromatase activity. Co-incubation of control liver microsomeswith C8 in the aromatase assay for 2 hr dose dependently reducedthe apparent aromatase activity. This inhibition of aromatasein vitro but increase in vivo was further investigated usingcultured rat hepatocytes. Decreases in aromatase activity werefound after up to 42 hr of treatment with C8, but the enzymeactivity was increased almost 2-fold after 66 hr. The resultsof this study suggest that the increased serum concentrationof estradiol produced by C8 in rats is at least partly due toa direct effect on the liver to increase synthesis of estradiolthrough induction of aromatase cytochrome P450 in the endoplasmicreticulum.     

Developmental Toxicity of Inhaled trans-1,2-Dichloroethylene in the Rat

The developmental toxicity of trans-1,2-dichloroethylene (t-DCE),a component of certain Freon cleaning agents, was examined inpregnant rats. t-DCE was administered by inhalation 6 hr dailyon Days 7–16 of gestation (the day copulation was confirmedwas termed Day 1 of gestation) at exposure levels of 0, 2000,6000, or 12,000 ppm. The offspring were then examined on Day22 of gestation. Overt maternal toxicity was expressed as asignificant reduction in weight gain at 12,000 ppm and in feedconsumption at 6000 and 12,000 ppm. During the exposure period,lacrimation and stained periocular hair, and signs of occularirritation, were observed in all groups. In addition, increasedincidences of alopecia, lethargy, and salivation were observedin the high-dose dams. Significant increases in the mean numberof resorptions per litter were seen in the litters of dams exposedto 6000 and 12,000 ppm of t-DCE; however, these values are withinthe range of historical controls and not considered to be treatmentrelated. The mean combined and female fetal weights were significantlyreduced in the litters of dams exposed to the highest concentration(12,000 ppm) of t-DCE. Marginal effects on feed consumption,unaccompanied by other changes and reflective of the patternseen at higher doses, were seen at 2000 ppm. Thus, marginalmaternal toxicity was seen at 2000 ppm and exposures to 6000ppm t-DCE or higher caused frank maternal toxicity while thefetus was affected only at 12,000 ppm. Therefore, t-DCE is notconsidered to be uniquely toxic to the rat conceptus.     

Investigation of the Effects of Benomyl on Rat Nasal Mucosa

Benomyl [methyl 1-(butylcarbamoyl)-2-benzimidazolecarbamate,CAS Registry No. 17804-35-2] is a widely used agricultural fungicide.Previously, olfactory epithelial lesions were produced followinga 45-day inhalation exposure to 50 and 200 mg/m³ benomyl. Thepresent study, part of a range-finding study for a two-generationreproduction study, was conducted to determine if the previouslyreported effects on the nasal mucosa are the result of systemictoxicity or attributable to the inhalation route of exposure.Groups of 10 7-week-old male Crl:CD BR rats were fed diets containing0, 5000, 10000, or 15000 ppm benomyl for 32 days. Individualbody weights and food consumption were determined weekly andon the last day of the study. After 32 days on test, rats wereeuthanatized by pentobarbital anesthesia and exsanguinationand were examined for gross alterations. The nasal cavity wasprocessed for pathological examination. Mean body weight gainwas statistically significantly decreased during the first weekof treatment and the overall test period (Days 0–32) atthe two highest dose levels. A significant decrease in foodconsumption also was seen during test interval Days 0–7for the two highest dose groups. In addition, statisticallysignificant decreases in food consumption were observed at theDay 7–14 interval for the 15,000 ppm dose group and atthe 21–28 and 28–32 intervals for the two highestdose groups compared with controls. No histopathological lesionswere noted in the nasal epithelium of any of the control orbenomyl-treated rats. These results suggest that the nasal cavityis not a target following dietary administration of benomyland the olfactory epithelial damage reported following inhalationexposure is specific to the route of exposure.     

Histopathology of Acute Toxic Responses in Selected Tissues from Rats Exposed by Inhalation to Methyl Bromide

Histopathology of Acute Toxic Responses in Selected Tissuesfrom Rats Exposed by Inhalation to Methyl Bromide. HURTT, M.E., MORGAN, K. T., AND WORKING, P. K. (1987). Fundam. Appl.Toxicol. 9, 352–365. To determine and characterize thehistological changes induced in selected tissues from the Fischer344 rat by acute inhalation exposure to methyl bromide (MeBr),groups of 10 male rats(11–13 weeks old) were exposed to0, 90, 175, 250, or 325 ppm MeBr 6 hr/day for 5 days. Animalswere anesthetized with phenobarbital then perfusion-fixed 1–2hr after the last exposure or in extremis (325 ppm, 4 days)with Karnovsky's fixative and selected tissues were processedfor light microscopy. With the exception of the nasal passages,tissues were selected on the basis of previous studies withmethyl chloride (MeCl). The nose was examined as part of ongoingresearch of nasal toxicity in this laboratory. The principalclinical signs, confined to the 250 and 325 ppm groups, werediarrhea, hemoglobinuna, and, in a few cases, gait disturbancesand convulsions. A dose-dependent vacuolar degeneration of thezona faseiculata of the adrenal glands, cerebellar granule celldegeneration, and nasal olfactory sensory cell degenerationwere seen in all concentration groups except at 90 ppm. Cerebralcortical degeneration and minor alterations in testicular histologywere seen only in the 325 ppm group. Hepatocellular degenerationwas confined to the 250 and 325 ppm groups. No changes wereseen in the kidneys or epididymides. This study demonstratesthat MeBr has similar target organs to MeCl suggesting thatsimilar mechanisms of toxicity may be operational. However,important differences in the nature of cellular responses tothese chemicals may facilitate studies on their mechanisms ofactions.     

Decreasing Epididymal Sperm Reserves Enhances the Detection of Ethoxyethanol-lnduced Spermatotoxicity

Decreasing Epididymal Sperm Reserves Enhances the Detectionof Ethoxyethanol-lnduced Spermatotoxicity. HURTT, M.E., ANDZENICK, H. (1986). Fundam. Appl. Toxicol, 7, 348-353. Currenttest strategies for assessing male reproductive toxicity maybe inadequate for estimating risk in humans. High levels ofsperm production and existence of large epididymal sperm reservesin most test species may impede the detection of spermatotoxicityat low doses. The current report reflects initial efforts toaddress these issues. An active schedule of copulation was employedto reduce cauda epididymal reserves in the rat. The detectionof spermatotoxicity in this animal relative to its nonmatedcounterpart was then compared following exposure to ethoxyethanol(EE). Adult, male Long-Evans hooded rats were assigned to a"mate" or "non-mate" condition, with the former mated everyother day (3-hr sessions) for 2 weeks prior to and then throughoutthe experiment. After 2 weeks, males from each group were randomlyassigned to receive either 0, 150, or 300 mg/kg (po) of EE,5 days/week for 6 weeks. Males were then sacrificed and organweights, testicular spermatid counts, and cauda epididymal spermcount and sperm morphology were obtained. EE produced a significantreduction in testicular weight and spermatid counts in matedand nonmated animals receiving 300 mg/kg. Significant decreaseswere also noted in epididymal sperm count and percentage normalmorphology. However, these effects were seen in the nonmatedanimals only at 300 mgsol;kg, whereas significant reductionsin both parameters were also obtained at 150 mg/kg in the malesmated bidaily. The data from this study suggest that bidailymatings, by reducing epididymal sperm reserves, can enhancethe detection of spermatotoxicity.     

Developmental Toxicity of Oral and Inhaled Vinyl Acetate in the Rat

Vinyl acetate (VA) is used almost exclusively as an industrialchemical in polymerization, copolymerization, or as a chemicalintermediate. The present studies were undertaken as part ofa collaborative effort by the VA producers of Western Europe,Japan, and the United States to provide animal toxicology datafor risk assessment. To assess the potential of VA causing developmentaltoxicity in rodents, groups of 23 or 24 Crl:CD(SD)BR rats weregiven 0, 200, 1000, or 5000 ppm VA in drinking water or exposed6 hr/day to 0, 50, 200, or 1000 ppm VA vapors on Days 6–15of gestation (both routes approximating 0, 25, 100, or 500 mg/kg/day).Administration of VA in the drinking water produced no evidenceof maternal or developmental toxicity. A significantly loweredwater intake was observed in dams from the 5000 ppm VA groupand probably reflected unpalatability of the VA water solutionat the highest dose level. In the inhalation study, maternaltoxicity was evident by a marked reduction in weight gain ofdams exposed to 1000 but not 200 or 50 ppm VA. Fetal toxicitywas evident by a statistically significant decrease in meanfetal weight and mean crown-rump length in fetuses from the1000-ppm VA group. In addition, there was a statistically significantincrease in the incidence of minor skeletal alterations in fetusesfrom dams exposed to 1000 ppm VA. Delayed ossification was themain skeletal alteration. In summary, pregnant rats were relativelyinsensitive to the effects of VA administered in the drinkingwater at a concentration level as high as 5000 ppm. However,VA did adversely affect both the dam and the conceptus at aninhaled concentration of 1000 ppm, but not at lower exposurelevels. These results indicate that VA is not uniquely toxicto the conceptus. The no-observed-effect level for the dam andconceptus under these experimental conditions was greater than5000 ppm for the drinking water study and was 200 ppm for theinhalation study.     

Evaluation of Spermatogenesis and Sperm Quality in the Rat following Acute Inhalation Exposure to Methyl Bromide

Evaluation of Spermatogenesis and Sperm Quality in the Rat followingAcute Inhalation Exposure to Methyl Bromide. HURTT, M. E., ANDWORKING, P. K. (1988). Fundam Appl Toxicol. 10, 490–498.Methyl chloride (MeCl) and methyl bromide (MeBr) have similartarget organ specificities in the male F-344 rat, and MeCl isa known reproductive toxicant in that species. Recently, bothacute and subchronic inhalation exposures to MeBr were foundto produce varying degrees of testicular alteration (S. L. Eustis,S. B. Haber, R. T. Drew, and R. S. H. Yang, 1986, Toxicologist,6, 54; N. Kato, S. Morinobu, and S. Ishizu, 1986, Ind. Health,24, 87–103; M. E. Hurtt, K. T. Morgan, and P. K. Working,1987, Fundam. Appl. Toxicol, 9, 352–365). The presentstudy examined the reproductive effects of MeBr in the maleF-344 rat. Adult males (11– 13 weeks) were exposed byinhalation to 0 or 200 ppm MeBr 6 hr/day for 5 days (first dayof exposure = Day 1). Ten animals from each group were anesthetizedwith pentobarbital and terminated on Days 1,3,5, and 8. Additionally,five males from each group were killed on Days 6, 10, 17, 24,38, 52, and 73. Plasma testosterone concentration was reducedduring and immediately following exposure (Days 1, 3, 5, and6), but returned to control levels by Day 8. Nonprotein sulfhydryl(NPSH) content of the liver and testis was reduced during exposurebut returned to control levels by Day 8 (3 days postexposure).No other reproductive indices, including testis weight, dailysperm production, cauda epididymal sperm count, sperm morphology,percentage motile sperm, linear sperm velocity, and epididymaland testicular histology, were affected by MeBr exposure atany time point examined. Thus, although MeBr causes a transientdecrease in plasma testosterone and testicular NPSH concentrationsduring acute exposure, it has no lasting effect on sperm qualityor spermatogenesis in the F-344 rat.     

13-Week Inhalation Toxicity Study of Dimethylformamide (DMF) in Cynomolgus Monkeys

This study was conducted to assess the subchronic inhalationtoxicity of dimethylformamide (DMF) in the cynomolgus monkey.Particular attention was paid to the liver since DMF has beenshown to produce liver damage in rodents, dogs, and humans.Groups of three cynomolgus monkeys/sex/group received whole-bodyexposures for 6 hr/day, 5 days/week for 13 weeks to0, 30, 100,or 500 ppm DMF. Evaluations for toxicity included body and organweights, clinical observations, hematology, serum chemistry,urinalysis, and gross and microscopic examinations. Clinicallaboratory evaluations were conducted twice prior to the startof the study at exposure weeks 2, 4, 8, and 12 and at necropsy.Semen, collected from male monkeys three times prior to thestart of the study and weekly during the course of the study,was analyzed for sample volume, sperm count, motility, and morphology.In addition, daily vaginal swabs were obtained from all femalesprior to exposure to determine mean menses cycle length. Althoughthere was a slight trend toward increased cycle length, thistrend could not be definitely attributed to compound exposure.Based on extensive monitoring of the monkeys' clinical condition,semen quantity and quality, and clinical and pathological evaluations,no exposure-related adverse health effects were detected followingexposure to concentrations of DMF ranging from 30 to 500 ppmfor 13 weeks.