K.D. Nitschke

Three-generation reproduction study of rats given 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the diet.

Abstract A three-generation reproduction study was conducted to evaluate the effects of chronic, low-level ingestion of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Sprague-Dawley rats were maintained continuously on diets providing dose levels of 0, 0.001, 0.01, or 0.1 μg TCDD/kg/day. No significant toxicity was noted in the f0 rats of either sex during the 90 days of TCDD ingestion prior to mating. Significant decreases in fertility and neonatal survival were observed in the f0 generation rats receiving 0.1 μg TCDD/kg/day; these effects precluded continuation of this high dose level in subsequent generations. At 0.01 μg TCDD/kg/day, fertility was significantly decreased in the f1 and f2 but not f0 generations. Other indications of toxicity seen at 0.01 μg TCDD/kg/day included decreases in litter size at birth, gestation survival (proportion of pups born alive), and neonatal survival and growth. Among the rats receiving 0.001 μg TCDD/kg/day, no effect on fertility, litter size at birth, or postnatal body weight was observed in any generation. No consistent effect on neonatal survival was observed at 0.001 μg TCDD/kg/day. In summary, the reproductive capacity of rats ingesting TCDD was clearly affected at dose levels of 0.01 and 0.1 μg TCDD/kg/day, but not at 0.001 μg TCDD/kg/day, through three successive generations.

Methylene chloride: a two-year inhalation toxicity and oncogenicity study in rats and hamsters.

Abstract Methylene Chloride: A Two-Year Inhalation Toxicity and Oncogenicity Study in Rats and Hamsters. BUREK, J. D., NITSCHKE, K. D., BELL, T. J., WACKERLE, D. L., CHILDS, R. C., BEYER, J. E., DITTENBER, D. A., RAMPY, L. W., AND MCKENNA, M. J. (1984). Fundam. Appl. Toxicol. 4, 30–47. A long-term study was conducted to determine the possible chronic toxicity and oncogenicity of methylene chloride. Rats and hamsters were exposed by inhalation to 0, 500, 1500, or 3500 ppm of methylene chloride for 6 hr per day, 5 days a week, for 2 years. No exposure-related cytogenetic effects were present in male or female rats exposed to 500, 1500, or 3500 ppm. Females rats exposed to 3500 ppm had an increased mortality rate while female hamsters exposed to 1500 or 3500 ppm had decreased mortality rates. Carboxyhemoglobin values were elevated in rats and hamsters exposed to 500, 1500, or 3500 ppm with the percentage increase in hamsters greater than in rats. Minimal histopathologic effects were present in the livers of rats exposed to 500, 1500, or 3500 ppm. Decreased amyloidosis was observed in the liver and other organs in hamsters exposed to 500, 1500, or 3500 ppm. While the number of female rats with a benign tumor was not increased, the total number of benign mammary tumors was increased in female rats in an exposure-related manner. This effect was also evident in male rats in the 1500- and 3500-ppm exposure groups. Finally, male rats exposed to 1500 or 3500 ppm had an increased number of sarcomas in the ventral neck region located in or around the salivary glands. Therefore, in this 2-year study, some effects were observed in male and female rats exposed to 500, 1500, or 3500 ppm of methylene chloride. In contrast, hamsters exposed to the same exposure concentrations had less extensive spontaneous geriatric changes, decreased mortality (females), and lacked evidence of definite target organ toxicity.

Teratogenicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin in CF-1 mice☆

Abstract The effect of TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) on the developing embryo and fetus of CF-1 mice has been evaluated. Pregnant CF-1 mice were given TCDD by oral gavage on Days 6 through 15 of gestation at dosages of 0, 0.001, 0.01, 0.1, 1, and 3 μg/kg/day. Little or no maternal toxicity was observed at any dosage. Cleft palate and dilated renal pelvis were found at 3.0 μg/kg/day. Cleft palate was found at 1.0 μg/kg/day. No malformations were found at the intermediate dosages of 0.1 or 0.01 μg/kg/day. Teratogenic effects observed, i.e., cleft palate and dilated renal pelvis, were comparable to those seen in studies utilizing other strains of mice. The incidence of malformations was not statistically significant at the 0.1-μg/kg/day dosage and below. The experimental nonteratogenic dosage for TCDD in the developing embryo and fetus of CF-1 mice was estimated to be 0.1 μg/kg/day.

Comparison of the teratogenic potential of inhaled ethylene glycol monomethyl ether in rats, mice, and rabbits.

Studies to assess the effects of inhaled ethylene glycol monomethyl ether (EGME) on embryonal and fetal development were conducted on groups of Fischer 344 rats, CF-1 mice, and New Zealand White rabbits. Rabbits and rats were exposed to vapor concentrations of 0, 3, 10, or 50 ppm for 6 hr/day on Days 6 through 18, or Days 6 through 15 of gestation, respectively; mice were exposed to 0, 10, or 50 ppm on Days 6 through 15 of gestation. Exposure of pregnant rabbits to 50 ppm produced significant increases in the incidence of malformations, minor variations, and resorptions, as well as a decrease in fetal body weight. Rats and mice exposed to 50 ppm showed no evidence of a teratogenic effect, although indications of slight fetotoxicity were observed in both species. Transient decreases in maternal body weight gain among rats, mice, and rabbits exposed to 50 ppm were the only consistent signs of maternal effects. No significant treatment-related effects on fetal development were observed in any of the species tested at 10 ppm of EGME or below.

In vitro kinetics of styrene and styrene oxide metabolism in rat, mouse, and human

Styrene oxide (SO), a labile metabolite of styrene, is generally accepted as being responsible for any genotoxicity associated with styrene. To better define the hazard associated with styrene, the activity of the enzymes involved in the formation (monooxygenase) and destruction of SO (epoxide hydrolase and glutathione-S-transferase) were measured in the liver and lungs from naive and styrene-exposed male Sprague-Dawley rats and B6C3F1 mice (three daily 6-h inhalation exposures at up to 600 ppm styrene) and Fischer 344 rats (four daily 6-h inhalation exposures at up to 1000 ppm styrene), and in samples of human liver tissue. Additionally, the time course of styrene and SO in the blood was measured following oral administration of 500 mg styrene/kg body weight to naive Fischer rats and rats previously exposed to 1000 ppm styrene. The affinity of hepatic monooxygenase for styrene, as measured by the Michaelis constant (Km), was similar in the rat, mouse, and human. Based on theVmax for monooxygenase activity and the relative liver and body size, the mouse had the greatest capacity and humans the lowest capacity to form SO from styrene. In contrast, human epoxide hydrolase had a greater affinity (i. e., lowerKm) for SO than epoxide hydrolase from rats or mice while the apparent Vmax for epoxide hydrolase was similar in the rat, mouse, and human liver. However, the activity of epoxide hydrolase relative to monooxygenase activity was much greater in the human than in the rodent liver. Hepatic glutathione-S-transferase activity, as indicated by theVmax, was 6- to 33-fold higher than epoxide hydrolase activity. However, the significance of the high glutathione-S-transferase activity is unknown because hydrolysis, rather than conjugation, is the primary pathway for SO detoxification in vivo. Human hepatic glutathione-S-transferase activity was extremely variable between individual human livers and much lower than in rat or mouse liver. Prior exposure to styrene had no effect on monooxygenase activity or on blood styrene levels in rats given a large oral dose of styrene. In contrast, prior exposure to styrene increased hepatic epoxide hydrolase activity 1.6-fold and resulted in lower (0.1>P>0.05) blood SO levels in rats given a large oral dose of styrene. Qualitatively, these data indicate that the mouse has the greatest capacity and the human the lowest capacity to form SO. In addition, human liver should be more effective than rodent liver in hydrolyzing low levels of SO. Quantitative evaluation of the species differences in enzyme levels are being evaluated with the development of a physiologically based pharmacokinetic model for styrene that includes SO.

Methylene chloride: a 2-year inhalation toxicity and oncogenicity study in rats

Male and female Sprague-Dawley rats were exposed to 0, 50, 200, or 500 ppm methylene chloride for 6 hr/day, 5 days/week for 2 years. Blood carboxyhemoglobin levels were elevated in a dose-dependent (less than linear) manner in rats exposed to 50-500 ppm methylene chloride. Histopathologic lesions related to methylene chloride exposure were confined to the liver and mammary tissue of rats. An increased incidence of hepatocellular vacuolization was observed in male and female rats exposed to 500 ppm methylene chloride. Female rats exposed to 500 ppm methylene chloride also had an increased incidence of multinucleated hepatocytes and number of spontaneous benign mammary tumors/tumor-bearing rat (adenomas, fibromas, and fibroadenomas with no progression toward malignancy); the incidence of benign mammary tumors in female rats exposed to 50 or 200 ppm methylene chloride was comparable to historical control values. No increase in the number of any malignant tumor type was observed in rats exposed to concentrations as high as 500 ppm methylene chloride. Additional groups of female rats were exposed to 500 ppm methylene chloride for the first 12 months or the last 12 months of the 24-month study. The response observed in female rats exposed to 500 ppm for the first 12 months was the same as that observed in female rats exposed to 500 ppm for 2 years. Conversely, the response observed in female rats exposed to 500 ppm during the last 12 months of the study was similar to that observed in control animals. Based upon the results of this study, the no-adverse-effect level for chronic inhalation exposure of Sprague-Dawley rats was judged to be 200 ppm methylene chloride.

Embryotoxicity and fetotoxicity of inhaled or ingested vinylidene chloride in rats and rabbits.

Abstract The teratogenic potential of inhaled or ingested vinylidene chloride was evaluated in Sprague-Dawley rats and New Zealand white rabbits. Both species were exposed to the test material by inhalation for 7 hr/day at concentrations of 20 (rats only), 80, or 160 ppm. For the ingestion study, rats were given drinking water containing 200 ppm vinylidene chloride. Rats were given vinylidene chloride from the 6th to the 15th days of gestation and rabbits on the 6th to the 18th days. A teratogenic effect was not seen in rats or rabbits inhaling concentrations of up to 160 ppm vinylidene chloride for 7 hr/day or in rats given drinking water containing 200 ppm vinylidene chloride. Toxicity to both the dams and their developing embryos was observed among the rats inhaling 80 or 160 ppm and among the rabbits inhaling 160 ppm. At exposure levels which caused little or no maternal toxicity (20 ppm in rats and 80 ppm in rabbits), there was no effect on embryonal or fetal development. Among the rats given drinking water containing 200 ppm VDC, there was no evidence of toxicity to the dams or their offspring.

Cleft palates in CF-1 mice after deprivation of water during pregnancy.

Abstract Pregnant CF-1 mice were deprived of water for 48-hr intervals between Days 11 and 15 of gestation. Water deprivation on Days 12 and 13, or on Days 13 and 14 of gestation resulted in a high incidence of cleft palate in the offspring of CF-1 mice. Water deprivation caused a significant decrease in food consumption, which by itself was adequate to cause an increase in the incidence of cleft palate comparable to that observed in mice deprived of water. Therefore, the teratogenic effect resulting from deprivation of water is perhaps medicated through a decrease in the food consumption in CF-1 mice. These results also demonstrate the importance of being attentive to the housing conditions of mice as well as the importance of measuring food and water consumption of mice in teratological studies.

Inhalation toxicity of sulfuryl fluoride in rats and rabbits

The inhalation toxicity of the structural fumigant sulfuryl fluoride (SO2F2) was evaluated in rats and rabbits. Exposures for a preliminary 2-week study were 6 hr/day, 5 days/week, to 0, 100, 300, or 600 ppm SO2F2. Nine of ten rats at 600 ppm died or were moribund between the second and sixth exposures. Extensive kidney lesions were present in all rats exposed to 600 ppm, whereas only minimal renal changes were noted in rats at 300 ppm. Upper and lower respiratory tissues were inflamed in the single rat that survived the 2-week exposure to 600 ppm. Rabbits exposed to 600 ppm SO2F2 were hyperactive and one animal had a convulsion. Exposure to 300 or 600 ppm for 2 weeks resulted in vacuolation and/or malacia in the cerebrum of all rabbits and most of these rabbits also had moderate inflammation of nasal tissues; a few rabbits at 600 ppm had inflammation of the trachea or bronchi. A subsequent 13-week study evaluated rats and rabbits exposed to 0, 30, 100, or 300 ppm SO2F2 (337 ppm TWA for rabbits). Rabbits initially were exposed to a high concentration of 600 ppm; however, convulsions were noted in two animals after nine exposures and the concentration subsequently was reduced to 300 ppm. Vacuolation and/or malacia were observed in the cerebrum of all rabbits at the highest concentration; one rabbit exposed to 100 ppm also had cerebral vacuolation. Rabbits at the highest concentration, as well as one rabbit exposed to 100 ppm, had inflammation of the nasal tissues. Rats exposed to 300 ppm SO2F2 for 13 weeks had mottled incisor teeth, minimal renal effects, pulmonary histiocytosis, inflammation of nasal tissues, and cerebral vacuolation. Also, rats exposed to 100 ppm SO2F2 for 13 weeks had mottled teeth. Fluoride toxicity was suggested by mottled teeth in rats as well as elevation of serum fluoride levels in rats and rabbits exposed to SO2F2 for 13 weeks. Although repeated exposure of rats and rabbits to 100-600 ppm SO2F2 resulted in toxicity of the kidneys (rats only), brain, and respiratory system, no effects were detected in animals exposed to 30 ppm for 13 weeks.

Incapacitation and Treatment of Rats Exposed to a Lethal Dose of Sulfuryl Fluoride

Rats exposed to 4000 ppm sulfuryl fluoride (VIKANE gas fumigant, SO2F2) were incapacitated within 45 min and died within several hours after exposure. Exposure to higher concentrations resulted in a shorter time to incapacitation and death occurred within minutes. Treatment with calcium gluconate before exposure to 4000 ppm SO2F2 for 45 min resulted in 80% survival. However, calcium gluconate did not alleviate SO2F2-induced convulsions. Administration of phenobarbital before or after exposure to 4000 ppm SO2F2 for 45 min effectively reduced the frequency and severity of convulsions and resulted in survival of all animals. Exposure of rats to 10,000 ppm SO2F2 for 15 min followed by treatment with phenobarbital reduced the frequency of convulsions and delayed death, but did not prevent death. Diazepam was less effective than phenobarbital while diphenylhydantoin had no beneficial effect and, in fact, made the convulsions more severe and longer in duration. The results of this study indicate that phenobarbital was effective in ameliorating the acute toxic effects of an overexposure to SO2F2 in rats.