An F1-extended one-generation reproductive toxicity study in Crl:CD(SD) rats with 2,4-dichlorophenoxyacetic acid.

2,4-Dichlorophenoxyacetic acid (2,4-D) was assessed for systemic toxicity, reproductive toxicity, developmental neurotoxicity (DNT), developmental immunotoxicity (DIT), and endocrine toxicity. CD rats (27/sex/dose) were exposed to 0, 100, 300, 600 (female), or 800 (male) ppm 2,4-D in diet. Nonlinear toxicokinetic behavior was shown at high doses; the renal clearance saturation threshold for 2,4-D was exceeded markedly in females and slightly exceeded in males. Exposure was 4 weeks premating, 7 weeks postmating for P1 males and through lactation for P1 females. F1 offspring were examined for survival and development, and at weaning, pups were divided in cohorts, by sex and dose, and by systemic toxicity (10), DNT (10), DIT (20), and reproductive toxicity (≥ 23). Remaining weanlings were evaluated for systemic toxicity and neuropathology (10-12). Body weight decreased during lactation in high-dose P1 females and in F1 pups. Kidney was the primary target organ, with slight degeneration of proximal convoluted tubules observed in high-dose P1 males and in high-dose F1 males and females. A slight intergenerational difference in kidney toxicity was attributed to increased intake of 2,4-D in F1 offspring. Decreased weanling testes weights and delayed preputial separation in F1 males were attributed to decreased body weights. Endocrine-related effects were limited to slight thyroid hormone changes and adaptive histopathology in high-dose GD 17 dams seen only at a nonlinear toxicokinetic dose. 2,4-D did not cause reproductive toxicity, DNT, or DIT. The "No Observed Adverse Effect Level" for systemic toxicity was 300 ppm in both males (16.6 mg/kg/day) and females (20.6 mg/kg/day), which is approximately 6700- to 93 000-fold higher than that reported for 2,4-D exposures in human biomonitoring studies.

Developmental sensitivity to the induction of great vessel malformations by N-(2-aminoethyl)ethanolamine.

N-(2-Aminoethyl)ethanolamine (AEEA) induced malformations of the great vessels in the offspring of rats treated during gestation and early lactation (Schneider et al., 2012. Birth Defects Res B Dev Reprod Toxicol [in press]). The aim of this study was to determine if in utero exposure alone was sufficient to induce these malformations or whether a peri-postnatal exposure or physiological component was required. Three groups of five time-mated female Wistar Han rats were administered AEEA (250 mg/kg/day) by gavage from gestation day (GD) 6 to GD 19 (groups 1 and 2) or from GD 6 to postnatal day 3 (group 3). Animals were euthanized on GD 21 (group 1) or postnatal day 4 (groups 2 and 3), and the hearts of the offspring were examined for changes to the great vessels. The incidence of malformations in group 1 was 91.1%, and primarily consisted of high aortic arch and abnormal carotid course. One fetus had an aortic aneurysm. All fetuses in groups 2 and 3 were malformed, primarily exhibiting abnormal carotid course and aneurysms, which mainly affected the aorta, ductus arteriosus, and pulmonary trunk. The incidence of high aortic arch was lower relative to group 1. Aneurysms were more prevalent in group 3 compared to group 2. These findings indicate that exposure to AEEA during gestation alone was sufficient to induce malformations of the great vessels and aneurysms, which may be triggered by physiological changes that occur during or after birth, but that the critical period of susceptibility to AEEA-induced aneurysms in the rat extends beyond gestation into the early postnatal period.

Role of iodine in the toxicity of diiodomethyl-p-tolylsulfone (DIMPTS) in rats: ADME.

Metabolism of diiodomethyl-p-tolylsulfone (DIMPTS) was investigated in rats to determine the role of iodide in its toxicity. Fischer 344 (F-344) (5 or 50mg/kg) or Sprague Dawley (SD) (5mg/kg) rats were gavaged with (14)C-DIMPTS or dermally applied with 5mg/kg (F-344 only) and absorption, distribution, metabolism and excretion (ADME) determined. Additional experiments were conducted with its deiodinated analog (methyl-p-tolylsulfone, MPTS) in female F-344 rats (20mg/kg) for comparison. Orally administered (14)C-DIMPTS was rapidly absorbed and eliminated in urine (92%). The elimination t(½) was 1-4h. Dermally applied (14)C-DIMPTS remained undetectable in plasma with bioavailability ≈ 7%, only 5-7% of the dose was recovered in urine. DIMPTS liberated one or both of its iodine atoms upon absorption. The rate of elimination of the liberated iodide from the systemic circulation was 2- to 3-fold slower in SD than F-344 rats, which resulted in higher bioavailability of iodide to SD rats. DIMPTS was primarily oxidized at the benzylic methyl moiety forming the corresponding benzoic acid. Glutathione conjugation on the sulfonyl methyl group, via displacement of I(-) was also observed. Overall 67-80% of the total iodine atoms were metabolically released from DIMPTS. The MPTS was rapidly absorbed from the GI tract, metabolized and eliminated in urine similar to that of DIMPTS. These data were compared to iodide toxicokinetic results of a reproductive toxicity study for DIMPTS (80 mg/kg/day) and MPTS (32 mg/kg/day), where DIMPTS was toxic to dams and pups, while MPTS caused no toxicity. These data show that the liberated iodide is the ultimate toxicant of DIMPTS, which is readily transported to pups through milk, while the methyltolylsulfone backbone structure (MPTS) of DIMPTS is relatively nontoxic.

Role of iodine in diiodomethyl-p-tolylsulfone induced reproductive toxicity in rats: proposed mode of action.

The biocide diiodomethyl-p-tolylsulfone (DIMPTS) caused dystocia, decreased neonatal survival and hypothyroidism in rat reproduction studies resembling the effects caused by iodine. One molecule of DIMPTS contains two iodine moieties that are hydrolyzed upon ingestion and systemically absorbed, suggesting iodine toxicity as a probable mode of action for the effects observed in rats. This study compared the effects induced by DIMPTS and an equimolar concentration of its de-iodinated analogue, methyl-p-tolylsulfone (MPTS). Groups of 20 female Sprague Dawley rats were fed diets supplying 80 mg DIMPTS/kg/day, 32 mg MPTS/kg/day or control feed from prior to breeding through lactation and gonadal function, mating performance, conception, gestation, parturition, lactation, survival, growth and development of pups evaluated through postnatal day 7. Serum thyroid hormones and iodine levels in milk and sera were also determined. Females given DIMPTS had increased incidence of vulvar discharge and dystocia, decreased litter size, decreased body weights and feed consumption, increased thyroid weights, thyroid follicular cell hypertrophy with decreased colloid, decreased triidothyronine, and increased thyroid stimulating hormone levels. DIMPTS pups had decreased neonatal survival and body weights. These effects were associated with elevated levels of iodine in milk and sera. In contrast, MPTS did not produce similar effects in adult females or their offspring. These data support the hypothesis that the dystocia, altered neonatal survival and hypothyroidism following repeated dietary administration of DIMPTS were due to excessive iodine released from DIMPTS during absorption and metabolism.

Society of Toxicologic Pathology position paper: organ weight recommendations for toxicology studies.

The evaluation of organ weights in toxicology studies is an integral component in the assessment of pharmaceuticals, chemicals, and medical devices. The Society of Toxicologic Pathology (STP) has created recommendations for weighing organs in GLP general toxicology studies lasting from 7 days to 1 year. The STP recommends that liver, heart, kidneys, brain, testes, and adrenal glands be weighed in all multidose general toxicology studies. Thyroid gland and pituitary gland weights are recommended for all species except mice. Spleen and thymus should be weighed in rodent studies and may be weighed in non-rodent studies. Weighing of reproductive organs is most valuable in sexually mature animals. Variability in age, sexual maturity, and stage of cycle in non-rodents and reproductive senescence in female rodents may complicate or limit interpretation of reproductive organ weights. The STP recommends that testes of all species be weighed in multidose general toxicology studies. Epididymides and prostate should be weighed in rat studies and may be weighed on a case-by-case basis in non-rodent and mouse studies. Weighing of other organs including female reproductive organs should be considered on a case-by-case basis. Organ weights are not recommended for any carcinogenicity studies including the alternative mouse bioassays. Regardless of the study type or organs evaluated, organ weight changes must be evaluated within the context of the compound class, mechanism of action, and the entire data set for that study.

The Tg.AC Transgenic Mouse as a Screening Tool for Anticarcinogens: Broccoli Juice Protected Against 12-O-Tetradecanoylphorbol-13-Acetate (TPA) But Not Benzo[a]Pyrene (B[a]P)-Induced Skin Tumors.

The Tg.AC mouse model was used to assess the utility of a short-term screening assay to evaluate compounds with suspected carcinogenic/anticarcinogenic activity. Crude broccoli juice (BROC) was evaluated for antitumorigenic effects against 12-Otetradecanoylphorbol-13-acetate (TPA) and benzo[a]pyrene (B[a]P)-induced tumors. Groups of female mice were dosed three times a week (200 muL/mouse) with one of the following: acetone vehicle control, TPA (2.5 mug/mouse), B[a]P (250 mug/mouse), BROC/TPA, or BROC/B[a]P. BROC (200 muL, 1:1 acetone) was dermally administered 1 h prior to the administration of either TPA or B[a]P to evaluate anticarcinogenic activity. Papilloma numbers were recorded weekly for each mouse. Following 13 weeks of treatment, samples from the dermal test site from all mice were examined histologically. B[a]P-induced tumors were evaluated for transgene expression by RT-PCR and immunohistochemically for cyclin D1 and p53 proteins. TPA and B[a]P induced tumors in all surviving mice. BROC showed effective antitumorigenic activity against TPA but not B[a]P. Tumor development was distinct between TPA (small, benign papillomas) and B[a]P (large, ulcerated, squamous cell carcinomas). The transgene v-Ha-ras, cyclin D1, and p53 proteins were highly expressed in B[a]P tumors where progression to malignancy was rapid (< 13 weeks). The effects induced by B[a]P appeared to cooperate with transgene expression to enhance conversion to malignancy and could serve as a phenotypic indicator for genotoxic versus nongenotoxic carcinogens. The model distinguished differences in tumor response for carcinogenic and anticarcinogenic agents. The Tg.AC mouse model offers a potentially useful screen for identifying new anticarcinogenic agents and directing future mechanistic evaluations.

Recommended tissue list for histopathologic examination in repeat-dose toxicity and carcinogenicity studies: a proposal of the Society of Toxicologic Pathology (STP).

The Executive Committee of the Society of Toxicologic Pathology (STP) appointed an ad hoc task force to devise and recommend a standard list of tissues to be evaluated histopathologically in repeat-dose toxicity and carcinogenicity studies that are used to support the registration of new pharmaceutical products. The recommended tissue list is intended to be a minimum core list that can be used for all types of repeat-dose toxicity and carcinogenicity studies, regardless of route of administration, species or strain of mammalian laboratory animal, duration, or class of drug to be tested. The resulting recommendations of the task force, presented here, were subsequently reviewed by the STP membership and endorsed by the STP Executive Committee.