John R. Latendresse

Adrenal Gland: Structure, Function, and Mechanisms of Toxicity:

The adrenal gland is one of the most common endocrine organs affected by chemically induced lesions. In the adrenal cortex, lesions are more frequent in the zona fasciculata and reticularis than in the zona glomerulosa. The adrenal cortex produces steroid hormones with a 17-carbon nucleus following a series of hydroxylation reactions that occur in the mitochondria and endoplasmic reticulum. Toxic agents for the adrenal cortex include short-chain aliphatic compounds, lipidosis inducers, amphiphilic compounds, natural and synthetic steroids, and chemicals that affect hydroxylation. Morphologic evaluation of cortical lesions provides insight into the sites of inhibition of steroidogenesis. The adrenal cortex response to injury is varied. Degeneration (vacuolar and granular), necrosis, and hemorrhage are common findings of acute injury. In contrast, chronic reparative processes are typically atrophy, fibrosis, and nodular hyperplasia. Chemically induced proliferative lesions are uncommon in the adrenal cortex. The adrenal medulla contains chromaffin cells (that produce epinephrine, norepinephrine, chromogranin, and neuropeptides) and ganglion cells. Proliferative lesions of the medulla are common in the rat and include diffuse or nodular hyperplasia and benign and malignant pheochromocytoma. Mechanisms of chromaffin cell proliferation in rats include excess growth hormone or prolactin, stimulation of cholinergic nerves, and diet-induced hypercalcemia. There often are species specifi city and age dependence in the development of chemically induced adrenal lesions that should be considered when interpreting toxicity data.

Trichloroethylene, Trichloroacetic Acid, and Dichloroacetic Acid: Do They Affect Fetal Rat Heart Development?

Trichloroethylene (TCE), trichloroacetic acid (TCA), and dichloroacetic acid (DCA) are commonly found as groundwater contaminants in many regions of the United States. Cardiac birth defects in children have been associated with TCE, and laboratory studies with rodents report an increased incidence of fetal cardiac malformations resulting from maternal exposures to TCE, TCA, and DCA. The objective of this study was to orally treat pregnant CDR(CD) Sprague-Dawley rats with large bolus doses of either TCE (500 mg/kg), TCA (300 mg/kg), or DCA (300 mg/kg) once per day on days 6 through 15 of gestation to determine the effectiveness of these materials to induce cardiac defects in the fetus. All-trans retinoic acid (RA) dissolved in soybean oil was used as a positive control. Soybean oil is commonly used as a dosing vehicle for RA teratology studies and was also used in this study as a dosing vehicle for TCE. Water was used as the dosing vehicle for TCA and DCA. Fetal hearts were examined on gestation day (GD) 21 by an initial in situ, cardiovascular stereomicroscope examination, and then followed by a microscopic dissection and examination of the formalin-fixed heart. The doses selected for TCA and DCA resulted in a modest decrease in maternal weight gain during gestation (3% to 8%). The fetal weights on GD 21 in the TCA and DCA treatment groups were decreased 8% and 9%, respectively, compared to the water control group and 21% in the RA treatment group compared to soybean oil control group. The heart malformation incidence for fetuses from the TCE-, TCA-, and DCA-treated dams did not differ from control values on a per fetus or per litter basis. The rate of heart malformations, on a per fetus basis, ranged from 3% to 5% for TCE, TCA, and DCA treatment groups compared to 6.5% and 2.9% for soybean oil and water control groups. The RA treatment group was significantly higher with 33% of the fetuses displaying heart defects. For TCE, TCA, and DCA treatment groups 42% to 60% of the litters contained at least one fetus with a heart malformation, compared to 52% and 37% of the Utters in the soybean oil and water control groups. For the RA treatment group, 11 of 12 litters contained at least one fetus with a heart malformation. Further research is needed to quantify the spontaneous rates of heart defects for vehicle control rats and to explain the disparity between findings in the present study and other reported findings on the fetal cardiac teratogenicity of TCE, TCA, and DCA.

Pathologic Effects of Butylated Triphenyl Phosphate-Based Hydraulic Fluid and Tricresyl Phosphate on the Adrenal Gland, Ovary, and Testis in the Fischer-344 Rat

Triaryl phosphates including tricresyl phosphate (TCP) and butylated triphenyl phosphates (BTPs) are used in the commercial manufacture of plastics, lubricants, and hydraulic fluids. Recent reports implicate these compounds as endocrine and reproductive toxicants in rodents. The objectives of this study were to develop and characterize a rat model to investigate the mechanism(s) of toxicity of triaryl phosphate-based hydraulic fluids and to elucidate potential mechanistic pathways of toxicity through studies of structural/ functional relationships. Groups of male and female rats received daily oral doses of either sesame oil alone or 1.7 g/kg of BTP or 0.4 g/kg TCP in sesame oil vehicle or 2.8 g/kg neat BTP for 20, 40, and 60 days. Light microscopic, morphometric, ultrastructural, and histochemical studies revealed hypertrophy and cholesteryl lipidosis of adrenocortical (both sexes) and ovarian interstitial cells that were progressive with duration of exposure. Minimal degeneration was observed in the adrenal cortex and ovary. TCP caused the most severe lesions in both the adrenal gland and ovary, but the morphologic and histochemical changes produced were similar for both compounds, suggesting a common mechanism of toxicity. Decreased testicular weight and degeneration of seminiferous tubules were detected only in TCP-treated rats. The Fischer-344 rat model using TCP and BTP administered by gavage is a valuable system to study mechanisms of endocrine and reproductive toxicity induced by triaryl phosphate-based hydraulic fluids.

Dissimilar characteristics of N-methyl-N-nitrosourea-initiated foci and tumors promoted by dichloroacetic acid or trichloroacetic acid in the liver of female B6C3F1 mice

Dichloroacetic acid (DCA) and trichloroacetic acid (TCA) are metabolites of the industrial solvent and environmental contaminant trichloroethylene (TCE), as well as contaminants of chlorinated drinking water. Human exposure to these chemicals is of concern as all three have been shown to increase liver tumor incidence in mice. Differences in dose-response curves, progression to cancer, and postexposure regression of lesions suggest that TCA and DCA work through different mechanisms. The purpose of this study was to further characterize the proliferative hepatocellular lesions promoted by TCA and DCA using biomarkers of cell growth, differentiation, and metabolism in liver sections to better delineate the distinctions in the mechanism of the two chloroacetates. Fifteen-day-old female mice were initiated with 25 mg/kg N-methyl-N-nitrosourea. The initiated mice were administered DCA or TCA (20.0 mmol/L) in drinking water from age 49 days until euthanasia at age 413 days. The pathologic assessment showed that the foci of altered hepatocytes and tumors occurring in the animals promoted with DCA were eosinophilic and positive immunohistochemically for TGF-α, c-jun, c-myc, CYP 2E1, CYP 4A1, and glutathione S-transferase-π (GST-π). The DCA lesions also were essentially negative for c-fos and TGF-β, but nontumor hepatocytes were consistently TGF-β-positive. In contrast, tumors promoted by TCA were predominantly basophilic, lacked GST-π, and stained variably; usually, more than 50% of the tumor hepatocytes were essentially negative for the other biomarkers. This study demonstrates some striking differences in certain molecular biomarkers of cell growth, differentiation, and metabolism between DCA and TCA. The results also suggest some potential growth signal transduction pathways that may contribute to the DCA promotion of tumors, further support the premise that these two chloroacetates promote hepatocarcinogenesis in different ways, and provide a rational basis for a similar comparison with TCE. Such a comparison should give some insight as to whether DCA, TCA, or both are playing a significant role in the murine liver carcinogenesis of the parent compound, TCE.

Reproductive Toxicity of Butylated Triphenyl Phosphate and Tricresyl Phosphate Fluids in F344 Rats

The effects of tricresyl phosphate (TCP) and butylated triphenyl phosphate (BTP)-based hydraulic fluid on reproduction were studied in F344 rats using a modification of the National Toxicology Programs Continuous Breeding Protocol. Groups of breeding pairs received single daily oral doses of an equal volume of either 0, 0.6, 1.0 g BTP/kg or 0.4 TCP/kg in sesame oil or 1.7 g neat BTP/kg for up to 135 days. A naive control group allowed to breed, but not dosed or handled daily, demonstrated that daily dosing and handling of the rats had no effect on reproduction. The fertility index and number of litters born were significantly decreased in rats exposed to 1.0 and 1.7 g BTP/kg and 0.4 g TCP/kg. The number of pups per litter was significantly decreased in the TCP group. A crossover mating experiment using 0.4 g TCP/kg/day and 1.0 g BTP/kg/day groups, each mated with vehicle controls, demonstrated that TCP caused 100% infertility in male rats but did not affect reproduction in females. BTP caused a significant decline in reproduction in female rats characterized by low mating and fertility indices, decreased number of litters, and abnormal estrous cycles. Fertility was decreased in the BTP-dosed male rats. Both sexes of rats in the crossover experiment with TCP and BTP had significant decreases in terminal body weights and increases in adrenal gland and liver weights. Only TCP-dosed male rats had significantly decreased testicular and epididymal weights.(ABSTRACT TRUNCATED AT 250 WORDS)

Alterations in alveolar clearance after 4-ipomeanol-induced necrosis of Clara and ciliated cells in the terminal bronchiole of the rat

The administration of 4-ipomeanol, [0, 10 (LD), and 25 (HD) mg/kg, ip], to rats resulted in dose-dependent degeneration and necrosis of the nonciliated (Clara) and ciliated epithelial cells of the terminal bronchioles. More extensive necrosis of the terminal bronchiolar epithelium, with exposure of the basement membrane, was produced in the HD group. Repair of the terminal bronchiolar epithelium was complete within 10 days. Alveolar clearance of 51Cr labeled polystyrene latex microspheres was analyzed through 40 days postinstillation by nonlinear regression for a double exponential model. Alveolar clearance during phase 1 (Days 2 to 6) was delayed and significantly decreased in both the LD and HD groups. Alveolar clearance during phase 2 (Days 10 to 40) was significantly decreased only in the HD group. The decreased alveolar clearance in HD subjects was long term and did not correlate with the return of morphologically normal appearing Clara and ciliated cell structure.

Use of hematoxylin stain to enhance evaluation of heart malformations in the fetal rat

Fresh and formalin-fixed visceral microdissection techniques are valuable tools for studying cardiac teratology in the fetal rat, but are limited by the difficulty experienced in visualizing the minute structures needing to be evaluated. This paper describes a simple and quick staining technique used to enhance the examination procedure. A hematoxylin-saline mixture applied directly on endothelial-lined surfaces of the heart at different stages during microdissection greatly improved visualization of membrane-thin structures such as the pulmonary and aortic semilunar valves, right- and left-side atrioventricular valves, and atrial and ventricular septa. Hematoxylin staining of endocardial surfaces is a useful adjunct to the standard visceral microdissection technique.

Histopathologic effects of JP-4 aviation fuel on fathead minnows (Pimephales promelas)

The objective of this preliminary study using fathead minnows was to determine which tissues and organs are affected by acute exposure to the water soluble fraction (WSF) of JP-4 aviation fuel. The results suggest that the olfactory rosette and possibly the pseudobranch and gill of the fathead minnow are injured by acute exposure to WSF JP-4. These observations are in accordance with results other researchers have found studying the toxic effect of hydrocarbon fuels on teleosts. It is believed that these selected tissues should be examined when utilizing fish for environmental toxicological research involving hydrocarbons.