Toxicologic Pathology

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.

Knowledge of patterns of lesion distribution can provide insight into the relative roles played by regional tissue dose and local tissue susceptibility in toxic responses to xenobiotics in the nose and assist assessment of potential human risk. A consistent approach is needed for recording lesion distribution patterns in the complex nasal airways of rats and mice. The present work provides a series of diagrams of the nasal passages of the Fischer-344 rat and B6C3F1 mouse, designed for mapping nasal lesions. The diagrams present each of the major cross-sectional airway profiles, provide adequate space for nasal mucosal lesion recording, and are suitable for duplication in a commercial photocopier. Sagittal diagrams are also provided to permit transfer oflesion location data observed in transverse sections onto the long axis of the nose. The distribution of lesions induced by a selected range of xenobiotics is presented. Approaches to application of the diagrams and interpretation of results obtained are discussed in relation to factors responsible for lesion distribution in the nose and their relevance to interspecies extrapolation. A modified approach to anatomical classification of the ethmoturbinates of the rodent is also presented.

We attempted to establish an animal model for renal pelvic and ureteral carcinomas. In Experiment 1, Sprague-Dawley (SD)/cShi and SD/gShi strains of male and female rats and Fischer-344 (F-344) DuCrj and LEW/Crj strains of male rats, 6 wk old, were given 0.05% N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) for 12 wk and then treated without chemical for 10 wk. Only the SD/cShi strain. which has spontaneous hydronephrosis and hydroureter, showed high incidences of carcinomas in the renal pelvises and ureters, whereas the other strains did not. In Experiment 2, SD/cShi and Crj:CD strains of male and female rats. 6 wk old, were exposed to 0.05% BBN for 20 wk. Both sexes of SD/cShi rats had high incidences of bilateral carcinomas in renal pelvises and/or ureters associated with urinary bladder carcinomas, whereas both sexes of Crj:CD rats had urinary bladder tumors only. These results indicate that the SD/cShi strain may be a suitable animal for the induction of renal pelvic and ureteral carcinomas that appear to depend on urine stagnation secondary to hydronephrosis and hydroureter.

Histological and ultrastructural changes were investigated in lung, liver, and heart of mice given interleukin-2 (IL-2), either alone or in combination with other cytokines. IL-2 induced a vascular leak syndrome (VLS) of a moderate degree with infiltration of lymphoid cells, moderate endothelial damage. mild hepatic parenchymal damage, and minimal myocardial alterations. Interferon-a (IFN-α) produced infiltration mainly of monocytes/macrophages in liver and heart; endothelial cell damage was absent in lung and heart and minimal in liver. Interleukin-1α (IL-1α) caused an increased number of neutrophils in liver and lung; VLS and parenchymal cell and endothelial damage were not found. The VLS and the cellular damage caused by the combination of IL-2 and IFN were much more severe than those produced by IL-2 alone. In animals treated with IL-2, IFN-α, and IL-1α, VLS was minimal and parenchymal and endothelial cell damage were less severe than after IL-2 alone or IL-2 plus IFN-α. Taken together, these observations show that IL-1α reduces ultrastructural changes produced by IL-2 and IFN-α. This reduction may be clinically useful in the treatment of neoplasms.

Effects of in vivo exposure to the synthetic polyprenoic acid, 3,7,11,15-tetramethyl-2,4,6,10,14-hexadecapentaenoic acid (E-5166) using 6 groups of rats treated simultaneously or individually with phenobarbital (PB) or with p,p'-dichlorodiphenyltrichloroethane (DDT) on the gap junctions of hepatocytes were examined by freeze-fracture analysis. Experimental groups were as follows: Group 1, basal diet alone; Group 2, E-5166 treatment; Group 3, 0.05% PB diet; Group 4, PB diet and E-5166 treatment; Group 5, 0.05% DDT diet; and Group 6, DDT diet and E-5166 treatment. E-5166 was given by gavage (40 mg/kg body weight, 3 times/ wk). Experimental diets and E-5166 were started at 6 wk of age. All animals were sacrificed at 2 wk after the start of the experiments. The density of gap junctions in PB- or DDT-treated rats was higher and that in E-5166-treated rats was lower than that in controls. The density of gap junctions in rats given PB and E-5166-/or DDT and E-5166-treated rats was slightly increased when compared to controls. The average area of gap junctions in DDT- or PB and E-5166-treated rats was significantly smaller than in controls (p < 0.005), although no statistical differences were found among PB-, E-5166-, and DDT and E-5166-treated rats and controls. Unit area of gap junctions to membrane area in rats given E-5166, DDT, or PB and E-5166 was lower than in controls. However, the unit area of gap junctions in DDT and E-5166-treated rats did not differ from in controls. These findings indicate that E-5166 synergistically reduces the gap junctions with PB but has an inhibitory effect on DDT that decreases the gap junctions.

Groups of 10 male Hartley guinea pigs were exposed to 3.0, 2.0, 1.0, or 0.1% (v/v) 1,1-Dichloro-2,2,2-trifluoroethane (HCFC-123) or 1.0% (v/v) halothane by inhalation for 4 hr. A sixth group of 10 guinea pigs received only air. All animals were sacrificed 48 hr postexposure. Gross and histopathologic examination of the liver, heart, and kidney and routine hematology and clinical chemistry analyses [including isocitrate dehydrogenase (ICDH)] were done on all guinea pigs. Lesions related to HCFC-123 and halothane exposure were limited to the liver and included centrolobular vacuolar (fatty) change, multifocal random degeneration and necrosis, and centrolobular degeneration and necrosis. These lesions were observed in 90-100% of the exposed animals and were absent in the air-only controls. There was significant individual animal variation in susceptibility to both HCFC-123 and halothane, resulting in a spectrum of histologic lesions and clinical chemistry values within each exposure group. Alanine aminotransferase, aspartate aminotransferase, and ICDH were the most significant predictors of hepatocellular damage. Similarities in the response between halothane and HCFC-123 in this guinea pig model suggests that humans susceptible to halothane-induced hepatitis may be susceptible to HCFC-123 by a common mechanism of toxicity.

Mouse renal cell tumors (RCTs) were induced in male CBA mice by 5 subcutaneous injections of 8 mg 1,2-dimethylhydrazine (DMH)/kg body weight once a week. After a lag period of 2 yr kidneys were removed, and serial cryostat sections of the kidneys were histochemically analyzed for the following parameters: glycogen content, basophilia, and the activities of glycogen synthase (SYN), glycogen phosphorylase (PHO), glucose-6-phosphatase (G6Pase), glucose-6-phosphate dehydrogenase (G6PDH), hexokinase (HK), pyruvate kinase (PK), lactate dehydrogenase (LDH), malic enzyme (ME), succinate dehydrogenase (SDH), alkaline phosphatase (ALPase) and γ-glutamyltranspeptidase (GGT). RCTs displayed the same histochemical profile irrespective of their size and growth pattern. In comparison with the normal kidney epithelium, the neoplastic cells exhibited elevated activities of enzymes for glycolysis (HK, PK, LDH) and the pentose phosphate pathway (G6PDH), while negative G6Pase and low SDH activity were observed in these cells. The majority of RCTs showed high PHO activity and weak staining for SYN. Activities of ALPase and GGT were negative in most of the RCTs. Markedly enlarged cells with atypical nuclei were detected in some advanced RCTs. Higher activities of glycolytic and mitochondrial enzymes and G6PDH were found in these enlarged cells than in other tumor cells. Tubular preneoplastic lesions were similar to neoplastic lesions in morphological and histochemical characteristics. The present study revealed that a markedly elevated capacity for glycolysis and the pentose phosphate pathway occurred in RCTs in mice. A similar histochemical pattern in the few preneoplastic tubular lesions observed suggests that these metabolic aberrations emerge early during carcinogenesis, but additional studies on early stages of renal carcinogenesis are needed to substantiate this assumption.

Spontaneous hemangioendothelial cell hyperplasia of the heart was found in 12 of 3,910 female B6C3F ₁(C57BL/6CrSlc x C3H/HeSlc) mice that were used in chronic toxicology and carcinogenicity studies at the An-Pyo Center (Shizuoka, Japan). Histologically, the lesions appeared diffusely in the heart as widened vascular spaces lined by a single layer of endothelial-like cells with enlarged nuclei between the muscle fibers. These proliferating cells were identified as hemangioendothelial cells by immunohistochemical techniques, lectin-binding patterns, and electron microscopic examination. Morphological differences were found between this spontaneous hemangioendothelial hyperplasia and that which has been reported as a preneoplastic lesion of hemangiosarcoma. Spontaneous hemangioendothelial cell hyperplasia of the heart occurs rarely; however, it was interesting that spontaneous hemangioendothelial cell hyperplasia was only seen in females with adenocarcinomas of the mammary gland.

Aroclor-1254 (Ar-1254) and dichlorodiphenyltrichloroethane (DDT) were compared to phenobarbital (PB) for their ability to promote hepatocellular proliferative lesions to hepatocellular adenomas and carcinomas and to hepatoblastomas in D2B6F1 male mice initiated with N-nitrosodiethylamine (NDEA). Hepatocellular neoplasms developed in all mice given NDEA and were more numerous in mice fed promoters. Multiplicities decreased in the order Ar-1254 > PB > DDT, indicating that Ar-1254 was more potent than either PB or DDT at the dosage levels used. PB was the most effective of the 3 agents in stimulating the evolution of hepatocellular neoplasms to hepatoblastoma. The incidence of hepatoblastomas in the NDEA.PB group was 72% but was only 27% in NDEA-initiated, DDT-promoted mice and 33% in low-dose and only 9% in high-dose Ar-1254-promoted mice. In contrast, lesions resembling benign and malignant cholangiocellular neoplasms were frequently found within hepatocellular tumors in Ar-1254-promoted mice but not in mice fed PB or DDT, either alone or after NDEA. Some cystic glandular structures in Ar-1254-promoted mice contained mucous cells, argentaffin cells, and Paneth cells and thus constituted intestinal metaplasia. Hepatoblastoma and intestinal metaplasia/cholangiocellular tumor morphology appear to constitute different patterns of genetic programming induced by certain promoters in expanding clones of initiated hepatocytes, on favorable genetic backgrounds such as that of D2B6F1 male mice.

Myelolipomas occurring simultaneously in the adrenal and liver were found in a 2.7-yr-old, bred female common marmoset (Callithrix jacchus). The animal bore single adrenal and multiple hepatic myelolipomas. The adrenal myelolipoma consisted of mature adipose cells and focal collections of normal hematopoietic elements and was unencapsulated. In the liver, the myelolipomas, which were partially encapsulated, included a large amount of hematopoietic tissue and adipose cells that resembled normal bone marrow cells in various ways. Additionally, one of the multiple nodules contained several bony spicules with osteoblasts. Furthermore, there were invasive figures of hematopoietic cells, such as myeloblasts, erythroblasts, and megakaryocytes, in the hepatic sinusoids around the lesions. Thus, this case has some unusual morphological characteristics and is the first report, to our knowledge, on multiorganic myelolipomas in common marmosets.