Cancer chemopreventive efficacy of a chalcone, isoliquiritigenin (ISO), was assessed on the rat hepatocarcinogenesis associated with fibrosis caused by a choline-deficient, L-amino acid-defined (CDAA) diet, using glutathione S-transferase placental form (GST-P)-positive preneoplastic foci as the end point lesion, and on the rat superficial bladder carcinogenesis initiated by N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN). ISO, when given to F344 male rats at the doses of 12.5, 50, 100, and 200 ppm in the CDAA diet for 15 weeks, did not significantly affect the number and size of GST-P-positive foci and the % liver area occupied by the foci. However, it tended to decrease the grade of liver fibrosis. ISO, which was given in a basal diet at doses of 12.5, 25, and 100 ppm for 12 weeks to F344 male rats initiated by 0.05% BBN in drinking water for 12 weeks, also exhibited no clear chemopreventive effects on the development of urinary bladder tumors. Nevertheless, it tended to decrease the multiplicity of nodulo-papillary hyperplasia but not of transitional cell carcinoma (TCC), and differentiation grade of TCCs. The results indicate that ISO at least at the doses used in the present study, possesses no clear cancer chemopreventive efficacy on rat hepatocarcinogenesis caused by a CDAA diet and on rat superficial urinary bladder carcinogenesis by BBN.

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Oligodendrocytes and myelin in the corpus callosum of black tremor and normal hamsters aged 3, 7, and 14 weeks were ultrastructurally examined to determine myelination index (ratio of myelin thickness/diameter of axon), percentage of naked axons, and proportions of oligodendroglial subtypes (light, medium and dark). The mutant hamsters were remarkably hypomyelinated, with a low myelination index and a high proportion of naked axons, but the proportions of the three subtypes in the mutants were comparable with those of normal hamsters at 3 weeks of age, suggesting the low myelinogenetic activity of oligodendrocytes in the mutant. Furthermore, oligodendrocytes of the mutant hamsters appeared to mature earlier than those of age-matched controls, since the proportions of dark inactive oligodendrocytes at 7 and 14 weeks of ages were higher than those of the control hamsters.

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Theophylline, a phosphodiesterase inhibitor, is known to induce enlargement of the salivary glands. This enlargement has been thought to be associated with enhanced cellular levels of cyclic AMP as a result of inhibition of phosphodiesterase. In the present study, to clarify the sequential changes in salivary glands following the administration of theophylline, a four-day repeated dose study and a single dose study were carried out in male rats, and the parotid and submaxillary glands were examined histologically. The results from the glands were almost identical. In the repeated dose study, the increased organ weight just before the last administration was reduced at 4 hr and then restored at 8 hr after the last administration. In the single dose study, a slight but significant increase in organ weight was observed at 24 hr after administration, following a transient reduction. Changes in acinus histology were consistent with these organ weight changes. Acinar cells were loaded with excessive secretory granules just before the last administration in the repeated dose study. After the last administration, granules were reduced at 4 hr, and then reaccumulated at 8 hr. Such time-course variability in the number of secretory granules corresponded well to clinically observed salivation. These results suggest that inhibition of phosphodiesterase is primarily involved in the discharge of secretory granules, and that the salivary hypertrophy previously reported is an adaptive response caused by the repeated stimulation of saliva secretion.

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1-β-D-Arabinofuranosylcytosine (Ara-C), a cytidine analogue, inhibits DNA synthesis and is used for the treatment of myelogenous leukemia. On the other hand, it is also known that Ara-C has a teratogenic effect. In the present study, pregnant rats were treated with 250 mg/kg of Ara-C on day 13 of gestation, and fetal tissues and placentae were collected from 3 to 48 hours after treatment to examine the sequential histopathological changes induced by prenatal Ara-C treatment. The weights of fetuses and placentae and the thickness of the placental labyrinth zone were significantly reduced at 48 hours after treatment. In the fetal tissues such as central nervous system and mesenchymes and in the placental labyrinth zone, the number of pyknotic cells stained positively by the TUNEL method increased markedly after treatment with Ara-C. In most fetal tissues, the number began to increase from 3 to 6 hours and peaked at 9 to 12 hours after treatment. In the placental labyrinth zone, the number peaked at 6 hours after treatment. Electron microscopical features of the pyknotic cells consisted with the ultrastructural characteristics of apoptotic cells. In various fetal organs and placentae, abnormal induction of apoptosis is suggested to induce growth inhibition and have a certain relation to the malformations. Moreover, abnormal induction of apoptosis would have a deep connection with the cytotoxic effect of Ara-C which may affect proliferative cells in fetuses and placentae developing rapidly.

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Susceptibility to the promoting effects of sodium L-ascorbate (Na-AsA) on the development of pelvis and urinary bladder tumors in male and female SD/cShi rats, featuring spontaneous hydronephrosis, was investigated. Rats received 0.05% N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) in their drinking water for 4 weeks and subsequently given basal diet with or without a 5% Na-AsA supplement for 32 weeks. Histopathological examination revealed the promoting effect of Na-AsA on not only the development of urinary bladder tumors but also renal pelvic tumors in the animals of both sexes in this two-stage carcinogenesis experiment, the effect being more prominent in males. Administration of either BBN or Na-AsA alone also induced papillomas and papillary or nodular hyperplasia of renal pelvis or urinary bladder, respectively, in male but not female rats. However, the 5-bromo-2'-deoxyuridine-labeling index of urothelium in the pelvis and bladder increased slightly in male rats and significantly in female rats given Na-AsA alone for 8 weeks. N-butyl-N-(3-carboxypropyl)nitrosamine, which is a metabolite of BBN and proximate carcinogen, was found more in the urine of urinary bladder than that of renal pelvis. These results indicate that the urothelium of the renal pelvis and urinary bladder in SD/cShi rats is susceptible to promoting effects of Na-AsA in the present two stage model urinary tract carcinogenesis, with the urinary bladder of male rats as the most sensitive organ.

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Pregnant rats were administered flutamide at a daily dose of 10 mg/kg from gestation day 12 (GD12) to GD21, or 30 mg/kg on 2 successive days in the period from GD14 to 21 (GD14-15, GD16-17, GD18-19, and GD20-21). The effects on ventral prostate in male offspring were examined by RT-PCR on postnatal day 1 (PND1) for the 10 mg/kg group, and by receptor binding assay at PND76 or 78 and morphologically at PND7, 14, and 21 for all groups. RT-PCR demonstrated increase in mRNAs for the androgen receptor (AR) and the keratinocyte growth factor (KGF), but not transforming growth factor (TGF)-beta1 and beta2, the epidermal growth factor receptor (EGFR), and the vascular endothelial growth factor (VEGF). Prostate tissue showed a consistent reduction in the number of main ducts and ductal branchpoints, as well as the complexity of the terminal ductal network in the 10 mg/kg group, and the 30 mg/kg GD16-17 and GD18-19 groups. The effect on ductal architecture was severest with the 10 mg/kg regimen. The organ weights at PND76 or 78 were reduced in all flutamide treated groups. The value for the 10 mg/kg group was lowest, while the 30 mg/kg GD14-15 and GD20-21 groups were least affected. Receptor binding assays exhibited no significantly difference regarding maximum binding capacity (Bmax) and dissociation constant (Kd) between the control and any flutamide treated group. In conclusion, prenatal exposure to flutamide caused increased AR and KGF mRNA expression just after exposure, and an irreversible morphological change that is severest when the prostatic buds are developing in the ventral prostate. However the receptor binding capacity later in life was not affected.

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Intestinal type alkaline phosphatase (IALP) mRNA-specific cRNA probe was synthesized by non-cloning method for in situ hybridization (ISH) analysis. Total RNA was extracted from small intestine of male F344 rat. Reverse transcription-polymerase chain reaction (RT-PCR) was performed for IALP gene. T7 promoter adapter was directly added to IALP cDNA-specific PCR product by ligation reaction. IALP cDNA accompanied by T7 promoter adapter was then amplified by PCR using IALP gene-specific primer in combination with T7 promoter adapter primer sequence and used as a template of cRNA probe. Digoxigenin-labeled cRNA was synthesized by in vitro transcription method using RNA polimerase. ISH was performed using formalin-fixed, paraffin embedded small intestine tissue. As a result, the cytoplasm of mucosal epithelial cells was successfully stained with this cRNA probe. The present procedure is quite simple and a rapid way to prepare the cRNA probe and would be very useful for ISH analysis of formalin-fixed paraffin embedded tissue sample.

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Although Sprague-Dawley rats are most frequently used in numerous toxicological studies, there are only few reports dealing with the spontaneous occurrence of ectopic sebaceous glands in the oral tissue (Fordyce's granules) so far. Hence, to clarify the morphological characteristics of Fordyce's granules with their incidence in this strain, 110 male and 110 female CD(SD)IGS rats aged 19 to 112 weeks were examined microscopically. Fordyce's granules were identified in the upper molar gingiva, and most of them were located around the first and third molars. The granules consisted of one or more sebaceous glands without hairs and hair follicles, and had some lobules comprising acini with adipose vacuoles. The short excretory duct was lined with stratified squamous cells and drained to the gingival surface. The sebum constituting disintegrated acinar cells in the duct was also noted. In some cases, the granules were accompanied by cystic dilatation in the ducts and/or inflammatory reactions, without neoplastic or pre-neoplastic lesions. The overall incidence (9.1%) of Fordyce's granules in males was higher than that (0.9%) in females, and it somewhat increased with ascending ages, demonstrating sex- or age-specific difference. Based on the above findings, the nature of Fordyce's granules seen in CD(SD)IGS rats was similar to those of other strains and humans.

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Effects of maternal exposure to nonylphenol (NP) on growth and development of the female reproductive system and uterine carcinogenesis in Donryu rats were investigated. Dams were administered 0, 0.1, 10 or 100 mg/kg NP daily by gavage from gestation day 2 up to the day before weaning. The treatment with NP did not influence the reproductive ability of the dams. In their female offspring, there were no significant effects on the reproductive system such as uterine growth and development, vaginal opening, and hormonal secretion until puberty. Moreover, NP had no apparent influence on estrous cyclicity after maturation, morphology of the reproductive organs, and uterine carcinogenesis initiated by N-ethyl-N'-nitro-N-nitrosoguanidine. Regarding biotransfer of NP, the chemical was detected at low levels in the milk of dams given NP at 10 and 100 mg/kg/day in a dose-dependent manner, but not in the serum. In the offspring also, NP was not detected in the liver in any of the treated groups. Taken together, maternal exposure of rats to 0.1 - 100 mg/kg NP did not have any effects on the female reproductive system of offspring from puberty up to 15 months of age. NP at 10 mg/kg and 100 mg/kg doses was transferred from dams to their offspring via the milk, but with these doses no accumulate in the liver of offspring was evident.

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CB6F1-TgHras2 mice (rasH2 mice) are considered to be a promising model for short-term carcinogenicity studies. This study was conducted to obtain background data for rasH2 mice, and the data were compared with those for CB6F1-nonTgHras2 mice (nonTg mice). This study revealed that the survival ratio of rasH2 mice was 92% for males and 97% for females and the body weights were lower than those of nonTg mice while plasma AST and ALT levels in rasH2 males were higher than those in nonTg males. On histopathology, higher incidences of the following neoplastic lesions were observed in rasH2 mice: hemangiosarcoma in the spleen, bronchiolo-alveolar adenoma/carcinoma in the lung in both sexes, hepatocellular adenoma in the liver, squamous cell papilloma in the forestomach, and adenocarcinoma in the Harderian glands in males. For non-neoplastic lesions in rasH2 mice, the incidences of dilatation of the glands in the glandular stomach, inflammatory cell infiltration in the submucosa of the glandular stomach, myopathy of the tongue and skeletal muscle, eosinophilic bodies in the respiratory epithelium of the nasal cavity, and basophilic renal tubules in the kidney were higher than those in nonTg mice for both sexes. In this study, various tumors and characteristic non-neoplastic lesions were observed in the rasH2 mice; furthermore, the incidence of hepatocelluler adenoma was higher than that in previous reports. These findings will be useful when evaluating the short-term carcinogenicity study using rasH2 mice.

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A spontaneous granular cell tumor was found in the cecum of a 9-month-old female beagle. The tumor cells were distributed widely and multifocally throughout the cecal submucosa and basal portion of the lamina propria, but never formed macroscopic nodular lesions. Partial infiltration into the muscular layer was observed, but neither metastasis nor invasion to other tissues was detected. The tumor cells were oval to polygonal and were characterized by abundant eosinophilic and PAS positive granules in the cytoplasm. Immunohistochemically, the tumor cells were positive for vimentin, but negative for S-100 protein, NSE, desmin, lysozyme, and α1-antichymotripsin. Ultrastructurally, various numbers of lysosomal bodies containing lamellar and membranous structures were present in the cytoplasm. Neither basal lamina nor cell-to-cell communications were observed. This is the first report of canine granular cell tumor arising in the cecum and is of interest as it occurred in a young laboratory beagle.

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Urinary bladder rhabdomyosarcoma from a 9-month-old female Labrador retriever dog is described. Grossly reddish black multiple friable multinodular masses filled the lumen of the urinary bladder. Histologically, the masses consisted of round, fusiform and polygonal cells with various consistencies. There were also some strap-like cells with chained nuclei and cross-striations. It was diagnosed as a urinary bladder rhabdomyosarcoma. Additionally, the diagnosis was confirmed by immunoreactivity of tumor cells for canine myoglobin.

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An ovarian tumor was observed in a five-year-old, female cynomolgus monkey. The tumor occupied the entire left ovary, and normal ovarian tissue was not observed. The tumor was composed of endodermal, mesodermal and ectodermal cell components with well-differentiated tooth and bone formation. The present tumor was diagnosed as a mature ovarian teratoma.

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7,12-Dimethybenz[a]anthracene (DMBA) is an indirect carcinogen that enlists the host metabolism to produce its ultimate carcinogenic form, and it is known that this metabolism is conducted by cytochrome P450 1A1 and/or 1B1(CYP1). However, the time course of expression of rat liver CYP1 following DMBA administration has been unclear. After DMBA administration (100 mg/kg b.w.) to SD rats, expression of liver CYP1mRNA was observed at 6 hr and gradually increased with time to peak on days 1-2, but then disappeared on day 5. Expression of CYP1A1 protein was first observed at 12 hr, peaked on day 2 and decreased on day 5, while expression of CYP1B1 protein was first observed on day 2 and decreased on day 5. Expression levels of CYP1A1mRNA and protein were higher than those of CYP1B1. The present study suggests that high CYP1 enzyme production and DMBA metabolism might occur in rat liver substantially on day 2 following DMBA administration.

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