Early vascular changes in the heart were observed in 52-week-old rats given excessive doses of vitamin D₂ and cholesterol for 5 days. The body weight of treated rats was decreased. The serum cholesterol and calcium levels were significantly elevated (P<0.01) in the treated group. Microscopically, in the treated rat atrophy and necrosis of the myocardial fibers were seen in the myocardium accompanied with macrophage infiltrating into the necrotizing areas. The coronary arteries were irregular shape and arterial wall showed thin and fragmentary appearance. Microcorrosion casts revealed wavy courses of the right and left coronary arteries and spiral courses of the capillaries in the affected cardiovasculature. Endothelial impressions on the arterial surface were weakened. These results domonstrated that not only myocardial degeneration but also abnormal cardiovascular structures occurred within a short period after administration of excessive doses of vitamin D₂ and cholesterol. This implied that the abnormal courses of the cardiovasculature may in part contribute to the reduction of cardiac functions.

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The effects of estrogen on liver sinusoidal cells were investigated immunohistochemically and morphometrically. Male AKR/n mice were orally administered 30mg/kg/day of mestranol suspended in sesame oil for 7 days. Some of the mice were splenectomized before mestranol administration. Relative liver weight significantly increased from day l to day 7 in mestranol-administered mice with or without spleen. Relative spleen weight also significantly increased from day 3 to day 5 in these mice. Histologically, cellular increase was observed in both the sinusoid of the liver and the red pulp of the spleen. Almost all cells that increased in the sinusoid of the liver were positive for F4/80, and cells positive for PCNA also increased. These changes were observed in mestranol-administered mice with or without spleen. In the morphometrical study, the peak number of F4/80 positive cells in the liver was observed from day 5 and day 7 in normal and splenectomized mice administered mentranol, respectively. On day 5 and day 7, the number of F4/80 positive cells observed in the mestranol-administered splenectomized mice was significantly higher than that in mestranol-administered normal mice. The peak number of PCNA positive cells in the liver was observed on day 3 in mestranol-administered splenectomized mice, and on day 5 in mestranol-administered normal mice. The number of PCNA positive cells was significantly higher on day 3 in splenectomized rather than normal mice administered mestranol. These results suggest that the liver disturbances observed in women taking estrogen for a long time may be closely related to local proliferation of Kupffer cells in the sinusoid.

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Mustard Gas (MG), a potent carcinogen, has been elucidated to induce lung cancer, by means of long-term clinical observation in former workers in MG factory on Okunojima, Hiroshima prefecture. On this rare occasion, an opportunity to observe pulmonary carcinogenetic process in human was given. Histological research on bronchial epithelium revealed the presence of preneoplastic lesions including dysplasia and various types of hyperplastic or metaplastic lesions. Furthermore, recent progresses in research techniques on genetic abnormality have been applied to analyze specific genetic changes related to exposure to MG, in order to clarify multistep carcinogenesis of bronchial epithelium. The knowledge obtained through research of former workers in MG factory should prove useful to understand chemical carcinogenesis in humans.

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An individual pulmonary tumor induced by a chemical carcinogen often shows complex histologic features associated with different cellular and structural atypias which suggest tumor progression in the tumorigenesis. Analysis of their cellular characteristics is important to elucidate tumor cell origin and the relationship between tumor histology and cell origin. For this purpose, we examined lectin binding affinities in various types of pulmonary tumor induced by 3-methylcholanthrene (3-MC) in mice.
3-MC induced proliferative lesions of the A/J mouse lung were classified into four basic histological types : hyperplasia (HP), alveolar adenoma (AA), papillary adenoma (PA), and papillary adenocarcinoma (PC), and their combined type : PA in AA, PC in AA, PC in PA, etc. Frequency of PA, PC, and combined type tumors increased as a function of time after carcinogen administration. Binding affinities of cells in normal respiratory epithelia and 3-MC induced proliferative lesions to four perox-idase-conjugated lectins, Maclura pomifera agglutinin (MPA), Arachis hypogea agglutinin (PNA), Ricinus communis agglutinin (RCA), and wheat germ agglutinin (WGA) were examined. In normal epithelia, both Clara cells and type II alveolar cells showed strong affinity to MPA and WGA. Cells of HP and AA showed fairly strong affinity to all four lectins. PA and PC cells, however, lost their binding affinity to MPA, PNA, and RCA, but not to WGA. A distinct difference in lectin binding affinity between HP/AA and PA/PC was clearly shown in this experiment and the evidence obtained was supportive of progressive development of mouse pulmonary tumors.

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Lesions induced in the respiratory organs of male F344 rats by methanol engine exhaust were investigated histopathologically and electron-microscopically. In experiment 1, animals were exposed to the exhaust by the inhalation route for 8 hours/day for 7 days, and then sacrificed immediately or after recovery periods of 1, 4, and 12 weeks. The concentrations of the main components in the exhaust were carbon monoxide 98ppm, formaldehyde 6ppm, nitrogen monoxide 41ppm, nitrogen dioxide 12ppm, and methanol 16ppm. Histopathologically, lesions were found in the nasal cavity and the lungs of exposed animals at the end of the 7-day exposure. The lesions in the nasal cavity were erosion and/or hyperplasias/squamous cell metaplasias of the respiratory epithelium lining the naso- and/or maxillo-turbinate and neutrophil-infiltration into the submucosa. In the lungs, decrease or loss of cilia in the bronchial and/or bronchiolar epithelium, and reduction of apical blebs of Clara cells on the terminal/respiratory bronchioli were observed. These lesions became less prominent time-dependently after the cessation of exposure, and both nasal cavity and lung lesions were histologically no longer detectable in all animals after 4 week and I week recovery periods, respectively.
In experiment 2, rats inhaled high, medium or low concentration exhaust for 8 hours/day, 6days/week for 4, 8, or 12 wk, and were sacrificed at the end of each exposure period. The main components in the high-concentration exhaust were carbon monoxide 89.8ppm, formaldehyde 2.3ppm, nitrogen monoxide 21.8ppm, nitrogen dioxide 1.1ppm, and methanol 8.1ppm. Dilutions of about I in 5 and I in 25 (NOx ratio) were used for the medium- and low-concentration groups. Exposure-related histopathological changes were only found in the high-concentration group, and included hyperplasias/squamous cell metaplasias of the respiratory epithelium in the nasal cavity. The lesions were detected from the 4-week time point, with their incidences and extents increasing time-dependently. In the medium- and low-concentration groups, no histological changes caused by the exhaust-exposure were seen in any organs, including the nasal cavity and lungs at any timepoint.
From these results, it is concluded that methanol engine exhaust-induced lesions in the nasal cavity and lungs of rats are reversible, and that the no effect concentration of included formaldehyde is 0.55ppm when rats are exposed through inhalation for up to 12 weeks.

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Cigarette tobacco tar, extracted from “Mild Seven” (Japan Tobacco Industries), was administered by force into the mouse stomach with or without pretreatment by subcutaneous injection of 4-nitroquinoline 1-oxide (4NQO), the incidence, size, and histopathology of the pulmonary tumors induced were studied to elucidate the effects of tobacco tar adiministered from the gastrointestinal tract on murine pulmonary tumorigenesis. One hundred and fifty ddY mice were divided into five groups; non-treated control animals, vehicle control animals treated only with intragastric injection of corn oil, tar-treated animals, 4NQO-treated animals, and 4NQO+tar-treated animals. The animals received intragastric injections of tar dissolved in corn oil at a dosage of 30 mg/kg body weight twice a day 6 days a week for 43 weeks. 4NQO was subcutaneously injected at a dosage of 8 mg/kg body weight at the first day of the experiment. The animals were sacrificed at the 52nd week. The average incidences of pulmonary tumors and adenocarcinomas per animal were 0.70 and 0.23 in the non-treated group, 0.55 and 0.31 in the vehicle control group, 0.82 and 0.61 in the tar-treated group, 2.32 and 1.00 in the 4NQO-treated group, and 2.26 and 1.33 in the 4NQO+tar-treated group, respectively. Gastrointestinal adminstration of cigarette tobacco tar did not increase the incidence of pulmonary tumors but did enhance the tumor grade from adenoma to adenocarcinoma. Thus, this study clarified that cigarette tobacco tar through the gastrointestinal tract could function as a progressive agent in pulmonary tumorigenesis.

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We studied cell kinetics of a mouse sarcoma induced by a subcutaneous injection of 4-nitroquioline 1-oxide by an immunohistochemical method. A mouse bearing a sarcoma received intraperitoneal injections of two thymidine analogues, iododeoxyuridine (IdUrd) and bromodeoxyuridine (BrdUrd) 3 hr and 1 hr, respectively, before sacrifice, and BrdUrd/IdUrd double immunostaining using anti-BrdUrd monospecific antibody and anti-BrdUrd/IdUrd bi-specific antibody was performed to evaluate BrdUrd labeling index (LI), S-phase duration (ts), cell cycle time (Tc), and turn-over time (To) in the tumor cells and to characterize changes of these parameters in the different histological tumor patterns. Histologically, the sarcoma was composed of spindle cells and giant cells, and resembled malignant fibrous histiocytoma. The spindle cells formed two different histological patterns; fascicular and hemangiopericytomatous patterns. In the spindle tumor cells in the fascicular region, LI was 21.8%, is was 7.2 hr, Tc was 26.5 hr, and To was 3.1 hr. In the spindle cells in the hemangiopericytomatous region, LI was 14.2%, ts was 9.5 hr, Tc was 46.8 hr, and To was 66.8 hr. In the giant tumor cells, LI was 38.7%, ts was 24 hr, Tc was 25.4 hr, To was 31.7 hr. Thus, in this study, we revealed that the cell kinetics parameters were dependent on the histological tumor pattern and the tumor cell type in a sarcoma with heterogeneous histological components. The BrdUrd/IdUrd double immunostaining method is an easy and reliable method, and is useful to gain various cell kinetics parameters of animal tumors

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Repeated injections of thiophene induce vasogenic cerbellar degeneration and local calcium deposits in the cerebellar granule cell layer in rats. Using light and electron microscopy, we investigated the formation of cerebellar calcification between 1 hour and 40 days after thiophene administration. Alizarin red S staining was used on microslicer sections to demonstrate early pathological calcium deposition.
In the cerebellum of rats that developed truncal ataxia and/or seizure,
1) pathological calcium deposits appeared in the granule cell layer and the deep cerebellar nucleus from the early stage,
2) the calcium deposits were lamellated like the annual growth rings of trees in the granule cell layer but had disappeared in the deep cerebellar nucleus at 40 days after thiophene administration, and
3) ultrastructurally, needle-like crystal deposits of calcium were found in the postsynaptic endings of the cerebellar glomeruli, perikarya of granule cells, axoplasm of myelinated fibers and presynaptic endings of deep cerebellar nuclei. It is suggested that the cerebellar calcification induced by thiophene is initiated in granule cells through an excitotoxic mechanism and/or disorder of calcium metabolism.

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Effects of haloperidol on the breast were investigated histologically and hormonally in female dogs at each stage of estrous cycle. In immature-period, metestrus and anestrus, female dogs were treated orally with 3 mg/kg/day of haloperidol for 5 weeks. The levels of progesterone and prolactin in serum were determined frequently during the pre-dosing and dosing period. After the dosing period, the animals were autopsied and the mammary glands were examined microscopically. Levels of serum prolactin increased by administration of haloperidol at every stage of estrous cycle. Levels of serum progesterone were high in metestrus and were low in immature-period and anestrus. In metestrus, swelling of the breast, lactation, and prominent hyperplasia of mammary glands were observed. In immature-period, mammary glands and ducts were not observed. In anestrus, the mammary glands reduced in size and the mammary ducts became dilated. Thus the mammary change should be due to the difference of hormonal environment, especially levels of progesterone and prolactin, and this difference of hormonal environment was caused by administration of haloperidol or estrous cycle in each animal.

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We differentiated the histological changes induced by irradiation with ultraviolet-A (UVA) and -B (UVB) light in the auricular skin and eye in female Balb/c mice. The animals were irradiated (1.30mW/cm²) with UVA for 1, 3, or 7 days, or with UVB for 2, 4, or 8 hr or 1, 3, or 7 days. Irradiation with UVA and UVB induced inflammation in the auricular skin, but the early changes were different. UVA-irradiation caused neutrophil infiltration without apparent edema, while UVB-irradiation induced marked edema with degranulation of mast cells and enlargement of endothelial cells of blood vessels in the dermis. These lesions became more severe with time progression, with the auricle showing partial necrosis at the later stage. UVA-irradiation caused retinal degeneration with vesiculation of the photoreceptor outer segment as an early change. In contrast, UVB-irradiation initially induced degeneration of corneal cells, followed by degeneration of lens epithelial cells. Although the incidence of retinal changes induced by UVB-irradiation was much lower than that by UVA, the number of migrating cells in the photoreceptor segments with UVB significantly increased at the later stage.

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To determine whether promotion of thyroid proliferative lesion development is more effectively achieved by increasing dose treatment with a goitrogen than with continuous administration, male F344 rats initiated with N-bis(2-hydroxypropyl)nitrosamine (DHPN, 2, 800mg/kg body weight, single s.c. injection) were given water containing 0.1% sulfadimethoxine (SM) for 20 weeks (group 1), 0.025, 0.05, and 0.1 of SM for 8, 4 and 4 weeks, respectively (group 2), and 0.025, 0.05, 0.1 and 0.2% of SM for 8, 4, 4 and 4 weeks, respectively (group 3). Control rats (group 4) were maintained without further treatment for 20 weeks after DHPN-initiation. Both absolute and relative thyroid weights in group 3 were significantly increased as compared to group 1. Serum T3 and T4 levels were significantly decreased in groups, 1, 2, and 3 as compared to group 4, the serum T4 level in group 3 being also significantly decreased as compared to the group I value. Serum TSH was significantly elevated in groups 1, 2, and 3 and higher in the latter two cases than in group 1. The numbers of follicular cell hyperplasias in groups 2 and 3 and adenomatous tumors in group 3 were also significantly increased, as compared to group 1, along with BrdU labeling indices for follicular cells, hyperplasias, and adenomatous tumors. The present results indicate that step-wise increasing dose treatment with SM enhances production of thyroid proliferative lesions and their cell proliferative activity to a greater extent than continuous treatment despite a lower total intake and suggest that this approach may be more sensitive for detection of thyroid tumor promoters in rat thyroid two-stage carcinogenesis models.

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A malignant granulosa cell tumor and a benign granulosa/theca cell tumor were found in a 124-week-old Sprague-Dawaley female rat. These ovarian tumors were deemed to be estrogenically active because of the presence of severe adrenal peliosis. Pituitary prolactinoma and mammary adenocarcinoma were also found. Sex hormones were not measured, but the concomitant occurrence of ovarian, pituitary, and mammary tumors suggested that estrogen derived from ovarian tumors could have induced prolactinoma and that prolactin derived from the pituitary tumor could have been an important factor in producing the mammary adenocarcinoma.

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This paper describes spontaneous ovarian choriocarcinoma in two B6C3F₁ mice used in toxicity studies. One animal (case 1) was sacrificed at eight weeks of age, and the other one (case 2) was found dead at sixty-seven weeks of age. Macroscopically, a dark red dot measuring approximately I mm in diameter (case 1) and hematoma-like enlargement (case 2) were observed in the ovary. Microscopically, in case I, the neoplasm was fully packed with tumor cells and erythrocytes. The tumor cells were round to oval giant cells with abundant, faintly basophilic cytoplasm containing PAS positive granules. In case 2, some small and two large hematocysts were present. The hematocysts contained a large number of erythrocytes and a small number of tumor cells. The tumor cells were oval to elongated giant cells with abundant, basophilic cytoplasm. Immunohistochemically, tumor cells in both cases were positive or weakly positive for human placental lactogen (HPL), human chorionic gonadotrophin (HCG), and human specific beta-1 glycoprotein (SP-1). Both cases were considered to be originated from germ cells since those female animals were virgin.

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This paper describes a case of primary thymoma found in a 23-week-old young female SD rat. Grossly, the tumor was an encapsulated, grayish, firm mass measuring 20×15×15mm, which was found cranial to the heart in the thoracic cavity. Histologically, the neoplasm consisted of keratinpositive neoplastic epithelial cells and considerably abundant supporting nonneoplastic stroma, which were arranged in storiform and wavy patterns. The neoplastic cells were randomly distributed in the stroma in single or clusters sometimes forming pseudoductular structures. Spindle cells forming the stroma were immunopositive for muscle actin, suggesting features of myofibroblasts. No proliferating lymphocytes were observed in the tumor. Based on these findings, the present case was diagnosed as a benign thymoma, epithelial type with extensive development of collagen fibers. To our knowledge, such a unique thymoma has not been reported in young rats.

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