The aim of this study was to examine tumor necrosis factor-α (TNF-α) expression and Kupffer cell activation in hepatotoxicity caused by microcystin-LR (MCLR). Mice received a single intraperitoneal injection of 60.0 μg/kg MCLR and were killed at several time points within 24 hours. MCLR caused hemorrhage within 7 hours, followed by hepatocellular necrosis and apoptosis. A real-time quantitative reverse-transcription polymerase chain reaction demonstrated that the level of TNF-α mRNA was 2.3-fold higher than in the controls at 17 hours. The number of TNF-α-immunopositive nonparenchymal cells was 2.6- and 7.7-fold greater than that in the controls at 7 and 17 hours, respectively, and they frequently infiltrated into necrotic areas, probably in association with neutrophil recruitment. Some apoptotic hepatocytes were immunopositive for cleaved caspase-3. To inactivate Kupffer cells, mice were pretreated with a single intravenous injection of gadolinium chloride (GdCl₃, 2 mg/mouse). That brought about the apoptosis of sinusoidal cells, which indicated Kupffer cell depletion. GdCl₃ pretreatment attenuated MCLR-induced hepatocellular apoptosis by 68%, and likely decreased hepatocellular necrosis. These results suggest that Kupffer cell activation, represented by enhanced TNF-α expression, is involved in the progression of MCLR-induced hepatotoxicity in mice.

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In order to confirm the relationship between benign prostatic hyperplasia (BPH) regression and atrophic effect of chlormadinone acetate (CMA), the prostates of spontaneous BPH dogs administered CMA for 6 months were investigated histopathologically and immunohistochemically. Male beagle dogs (5-8 years old) were divided into four experimental groups; Group 1 consisted of untreated controls. Groups 2 to 4 received CMA 0.03, 0.1, and 0.3 mg/kg/day, p.o., respectively, for 6 months. In group 1, glandular hyperplasia of the prostate was clearly detected. In groups 2 to 4, CMA produced marked atrophy of the glandular epithelium. The interacinar fibro-muscular stroma was prominent. To evaluate the frequency of apoptosis, we counted the positive cells stained by the nick end labeling method. In group 1, the apoptotic index was 0.68 ± 0.03 %. In groups 2 to 4, apoptotic indices were 10.32 ± 0.96 %, 2.05 ± 0.88 %, and 1.15 ± 0.75 %, respectively. Apoptotic cell death was mainly observed in the glandular epithelial cells. Based on our data, regression of BPH after treatment with CMA may be apoptotic cell death.

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To investigate the susceptibility of heterozygous p53-deficient CBA mice [p53 (+/-) mice] to promotion of liver proliferative lesions in a two stage hepatocarcinogenesis model, 30 p53 (+/-) mice and 30 wild-type littermates [p53 (+/+) mice] received an i.p injection of 5 mg/kg of N-nitrosodimethylamine (DMN), and from one week later, each group was given free access to drinking water containing 0.05 or 0 % of phenobarbital (PB) for 26 weeks. The final body weights were significantly decreased in both p53 (+/-) and p53 (+/+) mice of the DMN+PB compared to the DMN alone groups and the liver weights were significantly increased. The survival rate was 67 and 73% in p53 (+/-) and p53 (+/+) mice of the DMN+PB group, respectively, and there were no deaths in any of the mice receiving DMN alone. The incidences of eosinophilic foci in the liver (90 and 54.6% in p53 (+/-) and p53 (+/+) mice, respectively) in the DMN+PB groups were significantly higher than those with DMN alone (6.7% in p53 (+/-) mice, 0% in p53 (+/+) mice). The incidences of clear cell foci and hepatocellular adenomas were 10.0 and 9.1%, and 60.0 and 27.3%, respectively, in p53 (+/-) and p53 (+/+) mice receiving DMN+PB. These lesions were not seen in the DMN alone group. The PCNA labeling indices for eosinophilic foci and hepatocellular adenomas in the DMN+PB group were significantly higher in p53 (+/-) than in p53 (+/+) mice. The present results suggest that p53 (+/-) CBA mice are very susceptible to promotion of the development of liver proliferative lesions by PB after DMN initiation.

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In order to standardize test procedures for evaluation of neurotoxicity of chemical exposure, a collaborative study with a common protocol based on the Organization for Economic Cooperation and Development (OECD) test guideline TG407 was conducted by eleven laboratories, using acrylamide and 3,3’-iminodipropionitrile (IDPN) as positive neurotoxicants. This report summarizes the results of this neuropathological evaluation of neurotoxicity, when acrylamide (3, 10, and 30 mg/kg) and IDPN (20, 50, and 100 mg/kg) were orally administered to rats for 28 days. The study revealed typical alterations, including degeneration of peripheral nerve fibers and Purkinje's cell necrosis with acrylamide, and axonal swelling in the brainstem and spinal cord with IDPN. In addition to the neurotoxicity, IDPN exerted toxic effects on the cerebral arteries, hyalinization of vessel walls being observed. In general, the neuropathological findings well demonstrated the neurotoxicity of both chemicals. These results indicate that neuropathological evaluation can play a crucial role in screening for potential neurotoxicants with risk for human health, the expected neurotoxic effects being detected in this collaborative study consistently in all the laboratories.

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We investigated the effects of granulocyte colony-stimulating factor (G-CSF) on the development of inflammation in the lung induced by intratracheal administration of bleomycin (BLM, 2 mg/200 μl) in rats. G-CSF (100 μg/kg/day) was subcutaneously administered for 7 days starting from 3 (inflammatory phase) or 14 days (fibrogenic phase) after BLM administration. The administration of G-CSF to BLM-treated rats in the inflammatory phase increased the total lung lesion per unit pulmonary parenchyma, the score of lung fibrosis, the peripheral neutrophil count and the number of neutrophils infiltrating the pulmonary lesion, as well as the severity of other histopathologic changes (edema, hemorrhage and proliferation of myofibroblasts in the alveolar lumen, hyperplasia of bronchial and alveolar epithelial cells, and bronchiolization). Although increases in the peripheral neutrophil count and the number of neutrophils infiltrating the pulmonary lesion were observed following treatment with G-CSF, there were no further increases on lung injury in the fibrogenic phase in BLM-treated rats. These findings suggest that the effects of neutrophils on the lung in the rats treated with G-CSF may be closely related not only to the number of neutrophils, but also to the inflammatory phase of BLM-induced pulmonary lesions.

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Nivalenol (NIV) is a potent toxic trichothecene mycotoxin produced by Fusarium nivale, and our previous study clarified first that NIV induces apoptosis in lymphoid organs of mice in a dose-dependent manner. In this study, the development of apoptosis and changes in the expression of apoptosis-related genes (fas, c-fos, c-jun, p53, bcl-2 and c-myc) mRNAs were examined in the thymus of mice for up to 12 hours after inoculation (HAI) of NIV (15 mg/kg b.w.) to elucidate the relationship between the molecular genetic regulatory mechanisms and NIV-induced lymphocyte apoptosis. The number of apoptotic lymphocytes evaluated by TUNEL method clearly increased from 3 HAI and peaked at 9 HAI. The DNA ladder formation by agarose gel electrophoresis was clearly observed at 6 and 9 HAI. The expression of c-fos and c-jun mRNAs clearly elevated from 0.5 HAI and peaked at 1 HAI. The expression of fas, p53, bcl-2 and c-myc mRNAs showed no significant changes. These results indicate that c-fos and c-jun may play an important role in NIV- induced lymphocyte apoptosis.

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A very uncommon myxoid liposarcoma was found in a 41-week-old Crj:CD(SD)IGS male rat. At necropsy, an encapsulated elastic mass was observed in the subcutis of the left cervical region. The cut-surface was whitish, translucent and myxomatous with focal hemorrhage. No metastasis to other tissues or organs was noted. Histopathologically, the tumor was composed of numerous capillaries, round to oval cells and spindle cells with abundant myxoid matrix. Most tumor cells contained vacuoles of various size in their cytoplasm. These vacuoles were positive for oil red-O staining. Immunohistochemically, tumor cells were positive for S-100 protein and vimentin, but negative for cytokeratin. Ultrastructurally, tumor cells contained a large number and various sizes of lipid droplets, rough endoplasmic reticulum, and small numbers of smooth endoplasmic reticulum, Golgi complexes, and mitochondria. Thus the tumor cells were considered as lipoblasts at various differential stages. We therefore diagnosed this tumor as a myxoid liposarcoma, the tumor being similar to myxoid liposarcoma in humans. This is the first case report of such a myxoid liposarcoma in rats including Crj:CD(SD)IGS rats.

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Unilateral renal dysplasia was found in a male Std: Syrian hamster at 13 weeks of age. At necropsy, the kidney was markedly reduced in size (0.1539 g), and had a granular surface and pale color. The kidney had renal papillae. Histologically, the kidney had a distinct cortico-medullary junction. In the cortex, small glomeruli, poorly differentiated tubular epithelia, and flattened tubular epithelia were observed. The poorly differentiated epithelia were single-or multi-layered, and had nuclei of varying size and shape, i.e. round, oval or kidney-shaped. They were described as having moth-eaten basement membranes. In the medulla, flattened tubular epithelia were observed. Ultrastructurally, the small glomeruli were composed of well-differentiated cells, therefore they may not have been primitive, but rather immature. The poorly differentiated epithelia did not have lumina or brush borders, and were concluded to be primitive tissues. This case was characterized by immature glomeruli and primitive cortical tubular epithelia, and likely represents a developmental disorder. In conclusion, this case was diagnosed as renal dysplasia, but the histological features did not resemble those of human renal dysplasia.

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Spontaneous endometriosis was found in a 15-year-old female rhesus monkey. Macroscopically, there was thickening of the intestinal wall, mesenterium, and the gastrocolic omentum. Sporadic nodules were noted in the serosa of the jejunum and ileum. Histopathologically, a large number of glandular tubules along with massive connective tissue was observed in the serosa and subserosa. The glandular tubules were covered with mono- or multilayered epithelial cells with ciliary structures protruding into the glandular lumen. The connective tissue was composed of mesh-like fiber, fibroblast-like cells, and tortuous blood vessels. The glandular tubules and the connective tissue were similar to the glands and interstitial tissue in the endometrium.

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During a Joint Society of Toxicologic Pathologists (STP)/International Federation of Societies of Toxicologic Pathologists (IFSTP) International Symposium, held between June 24 and 28, 2001, in Orlando, FL, USA, there was a session entitled as “Genetically Modified Foods-Hazard Identification and Risk Assessment”. The purpose of this session was to present and discuss the current situations in the US, European Union and Japan for the public concerns, safety assessments and regulations on genetically modified (GM) products used as food or food ingredients. Assuming the wide and fast growing of the usage of GM products, it is the duty for us, toxicologic pathologists, to supply reliable data on their safety and possible risks or hazards as a world-wide basis to not only governments or regulatory agencies but also general public of our countries.

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