Chronic progressive nephropathy (CPN) is a rodent-specific, age-related renal disease, particularly of male rats, characterized by a spectrum of distinct histological changes which may begin early in the animal's life and progress to end-stage renal disease in certain rat strains. Although CPN-related pathology is well known to most toxicological pathologists other features of CPN such as pathogenesis, modulating factors, proliferative nature, response to chemical exposure and relationship to tumorigenesis are less clearly acknowledged. CPN is generally regarded as a degenerative to atrophic disease with compensatory regenerative hyperplasia. The proliferative nature of CPN often becomes problematic in advanced to end-stage renal disease. At this stage, a number of tubule profiles may be mistaken for atypical tubule hyperplasia, the reported precursor lesion of tubule adenoma. CPN associated proliferative tubule profiles must be carefully separated from atypical tubule hyperplasia particularly in studies where chemical exposure has exacerbated CPN. Over the past several years increasing evidence has supported the hypothesis that CPN may be regarded as a type of "mode of action" during renal carcinogenesis in rodent bioassay studies. Retrospective studies of control and treated animals have consistently shown a relationship between the increased severity of CPN and the presence of atypical tubule hyperplasia and small, incipient renal adenomas. Understanding CPN-related tumorigenesis is important for human risk assessment interpretation. Since CPN is a rodent specific disease with no apparent similar human kidney disease condition, evidence that renal tumors may arise from an interaction with CPN could assist regulatory agencies in interpreting data from studies with exacerbated CPN.

View full abstract

Current advances in automation of pathology methods with focus on toxicologic pathology are dealt with. Areas of tissue can be separated and collected for further analysis by laser microdissection. Automatic stainers for histology and immunohistochemistry rapidly produce high standard quality stains of tissue sections. Confocal laser scanning microscopy allows visualization of fluorochromes with superior quality optical data. Introduction of field emission cathodes and low vacuum technologies to scanning electron microscopy enables examination of non-coated and non-dehydrated specimens. Energy filtering transmission electron microscopy yields high contrast images and provides novel specimen information. Digital image analysis offers systems to support users in specific evaluations. To facilitate orientation on the market, a selection of commercially available equipment is presented and the features of the devices are compared.

View full abstract

In order to clarify the metallothionein (MT) localization in the developing placenta, we histologically investigated the sequential MT expression in placentas and fetal livers using pregnant rats during gestation days (GDs) 9 - 21. The placentas were sampled and weighed on GDs 9, 11, 13, 15, 17, 19 and 21. In the early post implantation period, the expression of MT was slightly detected in the yolk sac and the primary decidual zone around the embryo. MT was then mainly present in the deciduas parietalis and yolk sac. After the deciduas parietalis ruptured, MT was subsequently detected in the yolk sac and deciduas basalis. MT continued to be detected in the yolk sac until GD 21, but it was reduced in the deciduas basalis in accordance with development of the fetal liver with elevated MT expression. In conclusion, the main expression site of MT changes from the maternal placenta to the fetal placenta, and then to the fetal liver in accordance with the fetal development. However, we speculate that the MT-positive cells in the placenta are positioned between the maternal and embryonic environments throughout the gestation period and always surround the embryo/fetus.

View full abstract

To investigate the early changes involved in glomerular calcification, 360 mM Na₂HPO₄ was administered to rats at 8 mL/kg once daily via the tail vein singly or repeatedly. Urinalysis was carried out on days 1, 3, 5 and 8 of dosing, and rats were sacrificed on days 2, 4 and 9 for histopathological and electron microscopic examination of the kidneys. Following single dosing, there were no gross or histological findings, but electron microscopy revealed a number of vacuoles scattered within the Bowman's space. On day 4, minimal and focal mineralization was observed within the parietal epithelial cells. On day 9, mineralization was minimal to mild and localized within the parietal epithelial cells and glomerular basement membrane. Hypertrophy and increased mitotic figures were also frequently observed in the parietal epithelial cells. Low-density lamellar structures with effacement of podocytes, an increased number of microvilli and large amounts of debris filling the Bowman's space were the main electron microscopic changes on days 4 and 9. Increased urinary protein excretion correlated well with the glomerular changes. Immunohistochemically, increased expression of desmin and decreased expression of podoplanin were evident in glomeruli on day 9. The numbers of PCNA-positive podocytes and parietal epithelial cells showed a tendency to increase on day 9. These results suggest that the onset of glomerular calcification is preceded by primary podocyte damage.

View full abstract

A unique conjunctive stricture was observed in the eye of a male 17-week-old Japanese White rabbit (Kbl:JW). The conjunctival membrane had proliferated centripetally and covered a large portion of the cornea. However, the membrane did not adhere to the cornea. Histopathologically, the inner epithelium of the conjunctival membrane appeared flattened, while the outer epithelium had become stratified and squamous. Goblet cells were observed on both sides of the epithelium. The lamina propria consisted of well-developed, vascularized collagen fiber. Myxoid change was seen near the tip of the membrane. In animals, these conjunctival membranes have been reported in a few dwarf rabbits, dogs and horses and have had various different terms applied to them due to their unknown etiology. Based on the conventions of human ophthalmology, such lesions should be regarded as pseudopterygia. Therefore, the present case was diagnosed as involving a pseudopterygium. The centripetal proliferation of the conjunctival membrane may be a characteristic finding in animal cases. Our goal is to encourage accumulation of such cases by researchers and practitioners working in the field of toxicology.

View full abstract

In a young adult (8 weeks old) Sprague-Dawley (SD) rat, a 7 mm-diameter hemispherical nodule with white spots was macroscopically identified on the visceral surface of the spleen. Microscopically, the nodule consisted of areas of enlarged white pulp. The periarteriolar lymphoid sheaths (PALS) area was expanded due to an increase in lymphocytes, but the marginal zone was relatively atrophied. Although minimal compressions and mitotic figures were sporadically observed, there were no structural alterations or atypical cells. Immunohistochemically, there were increased numbers of proliferating cells (Ki-67 positive cells) in the PALS and red pulp areas. In the PALS area, there were increased numbers of T-cells and macrophage/dendritic cells, while in the red pulp area, there were increased numbers of T-cells and B-cell/plasma cells. This lesion, which is believed to be hyperplastic and not neoplastic, was diagnosed as lymphoid hyperplasia, a condition considered to be uncommon in young adult SD rats.

View full abstract

We report pathological changes observed in the lungs of Phenylhydrazine (PHZ)-treated rats. Seven male SD rats were treated intraperitoneally with 50 mg/kg/day of PHZ for 3 days. PHZ-treated rats developed severe hemolytic anemia, and four out of the seven rats died after the final administration of PHZ; the three surviving rats were terminally sacrificed 24 hours after the final administration. Macroscopically, all lobes of the lungs of PHZ-treated rats were red or dark, and small white lesions were observed in the terminally sacrificed rats after fixation. Light microscopy revealed deposition of eosinophilic material in the alveolar septa of all PHZ-treated rats. This eosinophilic material was identified as fibrin (fibrin thrombus formation) in alveolar capillaries by immunohistochemistry and electron microscopy. This is the first report showing that PHZ induces acute thrombus formation in animals.

View full abstract

We examined the histopathological characteristics of ganglion-like cells in relation to C-cell proliferations of the thyroid gland in CD(SD)IGS, CD(SD) and F344 rats. The incidence of the focus being composed of ganglion-like cells was extremely low, between 0.5% and 2.5%, in each strain of aging (approximately 2 years old) rat. The lesions were histologically classified into 2 different types, the complex or ganglioneuroma types. The complex type was observed in 2/367 female F344 rats and 1/290 male CD(SD) rats, and the ganglioneuroma type was observed in 3/122 female CD(SD)IGS rats and 3/290 male and 2/247 female CD(SD) rats. The ganglion-like cells were characterized by abundant cytoplasm with large, eccentric, round nuclei and prominent nucleoli. They were stained positive by Nissl staining and weakly positive by immunohistochemical staining for neuron-specific enolase, but were negative for calcitonin, synaptophysin and neuronal nuclei. Based on these results, we considered ganglion-like cells to be a variation of C-cells, which originate from the neural crest, with a loss of its characterization.

View full abstract

Protocadherins are a major subfamily of the cadherin superfamily that play an important role in communication between adjacent cells. To clarify the involvement of Protocadherin 20 (Pcdh20) gene in rat lung carcinogenesis, we investigated the expression of Pcdh20 and its methylation status in rat lung adenocarcinomas induced by N-nitrosobis(2-hydroxypropyl)amine (BHP). Six-week old male Wistar rats were given 2000 ppm BHP in their drinking water for 12 weeks and were maintained without further treatment until they were sacrificed after 25 weeks. Total RNAs were extracted from 7 lung adenocarcinomas, one from each BHP-treated rat, and the expression levels of Pcdh20 were measured using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis. Four out of 7 tumors showed reduced expression of Pcdh20, compared with 3 normal lung tissues. For methylation analysis, bisulfite sequencing was performed. The 5' upstream region of Pcdh20 was methylated in 4 adenocarcinomas with reduced expression of Pcdh20, but was unmethylated in normal lung tissue. These results suggest that aberrant methylation of the Pcdh20 gene might be involved in the development of lung adenocarcinomas induced by BHP in rats.

View full abstract