Fibrous mastocytoma in a patient with generalized cutaneous mastocytosis.

A 57-year-old woman with cutaneous mastocytosis of 23 years duration developed a hyperpigmented abdominal plaque composed of confluent indurated papules that enlarged for a period of 1 year to 12 x 8 cm. Biopsy showed dermal infiltration by closely packed spindle-shaped mast cells, fibroblasts, collagen, and scattered lymphocytes, predominantly T-suppressor cells. Electron microscopy showed close contact between mast cells, fibroblasts, and lymphocytes. Piecemeal mast cell degranulation and extrusion of mast cell granules was seen, with rare mast cell granules in fibroblasts, and collagen fibers in peripheral and perinuclear endoplasmic reticulum of mast cells. the term Fibrous mastocytoma is suggested for this tumor-like dermal fibrosis, possibly induced by lymphokines.

Correlation of DNA ploidy levels with altered cytokeratin patterns in rat bladder tumors.

Flow cytometry analysis of primary bladder carcinoma cells is appropriate for demonstrating the association between the presence of aneuploid cells and increased malignant potential for transitional cell carcinoma (TCC). Our laboratory previously has reported a specific alteration in the cytokeratin (CK) pattern of experimentally induced bladder carcinomas in rats. Unique immunohistochemically detectable changes in CK expression were the loss of CK 13 expression in invasive TCC cells and the loss of CK 19 expression in induced TCC. We now report the correlation of these changes in CK expression with cellular aneuploidy of the induced bladder carcinomas. Bladder carcinomas were induced in rats by direct instillation of N-methyl-N-nitrosourea. Immunohistochemistry was performed on the induced tumors using four commercially available monoclonal antibodies specific for CK 18 (TR1031), CK 13 (K8.12), CK 19 (K4.62), and the CKs 5, 7, and 8 (K8.13). Flow cytometry data from the induced bladder carcinomas was analyzed and the DNA index, proliferative index, and percent aneuploid cells were calculated for each time point. The percent aneuploid cells and CK 19 staining were tested statistically and were shown to be negatively correlated. We therefore hypothesize that the combination of the loss of CK 19 as detected by the antibody K4.62 coupled with the presence of aneuploid cells in histopathologically diagnosed invasive TCC is a significant factor in predicting the prognosis for any given diagnosis.

Correlation of cytokeratin patterns with histopathology during neoplastic progression in the rat urinary bladder.

The discrimination of atypical (premalignant) cells from invasive neoplastic cells in primary bladder lesions is a major diagnostic problem in cytopathology and surgical pathology. We have used an animal model of urinary bladder carcinogenesis to determine the specific changes which occur in the expression of certain cytokeratins (CK) during the progression of lesions from regenerative hyperplasia and carcinoma in situ to transitional cell carcinomas. At sequential time points following exposure of the rat bladder epithelium to N-methyl-N-nitrosourea in vivo, immunohistochemical staining of CKs was evaluated in ethanol-fixed samples from the induced urothelial lesions using commercially available anti-CK mouse monoclonal antibodies. Specific changes were found in the expression of CKs 13, 18, and 19 during the neoplastic progression of induced urothelial lesions in the rat. These changes included the reciprocal loss of expression of CK 19 and the reappearance of CK 18 as malignant tumors developed. Invasive cells also did not express CK 13. Our results, based on the rat model, are similar to those reported by others on CK expression in human bladder tumors. Because these changes in CK expression occurred at specific points in the progression of urothelial lesions, the antibodies utilized in this study may be helpful in predicting the invasive potential of cells present in cytopathological specimens and tissue biopsies from human urothelial lesions.

Discrimination of cell types in primary transitional cell carcinoma by monoclonal anti-cytokeratin antibodies.

The morphologic discrimination between low and high grade malignant tumor cells arising in the urinary bladder is a major diagnostic problem in cytopathology. Using immunochemical peroxidase staining of cytokeratins (CKs) of human bladder exfoliative cytology specimens, we have been able to discriminate between transitional cell carcinoma cells, atypical cells and normal bladder cells. Commercially available monoclonal antibodies used in this study were: anti-CK 13 (Sigma K8.12), anti-CK 5, 7 and 8 (Sigma K8.13), anti-CK 19 (Sigma K4.62), and anti-CK 18 (Transformation Res. 1091). When normal bladder cells are stained with these antibodies, CK 18 is specific for superficial cells; CK 13 is specific for the basal type cells and CKs 5, 7, 8 and 19 are expressed by all urothelial cell types. Four cases diagnosed by cytopathological criteria as 'atypical' and 7 diagnosed as 'positive' (various grades) were used in this study. Cytologic diagnosis was confirmed by histopathology in 7 cases. Tissue was not available for histopathology in 4 cases. Malignant cells with a 'basal' or 'immature' phenotype preferentially stained with CK 13 and were associated with increased metastatic or malignant potential. Patients with higher grade tumors had more cells positive for CK 13 than did patients with atypical or lower grade malignancies. Patients with well-differentiated, low grade tumors predominantly shed cells that expressed CK 18 and CK 19. High grade, invasive bladder lesions were characterized by many cells expressing CK 13, and fewer cells expressing CK 19. The combination of diagnosis by morphologic criteria on Papanicolaou-stained specimens with immunochemical characterization of the same cells for CKs facilitate an accurate diagnosis of bladder lesions.

Isolation and primary culture of urothelial cells from normal human bladder.

Trypsinization (WT) was employed to disaggregate urothelial cells from normal human urinary bladder mucosa for the preparation of primary cultures. Urinary bladders of two male adults both 25 years old were obtained autopsy 1-2 h after death. The mucosa was incubated in HBSS containing 0.25% trypsin at 37 degrees C. Mean cell yield, viability, and attachment were 14.6 X 10(7), 76%, and 42.5% after WT. In histologic sections of treated mucosa, most of the urothelium was removed by WT. Following plating and attachment, cells obtained by WT formed a monolayer of flattened epithelial-like cells. When stained with polyclonal antikeratin antibody using the indirect immunoperoxidase technique, all of the cells were immunoreactive indicating an epithelial origin. In conclusion, based on morphology and immunocytochemistry, WT removed virtually all of the urothelial cells from the mucosa and no contamination by mesenchymal cells resulted from this procedure. Thus, WT is an appropriate technique for the isolation of urothelial cells and initiation of primary cultures of normal human urothelial cells for subsequent study.

Histomorphometry and cell kinetics of normal human bladder mucosa in vitro.

In this study, we have quantified the morphologic and kinetic parameters of explant cultured normal adult human urinary bladder mucosa. Quantitative parameters studied were urothelial height, cell density, labelling index and mitotic index. For these studies, urinary bladders from seven adults with no previous history of urologic disease were obtained at autopsy. Mucosal explants were maintained in rocking culture on Gelfoam rafts for up to 33 days using supplemented CMRL 1066 medium. Prior to sampling, cultures were treated with tritiated thymidine and colchicine to investigate tissue kinetics. Data was based on histologic autoradiograms. During culture, urothelial cells retained normal polarity. During the first week of culture, urothelial height increased and cell density decreased. DNA synthesis and mitotic activity occurred primarily among basal cells. DNA synthesis was first noted on day 2 of culture; mitotic activity began after 3 days of culture. Morphologically, human urothelium was well maintained; DNA synthesis and mitotic activity was variable but continued throughout culture.