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Abstract
A 7-year-old, intact female mixed-breed dog was presented for evaluation of hematuria. Physical examination revealed a suprapubic mass. Ultrasonographic examination showed a large lobular mass occupying the urinary bladder. At the owners' request, the dog was euthanatized and a postmortem examination was performed. Necropsy confirmed the presence of a lobular mass of about 5- to 6-cm diameter protruding into the lumen of the bladder. Histologically, the mass was composed of a large number of atypical lymphoid cells in the lamina propria and mucosal epithelium. Immunohistochemically, the neoplastic cells expressed CD3 but not CD79
or keratin and vimentin, supporting a diagnosis of T-cell lymphoma.
Key words: Dogs; epitheliotropism; urinary bladder; T-cell lymphoma.
Primary malignant lymphoma of the urinary bladder is extremely rare in humans.5,8,10 In veterinary medicine, only eight cases have been reported: two in dogs, two in cats, and four in cows.9 In this paper we describe the histologic and immunohistochemical features of a primary malignant lymphoma of the urinary bladder with epitheliotropism in a dog, a finding not previously reported in this species.
A 7-year-old, intact female, mixed-breed dog was presented to the Veterinary Medicine Faculty, University of Naples, for examination after the owners complained that their pet had constant hematuria. The dog had no other clinical history. Physical examination revealed a suprapubic mass. Plain abdominal radiographs and ultrasonographic examination showed the presence of a large lobular mass of uniform density in the area of the urinary bladder. No other abnormalities could be seen. A tentative diagnosis of a tumor was made. In view of the poor prognosis, the owners requested euthanasia rather than an exploratory laparotomy.
At postmortem examination, abnormalities were confined to the urinary bladder. The bladder was grossly abnormal and the whole organ was removed. The gross pathologic examination confirmed the presence of a large, lobular, 5- to 6-cm-diameter, light tan mass protruding from the ventral mucosal surface, with no extension to the trigone evident. On the cut surface, the bladder mucosa was markedly thickened with prominent rugae and multiple foci of hemorrhage and ulcers. No evidence of metastasis or other gross abnormalities were found.
Representative areas of the tumor were fixed in 10% formalin for 48 hours and then processed for paraffin embedding. Four-micrometer sections were mounted on poly-L-lysine coated glass slides and incubated at 37 C overnight to optimize tissue adhesion to the slide. The sections were dewaxed in xylene, dehydrated in graded alcohols, and stained with hematoxylin and eosin. For immunohistochemical analysis some sections were washed three times for 5 minutes each time in 0.01 M phosphate-buffered saline (PBS), pH 7.2–7.4. Endogenous peroxidase activity was blocked with hydrogen peroxide 0.3% in absolute methanol for 30 minutes. Before the immunohistochemical procedure (a streptavidin–biotin peroxidase method) with a commercial kit (LSAB Kit, Dako, Milan, Italy) some sections were treated with 0.1% trypsin (Sigma, Milan, Italy) for 1 hour at 37 C for detection of CD3 and some were incubated twice for 5 minutes at 700 W in citrate buffer (pH 6.0) in a microwave oven for detection of CD79 and keratin. Then the sections were incubated overnight at 4 C with one of the following antibodies diluted in PBS containing 0.1% bovine serum albumin (BSA, Sigma) and sodium azide 0.05%: CD3 (T-cell marker, diluted 1:50, Dako), CD79 c
(clone HM57, diluted 1:50, Dako), keratin (Keratin cocktail CK22, prediluted, Biomeda, Milan, Italy), vimentin (clone V9, diluted 1:50, Dako). Biotinylated anti-mouse and rabbit immunoglobulins (LSAB Kit, Dako), diluted in PBS, were used as secondary antibody, and were applied for 30 minutes. After washing in PBS, the sections were incubated in streptavidin conjugated to horseradish peroxidase in Tris-HCl buffer containing sodium azide 0.015% (LSAB Kit, Dako) for 30 minutes and finally exposed to the 3,3'-diaminobenzidine tetrahydrochloride chromogen (DAB, Sigma) containing hydrogen peroxide 0.03%, and counterstained with Harris' hematoxylin, dehydrated, cleared, and coverslipped with Eukitt (Bio-Optica, Milan, Italy). Sections of a normal dog lymph node were used as positive controls and to ensure specificity of the antibodies. Negative controls consisted of sections incubated with PBS.
Histologically, the mucosa and submucosa of the urinary bladder were massively infiltrated by a large number of medium to large atypical lymphoid cells. The cells were usually round with scant pale eosinophilic or large optically clear cytoplasm. Nuclei were small, round to irregularly convoluted with finely aggregated and uniformly distributed chromatin, and contained rare nucleoli. Mitotic figures were frequent (two to four per high-power field). The neoplastic cells invaded the mucosal epithelium, with effacement of the stromal–epithelial junction, which was often eroded and replaced by the atypical lymphoid cell population expressing CD3 (Fig. 1). The intraepithelial lymphocytes were often surrounded by a halo of clear cytoplasm. A great number of histiocytic cells were also scattered throughout the neoplastic tissue imparting a "starry sky" appearance. The neoplastic cells were positive for the T-cell marker anti-human CD3 (Fig. 2) but not for vimentin, keratin, and CD79, supporting a diagnosis of T-cell lymphoma. No evidence was found of bacterial infection.
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Acknowledgments
We wish to thank Mr. R. Ilsami for his technical assistance.
References
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  V. E. Valli, W. Vernau, L.-P de Lorimier, P. S. Graham, and P. F. Moore Canine Indolent Nodular Lymphoma Vet. Pathol., May 1, 2006; 43(3): 241 - 256. [Abstract] [Full Text] [PDF]
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