A Case of Feline T-cell Lymphoma with Tropism for Striated Muscle and Peripheral Nerve.

An 11-year-old female American shorthair cat was presented with a 3-month history of hindlimb ataxia and knuckling of the left forelimb. Clinical abnormalities included weight loss, hyperaesthesia of the neck and back, cardiac murmur and systemic muscle atrophy. The cat died 10 days after the initial presentation and a necropsy examination was performed. Grossly, extensive pale lesions were seen in the wall of the left ventricle and the septum of the heart. There were no detectable masses in the heart, skeletal muscles or peripheral nerves. Histopathological examination revealed diffuse, extensive infiltration of atypical lymphoid cells in the heart; the cardiac muscles were markedly degenerate and atrophic and were replaced by the neoplastic cells. Neoplastic cells with similar morphology were seen in all specimens of the skeletal muscles and peripheral nerves. Clonality analysis of the paraffin wax-embedded heart tissue revealed a monoclonal rearrangement of the gene encoding the T-cell receptor γ chain. Based on these findings, the case was diagnosed as T-cell lymphoma with tropism for striated muscle and peripheral nerve.

Participation of Tumor-Associated Myeloid Cells in Progression of Amelanotic Melanoma (RMM Tumor Line) in F344 Rats, with Particular Reference to MHC Class II- and CD163-Expressing Cells.

Tumor progression is often influenced by infiltration of myeloid cells; depending on the M1- or M2-like activation status, these cells may have either inhibitory or promoting effects on tumor growth. We investigated the properties of tumor-associated myeloid cells in a previously established homotransplantable amelanotic melanoma (RMM tumor line) in F344 rats. RMM tumor nodules were allowed to reach the sizes of 0.5, 1, 2 and 3 cm, respectively. Immunohistochemistry and flow cytometry was performed for macrophage markers CD68 and CD163, and for the antigen-presenting cell marker, MHC class II. Although no significant change was observed in the number of CD68+ and CD163+ macrophages during RMM progression, the number of MHC class II+ antigen-presenting cells was reduced in 3 cm nodules. Real-time RT-PCR of laser microdissection samples obtained from RMM regions rich in MHC class II+ cells demonstrated high expressions of M1-like factors: IFN-γ, GM-CSF and IL-12a. Furthermore, fluorescence-activated cell sorting, followed by real-time RT-PCR for CD11b+ MHC class II+ (myeloid antigen-presenting cells), CD11b+ CD163+ (M2 type myeloid cells), CD11b+ CD80+ (M1 type myeloid cells) and CD11b+ CD11c+ (dendritic cells) cells was performed. Based on the levels of inflammation- and tumor progression-related factors, MHC class II+ antigen-presenting cells showed polarization towards M1, while CD163+ macrophages, towards M2. CD80+ and CD11c+ myeloid cells did not show clear functional polarization. Our results provide novel information on tumor-associated myeloid cells in amelanotic melanoma, and may become useful in further research on melanoma immunity.

Cutaneous Histiocytic Sarcoma with Regional Lymph Node Metastasis in a Netherland Dwarf Rabbit (Oryctolagus cuniculus).

A 10-year-old male Netherland dwarf rabbit (Oryctolagus cuniculus) was presented with a red nodular mass (1 cm in diameter) with ulceration and hair loss in the skin of the left upper lip. Cytological examination revealed atypical round cells. The mass was excised surgically. Histologically, the mass was composed of large round to polyhedral neoplastic cells with marked cytological atypia. The neoplastic cells were often binucleated or multinucleated. Immunohistochemically, the neoplastic cells were intensely positive for Iba1 and vimentin, but fewer neoplastic cells expressed E-cadherin. Nuclear immunoreactivity for Ki67 was detected in approximately 41% of the neoplastic cells. Metastasis to the left cervical lymph nodes was detected 6 months after the surgical excision. Based on clinical, histopathological and immunohistochemical findings, the present case was diagnosed as cutaneous histiocytic sarcoma. To the authors' knowledge cutaneous histiocytic disease has not been reported previously in lagomorphs.

Metastatic Liposarcoma in a South African Fur Seal (Arctocephalus pusillus).

A 14-year-old female South African fur seal (Arctocephalus pusillus) was presented with a large skin mass on the right shoulder. At necropsy examination, multiple white nodules were found in the lungs, liver, spleen and right axillary lymph nodes. Histologically, the skin mass was composed of round to polygonal neoplastic cells with round to oval nuclei and variably sized cytoplasmic vacuoles. Cellular and nuclear atypia were prominent. Immunohistochemically, the neoplastic cells expressed vimentin, but not cytokeratins, S100 protein, adipophilin or desmin. The cytoplasmic lipid droplets stained positively with oil red O. Metastasis was seen in the lungs, liver, spleen and right axillary lymph nodes, with similar morphological features to the skin mass. Based on these findings, a diagnosis of a pleomorphic liposarcoma with systemic metastasis was made. No previous reports of metastatic liposarcomas have been published in marine mammals.

Multiple Histiocytic Foam Cell Nodules in the Tongue of Miniature Dachshund Dogs.

Miniature dachshund dogs are a common breed in Japan and are known to be predisposed to granulomatous diseases. Here we report the pathologic features of multiple lingual nodules in 7 miniature dachshunds. Seven dogs had multiple nodules of variable sizes mainly on the ventral and lateral surface of the tongue. In addition, 1 dog also had masses on the left oral mucosa. Three cases had recurrence after surgical resection. Histologically, the lingual nodules were composed of aggregates of foam cells with clear vacuolated cytoplasm that were negative for oil red O, PAS, and alcian blue. They stained positively for CD204 (macrophage scavenger receptor) and MHC class II and negatively for Iba-1, E-cadherin, adipophilin, cytokeratins, S-100, and nestin. These findings indicate that the multiple lingual nodules in miniature dachshunds are an unusual, unique lesion consisting of macrophage-derived foam cells, which does not correspond to canine lingual diseases reported to date.

Myoepithelioma of the gland of the third eyelid in a dog.

A 12-year-old female miniature dachshund was presented with a tan-white, firm mass (4 × 3 × 2 cm) occupying the left medial canthus. The mass compressed and displaced the left eye dorsally, and it was surgically removed. Microscopically, the mass was composed of interlacing bundles of spindle cells with clear cytoplasm and a small number of atypical glandular epithelial cells. Immunohistochemically, the spindle cells expressed p63, α-smooth muscle actin and calponin, and were negative for cytokeratin AE1/AE3. The glandular epithelial cells expressed cytokeratin AE1/AE3. Based on these findings, this case was diagnosed as a myoepithelioma of the gland of the third eyelid.

A collision tumour consisting of malignant trichoblastoma and melanosarcoma in a rabbit.

A 7-year-old mixed breed neutered female rabbit (Orytolagus cuniculus) developed a solitary black nodular mass (1 cm in diameter) in the skin of the right flank. Microscopically, the mass consisted of an admixture of neoplastic trichoblasts and melanocytes. The former were arranged as solid, trabecular, island-like and gland-like structures and the cells had oval nuclei with prominent nucleoli and lightly eosinophilic scant cytoplasm. The latter population exhibited prominent nuclear atypia and high mitotic index in the clusters of a few cells or single cells. Immunohistochemically, the neoplastic trichoblasts expressed cytokeratins and E-cadherin, while the neoplastic melanocytes expressed vimentin, S100 protein, melan-A and melanoma antigen. A diagnosis of collision tumour involving malignant trichoblastoma and melanosarcoma was made.

Littoral cell angiosarcoma in a dog.

This report describes the microscopical and immunohistochemical characteristics of littoral cell angiosarcoma in a 12-year-old, neutered female, beagle dog. The dog succumbed to metastatic disease 3 months after diagnosis of a mid-splenic mass. The tumour was characterized by two histological patterns: anastomosing microvascular channels and microvascular papillary fronds. The neoplastic cells expressed both endothelial and histiocytic markers and were erythrophagocytic. Immunohistochemical findings consistent with malignancy were CD34 expression and high Ki67 nuclear immunoreactivity.

Immunophenotypical characterization of macrophages in rat bleomycin-induced scleroderma.

Scleroderma is a skin disorder characterized by persistent fibrosis. Macrophage properties influencing cutaneous fibrogenesis remain to be fully elucidated. In this rat (F344 rats) model of scleroderma, at 1, 2, 3, and 4 weeks after initiation of daily subcutaneous injections of bleomycin (BLM; 100 µl of 1 mg/ml daily), skin samples were collected for histological and immunohistochemical evaluations. Immunohistochemically, the numbers of cells reacting to ED1 (anti-CD68; phagocytic activity) and ED2 (anti-CD163; inflammatory factor production) began to increase at week 1, peaked at week 2, and decreased thereafter. In contrast, the increased number of cells reacting to OX6 (anti-MHC class II molecules) was seen from week 2 and remained elevated until week 4. α-Smooth muscle actin-positive myofibroblasts were increased for 4 weeks. Double labeling revealed that galectin-3, a regulator of fibrogenic factor TGF-ß1, was expressed in CD68+, CD163+, and MHC class II+ macrophages and myofibroblasts. mRNA expression of TGF-ß1, as well as MCP-1 and CSF-1 (both macrophage function modulators), were significantly elevated at weeks 1 to 4. This study shows that the increased number of macrophages with heterogeneous immunophenotypes, which might be induced by MCP-1 and CSF-1, could participate in the sclerotic lesion formation, presumably through increased fibrogenic factors such as galectin-3 and TGF-ß1; the data may provide useful information to understand the pathogenesis of the human scleroderma condition.

Heterogeneity of macrophage populations and expression of galectin-3 in cutaneous wound healing in rats.

The aim of this study was to investigate the properties of macrophages that infiltrated the sites of cutaneous wound healing in rats between 1 and 26 days post wounding (dpw). During the inflammation phase (1-3 dpw), ED1(+) (CD68(+)) macrophages with enhanced lysosomal activity dominated. From 5 to 7 dpw there was formation of granulation tissue as indicated by the presence of myofibroblasts expressing α-smooth muscle actin. At this stage, ED2(+) (CD163(+)) macrophages, capable of producing inflammatory factors, were dominant. The majority of ED1(+) macrophages expressed galectin-3, a regulator of fibrosis. Corresponding to the increased numbers of ED1(+) and ED2(+) macrophages at 3-9 dpw, there was increased expression of genes encoding transforming growth factor-ß1 (a major fibrogenic factor), monocyte chemoattractant protein-1 and colony stimulating factor-1. These macrophage-related factors might contribute to inflammation and formation of granulation tissue. OX6(+) macrophages expressing class II molecules of the major histocompatibility complex became predominant in the healing stages (15-26 dpw), indicating important roles for antigen-presenting cells in tissue remodelling. The OX6(+) macrophages were most likely derived from ED1(+) macrophages. The results of this study show that infiltration of phenotypically- and functionally-distinct macrophage populations characterizes different stages of the wound healing process.

Cardiac hamartoma in a young squirrel monkey who died suddenly.

A case of cardiac hamartoma in a 2-month-old squirrel monkey is reported. The monkey showed a loss of appetite and died suddenly. Microscopically, an encapsulated nodular lesion was found at the right atrial wall. The lesion consisted of irregularly shaped, slender myocytes intermingled with a few fibroblasts and collagen fibers. Neither nuclear atypia nor inflammatory cell infiltrate was seen. The constituting cells had stratified striations in the cytoplasm and reacted immunohistochemically for desmin, indicating the nature of myocytes. Based on the above findings, a diagnosis of cardiac hamartoma was made. This is the first case of cardiac hamartoma in this species.

Malignant seminoma with multiple visceral metastases in a guinea fowl (Numida meleagris) kept in a zoo.

A case of seminoma found in an adult guinea fowl (Numida meleagris) that has been exhibited in a zoo is reported. The right testis was extremely enlarged and replaced by round and polyhedral pleomorphic neoplastic cells showing nest, sheet, and diffuse growth patterns. The neoplastic cells had acidophilic cytoplasm and hyperchromatic and eccentrically placed nuclei. Metastatic lesions composed of diffuse growth of neoplastic cells similar to those of the primary tumor were seen in the liver, lungs, kidneys, and heart, and neoplastic emboli were often detected within blood vessels of these organs, indicating hematogenous metastasis. This is the first report of malignant seminoma with multiple metastases in the visceral organs in the guinea fowl.

Vasoformative disorder, resembling littoral cell angioma, of the spleen in a geriatric Japanese macaque (Macaca fuscata).

A 30-year-old female Japanese macaque showed marked splenomegaly. The enlarged spleen consisted of neoplastic proliferation of anastomosing vascular channels resembling morphologic structures of red pulp sinuses; occasionally, papillary fronds were seen in dilated channels. Immunohistochemically, the lining cells reacted to both endothelial cell (von Willebrand factor) and macrophage (macrophage scavenger receptor class A) markers, indicating features of littoral cells of the spleen. Based on the pathologic characteristics, particularly the presence of neoplastic cells with macrophage/histiocyte-like attributes, this tumor was regarded as littoral cell angioma; this is a rare benign splenic vascular tumor.

Macrophage populations and expressions of regulatory proinflammatory factors in the rat meninx under lipopolysaccharide treatment in vivo and in vitro.

Macrophages play important roles in host defense mechanisms. In the brain, besides microglial cells, meningeal macrophages are present. However, the pathobiological characteristics of meningeal macrophages in rats remain to be investigated. In normal meninx, immunohistochemically, macrophages reacting to CD163 (macrophage scavenger receptor) and major histocompatibility complex (MHC) class II-expressing cells (involving activated macrophages or dendritic cells) were sporadically seen without age-dependent changes. Injection of lipoplysaccharide (LPS) (5 microg; Escherichia coli) into the cerebrum increased the number of anti-CD68-positive macrophages (with greater phagocytic activity) in the meninx, with a peak at 12 h during observation period until 48 h; MHC class II-expressing cells showed a gradual increase in number from 3 h after injection; however, anti-CD163-positive macrophages did not show significant change. In in vitro studies, LPS (0, 0.02, 0.05, 0.5, 5, 50 and 100 microg/ml) was added to KMY-1 or KMY-2 cells, both of which had been established from a rat malignant meningioma. KMY-1 originally reacted to CD163, but LPS addition at 0.5 microg/ml and greater concentrations decreased the anti-CD163-positive cell number and instead increased the anti-CD68-positive cell number. LPS-treated KMY-2 increased the anti-CD163-positive cell number at 0.05 and 0.5 microg/ml. By RT-PCR methods, LPS (0, 0.5, 5, 50, and 100 microg/ml)-treated KMY-1 and KMY-2 showed an increase in mRNA of monocyte chemoattractant protein-1 (MCP-1, a chemokine), and LPS-treated KMY-2 increased mRNA of nerve growth factor (NGF, an immunological effecter). Collectively, under LPS treatment, macrophages with heterogeneous functions appear in rat meninx; rat meninx-forming cells may be involved in pathogenesis of meningeal inflammation by expressing different immunophenotypes and by producing regulatory proinflammatory factors such as MCP-1 and NGF.

Cutaneous rhabdoid tumor in a cat.

Rhabdoid tumor is a highly aggressive neoplasm of unknown cellular origin in humans, usually occurring in the kidney and central nervous system of infants or children. In older patients, it occurs rarely in other organs, including the skin and soft tissues. A subcutaneous mass in a 13-year-old male mixed-breed cat was composed of nests or sheets of round to polygonal cells with glassy eosinophilic cytoplasmic inclusions. Immunohistochemically, many neoplastic cells expressed vimentin, localized to the cytoplasmic inclusions, whereas the cytoplasm of some neoplastic cells was diffusely positive for neuron-specific enolase, neurofilament, or S-100 protein. By electron microscopy, the cytoplasmic inclusions were found to be composed of aggregates of intermediate filaments. These findings are quite similar to the histologic, immunohistochemical, and ultrastructural features of human rhabdoid tumors and the few rhabdoid tumors reported in animals.

Pancreatic metaplasia in the gastro-achlorhydria in WTC-dfk rat, a potassium channel Kcnq1 mutant.

The WTC-deafness Kyoto (dfk) rat is a new mutant characterized by deafness and abnormal, imbalanced behavior. WTC-dfk rats carry an intragenic deletion at the Kcnq1 gene; KCNQ1 plays an important role in K(+) homeostasis, and the mutation of Kcnq1 causes a cardiac long QT syndrome in humans. Here, we studied stomach lesions in these WTC-dfk rats. The most characteristic pathologic feature in the stomach was the appearance of hypertrophic gastric glands in the stomach body. The hypertrophic cells had many eosinophilic granules in their cytoplasm, and these granules were stained red with Azan stain; stained positively for trypsinogen, amylase, and chymotrypsin; and did not stain positively for pepsinogen when using immunohistochemical analysis. These staining results suggested a metaplasia toward a pancreatic acinar cells. Extensive fibrosis was found in the bottom part of the mucosa of 34-week-old WTC-dfk rats, suggesting a progression of stomach lesions with aging. Although cells that were positive for proliferating cell nuclear antigen were restricted in the area of the glandular neck in WTC control rats, positive cells in WTC-dfk rats were scattered throughout the mucosa. The parietal cells in WTC-dfk rats were negative for KCNQ1 immunohistochemical analysis. These findings indicate that a deficiency in rat Kcnq1 provokes an abnormal proliferation and differentiation of gastric glandular cells.

Neoplastic and non-neoplastic cell lines from a malignant peripheral nerve sheath tumour of the cervix of a rat.

A homotransplantable tumour (LY) and cell lines (LY-PPB6 and LY-H12) were established from a spontaneous malignant peripheral nerve sheath tumour (PNST) of the uterine cervix of an F344 rat. Primary and LY tumours consisted of oval or spindle-shaped cells arranged in a flatfield or streaming fashion, and indistinct nuclear palisades were seen. Immunohistochemically, neoplastic cells reacted to vimentin, S-100 protein, neuron-specific enolase (NSE), myelin basic protein (MBP), and glial fibrillary acidic protein (GFAP) in varying degrees, indicating neurogenic derivation. LY-PPB6-induced tumours in syngeneic rats developed cellular whorling patterns reacting particularly strongly to S-100 protein, NSE, MBP and GFAP. Nerve growth factor (NGF) mRNA expression was shown in LY-PPB6 cells by the reverse transcription-polymerase chain reaction (RT-PCR). By contrast, LY-H12 had a normal chromosomal number of 42, and did not produce tumours when injected into syngeneic rats. LY-H12 cells reacted to vimentin and alpha-smooth muscle actin (alpha-SMA), and the alpha-SMA-positive cell number was increased dose-dependently by the addition of transforming growth factor (TGF)-beta1, indicating a myofibroblastic nature. LY-PPB6 cells were neoplastic with properties of PNS cells, whereas LY-H12 cells were non-neoplastic stromal cells showing myofibroblastic differentiation. As TGF-beta1 mRNA expression was shown in both LY-PPB6 and LY-H12 cells by the RT-PCR, the myofibroblastic phenotype of LY-H12 cells may be mediated by paracrine or autocrine signalling in tumour tissues. LY-PPB6 and LY-H12 may prove useful for studies on the pathobiological nature of neoplastic cells and interactions between neoplastic and stromal cells in PNSTs.

Characterization of a rat subcutaneous malignant fibrous histiocytoma and its tumor lines, with reference to histiocytic features.

Malignant fibrous histiocytoma (MFH) is regarded as soft tissue-derived undifferentiated pleomorphic sarcoma, of which the histogenesis remains to be proven. To investigate the cellular characteristics, a homotransplantable tumor line (KJ) was established from a spontaneous MFH that developed in the subcutis of an aged F344 rat. KJ tumors have been produced in syngeneic rats by serial subcutaneous implantation of tissue fragments. The original and KJ tumors consisted of oval and fusiform cells arranged in interlacing bundles with fibrous stroma. Occasional giant cells with bizarre nuclei were observed. Enzyme/immunohistochemically, neoplastic cells reacted to ED1 and ED2 (antibodies specific for rat histiocytes/macrophages), and showed a positive reaction to vimentin and lysosomal enzyme markers such as acid phosphatase (ACP) and nonspecific esterase (Non-SE). Electron microscopically, neoplastic cells possessed lysosomal granules in cytoplasm. A cloned cell line (KJ-A) was isolated from a KJ tumor. KJ-A cells showed positive reactions to ED1, ED2, ACP, and Non-SE, and had cytoplasmic lysosomal granules. Tumors induced by KJ-A cells exhibited histologic and enzyme/immunohistochemical findings similar to those of KJ tumors. Lipopolysaccharide (LPS) treatment increased the number of ED1-positive cells and the expression of tumor necrosis factor-alpha mRNA by reverse transcription-polymerase chain reaction. Collectively, it is likely that rat MFH cells originally possess histiocyte/macrophage-like features that may be enhanced by LPS. Because tumor lines are useful for in vivo and in vitro studies concerning different characteristics of the original neoplasms. KJ and KJ-A should prove useful for studies concerning the morphogenesis of MFH.