Extranodal gammadelta-T-cell lymphoma in a dog with leishmaniasis

An 8-year-old intact male mongrel dog with alopecia and weight loss was referred to the Veterinary Faculty of Naples. The dog had pale mucous membranes, enlarged prescapular lymph nodes, and splenomegaly. Laboratory abnormalities included anemia, thrombocytopenia, and hyperglobulinemia. Bone marrow aspirate smears contained numerous Leishmania amastigotes and an immunofluorescent antibody titer was strongly positive (1:1280) for leishmaniasis. The dog was treated with a combination of meglumine antimoniate and allopurinol for 60 days and showed clinical improvement. Two months after the end of treatment the dog was again referred because of relapse of leishmaniasis and the presence of a firm subcutaneous mass on the medial right thigh. Based on cytologic examination of fine needle aspirates of the mass, a diagnosis of large-cell lymphoma was made. Flow cytometry of tumor cells revealed gammadelta-T-cell lymphoma with a CD5+, CD3+, TCRgammadelta+, CD4-, CD8-, CD45RA+ immunophenotype. Using nested PCR, amastigotes were not detected in the neoplastic tissue. An association between leishmaniasis and hematopoietic tumors has been described rarely. gammadelta-T cells may be involved in the host response to this parasite, and prolonged antigenic stimulation and chronic immunosuppression (typical of leishmaniasis) play a crucial role in the etiopathogenesis of T-cell lymphoma.     

Cutaneous lymphoma in a juvenile dog

Abstract: An 18-month-old male Doberman Pinscher was referred to the Veterinary Medical Teaching Hospital of the College of Veterinary Medicine for an erythemic nodular mass on the right forelimb. The mass was diagnosed as cutaneous lymphoma, based on cytologic examination of a mass aspirate and histopathology. Using immunohistochemistry the neoplastic cells were positive for CD3 but negative for CD79a, E-cadherin, and pancytokeratin, confirming their origin as T lymphocytes. No tumor recurrence was noted 18 months after surgery. To our knowledge, this is the first report of a solitary nodular form of cutaneous lymphoma in a young dog.     

Gamma delta T‐cell large granular lymphocyte lymphoma in a dog

A 2‐year and 6‐month‐old female neutered Labrador Retriever with Horner syndrome, megaesophagus, and a mediastinal mass was referred to the Queen Mother Hospital for Animals of the Royal Veterinary College. A large granular lymphocyte (LGL) lymphoma was diagnosed on cytology; flow cytometric analysis revealed a γδ T‐cell phenotype (CD3+, CD5+, CD45+, TCRγδ+, CD4?, CD8?, CD34?, CD21?). Chemotherapy was started with a combination of lomustine, vincristine, procarbazine, and prednisolone, followed by bleyomicin. Euthanasia was elected by the owners, due to progressive deterioration and lack of quality of life, 28 days after diagnosis. This is the first cytologic and immunophenotypic characterization of a canine γδ T‐cell lymphoma with LGL morphology and probably of mediastinal origin. The role of chemotherapy in delaying the disease progression remains unknown.     

Age-related changes in leukocytes and T cell subsets in peripheral blood of Japanese Black cattle

It is well known that the immune system changes with age during development and maturation in Holstein cattle. But age-related changes in leukocytes and T cell subsets in peripheral blood of Japanese Black cattle still remain unclear. The aim of the present study was to investigate comparative changes of leukocytes (granulocytes, monocytes, B cells and T cells) and T cell subsets (CD4⁺, CD8⁺, γδ, CD8⁺γδ and WC1⁺γδ T cells) in Japanese Black cattle aged 0.5, 1, 2, 6, 18 and 36–41 (adult) months on flow cytometry using specific monoclonal antibodies for the cell surface markers. T cell proportion was approximately 40% in 2-month-old cattle and decreased to 20.6% in adults. In contrast, B cell proportion significantly increased from 7.4% to 28.2% with age. In T cell subsets the percentage of CD4⁺ T cells significantly increased from 40.5% to 60%, but that of WC1⁺γδ T cell subset significantly decreased with age. The percentages of CD8⁺ and CD8⁺γδ T cells did not change. The present study details the proportional changes in leukocyte and T cell subsets with age in the peripheral blood of Japanese Black cattle and these findings are similar to those described for Holstein cattle.     

Differential cytokine expression in T cell subsets from bovine peripheral blood

Although distinct cytokine expression in T cell subsets is well understood in mice and humans, limited information is available on bovine T cell subsets. In the present study, we analyzed the mRNA expression of 10 kinds of cytokines and CD25 expression in CD4⁺, CD8⁺, WC1⁺ and WC1^-γδ T cell subsets in bovine peripheral blood by Concanavalin A (Con A) stimulation. CD25 expression was significantly increased in CD4⁺, CD8⁺ and WC1⁺γδ T cells, but not in WC1^-γδ T cells by Con A stimulation. In CD4⁺ T cells, the mRNAs of Interleukin (IL)-2, IL-6, IL-10, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, TNF-β and transforming growth factor (TGF)-β were expressed in control cultures, and IL-3, IL-4 and granulocyte-macrophage colony stimulating factor (GM-CSF) were newly expressed when the cells were stimulated with Con A. CD8⁺ T cells expressed the mRNAs of IL-6, TNF-α, TNF-β and TGF-β in control cultures, and newly expressed those of IL-2, IFN-γ and GM-CSF, but did not express those of IL-3, IL-4 or IL-10 after Con A stimulation. The cytokine expression profile of WC1⁺γδ T cells was similar to that of CD8⁺ T cells. However, WC1^-γδ T cells did not express any cytokine mRNA except TGF-â mRNA. These results will contribute to elucidate the participation of T cell subsets in immune responses against infectious disease in cattle.     

Cutaneous presentation of canine intravascular lymphoma (malignant angioendotheliomatosis)

A 9-year-old female spayed Boxer dog presented with variably sized, firm, black, raised, exudative subcutaneous masses on her head, neck and trunk, that tended to fluctuate in size and frequently ulcerate. Skin biopsy showed that the dermis was expanded by a densely cellular mass of proliferative capillaries distended with large pleomorphic neoplastic round cells mixed with fibrin and erythrocytes. Intravascular lymphoma was diagnosed and immunostains were compatible with a CD8⁺ T lymphocyte histogenesis (CD3⁺/CD79a⁻/TCRαβ⁺/CD8α⁺). Post-mortem examination, four months after diagnosis, revealed neoplastic T-cells within meningeal arteries. We are unaware of other reports of a cutaneous presentation and ante-mortem diagnosis of intravascular lymphoma in the dog. Additionally, this vasoproliferative form of intravascular lymphoma has not been previously described in dogs.     

B‐cell lymphoma with plasmacytoid differentiation,atypical cytoplasmic inclusions,and secondary leukemia in a dog

A 7‐year‐old male castrated Jack Russell Terrier was presented to the oncology service at the University of California–Davis Veterinary Medical Teaching Hospital for evaluation of suspected lymphoma. The dog had several enlarged lymph nodes and moderate lymphocytosis. Aspirates of an enlarged inguinal lymph node contained a bimorphic population of large immature lymphocytes and smaller cells with plasmacytoid features. Both cell types often contained a single large cytoplasmic inclusion that varied from clear to pale pink to sky blue. Cytologic changes were interpreted as most consistent with lymphoid neoplasia. Based on the predominantly mature cell morphology and some morphologic heterogeneity, the peripheral lymphocytosis was interpreted as most likely reactive in nature. However, the immunophenotype of the cells (CD20+, CD21+, CD79a+, MUM‐1+, and MHCII+) and clonality assays showed that tissue and blood lymphocytes were neoplastic B cells with clonal identity despite their different morphologic appearances. The cytoplasmic inclusions were positive with periodic acid‐Schiff and were immunoreactive for IgM and IgG. By transmission electron microscopy, inclusions consisted of aberrant rough endoplasmic reticulum; a few small Russell bodies were also noted. A final diagnosis of high‐grade B‐cell lymphoma with plasmacytoid differentiation, atypical cytoplasmic inclusions, and secondary leukemia was made. Chemotherapy was initiated, but the dog was euthanized due to severe and uncontrolled seizures 9 months after the initial diagnosis. This case extends the morphologic repertoire of canine plasmacytoid neoplasms and emphasizes their continuum with multicentric lymphoma. This case also demonstrates the need for advanced diagnostic techniques in establishing blood involvement in lymphoma in some instances.     

Cervical thymoma originating in ectopic thymic tissue in a cat

Abstract: An 11‐year‐old female spayed domestic shorthair cat was referred to The Ohio State University Veterinary Teaching Hospital (OSU‐VTH) for evaluation of a 6 × 4 × 3.5 cm mass in the left midcervical region causing increased respiratory sounds and lateral deviation of the trachea. A fine needle aspirate of the mass was obtained before referral and the cytology results were compatible with a reactive lymph node. Immunocytochemistry showed increased numbers of CD3+ T lymphocytes and small numbers of CD20+ and CD79a+ medium to large lymphocytes. Differential diagnoses from the referral pathologist were T‐cell‐rich B‐cell lymphoma and feline Hodgkin's‐like lymphoma. A subsequent fine needle aspirate performed at the OSU‐VTH showed similar results. On flow cytometry the majority of cells were CD3+ T lymphocytes that were double positive for CD4 and CD8 (73%), compatible with either a double‐positive (CD4+CD8+) T‐cell lymphoma or lymphocytes from ectopic thymic tissue. The mass was surgically removed. Histopathology and immunohistochemistry of the mass revealed a predominant population of CD3+ small lymphocytes and small numbers of medium to large lymphocytes with moderate anisocytosis and anysokaryosis. A population of cytokeratin‐positive epithelial cells surrounded small microcystic structures filled with eosinophilic material and structures interpreted as Hassall's corpuscles. These findings were consistent with thymic tissue and a diagnosis of ectopic thymoma was made. PCR results for lymphocyte antigen receptor rearrangement (PARR) were negative. The cat had no evidence of disease 16 months after removal of the mass. To our knowledge this is the first report of an ectopic cervical thymoma in a cat. The clinical and diagnostic features of this unusual case will be useful in helping veterinarians and pathologists obtain a presurgical diagnosis and establish a prognosis for similar lesions.     

Canine leishmaniasis in Northern California—A case report

A 3‐year‐old dog was referred to the Veterinary Medical Teaching Hospital of the University of California‐Davis for further evaluation of episodes of epistaxis of 1‐year duration and peripheral lymphadenopathy. The patient had a history of atopic dermatitis with no travel history outside of California. Hyperglobulinemia with a polyclonal gammopathy was noted on serum protein electrophoresis. Microscopic evaluation of a bone marrow aspirate sample revealed many free and intra‐cellular amastigotes of Leishmania sp. that was further confirmed by qPCR as L infantum. This is, to the best of our knowledge, the first reported case of canine leishmaniasis in the state of California. The patient is believed to have been vertically infected from the dam who is from Serbia and remained subclinical until presentation. Because the clinical progression of leishmaniasis is variable, it is important that precautions be discussed with owners acquiring puppies with dams from endemic regions of leishmaniasis to prevent zoonotic exposure in states where competent vectors are present.     

Cutaneous T-cell lymphoma with Sézary syndrome in a dog

An 8-year-old female spayed Cocker Spaniel mix breed dog was presented with generalized erythroderma, scaling and alopecia. Radiographs of the thorax demonstrated a discrete lung mass which was aspirated using ultrasound guidance and cytological analysis revealed large abnormal lymphocytes. Similar cells were observed in the peripheral blood and in skin biopsies. The cells in the skin biopsies were epidermotropic, indicative of an uncommon cutaneous lymphoma termed cutaneous T cell lymphoma (CTCL), sometimes also called mycosis fungoides. Immunohistochemical staining of a skin biopsy was positive for the CD3 antigen demonstrating that the lymphocyte infiltrate was of a T-cell lineage. The presence of neoplastic lymphocytes in the epidermis and peripheral blood indicate that this is a rare variant of Cutaneous Epidermotropic Lymphoma (CEL) called Sézary syndrome based on nomenclature used in the human literature. An unusual feature of this dog, not seen in previous cases, was the presence of a discrete neoplastic lung mass.     

Mediastinal lymphoma in a young Turkish Angora cat

An 8-month old intact male Turkish Angora cat was referred to the Veterinary Medical Teaching Hospital (VMTH), Seoul National University, for an evaluation of anorexia and severe dyspnea. The thoracic radiographs revealed significant pleural effusion. A cytology evaluation of the pleural fluid strongly suggested a lymphoma containing variable sized lymphocytes with frequent mitotic figures and prominent nucleoli. The feline leukemia virus and feline immunodeficiency virus tests were negative. The cat was euthanized at his owner''s request and a necropsy was performed. A mass was detected on the mediastinum and lung lobes. A histopathology evaluation confirmed the mass to be a lymphoma. Immunohistochemistry revealed the mass to be CD3 positive. In conclusion, the cat was diagnosed as a T-cell mediastinal lymphoma.     

Cellular immunophenotyping of exfoliative dermatitis in canine leishmaniosis (Leishmania infantum)

Lymphocyte subsets, major histocompatibility complex (MHC)-II expressing cells and number of amastigotes in the epidermis and dermis were investigated immunohistochemically in 48 dogs with patent leishmaniosis, with or without exfoliative dermatitis (ED) to study the immunopathogenesis of this common cutaneous form of the disease. Skin biopsies were obtained and compared for ED sites (group A, n = 26), normal-appearing skin from the same animals (group B, n = 24), and leishmanial dogs not exhibiting ED (group C, n = 22), and normal controls (group D, n = 22). The CD3+, CD45RA+, CD4+, CD8+ (CD8a+), CD21+, and MHC-II+ cells and leishmania amastigotes were identified immunohistochemically and counted with the aid of an image analysis system. Pyogranulomatous to granulomatous dermatitis, expressed in various histopathological patterns, was noticed in all groups A and B and in half of group C dogs. In the epidermis, the low number of T-cells and their subsets did not differ significantly between groups A and B, but CD8+ outnumbered CD4+ lymphocytes in both groups. MHC-II+ expression on epidermal keratinocytes was intense in the skin with and without lesions from dogs with ED but not in group C dogs. CD3+, CD8+ and MHC-II+ cells were fewer in group C compared to group A and B dogs. In the dermis, CD3+ cells in group A animals were mainly represented by the CD8+. CD45RA+ and CD21+ cells were also seen in high numbers. MHC-II expression, potentially in lymphocytes, fibroblasts, dendritic cells, and macrophages was intense. The numbers of all cellular subpopulations in the dermis were significantly different between the groups, being highest in group A and lowest in group D. In sebaceous adenitis sites, CD4+ outnumbered CD8+ cells in contrast to the neighbouring dermis and the epidermis. The number of CD21+ and CD45RA+ cells was much lower in the inflamed sebaceous glands compared to the dermis. Finally, the number of amastigotes in the normal-appearing skin was significantly higher in the ED dogs (group B) than in those not exhibiting this cutaneous form of the disease (group C).     

Flow cytometric immunophenotype of canine lymph node aspirates

Increasing availability of reagents able to distinguish subtypes of lymphocytes and other leukocytes has enabled greater understanding of lymphocyte biology and pathology in the dog. Lymphocytes in circulation most commonly are subjected to immunophenotypic assessment by flow cytometry, but needle aspirates of lymph nodes can be similarly suitable for immunophenotypic examination. In this investigation, the feasibility of immunophenotyping samples obtained by needle aspiration of lymph nodes from 32 dogs with no physical abnormalities and 6 dogs with lymphoma was determined. In addition, samples from 6 dogs were stored overnight at 4 degrees C and reanalyzed 24 hours later. For each sample, stained smear preparations were examined microscopically for lymphocyte morphology, neoplasia, and the presence of inflammatory cells. Expression of antigens on a corresponding sample of aspirated cells was determined by flow cytometric detection of antibody binding on a minimum of 10,000 events. The distribution of data was determined with Anderson-Darling tests, and reference intervals incorporating the central 95% of values were established. Adequate samples were obtained from 30 of 32 clinically normal dogs. Immunophenotypic results after 24 hours of storage were consistent with those obtained immediately after sampling. Reference intervals for lymphocyte subsets from normal dog lymph nodes were similar to the proportions of CD3+, CD4+, CD8+, and CD21+ lymphocytes found in blood. Aspirates of enlarged lymph nodes from dogs with lymphoma were readily classified by this technique. Aspiration of lymph nodes from dogs for comprehensive analysis by flow cytometry is feasible and applicable to immunophenotyping of lymphoma.     

Visceral leishmaniasis with cardiac involvement in a dog: a case report

A dog presented with cutaneous nodules, enlarged lymph nodes and oedema in limbs, face and abdomen. The diagnosis of visceral leishmaniasis was established by identification of Leishmania amastigotes within macrophages from skin and popliteal lymph node biopsies. At necropsy, lesions were found in different organs, but it was particularly striking to observe large areas of pallor in the myocardium. Histological examination revealed an intense chronic inflammatory reaction in many organs, and numerous macrophages were found to contain amastigote forms of Leishmania. The inflammatory reaction was especially severe in the heart, where large areas of the myocardium appeared infiltrated with huge numbers of mononuclear immune cells, causing cardiac muscle atrophy and degeneration. Despite the severe inflammation, the number of parasitized macrophages was low in the myocardium, as revealed by immunohistochemical staining of Leishmania amastigotes. Because cardiac involvement is not usually described in this condition, this dog represents a very rare case of canine visceral leishmaniasis with affection of the myocardium.     

Progression of an orbital T-cell rich B-cell lymphoma to a B-cell lymphoma in a dog

An 11-year-old Shetland Sheepdog was presented for exophthalmos caused by a locally extensive, poorly defined mass located behind the right eye. The primary orbital mass was identified by light microscopy and immunohistochemistry as a T-cell rich B-cell lymphoma (TCRBCL) composed predominantly of BLA.36-positive large neoplastic lymphoid cells admixed with fewer CD3- and CD79a-positive small lymphocytes. The dog was treated for lymphoma, but 6 months after presentation it was euthanatized for suspected hepatic and gastrointestinal metastasis. Gross findings revealed an enlarged liver with multiple well-demarcated, randomly distributed 0.1-1.5-cm white nodules, five firm white submucosal jejunal nodules, and ileocecal, mediastinal, and hilar lymphadenopathy. Metastatic liver lesions consisted of sheets of monomorphic large neoplastic lymphoid cells that effaced and expanded portal and centrilobular zones. These cells were morphologically similar to the large neoplastic cells of the original orbital tumor and were CD3-negative and variably BLA.36-positive, consistent with B-cell lineage. Similar cells comprised the jejunal nodules and effaced the lymph nodes. The progression of TCRBCL to a diffuse B-cell lymphoma in this case is consistent with reported human cases and has not been previously reported in the dog.     

Cellular immune response of lymph nodes from dogs following the intradermal injection of a recombinant antigen corresponding to a 66 kDa protein of Echinococcus granulosus

A recombinant polypeptide (referred to as EgA31), which represents a 66kDa protein, was prepared from an Echinococcus granulosus cDNA library. In order to assess its potential to induce cellular immune responses, dog popliteal and prescapular lymph nodes were sensitized with this recombinant polypeptide. Subpopulations of lymphocytes were then analyzed by flow cytometry and immunohistochemistry on lymph node sections. Five days after the sensitization, the paracortical areas of the lymph nodes appeared hypertrophic, the number of CD3+, CD4+, CD8+ and CD5+ cells increased, the number of B-cells began to augment and some secondary follicles occurred, and a number of CD4+ cells appeared in germinal centers. Many large secondary follicles and a significantly augmented number of CD5+ cells in cords of medullae were observed 10 days after the sensitization. These active cellular responses strengthen the interest for further studies on the development of a vaccine with this recombinant polypeptide.     

High-grade canine T-cell lymphoma/leukemia with plasmacytoid morphology: a clinical pathological study of nine cases.

The aim of this study is to determine the clinical, morphological, and immunophenotypical presentation of 9 cases of a particular type of canine T-cell lymphoma/leukemia. The morphological presentation was a diffuse infiltration of small, medium-sized, or large blast cells with eccentric nuclei, hyperbasophilic cytoplasm, and a juxtanuclear, pale cytoplasmic area, giving a plasmacytoid appearance and suggesting a B-cell morphology. Surprisingly, all 9 cases were of T-cell phenotype (CD3+). Among the 7 immunophenotyped cases, 4 were CD4-/CD8+, 2 CD8+/CD4+, and 1 CD4+/CD8-. The median Ki-67 index was 65.7%, which placed this lymphoma in the high-grade group. This type of lymphoma/leukemia was found in dogs between 1 and 11 years of age, with a median age of 5.8. The male-female ratio was 0.8 for a reference population of 1.04. The most significant clinical findings were lymphadenopathy either generalized or localized in all cases, a mediastinal mass in 4 cases, bone marrow involvement in 7 cases, hypercalcemia in 4 cases, along with an aggressive clinical course and a poor response to chemotherapy in all cases, with a median disease-free survival time of 3 months.     

B‐cell lymphoma with Mott cell differentiation in two young adult dogs

Abstract: Two young adult dogs with gastrointestinal signs were each found to have an intra‐abdominal mass based on physical examination and diagnostic imaging. On exploratory laparotomy, small intestinal masses and mesenteric lymphadenopathy were found in both dogs; a liver mass was also found in dog 1. Cytologic and histologic examination of intestinal and liver masses and mesenteric lymph nodes revealed 2 distinct lymphoid cell populations: lymphoblasts and atypical Mott cells. With Romanowsky stains, the atypical Mott cells contained many discrete, clear to pale blue cytoplasmic inclusions consistent with Russell bodies that were positive by immunohistochemistry for IgM and CD79a in both dogs and for IgG in dog 2. The Mott cells and occasional lymphoblasts stained strongly positive with periodic acid‐Schiff. Using flow cytometric immunophenotyping in dog 1, 60% of peripheral blood mononuclear cells and 85% of cells in an affected lymph node were positive for CD21, CD79a, IgM, and MCH II, indicative of B‐cells. With electron microscopy, disorganized and dilated endoplasmic reticulum was seen in Mott cells in tumors from both dogs. Antigen receptor gene rearrangement analysis of lymph node and intestinal masses indicated a clonal B‐cell population. Based on cell morphology, tissue involvement, and evidence for clonal B‐cell proliferation, we diagnosed neoplasms involving Mott cells. To the authors' knowledge, this is the second report of Mott cell tumors or, more appropriately, B‐cell lymphoma with Mott cell differentiation, in dogs. More complete characterization of this neoplasm requires further investigation of additional cases. This lymphoproliferative disease should be considered as a differential diagnosis for canine gastrointestinal tumors.     

Clinicopathological study of canine transmissible venereal tumour in leishmaniotic dogs

Introduction : Canine transmissible venereal tumour is occasionally observed in leishmaniotic dogs, and Leishmania amastigotes can be harboured in canine transmissible venereal tumour cells. Objectives : The aim of this paper was to investigate the clinicopathological significance of the association of both diseases. Methods : Nineteen dogs affected by canine transmissible venereal tumour and canine leishmaniasis were studied retrospectively. Results : In these dogs, the tumour manifested a large size and often aggressive behaviour (42%) and no predictive sign of spontaneous regression was observed. Sporadic Leishmania amastigotes were found within the canine transmissible venereal tumour in three cases, probably transported by infected macrophages often infiltrating the tumour. A high Leishmania parasitisation of canine transmissible venereal tumour was observed in two other cases and verified by immunohistochemistry. Clinical Significance : Canine transmissible venereal tumour is a tumour of the dog able to harbour a large number of Leishmania parasites. Alternatively, the systemic disease (canine leishmaniasis) may lower the immune defence against malignancy (canine transmissible venereal tumour).     

Identification of Leishmania donovani amastigotes in canine tissues by immunoperoxidase staining

This paper describes the demonstration of Leishmania donovani amastigotes in canine tissues by immunoperoxidase staining. An indirect immunoperoxidase method was applied to the organs of 20 dogs in which leishmaniasis was previously diagnosed. Haemosiderin pigment was eliminated with 5 per cent oxalic acid. Amastigotes of L donovani appeared as dark brown stained bodies which contrasted with haematoxylin stained host cells. No positively stained amastigotes could be seen in any of the sections incubated with control serum. The organs which more frequently showed leishmanids were: skin (macrophages and fibroblasts), liver, spleen, lymph nodes and bone marrow. In a few cases amastigotes were seen in kidneys, gut, adrenal glands, eyes and testicles. This technique is simple to perform, gives consistent results and allows unequivocal histopathological diagnosis of canine leishmaniasis.