Clinical Utility of Circulating Cell-Free DNA as a Liquid Biopsy in Cats With Various Tumours.

Only a limited number of tumour biomarkers are currently available in veterinary medicine, particularly in cats. Cell-free DNA (cfDNA) is an extracellular DNA fragment released upon cell death and is considered a minimally invasive biomarker for the diagnosis and monitoring of various human malignancies. This study aimed to clarify the utility of circulating cfDNA as a liquid biopsy for various feline tumours. Plasma samples were collected from 44 cats with various tumours, 24 cats with other diseases and 10 healthy controls. A follow-up study was conducted in three tumour-bearing patients. All cfDNA concentrations were quantified via real-time polymerase chain reaction (PCR), which provided short and long fragments of a newly identified feline LINE-1 gene. We found that cfDNA levels were significantly higher in cats with various tumours than in those with other diseases or healthy controls. The cfDNA concentration was not correlated with serum amyloid A (SAA) levels. Cats with tumours exhibited elevated cfDNA levels that predicted tumour-bearing with a sensitivity and specificity of 50.5% and 91.2%, respectively (AUC 0.736; p < 0.001). In lymphoma cases, cats with high cfDNA levels had significantly shorter survival times than those with low cfDNA levels (median: 33 days vs. 178 days; p = 0.003). In addition, the cfDNA levels of the three patients correlated with clinical status during follow-up. Collectively, these findings indicate the potential of cfDNA as a useful biomarker for the diagnosis, therapeutic monitoring and prognostic assessment of tumours in cats.

Histologic, immunohistochemical, and in situ hybridization study of myxoid stroma in feline oral squamous cell carcinoma.

Oral squamous cell carcinoma (oSCC) is a highly invasive malignant neoplasm in cats. Recently, tumor stroma, known as tumor microenvironments, have been considered to play an essential role in tumor progression. However, their role in feline squamous cell carcinoma (SCC) remains unclear. This study aimed to reveal the cancer microenvironment of feline oSCC and evaluate the pathological mechanisms of progression. We used 19 samples from 17 cats with oSCC, which were examined using light microscopy, immunohistochemistry, and in situ hybridization (RNAscope®). Feline oSCCs had two types of stroma, namely fibrotic and myxoid stromal reaction patterns, which were easily distinguished using hematoxylin-eosin staining. The myxoid stroma was rich in hyaluronic acid, which seems to be produced by neoplastic cells. Furthermore, the presence of myxoid stroma was correlated with histological parameters, including the appearance of cancer-associated fibroblasts and tumor budding. Periostin protein expression was also frequently observed in the stroma of feline oSCC and was significantly more common in the myxoid stromal reaction pattern group than in the fibrotic group. Positive signals for periostin mRNA were detected in stromal cancer-associated fibroblasts. This study indicates that the interaction between neoplastic cells and stromal reaction pattern components, such as hyaluronic acid and periostin, may be involved in tumor malignancy. Therefore, we propose that focus be placed not only on the tumor tissue but also on the characterization of the stroma for analyzing feline oSCC.

LINE-1 Methylation Status in Canine Splenic Hemangiosarcoma Tissue and Cell-Free DNA.

Splenic hemangiosarcoma is one of the most common malignant tumors in dogs, and early diagnosis is of great importance for achieving a good prognosis. DNA methylation plays an important role in cancer development. Long interspersed nuclear element 1 (LINE-1) is the most abundant repetitive element in the genome. LINE-1 hypomethylation has been shown to be related to carcinogenesis in humans, and it has been used as a novel cancer biomarker. This study aimed to evaluate the methylation status of LINE-1 in tumor tissue and circulating cell-free DNA and assess its clinical significance in canine splenic hemangiosarcoma. Genomic DNA was isolated from splenic masses of 13 dogs with hemangiosarcoma, 11 with other malignant tumors, and 15 with benign lesions. LINE-1 methylation was quantified using methylation-sensitive and -insensitive restriction enzyme digestion followed by real-time polymerase chain reaction. Additionally, blood samples were collected from eight patients to isolate cell-free DNA to determine LINE-1 methylation status changes during the treatment course. LINE-1 methylation in tumor samples was significantly lower in patients with hemangiosarcoma than in those with other malignant tumors and benign lesions. Non-significant but similar results were observed for the cell-free DNA samples. Our results demonstrate that LINE-1 methylation status is a potential biomarker for splenic hemangiosarcoma.

Clinical utility of liquid biopsy in canine oral malignant melanoma using cell-free DNA.

Introduction: Cell-free DNA (cfDNA), an extracellular free DNA released into the bloodstream by cells, is a potentially useful noninvasive marker to detect human malignancies and monitor response to treatment. In the present study, we evaluated the utility of circulating cfDNA in canine patients with oral malignant melanoma (OMM) in assessing therapeutic response and clinical outcomes. Methods: Plasma samples were collected from 12 dogs with OMM and 9 healthy controls. cfDNA concentration was quantified by real-time PCR resulting in short (99bp) and long (218bp) fragments of long interspersed nuclear element-1 (LINE-1), and the DNA integrity index (DII) was then calculated (218/99). A follow-up study was conducted on 6 dogs with OMM, and the plasma cfDNA and DII were quantified throughout disease progression. Results: Although cfDNA levels obtained from dogs with OMM were not significantly different compared to those obtained from healthy controls, the DII was significantly lower in dogs with OMM than in healthy controls. The DII tended to decrease as the disease stage progressed. Moreover, changes in cfDNA concentration and DII along the clinical course were observed when major changes, such as metastasis or apparent tumor progression, were observed. Discussion: The results of our study suggest that measurements of serum cfDNA and DII using LINE-1 might be valuable new biomarkers for monitoring OMM progression in dogs. This preliminary study demonstrated the potential clinical utility of monitoring plasma cfDNA in canine patients with OMM.

Hemophagocytic syndrome in a cat with immune-mediated hemolytic anemia.

A 10-year-old spayed female domestic short-haired cat presented with depression, anorexia, and tachypnea. A complete blood count revealed moderate regenerative anemia, severe leukopenia, and mild thrombocytopenia. Antibodies against feline immunodeficiency virus (FIV) were also detected. Abdominal radiography and ultrasonography revealed severe splenomegaly. Cytologic evaluation of the spleen revealed macrophagic infiltration with hemophagocytosis. Bone marrow aspiration revealed erythroid hyperplasia with no other abnormalities. A presumptive diagnosis of hemophagocytic syndrome secondary to immune-mediated hemolytic anemia was made based on a positive direct Coombs test result. Blood transfusion, prednisolone, and immunosuppressive treatments were performed; however, the blood abnormalities did not improve. The cat was then administered prednisolone and chlorambucil, followed by splenectomy. Leukopenia immediately recovered, and packed cell volume increased slightly. However, the blood abnormalities recurred, and the cat died. To the best of our knowledge, this is the first report of hemophagocytic syndrome secondary to immune-mediated disease in an FIV-positive cat.

Prognostic role of lymphocyte to monocyte ratio in feline high-grade lymphomas.

The lymphocyte to monocyte ratio (LMR) is a useful prognostic marker of various cancers in human and canine patients. This study aimed to determine whether this ratio could predict disease outcomes in cats with high-grade lymphoma. Medical records of 33 cats diagnosed with high-grade lymphoma were retrospectively analyzed. The prognostic influence of LMR and other clinicopathological data on the time to progression (TTP) and overall survival (OS) was studied using the Kaplan-Meier curves. The optimal cutoff value of this ratio was 3.4, which corresponded to the maximum sensitivity (1.000) and specificity (0.611) of the LMR for predicting median OS days, using receiver operating characteristic analysis. A univariate analysis demonstrated that cats with a low LMR had significant reductions in both TTP [hazard ration (HR) = 3.403, 95% confidence interval (CI): 1.502 to 8.720; P = 0.003] and OS (HR = 3.418, 95% CI: 1.433 to 9.449, P = 0.005). In multivariate analysis, independent predictors of OS included LMR (HR = 2.889, 95% CI: 1.048 to 8.843, P = 0.040), clinical stage (HR = 0.330, 95% CI: 0.118 to 0.960, P = 0.042), and age (HR = 4.151, 95% CI: 1.574 to 11.888, P = 0.004).

Rôle pronostique du rapport lymphocytes sur monocytes dans les lymphomes félins de grade élevé. Le rapport lymphocyte sur monocytes (LMR) est un marqueur pronostique utile de divers cancers chez les patients humains et canins. Cette étude visait à déterminer si ce rapport pouvait prédire l'issue de la maladie chez les chats atteints d'un lymphome de grade élevé. Les dossiers médicaux de 33 chats diagnostiqués avec un lymphome de grade élevé ont été analysés rétrospectivement. L'influence pronostique de la LMR et d'autres données clinicopathologiques sur le temps de progression (TTP) et la survie globale (OS) a été étudiée à l'aide des courbes de Kaplan-Meier. La valeur seuil optimale de ce rapport était de 3,4, ce qui correspondait à la sensibilité (1,000) et à la spécificité maximales (0,611) du LMR pour prédire les jours de OS médians, en utilisant l'analyse des caractéristiques de fonctionnement du récepteur. Une analyse univariée a démontré que les chats avec un faible LMR présentaient des réductions significatives du TTP [rapport de risque (HR) = 3,403, intervalle de confiance (IC) à 95 % : 1,502 à 8,720; P = 0,003] et de la OS (HR = 3,418, IC à 95 % : 1,433 à 9,449, P = 0,005). Dans l'analyse multivariée, les prédicteurs indépendants de la OS comprenaient le LMR (HR = 2,889, IC à 95 % : 1,048 à 8,843, P = 0,040), le stade clinique (HR = 0,330, IC à 95 % : 0,118 à 0,960, P = 0,042) et l'âge (HR = 4,151, IC à 95 % : 1,574 à 11,888, P = 0,004).(Traduit par Dr Serge Messier).

Acute myelomonocytic leukemia negative for alpha-naphthyl acetate esterase stain in a Holstein cow.

A 4-year, 7-month-old Holstein cow presented with anorexia. Physical examination revealed masses in the interscapular region and vagina. Blast cells were detected in the masses and peripheral blood by fine needle aspiration cytology and hematological examination. By bone marrow aspiration, blast cells constituted up to 24.2% of all nucleated cells, and 22% and 2% of non-erythroid cells stained positive for myeloperoxidase and alpha-naphthyl acetate esterase (ANAE), respectively. Pathological examination revealed the mass lesions consisted of a proliferation of tumor cells, which were positive for monocytic markers (HLA-DR and Iba-1). The cow was diagnosed with acute myelomonocytic leukemia (AMML). Even when tumor cells are ANAE-negative, AMML cannot be completely ruled out and should be considered when diagnosing cattle with leukemia/lymphoma.

Cardiomyopathy in a dog with multicentric lymphoma following treatment with several anthracyclines.

Background: Canine lymphoma is one of the most frequently occurring malignant neoplasms in dogs. Anthracycline-based chemotherapy for the treatment of canine lymphoma is very effective; however, there is not enough evidence for the development of cardiac toxicity using several anthracyclines as chemotherapeutic agents. Case Description: An 8-year-old, castrated, mixed-breed dog was diagnosed with multicentric lymphoma and received multi-agent chemotherapy. Complete remission was achieved, but the patient had a relapse of lymphoma. After third-line chemotherapy with epirubicin, the patient was diagnosed with dilated cardiomyopathy. The total cumulative doses of doxorubicin, mitoxantrone, and epirubicin were 125, 8, and 125 mg/m2, respectively. Although the patient was treated with cardiac drugs and clinically stabilized, the patient had a relapse of lymphoma and died shortly after the diagnosis of cardiomyopathy. Conclusion: The patient was suspected to have anthracycline-induced cardiomyopathy. Further studies are required to establish prevention and management strategies for dogs receiving potentially cardiotoxic therapies, such as anthracyclines.

A clinical case of acute myelomonocytic leukemia in a Holstein cow.

A 2-year, 3-month-old Holstein cow presented with anorexia and enlarged superficial lymph nodes. Fine needle aspiration cytology of the superficial lymph nodes revealed large blast cells. Hematological examination revealed anemia, neutropenia, and blast cells in peripheral blood. Blast cells were the predominant cell type in bone marrow aspirates. Of the non-erythroid cells, 26%, 58%, and 18% were positive for myeloperoxidase, α-naphthyl acetate esterase, and naphthol AS-D chloroacetate esterase, respectively. Pathological examination revealed the proliferation of neoplastic cells, which were positive for monocytic markers, in the affected lymph nodes. The cow was diagnosed with acute myelomonocytic leukemia based on these findings. This report highlights the importance of performing bone marrow aspiration cytology and cytochemical staining when diagnosing bovine myeloid leukemia.

Acute monoblastic leukemia in a feline leukemia virus-negative cat.

A 12-year-old female domestic short-haired cat was presented due to weight loss, anorexia, and tachypnea. Complete blood count revealed severe anemia, leukocytosis with massive undifferentiated blast cells, and thrombocytopenia. Bone marrow aspiration showed acute myeloid leukemia, subclassified as monoblastic leukemia (M5a) based on the outcomes of the cytochemistry examinations. The SNAP feline leukemia virus (FeLV) and feline immunodeficiency virus (FIV) test using whole blood was negative. In addition, FeLV/FIV proviral polymerase chain reaction test using bone marrow aspirate was also negative. Although the cat was treated with doxorubicin, cytosine arabinoside, and prednisolone, anemia did not improve without blood transfusion. The owner declined further treatment after 2 months, and the cat died a few days later.

Intramedullary spinal nephroblastoma in a mixed breed dog.

A 1-year-old male mixed breed dog presented for the evaluation of progressive hindlimb paresis. Neurological examination indicated a spinal cord lesion between the 3rd thoracic and 3rd lumbar vertebrae. Magnetic resonance imaging (MRI) revealed an intramedullary spinal cord lesion located at the level of the 1st and 2nd lumbar vertebrae. Following cytoreductive surgery of the mass, palliative radiation therapy was administered. A diagnosis of nephroblastoma was made based on histological examination. After radiation therapy, the disappearance of the spinal lesion was confirmed by MRI. The dog was improved from gait abnormality and alive at 16 months postoperatively, with slight signs of neurological dysfunction.

Quantitative analysis of the BRAF V595E mutation in plasma cell-free DNA from dogs with urothelial carcinoma.

Circulating tumor DNA (ctDNA), which carries tumor-specific mutations, is an emerging candidate biomarker for malignancies and for monitoring disease status in various human tumors. Recently, BRAF V595E mutation has been reported in 80% of dogs with urothelial carcinoma. This study investigates the BRAF V595E allele concentration in circulating cell-free DNA (cfDNA) and assesses the clinical significance of BRAF-mutated ctDNA levels in canines with urothelial carcinoma. A total of 15 dogs with urothelial carcinoma were included. cfDNA concentration was measured using a real-time polymerase chain reaction (PCR) of the LINE-1 gene. To measure the concentration of the mutated BRAF gene in cfDNA, allele-specific real-time PCR with a locked nucleic acid probe was performed. BRAF mutations were detected in 11 (73%) of the 15 tested tumor samples. BRAF-mutated ctDNA concentrations were significantly higher in dogs with the BRAF mutation (14.05 ± 13.51 ng/ml) than in wild-type dogs (0.21 ± 0.41 ng/ml) (p = 0.031). The amount of BRAF-mutated ctDNA in plasma increased with disease progression and responded to treatment. Our results show that BRAF-mutated ctDNA can be detected using allele-specific real-time PCR in plasma samples of canines with urothelial carcinoma with the BRAF V595E mutation. This ctDNA analysis may be a potentially useful tool for monitoring the progression of urothelial carcinoma and its response to treatment.

Quantification of plasma cell-free DNA levels in dogs with various tumors.

Circulating cell-free DNA (cfDNA) is extracellular DNA released into the bloodstream by apoptotic or necrotic tumor cells, with cfDNA determination proposed as a noninvasive, sensitive marker for the diagnosis of human cancer. We evaluated cfDNA quantification as a diagnostic and prognostic tool in dogs with various tumors. We quantified plasma cfDNA concentration by absolute real-time PCR of long interspersed nuclear elements in 50 dogs with malignant tumors, 13 dogs with benign tumors or nodules, and 11 healthy controls. Six patients with malignant tumors were followed-up, and plasma cfDNA was quantified throughout disease progression. We found that plasma cfDNA concentrations were significantly elevated in dogs with malignant tumors compared with dogs with benign nodules or healthy controls. The DNA integrity index (the ratio between long and short cfDNA fragments) was significantly lower in dogs with malignant tumors compared to healthy controls. Significantly higher cfDNA levels and a lower DNA integrity index were observed in dogs with lymphoma or leukemia, hemangiosarcoma, and distant metastasis; cfDNA levels correlated well with clinical stage and tended to increase during or before periods of disease progression, suggesting potential efficacy of cfDNA for the detection of distant metastasis and to monitor the clinical stage of neoplasia.

CD3 and CD20 co-expression in a case of canine peripheral T-cell lymphoma with prominent cardiac and peripheral nerve involvement.

An 8-y 9-mo-old male Pug dog was presented because of anorexia, hindlimb ataxia, vomiting, and progressive weight loss. Clinical examinations revealed atrophic hindlimb muscles with decreased postural reaction, enlargement of the cardiac silhouette and megaesophagus on radiograph, and reduced cardiac contractility on ultrasonography. The dog died 10 d after the initial examination, and an autopsy was performed. Grossly, the heart was enlarged, with multifocal-to-coalescing extensive plaque-like areas of discoloration on the epicardial surface. On cross-section, the ventricles were moderately dilated, and discoloration extended into the myocardium. Peripheral lymph nodes were of normal size. Histologically, atypical lymphoid cells replaced the myocardium and also extensively infiltrated peripheral nerve bundles in various organs. With immunohistochemistry, the neoplastic cells exhibited strong immunoreactivity for CD3 and CD20, and were negative for CD8, granzyme B, CD79α, and Pax5. Double-label immunofluorescence confirmed co-expression of CD3 and CD20 by the neoplastic cells. Molecular clonality analysis presented a clonal T-cell receptor gamma gene rearrangement. The case was diagnosed as a CD3+/CD20+ peripheral T-cell lymphoma with prominent cardiac and peripheral nerve involvement, indicating neurolymphomatosis.

Evaluation of costimulatory molecules in dogs with B cell high grade lymphoma.

B cell high grade lymphoma is the most common hematopoietic malignancy in dogs. Although the immune checkpoint molecules, programmed death-1 (PD-1) and cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4), and immune checkpoint inhibitors have been evaluated for the treatment of various human lymphoid malignancies, the expression of those molecules and their relationship with prognosis remain unknown in canine lymphoma. The objective of this study was to evaluate the expression of costimulatory molecules on peripheral blood lymphocytes and tumor infiltrating lymphocytes, in addition to associated ligand expression in the lymph nodes of patients with B cell multicentric high grade lymphoma. Eighteen patients diagnosed with B cell high grade lymphoma and nine healthy control dogs were enrolled. Flow cytometric analysis revealed that the expression of PD-1 on CD4+ peripheral and tumor infiltrating lymphocytes and CTLA-4 on CD4+ peripheral lymphocytes was significantly higher in the lymphoma group than in the control group. The expression level of CD80 mRNA was significantly lower in the lymphoma group than in the control group. In contrast, there were no significant differences in PD-L1, PD-L2, and CD86 expression between the groups. Dogs with CTLA-4 levels below the cutoff values, which were determined based on receiver operating characteristic curves, on peripheral CD4+, CD8+, and tumor infiltrating CD4+ lymphocytes had significantly longer survival than dogs with values above the cutoff. Although it is uncertain whether the expression of immune checkpoint molecules affect the biological behavior of canine lymphoma, one possible explanation is that PD-1 and CTLA-4 might be associated with the suppression of antitumor immunity in dogs with B cell high grade lymphoma, particularly through CD4+ T cells.

Lissencephaly in a Pekingese.

A 1-year-old neutered male Pekingese was presented for evaluation and further treatment of cluster seizures. The dog had behavioral abnormalities, and a prosencephalic lesion was suspected following neurological examination. The dog showed signs of learning difficulty. Magnetic resonance imaging of the brain revealed a remarkably smooth cerebral cortex with a reduced number of gyri, as well as a cystic lesion associated with the quadrigeminal cistern. A diagnosis of lissencephaly, concurrent with a quadrigeminal cisternal cyst, was made. High-dose and multiple anticonvulsants were necessary to control the seizures. This is the first report of lissencephaly in a Pekingese.

Enhancement of abdominal wall defect repair using allogenic platelet-rich plasma with commercial polyester/cotton fabric (Damour) in a canine model.

Platelet-rich plasma (PRP) has an important role in musculoskeletal surgery; however, it has been underutilized for accelerating the healing of abdominal wall defects in veterinary practice. Therefore, the aim of this study was to evaluate the use of commercial polyester/cotton fabric (Damour) as a new composite mesh for the repair of experimentally induced abdominal wall defects in canine models, and to investigate the possible role of PRP for improving such repair and reducing allied complications. For this purpose, abdominal wall defects were created in 24 healthy mongrel dogs and then repaired with mesh alone (control group) or mesh and allogenic PRP (PRP group). Dogs were euthanized after 2 or 4 months for gross examination of implantation site, detection of adhesion score and hernia recurrence. Moreover, tissue samples were collected for histological and gene expression analyses for neovascularization, collagen formation and tissue incorporation. Hernia recurrence was not recorded in PRP-treated dogs that also displayed significantly more neovascularization and less severe adhesion to the underlings (1.08 ± 0.51) in comparison to control group (2.08 ± 0.99). Histological and molecular evaluation confirmed the gross findings that collagen deposition, new vessel formation, and overexpression of angiogenic and myofibroplastic genes (COL1α1, COL3α1, VEGF and TGFß1) were observed more frequently in the PRP group, at both time points. In conclusion, we found that addition of allogenic PRP to Damour mesh enhanced neovessel formation, and increased tissue deposition and incorporation, with subsequent reduction of peritoneal adhesion and recurrence rate.

Evaluation of Costimulatory Molecules in Peripheral Blood Lymphocytes of Canine Patients with Histiocytic Sarcoma.

Histiocytic sarcoma is a rapidly progressive and fatal neoplastic disease in dogs. It is unclear whether costimulatory molecules, including CD28, cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4), and programmed death-1 (PD-1), are expressed on peripheral blood lymphocytes (PBLs) of canine patients with histiocytic sarcoma. The objective of this study was to evaluate the expression of CD28, CTLA-4, and PD-1 molecules on PBLs of patients with histiocytic sarcoma, patients with other tumors, and healthy controls. Twenty-six dogs were included in the study, with eight, ten, and eight dogs in the histiocytic sarcoma, other tumor, and healthy control groups, respectively. PBLs and serum were prospectively obtained from patients diagnosed histopathologically with histiocytic sarcoma, other tumors and healthy controls. The surface expression of CTLA-4, CD28, and PD-1 on T lymphocytes was examined using flow cytometric analysis. Serum samples were frozen at -30°C until serum interferon-γ (IFN-γ) was measured by enzyme-linked immunosorbent assay. The expression level of CTLA-4 on CD4+ lymphocytes was significantly higher in the histiocytic sarcoma group than in the control group. The expression of CTLA-4 on CD8+ lymphocytes was significantly higher in the histiocytic sarcoma group than in the other two groups. In addition, the expression of PD-1 on CD8+ lymphocytes was significantly higher in the histiocytic sarcoma group than in the control group. However, no significant differences in CD28 expressions and serum IFN-γ levels were observed. The present results provided evidence showing that the expression levels of CTLA-4 on both CD4+ and CD8+ lymphocytes and PD-1 on CD8+ lymphocytes in peripheral blood obtained from dogs with histiocytic sarcoma were upregulated. The overexpressions of CTLA 4 and PD-1 suggested that antitumor immunity may be suppressed in dogs with histiocytic sarcoma.

Ultrasonographic evaluation of depth-width ratio (D/W) of benign and malignant mammary tumors in dogs.

Depth-width ratio (D/W) is the only quantitative item in the criteria recommended by the Japanese Ultrasound Society for the evaluation of breast tumors in humans. However, the usefulness of the D/W has not been evaluated in dogs. Eighty-six mammary masses in 34 female dogs underwent ultrasonographic examination to determine the D/W and other characteristics. Results of ultrasonographic and histopathologic examinations were compared. The D/W of malignant tumors was significantly greater than that of benign tumors, and it had a sensitivity of 56.3% and a specificity of 92.9% for the diagnosis of malignancy when the threshold of D/W was 0.7. In addition, irregular margin, polymorphous shape and heterogeneous internal echographic characteristics were correlated with malignancy.

Overexpression of interleukin 2 receptor, thymidine kinase and immunoglobulin-associated alpha-1 messenger RNA in a clinical case of enzootic bovine leukosis.

A 49-month-old Holstein cow with anorexia, tachypnea, enlarged peripheral lymph nodes, and difficulty standing up was suspected of bovine leukosis. Hematological examination revealed lymphocytosis with the presence of neoplastic cells. Increased total lactate dehydrogenase (LDH) activity, isozymes of LDH-2 and LDH-3 activities and thymidine kinase activity were observed. Cytological findings of fine needle aspiration of subiliac lymph nodes indicated lymphosarcoma. Histopathology and antibody analysis confirmed the diagnosis of enzootic bovine leukosis, a B-cell bovine lymphoma caused by bovine leukemia virus. Gene expressions known as biomarkers of hematopoietic neoplasia in human were also examined in the present case. Increased messenger RNA expression of interleukin 2 receptor, thymidine kinase, and immunoglobulin-associated alpha-1 was observed in the case animal.