Comprehensive genomic characterization of five canine lymphoid tumor cell lines.

BACKGROUND: Leukemia/lymphoma cell lines have been critical in the investigation of the pathogenesis and therapy of hematological malignancies. While human LL cell lines have generally been found to recapitulate the primary tumors from which they were derived, appropriate characterization including cytogenetic and transcriptional assessment is crucial for assessing their clinical predictive value. RESULTS: In the following study, five canine LL cell lines, CLBL-1, Ema, TL-1 (Nody-1), UL-1, and 3132, were characterized using extensive immunophenotyping, karyotypic analysis, oligonucleotide array comparative genomic hybridization (oaCGH), and gene expression profiling. Genome-wide DNA copy number data from the cell lines were also directly compared with 299 primary canine round cell tumors to determine whether the cell lines represent primary tumors, and, if so, what subtype each most closely resembled. CONCLUSIONS: Based on integrated analyses, CLBL-1 was classified as B-cell lymphoma, Ema and TL-1 as T-cell lymphoma, and UL-1 as T-cell acute lymphoblastic leukemia. 3132, originally classified as a B-cell lymphoma, was reclassified as a histiocytic sarcoma based on characteristic cytogenomic properties. In combination, these data begin to elucidate the clinical predictive value of these cell lines which will enhance the appropriate selection of in vitro models for future studies of canine hematological malignancies.

Omental leiomyosarcoma with unusual giant cells in a Beagle dog - Short communication.

A 10-year-old castrated male Beagle dog was presented with a 2-month history of intermittent vomiting and abdominal pain. The dog was referred to the Veterinary Teaching Hospital at Iwate University for further evaluation, and a splenic tumour was suspected on the basis of ultrasonography and computed tomography. Surgery identified a large, solid, light-pink mass on the greater omentum with blood-coloured ascites in the abdominal cavity, and resection was performed. Microscopically, the mass comprised spindle-shaped tumour cells and scattered osteoclast-like giant cells. Most spindle-shaped cells were positive for vimentin, desmin, and smooth muscle actin (α-SMA), whereas osteoclast-like giant cells were positive only for vimentin. On the basis of histopathological and immunohistochemical findings, a diagnosis of leiomyosarcoma was made. To the best of our knowledge, this represents the first report of leiomyosarcoma associated with osteoclast-like giant cells developing from the greater omentum in a dog.

MicroRNA-214 and MicroRNA-126 Are Potential Biomarkers for Malignant Endothelial Proliferative Diseases.

Malignant endothelial proliferative diseases including human angiosarcoma (AS) and canine hemangiosarcoma (HSA) are serious diseases with a grave prognosis. Establishing liquid biopsy-based biomarkers for screening has definite clinical utility; however, plasma miRNAs up- or down-regulated in these sarcomas have been unclear. For identifying possible diagnostic plasma miRNAs for these sarcomas, we investigated whether plasma miR-214 and miR-126, which miRNAs play important roles in angiogenesis and tumorigenesis, were elevated in malignant endothelial proliferative diseases. For this investigation, human angiosarcoma and canine hemangiosarcoma cell lines and clinical plasma samples of canine hemangiosarcoma were examined by performing miRNA qRT-PCR. We report here that human angiosarcoma and canine hemangiosarcoma cell lines over-secreted miR-214 and miR-126 via microvesicles; in addition, their levels in the plasma samples from canines with hemangiosarcoma were increased. Moreover, the surgical resection of primary tumors decreased the levels of plasma miR-214 and miR-126. Our findings suggest that these malignant endothelial proliferative diseases over-secreted miR-214 and miR-126, thus suggesting that these miRNAs have potential as diagnostic biomarkers for malignant endothelial proliferative diseases in canine and possible in human angiosarcoma.

Epidermoid cyst of the ileum in a miniature dachshund dog.

A 13-year-old castrated male miniature dachshund dog was presented with chronic vomiting and diarrhea. Contrast radiography and surgical exploration revealed an ileal cyst, which was excised. A diagnosis of epidermoid cyst was made from a combination of the clinical findings and histological examination of the surgical specimen.

Kyste épidermoïde de l'iléon chez un chien Dachsund miniature. Un chien Dachsund miniature mâle castré âgé de 13 ans a été présenté avec des vomissements et de la diarrhée chroniques. Une radiographie à contraste et une exploration chirurgicale ont révélé un kyste iléal, qui a été excisé. Un diagnostic de kyste épidermoïde a été posé à partir d'une combinaison de constatations cliniques et de l'examen histologique du spécimen chirurgical.(Traduit par Isabelle Vallières).

Presumptive acute lung injury following multiple surgeries in a cat.

A 12-year-old, 3.5-kg spayed female domestic shorthair cat had a tracheal mass identified as malignant B-cell lymphoma. The cat had tracheal resection and subsequently developed laryngeal paralysis. Due to multiple episodes of respiratory distress the cat subsequently had tracheal surgeries. Finally, the cat had a sudden onset of severe respiratory distress and collapsed. Computed tomography imaging and arterial blood gas analysis supported a diagnosis of acute lung injury.

Blessure pulmonaire aiguë présumée après plusieurs chirurgies chez un chat. Une chatte commune domestique stérilisée âgée de 12 ans qui pesait 3,5 kg avait une masse trachéale identifiée comme un lymphome B malin. La chatte a eu une résection trachéale et a ensuite développé une paralysie laryngienne. En raison de plusieurs épisodes de détresse respiratoire, la chatte a ensuite subi des chirurgies trachéales. Enfin, la chatte a souffert soudainement d'une détresse respiratoire et s'est effondrée. Une tomographie par ordinateur et une analyse des gaz du sang artériel a appuyé un diagnostic de blessure pulmonaire aiguë.(Traduit par Isabelle Vallières).

Phenotypic and molecular characterization of novel pulmonary adenocarcinoma cell lines established from a dog.

Canine pulmonary adenocarcinoma (PAC) resembles human lung tumors in never-smokers, but it is rarer than human pulmonary adenocarcinoma. Therefore, research on canine PAC is challenging. In the present study, we successfully established various novel canine PAC cell lines from a single lesion in a dog, including two parent cell lines and fourteen cloned cell lines, and characterized their cellular properties in vitro. Several of these cell lines showed epithelial-mesenchymal transition (EMT)-like and/or cancer stem cell (CSCs)-like phenotypes. We additionally assessed the sensitivity of the cells to vinorelbine in vitro. Three clonal lines, two of which showed EMT- and CSC-like phenotypes, were resistant to vinorelbine. Furthermore, we evaluated the expression and activation status of EGFR, HER2, and Ras signaling factors. The findings indicated that the cell lines we established preserved the expression and activation of these factors to varying extents. These novel canine PAC cell lines can be utilized in future research for understanding the pathogenesis and development of treatments for canine PAC.

Expression profile of immunoregulatory factors in canine tumors.

Cancers utilize a variety of molecules to escape host immune responses. Better understanding the immune environment surrounding cancer may facilitate application of innovative cancer immunotherapies, such as immune checkpoint inhibitors, to dogs as well as humans. In this study, we screened the expression of 20 immune regulatory molecules in diverse canine tumors (n = 59). Quantitative RT-PCR (qPCR) analysis revealed that some immune regulatory molecules, such as LGALS9 (coding Galectin-9) and CD48, were expressed in most canine tumors, but other molecules, such as CD274 (coding PD-L1), IL4I1, PVR, TNFSF18, ICOSLG, and TNFSF4, were rarely expressed. NECTIN2 was highly expressed in epithelial tumors but was low in non-epithelial tumors. In contrast, VSIR and CD200 expressions were low in epithelial tumors but high in non-epithelial tumors. Interestingly, several tumors expressed distinctive immunoregulatory factors. Hepatocellular carcinomas expressed FGL1, mast cell tumors expressed PDCD1LG2 (coding PD-L2), transitional cell carcinomas expressed VTCN1 (coding B7x), and lymphomas and squamous cell carcinomas expressed CD70. Consistent with qPCR results, immunofluorescence staining confirmed that hepatocellular carcinomas expressed FGL-1 protein. Thus, this study reveals the expression profile of immunoregulatory molecules in canine tumors and opens the door to better understanding the relationship between canine tumors and host immunity.

Dysuria Associated with Non-Neoplastic Bone Hyperplasia of the Os Penis in a Pug Dog.

A three-year-old male Pug presented with a three-year history of urolithiasis and repeated urethral obstruction. Biochemical analysis, ultrasonography, and retrograde urethrocystography revealed probable portosystemic shunt and incomplete urethral obstruction due to uric acid ammonium calculi. Enhanced computed tomography (CT) revealed portosystemic shunt and proliferation of the osseous tissue of the os penis, which was surgically removed. Histopathologically, the excised osseous tissue comprised bland lamellar bone without atypia or inflammation. Hyperplasia of the os penis was diagnosed based on the image findings and histopathology. The dysuria improved postoperatively. This is the first report of dysuria associated with non-neoplastic bone hyperplasia of the os penis in a dog. Careful evaluation of the os penis by CT is needed for accurate diagnosis in case of repeated penile urethral obstruction.

Gallbladder agenesis in a Chihuahua.

A 4-year-old neutered male Chihuahua was presented with a history of anorexia and vomiting. Continuous elevation of liver enzymes was found on repeated blood examinations and the dog was referred to us for further evaluation. The absence of gallbladder was suspected on ultrasonography. Exploratory laparotomy and retrograde cholangiography confirmed gallbladder agenesis and a possible hypoplasia of the right medial and lateral liver lobes. Histologically, proliferation of bile ductules associated with portal fibrosis and pseudolobular formation were apparent in the liver lobes.

Antiemetic effect of oral maropitant treatment before the administration of brimonidine ophthalmic solution in healthy cats.

OBJECTIVES: The aim of this study was to investigate the antiemetic, behavioural and physiological effects of oral maropitant treatment before the administration of brimonidine ophthalmic solution in healthy cats. METHODS: Five cats received oral maropitant 8 mg or no treatment (control) 18 h before the administration of one drop of brimonidine solution in both eyes. Each cat was administered each of the two treatments, with a washout period of 1 week. The incidence of emesis, retching, sialorrhoea and lip-licking after brimonidine administration was recorded, while behavioural and physiological parameters, including heart rate, mean blood pressure, respiratory frequency and rectal temperature, were recorded before and 0, 30, 60, 90, 120, 180 and 240 mins after brimonidine administration. RESULTS: Emesis and retching were not observed when maropitant was administered. However, 4/5 cats exhibited vomiting and retching in the absence of maropitant pretreatment. The incidence of emesis and retching after brimonidine administration was significantly lower in the treatment group than in the control group. Sialorrhoea occurred in one cat in the control group, while all cats showed lip-licking after brimonidine administration. There were no significant differences in the incidence of sialorrhoea and lip-licking between the two groups. Although behaviour scores were comparable between the two groups, those obtained during heart rate, mean blood pressure and respiratory frequency measurements were significantly lower than the baseline scores; this indicated a sedative effect after brimonidine administration. The heart rate and mean blood pressure significantly decreased after brimonidine administration in both groups, while there were no intergroup differences in the heart rate, mean blood pressure, respiratory frequency and rectal temperature. CONCLUSIONS AND RELEVANCE: Oral maropitant treatment before the administration of brimonidine ophthalmic solution in cats can alleviate emesis and retching without affecting the sedative effects of brimonidine and important physiological parameters.

The Outcome and CT Findings of Low-Dose Intensity Modulated Radiation Therapy with SQAP in a Cat with Thymoma.

A 9-year-old male intact domestic cat weighing 4.6 kg was referred for tachypnea. A large mass was visible in computed tomography (CT) scans of the thoracic cavity. A histopathological evaluation of the mass was consistent with thymoma. The cat was treated with 2 × 8 Gy intensity modulated radiation therapy and sulfoquinovosyl acyl propanediol (SQAP). Post radiation therapy (RT), the tumor structure appeared cystic in the CT, and the tumor volume decreased by approximately 80% after aspiration than that before aspiration. The tumor was removed surgically. RT treatment with SQAP made it possible to treat the thymoma with a low total radiation dose.

Preparation of stable chitosan-carboxymethyl dextran nanoparticles.

Chitosan-carboxymethyl dextran nanoparticles (CHI-CMD NPs) were prepared by the formation of polyelectrolyte complex via the interaction of the positive charged amino group of chitosan and the negative charged carboxyl group of carboxymethyl dextran. Biocompatible organic reagents such as lactic acid, phosphate-buffered saline, and carbonate-bicarbonate buffer were used to make CHI-CMD NPs. The pH value of narrowly distributive CHI-CMD NP suspension was close to the physiological value of 6.8, which made it possible for it to be directly applied in an in vitro test without further additional operations. In the presence of sugars (30 mM), the colloidal stability of CHI-CMD NPs could be extended to 5 to 7 days when the particle size could be maintained within a nanometer scale, while the addition of xylose or lactose was found to most effectively extend the colloidal stability of CHI-CMD NPs to 10 days. Scanning electron microscopy, transmission electron microscopy, Fourier transform infrared spectroscopy, and X-ray diffraction analysis revealed that the structural integrity of CHI-CMD NPs was not destroyed after high steam pressure sterilization (121 degrees C, 30 min), which confirmed the autoclavable property of CHI-CMD NPs.

Aberrations of the FHIT gene and Fhit protein in canine lymphoma cell lines.

The fragile histidine triad (FHIT) gene is a tumor-associated gene, and aberrant FHIT gene and protein expression have been described in many types of human tumors. Furthermore, it has been reported that FHIT gene inactivation is induced by hypermethylation of 5' CpG islands in the gene or by genomic deletion around the open reading frame (ORF). In this study, we explored the aberrations in the canine FHIT gene and Fhit protein expression and assessed the methylation status and genomic deletions by using 5 canine lymphoma cell lines. We found that the decrease in the expression of the Fhit protein in canine lymphoma cell lines was similar to that in human tumors. The expression of the wild-type FHIT transcript was reduced in all 5 cell lines. However, we could not confirm the involvement of aberrant methylation events in the 5' CpG islands of the canine FHIT gene. We were able to identify homozygous or heterozygous deletions in the canine FHIT genes in all 5 cell lines. Moreover, a widespread genomic deletion of the FHIT gene, which included the ORF region, was detected in 1 cell line. In the present study, we detected aberrations in the FHIT gene and Fhit protein expression in all 5 canine lymphoma cell lines, and this phenomenon might be an important factor in promoting canine lymphoma.

Magnetic resonance imaging findings of an intradural extramedullary hemangiosarcoma in a dog.

An 11-year-old male Miniature Dachshund was referred for acute neurological deficits in the pelvic limbs. T2-weighted magnetic resonance imaging revealed that the spinal cord at the L1-2 intervertebral disc space was heterogeneously hyperintense in the sagittal plane and was mildly compressed from the ventral side by a small hypointense mass in the transverse plane. However, the lesion showed mass enhancement and severe spinal cord compression on post-contrast T1-weighted imaging. On three-dimensional myelography, a "golf tee sign" was observed around the mass. Therefore, we diagnosed an intradural extramedullary lesion. The mass was surgically removed and histologically diagnosed as a hemangiosarcoma. The "golf tee sign" observed on magnetic resonance myelography may be useful for distinguishing intradural extramedullary masses from intramedullary masses.

Effect of intramuscular administration of medetomidine and xylazine on tear flow measured by the Schirmer tear test I in healthy cats.

OBJECTIVES: This study aimed to investigate the effects of intramuscular medetomidine and xylazine on tear flow in healthy cats. METHODS: Five cats each received medetomidine 10, 20, 40 and 80 µg/kg IM; xylazine 1.0, 2.0, 4.0 and 8.0 mg/kg IM; and physiological saline (2.0 ml IM) in a randomised order separated by intervals of at least 1 week. The Schirmer tear test (STT) I was performed in both eyes before and 0.25, 0.5, 0.75, 1, 2, 3, 4, 5, 6, 7, 8 and 24 h after each dose. RESULTS: The STT I value decreased significantly at 0.5 and 1.0 h and at 0.75 and 1.0 h in both eyes after administration of medetomidine at 10 or 40 µg/kg. After administration of medetomidine 80 µg/kg, there was a significant decrease in the STT I reading at 0.75, 2 and 3 h in the left eye and 0.75, 1, 2 and 3 h in the right eye. The STT I value decreased significantly at: 0.5, 0.75, 1 and 2 h in the left eye and 0.75 h in the right eye after administration of xylazine 1.0 mg/kg; 0.5, 0.75, 1 and 2 h in the left eye and 0.5, 0.75, 1 and 3 h in the right eye after administration of xylazine 2.0 mg/kg; 0.5, 0.75, 1 and 2 h in both eyes after administration of xylazine 4.0 mg/kg; and 0.5, 0.75, 1, 2 and 3 h in the left eye and 0.75, 1, 2, 3 and 4 h in the right eye after administration of xylazine 8.0 mg/kg. CONCLUSIONS AND RELEVANCE: Both medetomidine and xylazine significantly decreased feline tear flow measured by STT I. Therefore, the ocular surface should be monitored carefully and protected appropriately in cats treated with these sedatives.

Mandibular Reconstruction by Using a Liquid Nitrogen-Treated Autograft in a Dog with an Oral Tumor.

A 10 yr old intact female German shepherd dog presented with a large peripheral odontogenic fibroma and malignant melanoma on her lower jaw. The tumor was resected with a unilateral subtotal rostral hemimandibulectomy. After the mandible was removed, it was devitalized intraoperatively by freezing it in liquid nitrogen. It was subsequently reimplanted. New bone tissue formed in the gap between the frozen bone and the host bone. The regenerated bone contained osteocytes, osteoblasts, and blood vessels. The cosmetic appearance of the dog was preserved. The dog had normal mastication. The malignant melanoma recurred rostral of the left canine tooth at 159 days after the reconstruction surgery. A subtotal hemimandibulectomy was consequently performed. This is the first reported case of mandibular reconstruction using a liquid nitrogen-treated autograft in a dog with oral tumors.

Circulating microRNA-214 and -126 as potential biomarkers for canine neoplastic disease.

Circulating microRNAs in the blood may provide diagnostic and prognostic information about canine neoplastic diseases, and their profiles may be conserved between human and canine species. We performed RT-qPCR to obtain the profiles of circulating plasma microRNA-214 and -126 in total 181 cases of canine neoplastic diseases and healthy controls. MicroRNA-214 levels were high in 2 epithelial tumours (thyroid and mammary carcinomas) and 4 non-epithelial tumours (osteosarcoma, histiocytic sarcoma, chondrosarcoma, and hemangiosarcoma). In contrast, microRNA-126 levels were high in 6 epithelial tumours (mammary, hepatocellular, squamous cell, thyroid, transitional cell carcinomas, and adenocarcinoma) and 4 non-epithelial tumours (osteosarcoma, mast cell tumour, melanoma, and hemangiosarcoma). The diagnostic potential of microRNA-214 was relatively high in sarcomas, whereas that of microR-126 was high in most types of the tumours. MicroRNA-214 and -126 were prognostic predictors in 2 groups (adenocarcinoma and non-epithelial tumours except for osteosarcoma) and 3 groups (epithelial tumours, adenocarcinoma, and melanoma), respectively. Additionally, the microRNA levels did not show a strong correlation with the other clinical parameters. In conclusion, circulating microRNA-214 and -126 have the potential to be diagnostic and prognostic biomarkers for canine neoplastic diseases. Furthermore, their profiles may be key references as well for exploring novel biomarkers for human cancers.

[Epidemiological survey of sexually transmitted male urethritis in Kyoto Prefecture].

The data of sexually transmitted urethritis in males have been collected at 24 institutes in Kyoto Prefecture since October, 2002. The data collected from January to December in 2004 are summarized herein. A total of 1,275 patients were diagnosed with urethritis during this period. Microbiological examinations isolated Neisseria gonorrhoeae alone in 368 (29%), Chlamydia tracomatis alone in 336 (26%), both in 85 (7%), and others in 453 (36%). Male patients under 20 years old tended to have Chlamydial urethritis, alone or combined with gonococcal infection, and had a predominant infectious source, a non-commercial-sexual-worker female partner, suggesting a profound problem in sexual life of adolescents. The urologist preferred to use quinolones as the first therapeutic modality against male urethritis. However, drug resistance of N. gonorrhoeae, especially against quinolones, has rapidly progressed, which was also observed by a sensitivity examination test. Antibiotics should be used adequately against male urethrits according to the recent guidelines.

Expression of NG2 proteoglycan in the degenerated intervertebral disc in dachshunds.

The pathogenesis of intervertebral disc (IVD) degeneration is not fully understood. The biomolecular signaling pathways involved in the IVD degeneration require further investigation. The aim of this study was to investigate the expression of NG2 proteoglycan in the degenerated IVD. IVD samples were obtained from 16 Dachshunds that were confirmed to have IVD herniation and subsequently underwent hemilaminectomy. The samples were subjected to histological and immunohistochemical (IHC) examinations. IHC revealed positive results for the expression of NG2 proteoglycan in all examined samples. The results showed the expression of NG2 proteoglycan by the degenerated IVDs.

Expression of a tumor-associated antigen, RCAS1, in canine mammary tumors.

Receptor-binding cancer antigen expressed on SiSo cells (RCAS1), one of novel cancer cell-surface antigens, is strongly expressed in invasive cancers. RCAS1 inhibits the in vitro growth of lymphocytes such as T cells and natural killer (NK) cells, and induces apoptotic cell death. We investigated the expression of RCAS1 in canine mammary tumor cell lines and tumor cells by immunohistochemistry, and also in situ deoxyribonucleic acid (DNA) fragmentation in tumor-infiltrating lymphocytes (TILs) by the terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate nick end labeling (TUNEL) method. All canine mammary tumor cell lines expressed RCAS1 at both the messenger ribonucleic acid (mRNA) and protein level. Immunohistochemically, RCAS1 was negative in 100% of normal mammary glands, but was expressed in 100% of malignant tumors examined. In most malignant mammary tumors, RCAS1 was localized in the cytoplasm with no polarity of expression. In benign mammary tumors, it was detected on the luminal surface of the tumor cell. RCAS1 expression or localization was significantly correlated with malignancy. In situ DNA fragmentation of CD3-positive TILs was observed in RCAS1-expressing tumors. RCAS1-expressing tumors, indicating a possible induction of apoptotic cell death in TILs through RCAS1 expression. These observations suggest that RCAS1 probably plays an important role in tumor progression and escape from immune surveillance in canine mammary tumors.