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BACKGROUND: Few epidemiological studies on respiratory medicine and the relationship between clinical signs and various respiratory diseases in cats have been reported. OBJECTIVES: This retrospective study aimed to investigate the prevalence and breed predisposition to feline respiratory diseases in Japan and determine the association between clinical signs, duration and type of respiratory diseases. METHODS: The medical records of cats with feline respiratory diseases were examined to obtain information on age, sex, breed, final diagnosis, clinical signs and duration. The odds ratios (ORs) and 95% confidence intervals were calculated to evaluate breed predispositions. Mann-Whitney U, Kruskal-Wallis and Dunn's tests were used to assess the duration of clinical signs. RESULTS: This study included 540 cats with 615 respiratory diagnoses. The American Shorthair breed was predisposed to bronchopneumonia (BP; OR: 5.0) and pulmonary tumour (PT; OR: 3.6), while the Russian Blue breed exhibited a predisposition to inflammatory lower airway diseases (OR: 3.4), BP (OR: 6.1) and interstitial lung diseases (OR: 11.1). Similarly, the Scottish Fold breed displayed predisposition to PTs (OR: 5.8). The duration of clinical signs among nasal diseases, nasopharyngeal diseases and lower tracheal/bronchial and pulmonary diseases differed significantly (p = 0.001, p = 0.012, p < 0.0001, respectively). CONCLUSIONS: The results suggest that some popular breeds in Japan are predisposed to feline respiratory diseases, especially the American Shorthair, Russian Blue and Scottish Fold breeds. The characteristics of occurrence, clinical signs and duration of each disease will aid in diagnosing, treating, preventing and elucidating the pathophysiology of feline respiratory disease.

This study aimed to investigate the association of respiratory rate (RR), oxygen saturation (SpO2), and blood findings with respiratory disease in dogs and to compare the examination findings in the chronic and acute phases. Dogs that visited a veterinary referral hospital with respiratory symptoms were classified into the chronic disease group (GC), and those that visited the emergency veterinary hospital were classified into the acute disease group (GA). In total, 704 and 682 dogs were included in GC and GA, respectively. The RR and SpO2 were significantly higher and lower, respectively, in patients with lung disease compared to other disease sites in both groups. White blood cell counts were significantly increased in patients with lung and pleural diseases in both groups. Respiratory alkalosis and respiratory acidosis were most common in GC and GA, respectively. The C-reactive protein levels were elevated in both groups, primarily in patients with lung disease. Associations between the results of several tests for understanding and diagnosing respiratory conditions and diseases were recognized, and differences in the trends of the chronic and acute phases were clarified. These tools may be used as adjuncts to other tests for the diagnosis and monitoring of treatment responses.

Advanced imaging techniques under general anesthesia are frequently employed to achieve a definitive diagnosis of canine nasal diseases. However, these examinations may not be performed immediately in all cases. This study aimed to construct prediction models for canine nasal diseases using less-invasive examinations such as clinical signs and radiography. Dogs diagnosed with nasal disease between 2010 and 2020 were retrospectively investigated to construct a prediction model (Group M; GM), and dogs diagnosed between 2020 and 2021 were prospectively investigated to validate the efficacy (Group V; GV). Prediction models were created using two methods: manual (Model 1) and LASSO logistic regression analysis (Model 2). In total, 103 and 86 dogs were included in GM and GV, respectively. In Model 1, the sensitivity and specificity of neoplasia (NP) and sino-nasal aspergillosis (SNA) were 0.88 and 0.81 in GM and 0.92 and 0.78 in GV, respectively. Those of non-infectious rhinitis (NIR) and rhinitis secondary to dental disease (DD) were 0.78 and 0.88 in GM and 0.64 and 0.80 in GV, respectively. In Model 2, the sensitivity and specificity of NP and SNA were 0.93 and 1 in GM and 0.93 and 0.75 in GV, respectively. Those of NIR and DD were 0.96 and 0.89 in GM and 0.80 and 0.79 in GV, respectively. This study suggest that it is possible to create a prediction model using less-invasive examinations. Utilizing these predictive models may lead to appropriate general anesthesia examinations and treatment referrals.

OBJECTIVES: This multicentre, retrospective observational study aimed to describe the clinical presentation, diagnostic methods, treatment and outcomes of cats with tracheal masses. METHODS: Eighteen cats from five academic or secondary/tertiary animal hospitals were included. RESULTS: The median age at diagnosis was 10.7 years (mean 9.5; range 1-17). There were nine castrated males, seven spayed females, one intact male and one intact female. Fourteen (78%) were domestic shorthairs, one (6%) was an Abyssinian, one (6%) was an American Shorthair, one (6%) was a Bengal and one (6%) was a Scottish Fold. The most common presenting complaints included chronic respiratory distress or dyspnoea (n = 14), followed by wheezing/gagging (n = 12), coughing (n = 5) and voice changes (n = 5). There was cervical tracheal involvement in 16/18, and two showed involvement of the intrathoracic trachea. The following methods were used for diagnosis: ultrasound-guided fine-needle biopsy (UG-FNB) and cytology (n = 8), bronchoscopic forceps biopsy and histopathology (n = 5), surgical resection and histopathology (n = 3), forceps biopsy via an endotracheal tube (n = 1) and histology of tissue sputtered from a cough (n = 1). Lymphoma was most often diagnosed (n = 15), followed by adenocarcinoma (n = 2) and squamous cell carcinoma (n = 1). Most lymphoma cases received chemotherapy with or without radiation according to various protocols, and partial (n = 5) or complete responses (n = 8) were noted. Kaplan-Meier survival data for cats with lymphoma revealed a median survival time of 214 days (95% confidence interval >149 days), which was significantly longer than that of other types of tumours (21 days). CONCLUSIONS AND RELEVANCE: Lymphoma was the most prevalent diagnosis, and showed a good response to chemotherapy with or without radiation therapy. Various diagnostic procedures were performed, and UG-FNB and cytology are good diagnostic procedures for cervical tracheal lesions. Owing to the variety of treatment protocols at different centres, it was impossible to compare outcomes.

BACKGROUND: Few studies have investigated the incidence of respiratory diseases based on anatomical sites or the relationship between breed and these diseases. OBJECTIVE: The objective of this study was to investigate the prevalence of canine respiratory diseases among dogs in Japan, with relationship to the breed. METHODS: We retrospectively reviewed the medical records of dogs with respiratory symptoms and calculated the odds ratio (OR) to evaluate the relationship between breed and disease. RESULTS: A total of 1050 dogs with respiratory symptoms were included in this study. Miniature dachshunds were the most common breed affected by respiratory diseases. Among tracheobronchial diseases, there was a significant association between some small breeds and tracheobronchial collapse, miniature dachshunds (OR: 4.44, 8.43, 95% confidence interval [CI]: 3.17-6.22, 4.33-16.0) and chronic bronchitis and bronchiectasis. Among nasal diseases, miniature dachshunds (OR: 27.2, 95% CI: 16.8-44.8) and golden retrievers (OR: 21.0, 95% CI: 6.43-69.3) were the most affected by non-infectious rhinitis and nasal aspergillosis, respectively. Brachycephalic obstructive airway syndrome was the most common disease among pharyngeal and laryngeal diseases, with a relationship with breed being found in some brachycephalic breeds, and Pomeranians (OR: 2.7, 95% CI: 1.42-5.17). CONCLUSIONS: Respiratory diseases in dogs are strongly correlated with popular breeds in Japan. Miniature dachshunds, in particular, are associated with many respiratory diseases, which may differ from international reports. Thus, this result may help in the early detection, prevention, treatment, and elucidation of the pathophysiology of canine respiratory diseases.

Canine lymphoma is the most common haematological malignancy in dogs and is typically treated with multidrug chemotherapy. Most cases are at risk of relapse after several courses of chemotherapy and the oncogenic mechanism remains unknown. This study was aimed at identifying genes expressed in canine lymphoma by cDNA microarray. We found elevated expression of Dishevelled, EGL-10 and pleckstrin (DEP) domain-containing 1B (DEPDC1B) in canine lymphoma cells compared with cells and tissues from healthy dogs. Canine DEPDC1B protein was detected in 13 of 41 lymphoma specimens by immunohistochemistry, but was not detected in lymph nodes from normal dogs. Immunoreactive DEPDC1B protein was also detected in several other types of canine tumour. This is the first report documenting the association of DEPDC1B with canine cancer and the results suggest that DEPDC1B might serve as a potential marker or therapeutic target for canine malignancies.

Nasal lymphoma (NL) is the most common nasal tumor in cats, and radiotherapy, chemotherapy, or a combination of these treatments have been described as the treatment for this disease. However, the previous studies included various machines and protocols of radiotherapy. Therefore, we aimed to retrospectively compare the prognosis among cases treated with palliative hypofractionated radiotherapy, chemotherapy, and a combination of them with united machine and protocol of radiotherapy. When compared overall survival and progression free survival, there was no significant difference among these three groups. The data of this study suggested that similar efficacy could be achieved by palliative hypofractionated radiotherapy, chemotherapy, or a combination of them.

Soluble interleukin-2 receptor (sIL-2r) is released directly from the surface of lymphocytes expressing interleukin-2 receptor alpha chain (CD25), and its serum concentration has been found to reflect the prognosis of human lymphoproliferative malignancies. In this study, we demonstrated the presence of sIL-2r in canine serum and developed a specific sandwich enzyme-linked immunosorbent assay (ELISA) to quantify the concentration of canine serum sIL-2r. In the immunoprecipitation (IP) assay, CD25 protein weighing approximately 45 kDa was detected in canine serum, smaller than the membrane-bound CD25 (approximately 55 kDa). To measure the concentration of serum sIL-2r in dogs, an ELISA system was developed. Serum sIL-2r levels were significantly higher in dogs with multicentric high-grade B-cell lymphoma before therapy than that in healthy dogs. Serum sIL-2r concentration was also found to be elevated in a proportion of dogs with other types of lymphoma. Changes in serum sIL-2r levels generally paralleled the changes in mass and lymph node size in dogs with high-grade B-cell lymphoma. This study demonstrated that serum sIL-2r level would be a marker to monitor tumour growth and regression in canine lymphoma.

The objective of this retrospective study was to report treatment outcomes in dogs with histiocytic sarcoma (HS) that were treated with nimustine (ACNU). This study evaluated data from 11 dogs including 5 with macroscopic tumors that were treated in the primary setting and 6 that underwent aggressive local therapy while being treated in the adjuvant setting. The median ACNU starting dose was 25 mg/m2 (range, 20-30 mg/m2; 3- to 5-wk intervals, 1-8 administrations). The median overall survival in the primary and adjuvant settings was 120 days (median progression-free survival [PFS], 63 days) and 400 days (median PFS, 212 days), respectively. Neutropenia was observed in eight cases (grade 1, n = 1; grade 2, n = 2; grade 3, n = 2; grade 4, n = 3) with nadir neutrophil count at 1 wk after ACNU administration. Mild gastrointestinal toxicity (grade 1-2) was observed in three cases. ACNU was well tolerated and showed a similar outcome to that seen for lomustine, which is a drug commonly used to treat canine HS, in terms of overall survival and PFS in the current study population. Further investigations will need to be undertaken to definitively determine if ACNU is an appropriate alternative to lomustine for the treatment of HS.

Meningiomas are the most common intracranial tumor in dogs and cats, and their surgical resection is often performed because they are present on the brain surface. Typical meningiomas show comparatively characteristic magnetic resonance imaging findings that lead to clinical diagnosis; however, it is necessary to capture not only macroscopic changes but also microstructural changes to devise a strategy for surgical resection and/or quality of removal. To visualize such microstructural changes, diffusion-weighted imaging (DWI) and diffusion tensor imaging (DTI) have been used in human medicine. The aim of this retrospective study was to investigate the different characteristics of the apparent diffusion coefficient (ADC) from DWI and fractional anisotropy (FA) from DTI of meningioma between dogs and cats. Statistical analyses were performed to compare ADC and FA values between the intratumoral or peritumoral regions and normal-appearing white matter (NAWM) among 13 dogs (13 lesions, but 12 each in ADC and FA analysis) and six cats (seven lesions). The NAWM of cats had a significantly lower ADC and higher FA compared to dogs. Therefore, for a comparison between dogs and cats, we used ADC and FA ratios that were calculated by dividing the subject (intra- or peritumoral) ADC and FA values by those of NAWM on the contralateral side. Regarding the intratumoral region, feline meningiomas showed a significantly lower ADC ratio and higher FA ratio than canine meningiomas. This study suggested that ADC and FA may be able to distinguish a meningioma that is solid and easy to detach, like as typical feline meningiomas.

A 2-year-old neutered female Shiba dog exhibited laboured breathing for 1 month. Computed tomography of the thoracic cavity revealed multiple nodules (2-5 mm diameter) in the lungs. Grossly, the lungs were firm and normal in shape. The nodules were grey-white in colour. Microscopically, the nodules were non-encapsulated and exhibited an irregular shape. They were composed of polygonal or spindle cells with indistinct cell borders arranged in sheets. The cells had large, round, hyperchromatic nuclei and abundant pale eosinophilic cytoplasm with no atypia. Intrapulmonary arterial emboli and infiltration into the bronchioles were observed. Immunohistochemically, the cells were positive for vimentin and negative for cytokeratin, glial fibrillary acidic protein and α-smooth muscle actin. Ultrastructurally, the cells displayed cytoplasmic processes, desmosomes and intermediate filaments. These findings led to a diagnosis of diffuse pulmonary meningotheliomatosis with sarcomatous transformation. To the best of our knowledge, this is the first report of diffuse pulmonary meningotheliomatosis in a dog.

OBJECTIVE To investigate epilepsy-related neuropathologic changes in cats of a familial spontaneous epileptic strain (ie, familial spontaneous epileptic cats [FSECs]). ANIMALS 6 FSECs, 9 age-matched unrelated healthy control cats, and 2 nonaffected (without clinical seizures)dams and 1 nonaffected sire of FSECs. PROCEDURES Immunohistochemical analyses were used to evaluate hippocampal sclerosis, amygdaloid sclerosis, mossy fiber sprouting, and granule cell pathological changes. Values were compared between FSECs and control cats. RESULTS Significantly fewer neurons without gliosis were detected in the third subregion of the cornu ammonis (CA) of the dorsal and ventral aspects of the hippocampus as well as the central nucleus of the amygdala in FSECs versus control cats. Gliosis without neuronal loss was also observed in the CA4 subregion of the ventral aspect of the hippocampus. No changes in mossy fiber sprouting and granule cell pathological changes were detected. Moreover, similar changes were observed in the dams and sire without clinical seizures, although to a lesser extent. CONCLUSIONS AND CLINICAL RELEVANCE Findings suggested that the lower numbers of neurons in the CA3 subregion of the hippocampus and the central nucleus of the amygdala were endophenotypes of familial spontaneous epilepsy in cats. In contrast to results of other veterinary medicine reports, severe epilepsy-related neuropathologic changes (eg, hippocampal sclerosis, amygdaloid sclerosis, mossy fiber sprouting, and granule cell pathological changes) were not detected in FSECs. Despite the use of a small number of cats with infrequent seizures, these findings contributed new insights on the pathophysiologic mechanisms of genetic-related epilepsy in cats.

Background: Leucine-rich glioma-inactivated (LGI) proteins play a critical role in synaptic transmission. Dysfunction of these genes and encoded proteins is associated with neurological disorders such as genetic epilepsy or autoimmune limbic encephalitis in animals and human. Familial spontaneous epileptic cats (FSECs) are the only feline strain and animal model of familial temporal lobe epilepsy. The seizure semiology of FSECs comprises recurrent limbic seizures with or without evolution into generalized epileptic seizures, while cats with antibodies against voltage-gated potassium channel complexed/ LGI1 show limbic encephalitis and recurrent limbic seizures. However, it remains unclear whether the genetics underlying FSECs are associated with LGI family genes. In the present study, we cloned and characterized the feline LGI1-4 genes and examined their association with FSECs. Conventional PCR techniques were performed for cloning and mutational analysis. Characterization was predicted using bioinformatics software. Results: The cDNAs of feline LGI1-4 contained 1674-bp, 1650-bp, 1647-bp, and 1617-bp open reading frames, respectively, and encoded proteins comprising 557, 549, 548, and 538 amino acid residues, respectively. The feline LGI1-4 putative protein sequences showed high homology with Homo sapiens, Canis familiaris, Bos taurus, Sus scrofa, and Equus caballus (92%-100%). Mutational analysis in 8 FSECs and 8 controls for LGI family genes revealed 3 non-synonymous and 14 synonymous single nucleotide polymorphisms in the coding region. Only one non-synonymous single nucleotide polymorphism in LGI4 was found in 3 out of 8 FSECs. Using three separate computational tools, this mutation was not predicted to be disease causing. No co-segregation of the disease was found with any variant. Conclusions: We cloned the cDNAs of the four feline LGI genes, analyzed the amino acid sequences, and revealed that epilepsy in FSEC is not a monogenic disorder associated with LGI genes.

Short chain fatty acids (SCFAs) play an important role in the maintenance of colonic homeostasis, and their depletion has been reported in various gastrointestinal disorders. Inflammatory colorectal polyps (ICRPs) are a recently recognized disease specific to miniature dachshunds (MDs), and fecal dysbiosis with a reduction of SCFA-producing bacteria has been reported with this disease. Therefore, this study was performed based on the hypothesis that a reduced SCFA concentration associates with the development of ICRPs. We recruited 11 ICR-Paffected MDs and 25 control MDs. Their fecal SCFA concentrations and bacterial proportions were quantified using high performance liquid chromatography and quantitative real-time PCR, respectively. The feces of ICRP-affected MDs contained lower amounts of propionic acid and lower proportions of Bifidobacterium than the feces of control MDs. Furthermore, fecal proportions of Bifidobacterium, Firmicutes and Lactobacillus exhibited significant positive correlations with fecal concentrations of total SCFAs and/or propionic acid; fecal Escherichia coli proportions correlated negatively with fecal concentrations of total SCFAs, as well as acetic, propionic and butyric acid. This result indicates an association between fecal dysbiosis and fecal SCFA concentrations; these phenomena may contribute to ICRP pathogenesis in MDs. Potential therapeutic targeting of the reduced propionic acid concentration using probiotics, prebiotics or SCFA enemas merits further study.

Objective: The familial spontaneous epileptic cat (FSEC) is thought to be a good genetic model of mesial temporal lobe epilepsy. In the current study, cerebral diffusion and perfusion magnetic resonance imaging (MRI) were used to confirm the functional deficit zone in the FSEC and evaluate the effect of a single seizure on different brain regions. Methods: Six FSECs and six healthy control cats were used in this study. MRI was performed in the interictal state (resting state for control) and postictal state immediately after the vestibular stimulation-induced generalized epileptic seizure (control cats received the same stimulation as the FSECs). The apparent diffusion coefficient (ADC), fractional anisotropy and perfusion parameters (i.e., relative regional cerebral blood volume (rCBV), relative regional cerebral blood flow (rCBF), and relative regional mean transit time (rMTT)) were measured in the hippocampus, amygdala, thalamus, and gray and white matter. Results: In the interictal state, the rCBV and rMTT in the hippocampus was significantly decreased in FSECs, compared to the control. In the postictal state, FSECs had a significantly decreased ADC and an increased rCBV, rCBF, and rMTT in the hippocampus, and an increased rMTT in the amygdala, compared to the interictal state. Conclusion: This study showed that FSECs had interictal hypoperfusion in the hippocampus, and postictal hypodiffusion and hyperperfusion in the hippocampus and/or amygdala. These findings suggested that the hippocampus and/or amygdala act as the functional deficit and expanded seizure-onset zones in FSECs.

Although MRI has become widely used in small animal practice, little is known about the validity of advanced MRI techniques such as diffusion-weighted imaging and diffusion tensor imaging. The aim of this retrospective analytical observational study was to investigate the characteristics of diffusion parameters, that is the apparent diffusion coefficient and fractional anisotropy, in dogs with a solitary intracranial meningioma or histiocytic sarcoma. Dogs were included based on the performance of diffusion MRI and histological confirmation. Statistical analyses were performed to compare apparent diffusion coefficient and fractional anisotropy for the two types of tumor in the intra- and peritumoral regions. Eleven cases with meningioma and six with histiocytic sarcoma satisfied the inclusion criteria. Significant differences in apparent diffusion coefficient value (x 10(-3) mm(2)/s) between meningioma vs. histiocytic sarcoma were recognized in intratumoral small (1.07 vs. 0.76) and large (1.04 vs. 0.77) regions of interest, in the peritumoral margin (0.93 vs. 1.08), and in the T2 high region (1.21 vs. 1.41). Significant differences in fractional anisotropy values were found in the peritumoral margin (0.29 vs. 0.24) and the T2 high region (0.24 vs. 0.17). The current study identified differences in measurements of apparent diffusion coefficient and fractional anisotropy for meningioma and histiocytic sarcoma in a small sample of dogs. In addition, we observed that all cases of intracranial histiocytic sarcoma showed leptomeningeal enhancement and/or mass formation invading into the sulci in the contrast study. Future studies are needed to determine the sensitivity of these imaging characteristics for differentiating between these tumor types.

OBJECTIVE To evaluate the usefulness of diffusion and perfusion MRI of the cerebrum in cats with familial spontaneous epilepsy (FSECs) and identify microstructural and functional deficit zones in affected cats. ANIMALS 19 FSECs and 12 healthy cats. PROCEDURES Diffusion-weighted, diffusion tensor, and perfusion-weighted MRI of the cerebrum were performed during interictal periods in FSECs. Imaging findings were compared between FSECs and control cats. Diffusion (apparent diffusion coefficient and fractional anisotropy) and perfusion (relative cerebral blood volume [rCBV], relative cerebral blood flow [rCBF], and mean transit time) variables were measured bilaterally in the hippocampus, amygdala, thalamus, parietal cortex gray matter, and subcortical white matter. Asymmetry of these variables in each region was also evaluated and compared between FSECs and control cats. RESULTS The apparent diffusion coefficient of the total amygdala of FSECs was significantly higher, compared with that of control cats. The fractional anisotropy of the right side and total hippocampus of FSECs was significantly lower, compared with that of control cats. The left and right sides and total hippocampal rCBV and rCBF were significantly lower in FSECs than in control cats. The rCBV and rCBF of the parietal cortex gray matter in FSECs were significantly lower than in control cats. CONCLUSIONS AND CLINICAL RELEVANCE In FSECs, diffusion and perfusion MRI detected microstructural changes and hypoperfusion (lowered function) in the cerebrum during interictal periods from that of healthy cats. These findings indicated that diffusion and perfusion MRI may be useful for noninvasive evaluation of epileptogenic foci in cats.

A 16-year-old castrated male mongrel cat presented with swelling under the left pinna and a 3 -month history of voice change. Laryngeal endoscopy revealed circumferential oedema around the arytenoid cartilages and hypersecretion of saliva. Histopathological examination of the mass around the left ear canal was considered the primary lesion that originated from cutaneous apocrine adenocarcinoma or parotid gland adenocarcinoma, and it metastasized to the larynx, lung and medial retropharyngeal lymph nodes. This report provides new insights into feline laryngeal diseases which could result in laryngeal metastasis with slight mucosal irregularity alone and without obvious radiographic abnormalities. Therefore, histopathological examination should be performed when a cat presents clinical signs such as stridor, dysphonia or voice change without any mass-forming laryngeal lesion.

The p16 gene acts as a tumor suppressor by regulating the cell cycle and is frequently inactivated in human and canine cancers. The aim of this study was to characterize genetic and epigenetic alterations of the p16 in feline lymphoid and non-lymphoid malignancies, using 74 primary tumors and 11 tumor cell lines. Cloning of feline p16 and subsequent sequence analysis revealed 11 germline sequence polymorphisms in control cats. Bisulfite sequencing analysis of the p16 promoter region in a feline lymphoma cell line revealed that promoter methylation was associated with decreased mRNA expression. Treatment with a demethylating agent restored mRNA expression of the silenced p16. PCR amplification and sequencing analysis detected homozygous loss (five tumors, 6.7%) and a missense mutation (one tumor, 1.4%) in the 74 primary tumors analyzed. Methylation-specific PCR analysis revealed promoter methylation in 10 primary tumors (14%). Promoter methylation was frequent in B cell lymphoid tumors (7/21 tumors, 33%). These genetic and epigenetic alterations were also observed in lymphoma and mammary gland carcinoma cell lines, but not detected in non-neoplastic control specimens. These data indicate that molecular alterations of the p16 locus may be involved in the development of specific types of feline cancer, and warrant further studies to evaluate the clinical value of this evolutionarily-conserved molecular alteration in feline cancers. (C) 2016 Elsevier Ltd. All rights reserved.

In order to investigate whether suppression of the p16 gene is mediated by histone 113 acetylation in 4 canine lymphoid tumor cell lines, the gene's acetylation status was examined. In 2 canine lymphoid tumor cell lines with low p16 mRNA expression (GL-1 and UL-1), the acetylation level was lower than that in CL-1 cells with high p16 mRNA expression. The expression of the p16 gene in these 2 cell lines was markedly restored after culture in the presence of a histone deacetylase inhibitors trichostatin A, indicating that p16 was inactivated by hypoacetylation. Findings obtained this study will add new insights and lead to the better understanding of the disease pathogenesis and future development of epigenetic therapeutic strategies.

Background: Epilepsy is the most common neurological disease in veterinary practice. However, contrary to human medicine, epilepsy classification in veterinary medicine had not been clearly defined until recently. A number of reports on canine epilepsy have been published, reflecting in part updated proposals from the human epilepsy organization, the International League Against Epilepsy. In 2015, the International Veterinary Epilepsy Task Force (IVETF) published a consensus report on the classification and definition of canine epilepsy. The purpose of this retrospective study was to investigate the etiological distribution, survival time of dogs with idiopathic epilepsy (IdE) and structural epilepsy (StE), and risk factors for survival time, according to the recently published IVETF classification. We investigated canine cases with epilepsy that were referred to our teaching hospital in Japan during the past 10 years, and which encompassed a different breed population from Western countries. Results: A total of 358 dogs with epilepsy satisfied our etiological study criteria. Of these, 172 dogs (48 %) were classified as IdE and 76 dogs (21 %) as StE. Of these dogs, 100 dogs (consisting of 65 with IdE and 35 with StE) were included in our survival study. Median survival time from the initial epileptic seizure in dogs with IdE and StE was 10.4 and 4.5 years, respectively. Median lifespan of dogs with IdE and StE was 13.5 and 10.9 years, respectively. Multivariable analysis demonstrated that risk factors for survival time in IdE were high seizure frequency (&gt;= 0.3 seizures/month) and focal epileptic seizures. Conclusions: Focal epileptic seizures were identified as a risk factor for survival time in IdE. Clinicians should carefully differentiate seizure type as it is difficult to identify focal epileptic seizures. With good seizure control, dogs with IdE can survive for nearly the same lifespan as the general dog population. Our results using the IVETF classification are similar to previous studies, although some features were noted in our Japanese canine population (which was composed of mainly small-breed dogs), including a longer lifespan in dogs with epilepsy and a larger percentage of meningoencephalomyelitis of unknown origin in dogs with StE.

Chronic gastrointestinal disease is associated with the alteration of gastrointestinal microbiota. Inflammatory colorectal polyps (ICRPs) are commonly observed in miniature dachshunds (MDs) in Japan and are characterized by multiple polyps that are restricted in the colorectal mucosa with severe neutrophil infiltration. This study was aimed to compare the fecal microbiota of ICRP-affected MDs with that of healthy MDs. High-throughput sequencing of amplicons derived from the V3-V4 region of the 16S rRNA gene was applied using the Illumina MiSeq system. Principal coordinates analysis revealed that fecal microbiota of ICRP-affected MDs was significantly altered compared with that of healthy MDs. Proportions of Fusobacteriaceae, Helicobacteraceae, Porphyromonadaceae, and Turicibacteraceae were significantly more abundant in ICRP-affected MDs, while those of Lachnospiraceae were significantly less abundant in ICRP-affected MDs compared with healthy MDs. These results suggest that the dysbiosis is associated with ICRPs and is a potential therapeutic target, though further investigations are needed. (C) 2016 Published by Elsevier Ltd.

Although regulatory T cells (Tregs) play an integral role in immunologic tolerance and the maintenance of intestinal homeostasis, their involvement in canine gastrointestinal diseases, including idiopathic inflammatory bowel disease (IBD) and intestinal lymphoma, remains unclear. Here we show altered numbers of forkhead box P3 (Foxp3)-positive Tregs in the intestine of dogs with IBD and intestinal lymphoma. IBD was diagnosed in 48 dogs; small cell intestinal lymphoma was diagnosed in 46 dogs; large cell intestinal lymphoma was diagnosed in 30 dogs; and 25 healthy beagles were used as normal controls. Foxp3-positive Tregs in the duodenal mucosa were examined by immunohistochemistry and immunofluorescence. Duodenal expression of interleukin-10 mRNA was quantified by real-time reverse transcription polymerase chain reaction. The number of Foxp3-positive lamina propria cells and the expression of interleukin-10 mRNA were significantly lower in dogs with IBD than in healthy dogs and dogs with intestinal lymphoma. The number of Foxp3-positive intraepithelial cells was higher in dogs with small cell intestinal lymphoma. Some large cell intestinal lymphoma cases had high numbers of Foxp3-positive cells, but the increase was not statistically significant. Double-labeling immu-nofluorescence showed that CD3-positive granzyme B-negative helper T cells expressed Foxp3. In small cell intestinal lymphoma cases, the overall survival of dogs with a high Treg density was significantly worse than that of dogs with a normal Treg density. These results suggest that a change in the number of Foxp3-positive Tregs contributes to the pathogenesis of canine IBD and intestinal lymphoma by disrupting mucosal tolerance and suppressing antitumor immunity, respectively.

Serum microRNAs (miRNAs) are mediators of cell-to-cell communication and alter the cellular microenvironment; they are stable for hours under certain conditions in body fluids despite the presence of RNases. Certain miRNAs have been found to be altered in the serum or plasma of humans with various cancers and may represent promising, non-invasive biomarkers for various diseases in humans and animals. The objective of this study was to determine the expression profile of circulating miRNAs in the serum of dogs with lymphoma. Serum samples were obtained from 61 dogs with lymphoma and 40 control dogs, and real-time reverse transcription-polymerase chain reaction was used for miRNA measurement. In order to select candidate genes, a comprehensive expression analysis was undertaken prior to validation of several candidate miRNAs. Of 277 miRNAs, five (let-7b, miR-223, miR-25, miR-92a, and miR-423a) were selected as candidates. The expression levels of four miRNAs (let-7b, miR-223, miR-25, miR-92a) were significantly reduced in the lymphoma group, whereas miR-423a levels were significantly increased compared to the controls. When the lymphoma cases were categorized into high- or low-grade as well as into their anatomic form, miR-25 levels were lower in the serum samples from the lymphoma group compared to those from the control group. Although the biological function of serum miRNAs still remains unclear, determining their functional roles in serum and tissues will contribute not only to the identification of potential biomarkers but also to the elucidation of the pathogenesis of canine lymphoma. (C) 2015 Elsevier Ltd. All rights reserved.

Polymerase chain reaction (PCR) amplification to detect immunoglobulin heavy chain (IgH) and T cell receptor gamma-chain (TCR gamma) gene rearrangements has recently become widely used as part of the diagnostic strategy for lymphoid tumors in dogs. In this study, we constructed a multicolor GeneScan analytical system to improve the sensitivity and resolution of the clonality analysis of antigen receptor gene rearrangements in dogs. We used 7 reactions per sample, with 2 PCR conditions, to amplify IgH/TCR gamma and control genes. By using multicolor-labeled primers, these 7 PCR products could be combined into 3 tubes before capillary electrophoresis. Clonal rearrangement of the IgH/TCR gamma genes was detected in 93.3% of dogs with multicentric lymphoma and 84.6% of dogs with gastrointestinal lymphoma. Detection sensitivity of the clonally expanded cells in the background of normal peripheral blood mononuclear cells was 1-10%. The multicolor GeneScan analytical system developed here may prove to be helpful for the diagnosis of lymphoid tumors in dogs. (C) 2015 Elsevier B.V. All rights reserved.

Inflammatory colorectal polyps (ICRPs) frequently occur in miniature dachshunds (MDs) in Japan. MDs with ICRPs develop multiple polyps with severe neutrophil infiltration that respond to immunosuppressive therapy. Therefore, ICRPs are thought to constitute a novel, breed-specific form of canine inflammatory bowel disease (IBD). Pattern recognition receptors (PRRs) play a key role in the distinction of pathogens from commensal bacteria and food antigens. Dysfunction resulting from genetic disorders of PRRs have been linked to human and canine IBD. Therefore, we analyzed the reactivity of PRRs in MDs with ICRPs. Twenty-six MDs with ICRPs and 16 control MDs were recruited. Peripheral blood-derived monocytes were obtained from each dog and then stimulated with PRR ligands for 6 and 24 hr; subsequently, messenger RNA (mRNA) expression levels and protein secretion of IL-1 beta were quantified using quantitative real-time PCR and ELISA, respectively. The levels of IL-1I3 mRNA and protein secretion after stimulation with a nucleotide-binding oligomerization domain 2 (NOD2) ligand were significantly greater in monocytes from ICRP-affected MDs than in those from control MDs. In addition, IL113 protein secretion induced by toll-like receptor (TLR) 1/2, TLR2 and TLR2/6 stimulation was also significantly greater in ICRP-affected MDs. These results suggest that reactivity against NOD2, TLR1/2, TLR2 and TLR2/6 signals is enhanced in ICRP-affected MDs and may play a role in the pathogenesis of ICRPs in MDs. Additional studies of the genetic background of these PRRs should be performed. KEYWORDS: inflammatory colorectal polyp, innate immunity, miniature dachshund, pattern recognition receptor, pro-inflammatory cytokine

Inflammatory colorectal polyps (ICRPs) frequently occur in miniature dachshunds (MDs) in Japan, typically form multiple polyps with severe neutrophil infiltration. ICRPs are speculated as a novel, breed-specific canine inflammatory bowel disease. Pattern recognition receptors (PRRs) play an important role in the differentiation of pathogens from commensal bacteria and food antigens, and polymorphisms of various PRRs have been shown to be associated with human and canine IBD. We recently reported that the reactivity of nucleotide-binding oligomerization domain 2 (NOD2), toll-like receptor (TLR) 1/2, TLR2, and TLR2/6 are greater in ICRP-affected MDs than that in controls. Therefore, this study was aimed to investigate single nucleotide polymorphisms (SNPs) of PRRs associated with ICRPs in MDs. Mutational analysis of canine NOD2, TLR1, TLR2, and TLR6 genes was performed with six ICRP-affected MDs, five control MDs, and five healthy beagles. The mutational analysis identified 13 non-synonymous SNPs in NOD2,TLR1, TLR2, and TLR6 genes, of which six SNPs in NOD2 exon 3 were further analyzed in an association study using 63 ICRP-affected MDs, 82 control MDs, and 237 control dogs of various breeds. Four of the SNPs (A1532G, T1573C, C1688G, and G1880A of the NOD2 gene) were in Hardy-Weinberg equilibrium and in complete linkage disequilibrium in MDs, and their minor allele frequencies were significantly lower in ICRP-affected MDs than in control MDs (0.016 vs. 0.140, P= 0.0002). The calculated inheritance model was an additive model (odds ratio = 0.10, 95% confidence interval = 0.02-0.45, P = 0.0001), which indicates that the haplotype with minor alleles in these SNPs (A, T, C, and Gin A1532G, T1573C, C1688G, and G1880A) possess a protective effect regarding the development of ICRPs. However, these SNPs were not specific for MDs, although the minor allele frequencies of these sNPs in control MDs were significantly lower than in other breed dogs. These results suggest that the identified four SNPs (A1532G, T1573C, C1688G, and G1880A in the NOD2 gene) may play a role in the pathogenesis of ICRPs in MDs. Because the majority of MDs and other breed dogs do not have the protective alleles, their absence may not be a specific cause of ICRPs in MDs but rather contribute to the development of inflammation. (C) 2015 Elsevier B.V. All rights reserved.

This study explored the hypothesis that inflammatory colorectal polyps (ICRPs) in miniature Dachshunds are more likely to occur ventrally in the colorectum. Angle-fixed colonoscopic images were collected from 11 miniature Dachshunds with ICRPs and randomly rotated. Macroscopic severity at 12 divided angles was scored by four veterinarians blinded to the rotation angle. Mean prevalence and severity scores of ICRPs were significantly higher ventrally than dorsally (P &lt; 0.01). (C) 2014 Elsevier Ltd. All rights reserved.

A strain of familial spontaneous epileptic cats (FSECs) with typical limbic seizures was identified in 2010. The electroencephalographic features suggested that an epileptogenic zone is present in the mesial temporal structures (i.e., amygdala and/or hippocampus). In this study, visual evaluations and quantitative analyses were performed by using 3D MR hippocampal volumetry in comparing FSECs with age-matched controls. Visual hippocampal asymmetries were seen in 8 of 14 (57.1%) FSECs. The FSEC group showed a significantly higher asymmetric ratio (4.15%) than the control group (0.99%). The smaller side of hippocampal volume (HV) (0.206 cm(3)) in FSECs was significantly smaller than the mean HV in controls (0.227 cm(3)). However, the means of left and right HVs and total HVs in FSECs showed no differences because the laterality of hippocampal atrophy was different in each individual. Therefore, since FSECs represent a true model of spontaneous epilepsy, hippocampal volumetry should be evaluated in each individual as well as in human patients. The significant asymmetry of HV suggests the potential for hippocampal atrophy in FSECs. (C) 2014 Elsevier B.V. All rights reserved.

Feline nasal tumours (NTs) are locally invasive and occasionally metastasise to distant sites. Although palliative hypofractionated radiotherapy (HRT) is used, its efficacy and long-term complications have not been adequately evaluated. The purpose of this study was to evaluate the efficacy of HRT in treating feline malignant NTs, including monitoring improvement in clinical signs, acute and late complications, and prognosis. The medical records of 65 cats with malignant NTs treated with HRT were included. Overall survival (OS) and progression-free survival (PFS) were calculated using the Kaplan-Meier method. The log-rank test and Cox proportional hazard model were used to evaluate factors that influenced OS and PFS. Clinical signs improved in 86.2% of cats following radiotherapy. Acute complications were observed in 58.5% of cats but were manageable and acceptable. Among late complications, cataract was most frequently observed (20.5%), and atrophy of the entire eyeball and osteochondroma at the irradiation site were each observed in two cats. The median OS and PFS in 65 cats were 432 days and 229 days, respectively. No significant difference between OS of cats with nasal lymphoma and that of cats with other tumours was observed. Despite some limitations due to the retrospective nature of the study, palliative HRT for feline NTs can be considered a useful treatment option because of the high incidence of improvement and more favourable prognosis, although it may be preferable not to use the hypofractionated regimen in young cats with lymphoma that are expected to survive for a long period. (C) 2014 Elsevier Ltd. All rights reserved.

Death-associated protein kinase (DAPK) is a 160-kD serine/threonine kinase known as a key molecule in interferon-gamma (IFN-gamma)-induced apoptosis and tumor suppression. Hypermethylation of the CpG island in DAPK inactivates the gene in a variety of human malignancies. This study aimed to detect the inactivation of DAPK in canine lymphoid tumor cells. The sequence of canine DAPK cDNA was obtained from normal dog peripheral blood mononuclear cells after reverse transcription polymerase chain reaction (RT-PCR). By rapid amplification of 5'-cDNA ends, the transcription initiation site of the DAPK gene was identified. The CpG island located upstream of the translation initiation site was identified by using a search algorithm. The methylation status of the CpG island was examined using bisulfite sequence analysis and methylation-specific PCR (MSP). The inactivation of DAPK gene was examined in 3 canine lymphoid tumor cell lines, GL-1 (B-cell leukemia), CLBL-1 (B-cell lymphoma), and CL-1 (T-cell lymphoma). DAPK mRNA expression was measured by real-time RT-PCR. IFN-gamma-induced apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) assay. The influence of demethylation was examined with 5-aza-2'-deoxycytidine (5-aza-dC). The methylation status in 14 dogs with various lymphoid tumors was screened by MSP. A 1926-bp CpG island containing 280 CpG repeats was identified upstream of the translation start site of canine DAPK. Bisulfate sequence analysis and MSP revealed hypermethylation of the CpG island in GL-1 cells, but not in CLBL-1 or CL-1 cells. The amount of DAPK mRNA was significantly smaller in GL-1 cells than CLBL-1 and CL-1 cells. IFN-gamma-induced apoptosis was detected in CLBL-1 and CL-1 cells but not in GL-1 cells. Treatment with 5-aza-dC significantly increased the amount of DAPK mRNA and IFN-gamma-induced apoptosis in GL-1 cells. These results revealed the inactivation of DAPK through methylation of its CpG island in GL-1 cells. MSP showed hypermethylation of the DAPK CpG island in 5 of 8 primary B-cell lymphoma samples, but not in any of the 6 primary T-cell lymphoid tumor samples obtained from canine patients. DAPK was inactivated through hypermethylation of its CpG island in canine B-cell lymphoid tumor cells. This study will lead to the use of canine B-cell lymphoid tumors as an animal model to evaluate the efficacy of demethylating agents. (C) 2014 Elsevier B.V. All rights reserved.

Objective-To examine the DNA methylation status of the ABCB1 gene in tumor cells of dogs with lymphoma. Animals-27 dogs with multicentric B-cell high-grade lymphoma (19 chemotherapy-sensitive dogs and 8 chemotherapy-resistant dogs). Procedures-The DNA methylation profile of the CpG island of the ABCB1 gene was analyzed by use of bisulphite sequencing and real-time methylation-specific PCR assay in lymphoma cells. Quantitative reverse transcriptase PCR assay of the ABCB1 gene was conducted to measure the amount of mRNA. Correlation between the amount of ABCB1 mRNA and the methylation rate was examined. Results-The CpG island of the ABCB1 gene was hypomethylated in most dogs in both the chemotherapy-sensitive and-resistant groups. No significant difference was detected in the methylation rate between the 2 groups, and no significant correlation was detected between the methylation rate and the mRNA expression level. Conclusions and Clinical Relevance-Expression of the ABCB1 gene was not suppressed by hypermethylation of its CpG island in most dogs with lymphoma regardless of their chemotherapy sensitivity status.

To investigate the epigenetic regulation of the p16 gene in canine lymphoid tumor cells, its methylation status was examined in four canine lymphoid tumor cell lines. In three canine lymphoid tumor cell lines (CLBL-1, GL-1, and UL-1) with low-level p16 mRNA expression, 20 CpG sites in the promoter region of p16 gene were consistently methylated although all of the CpG sites were not methylated in another cell line (CL-1) and normal lymph node cells. The expression level of p16 mRNA in these three cell lines was restored after cultivation in the presence of a methylation inhibitor, 5-Aza-2'-deoxycitidine, indicating inactivation of p16 gene via hypermethylation. This study revealed the inactivation of p16 gene through hypermethylation of its CpG island in a fraction of canine lymphoid tumor cells. (C) 2014 Elsevier Ltd. All rights reserved.

Although decreased intestinal IgA expression has been reported in dogs with inflammatory bowel disease (IBD), the mechanism underlying this decrease is unknown. Transmembrane activator and calcium-modulating cyclophilin-ligand interactor (TACI) and B cell-activating factor of the TNF family (BAFF) receptor (BAFF-R) are key receptors for T cell-independent IgA class switching by the binding of IgA-inducing cytokine a proliferation-inducing ligand (APRIL) and BAFF. Here we show decreased TACI and BAFF-R mRNA expression and hyper-methylation of their corresponding genes TNFRSF13B and TNFRSF13C, respectively in the duodenal mucosa of dogs with IBD. To examine whether DNA methylation of the TNFRSF13B and TNFRSF13C influences the mRNA expression of TACI and BAFF-R, respectively, we first analyzed methylation and mRNA expression levels in vitro using 2 canine B lymphoid cell lines, GL-1 and CLBL-1. Methylation profiles in the cells were examined by bisulfite sequencing and methylation-specific PCR (MSP) with primer pairs specific to methylated or unmethylated sequences. These methylation analyses revealed hyper-methylation of the CpG islands of both TNFRSF13B and TNFRSF13C in GL-1, but not in CLBL-1 cells. The mRNA expression levels of TACI and BAFF-R were significantly lower in GL-1 than in CLBL-1 cells. Treatment with 5-aza-2'-deoxycytidine significantly increased TACI and BAFF-R mRNA expression in GL-1 cells through demethylation of TNERSF13B and TNFRSF13C, respectively. These results suggest that the mRNA expression of TACI and BAFF-R is regulated through methylation of their genes in canine B cells. Quantitative real-time MSP showed significant hyper-methylation of the CpG islands of TNFRSF13B and TNFRSF13C in the duodenal mucosa of dogs with IBD. Furthermore, duodenal mRNA expression levels of TACI and BAFF-R were significantly lower in dogs with IBD than in healthy controls. The mRNA expression levels of TACI positively correlated with intestinal IgA expression, whereas the methylation level of its gene (TNFRSF13B) negatively correlated with IgA expression. The present results suggest the role of TACI in the regulation of mucosal IgA expression through epigenetic modifications. (C) 2014 Elsevier B.V. All rights reserved.

The prognostic significance of the inactivation of the p16, p15, and p14 genes has been reported in lymphoid malignancies in humans. To evaluate the relationship between inactivation of the p16, p15, and p14 genes and prognosis in canine high-grade lymphoma, primary tumor cell samples obtained from 71 dogs with high-grade lymphoma were examined for the expression levels of these genes. Quantitative and conventional reverse transcriptase-polymerase chain reaction (RT-PCR) analyses were used to measure the amounts of p16, p15, and p14 mRNAs. The methylation status of the CpG island of the p16 gene was evaluated using methylation-specific PCR. Overall survival (OS) was compared using the KaplanMeier method. The log-rank test and Cox proportional hazard model were used to evaluate factors that influenced OS. Of 62 dogs examined, p16, p15, and p14 mRNA levels were found to be undetectable in 21, 18, and 10 dogs, respectively. In 20/68 dogs analyzed, the CpG island of the p16 gene was shown to be methylated. The prognostic significance of inactivation of the p16, p15, and p14 genes as well as various conventional factors obtained from medical records was examined. p16 expression status and anatomic form/immunophenotype were found to correlate with OS in the dogs with high-grade lymphoma. p16 mRNA level over its cut-off value correlated with a poor prognosis; however, the expression levels of p15 and p14 mRNAs and p16 methylation status did not influence the prognosis in dogs with high-grade lymphoma. (C) 2013 Elsevier Ltd. All rights reserved.

The 6, p15 and p14 genes are widely known as tumor suppressor genes in human medicine. Although a large number of genetic and epigenetic aberrations in these genes have been reported in human malignancies, canine malignancies have not been well analyzed on the aberrations of these genes. In this study, the full-length complementary DNA (cDNA) of the canine p16 gene was cloned using the 5' and 3' rapid amplification of cDNA ends methods. Based on the sequence data, primers specific for p16,p15 and p14 were designed. Using these primers, the expression of p16, p15 and p14 mRNAs could be individually evaluated by reverse transcriptase polymerase chain reaction. Genomic aberrations were also examined using genomic polymerase chain reaction. Two of the 6 canine lymphoid tumor cell lines did not express detectable levels of p16,p15 and p14 mRNAs, and wide-ranging deletions in the p15-p14-p16 genomic locus were suspected. Wide-ranging deletions were also speculated in 2 of 14 dogs with T-cell lymphoid tumors. On the other hand, similar failure of amplification suggesting wide-ranging deletions were not observed in any of the 14 dogs with B-cell lymphoma. Deletion of the p15-p14-p16 genomic locus could be one of the molecular aberrations in canine lymphoid tumor cells.

Objectives To evaluate the efficacy of hypofractionated radiotherapy for canine nasal tumours, including the improvement in clinical signs, rate of complications and assessment of prognostic factors. Methods Medical records of 38 dogs with malignant nasal tumours were reviewed, and those treated with a weekly schedule of hypofractionated radiotherapy were included in the study. Acute and late side effects were defined as complications noted either within 1month or after 6months of irradiation, respectively. Progression-free interval and overall survival were calculated using the KaplanMeier method. Log-rank test and Cox proportional hazard model were also performed. Results Clinical signs improved in 30 of 36 dogs. Acute complications were seen in 28 of 36 dogs and were considered manageable. Late complications were observed in 17 of 30 dogs that survived 6months or longer, but severe side effects were not observed. The median progression-free interval and overall survival was 245days (95% CI: 127512days) and 512days (95% CI: 203820days), respectively. Age, breed and presence of dyspnoea were negatively correlated with overall survival. Clinical Significance These results suggest that hypofractionated radiotherapy could be a viable option for the treatment of nasal tumours in dogs that are not candidates for conventional multi-fractionated radiotherapy. Journal of Small Animal Practice (2013) 54, 80-86 DOI: 10.1111/jsap.12024

Fractalkine, also known as CX(3)CL1, is a unique chemokine that mediates inflammatory responses and is involved in the pathogenesis of several inflammatory disorders, including inflammatory bowel disease (IBD) in humans. In this study, we isolated cDNAs encoding canine fractalkine and its receptor CX(3)CR1, and assessed the biological activity of these molecules. The deduced amino acid sequence of the canine fractalkine cDNA showed 66% and 57% identity to human and mouse homologs, respectively. The N-terminal chemokine domain of the canine fractalkine showed 68% and 65% identity to human and mouse counterparts, respectively. The canine CX(3)CR1 amino acid sequence showed close homology to its human (83% identity) and mouse (81% identity) counterparts. Fractalkine and CX(3)CR1 mRNA were detected in all tissues in this study. Relatively higher expression levels of fractalkine mRNA were observed in the brain, medulla spinalis, small intestine, and mesenteric lymph nodes (MLNs), whereas higher expression levels of CX(3)CR1 mRNA were observed in the medulla spinalis, brain, liver, small intestine, and MLNs. The cross-reactivities of anti-human fractalkine antibody and anti-rat CX(3)CR1 antibody to canine proteins were confirmed using recombinant canine fractalkine and a cell line overexpressing canine CX(3)CR1, respectively. A transwell chemotaxis assay showed that the recombinant canine fractalkine induced migration in canine lymphoid cells expressing CX(3)CR1. The present study will be useful in understanding the canine immune system and the immunopathogenesis of canine inflammatory diseases. (C) 2011 Elsevier B.V. All rights reserved.