Role of beta1 integrins in adhesion of canine mastocytoma cells to extracellular matrix proteins

The interactions of tumor cells with the extracellular matrix (ECM) are a crucial step in invasion and metastasis. Integrins are adhesive molecules forming heterodimers with alpha and beta subunits that play a definitive role in these interactions. In this study, mastocytoma (mast cell tumor: MCT) cell-ECM interaction was investigated using 3 canine MCT cell lines: CM-MC (originating from cutaneous MCT), VI-MC (originating from intestinal MCT), and CoMS (originating from oral MCT). Flow cytometric analysis showed that all cells highly expressed the integrin beta1 and alpha1 through alpha5 subunits and that they moderately expressed the alpha6 subunit. In adhesion studies, CoMS weakly but spontaneously adhered to fibronectin (FN), which was enhanced by phorbol ester (TPA), while CM-MC and VI-MC required cell activation by TPA to adhere to FN. Anti-beta1 and alpha5 integrin antibodies strongly inhibited cell adhesion to FN in CM-MC and CoMS and moderately inhibited cell adhesion in VI-MC. Only VI-MC adhered to laminin (LN) under activation by TPA. Anti-beta1 integrin antibodies strongly inhibited cell adhesion to LN, but all anti-alpha integrin antibodies failed to inhibit cell adhesion to LN. No cells adhered to collagen types I and IV. Canine MCT cells from different origins expressed similar integrin patterns; however, there were some differences in adhesive behavior in response to various ECM proteins and activating stimuli.     

Survey of tortoises with urolithiasis in Japan

Urolithiasis is a disease often seen in tortoises at veterinary hospitals, however there have been no comprehensive research reports of tortoises with urolithiasis in Japan. In this study, we analyzed tortoises diagnosed with urolithiasis at three domestic veterinary hospitals. Based on medical records, we assessed the diagnostic method, species, sex, body weight, dietary history, husbandry, clinical signs, clinical examination, treatment for urolithiasis, and clinical outcome. The total number of cases in the 3 facilities was 101. As for species of tortoises, the most common was the African spurred tortoise (Centrochelys sulcata) with 42 cases (41.6%), followed by the Indian star tortoise (Geochelone elegans) with 30 cases (29.7%). Six other species were confirmed to have calculi. Almost all cases (99 cases, 98%) had a single calculus, and only 2 had multiple calculi. The prevalence of urolithiasis for the total number of tortoises having visited to one institution during the same period was 5.1%. Of the 86 cases that underwent calculi removal, 64 (74.4%) were successfully removed via the vent, and the efficacy of this method was confirmed. Nineteen cases (22%) were approached via plastronotomy, among which only 2 died postoperatively. In this study, we could not clarify the relationship between calculi formation and diets or other husbandry factors.     

Identification of tumor-initiating cells in a canine hepatocellular carcinoma cell line

Tumor-initiating cells (TICs) or cancer stem cells (CSCs), a small subset of tumor cells, are involved in tumor initiation, progression, recurrence and metastasis. In human hepatocellular carcinoma (HCC), TICs are enriched with cell surface markers and have the ability to self-renew and differentiate tumors at a high frequency. We established a canine HCC cell line, HCC930599, and analyzed it for stem and progenitor cell marker expression using flow cytometry. HCC930599 showed high CD44 and CD29, moderate CD90, and low CD133, CD34, CD24, CD117, and CD13 expression. CD90⁺CD44⁺ and CD90⁻CD44⁺ cells were characterized using the in vitro sphere assay and an in vivo transplant model. CD90⁺CD44⁺ cells acquired enhanced self-renewal capacity, proliferative activity and tumourigenicity compared with CD90⁻CD44⁺ cells, suggesting that TICs exist in the HCC930599 cell line and that CD90 is a marker for enriched TICs. Understanding TIC characteristics may help elucidate hepatic carcinogenesis and HCC therapy development.     

Shotgun proteomic analysis of porcine colostrum and mature milk

The epitheliochorial nature of the porcine placenta prevents the transfer of maternal immunity. Therefore, ingestion of the colostrum immediately after birth is crucial for neonatal piglets to acquire passive immunity from the sow. We performed a shotgun proteomic analysis of porcine milk to reveal in detail the protein composition of porcine milk. On the basis of the Swiss‐Prot database, 113 and 118 proteins were identified in the porcine colostrum and mature milk, respectively, and 50 of these proteins were common to both samples. Some immune‐related proteins, including interleukin‐18 (IL‐18), were unique to the colostrum. The IL‐18 concentration in the colostrum and mature milk of four sows was measured to validate the proteomic analysis, and IL‐18 was only detected in the colostrum (191.0 ± 53.9 pg/mL) and not in mature milk. In addition, some proteins involved in primary defense, such as azurocidin, which has never been detected in any other mammal's milk, were also identified in the colostrum.     

In vitro antiseptic susceptibilities for Staphylococcus pseudintermedius isolated from canine superficial pyoderma in Japan

Background –  Topical therapy, particularly with chlorhexidine, is becoming increasingly common as a treatment option for canine pyoderma; however, there are limited studies on the susceptibility of Staphylococcus pseudintermedius to chlorhexidine compounds. Objectives –  To determine the in vitro susceptibility of both meticillin‐resistant and meticillin‐susceptible S. pseudintermedius isolates to chlorhexidine and other antiseptic agents and the presence of multidrug efflux pump genes. Samples –  One hundred S. pseudintermedius isolates from 23 initial and 77 recurrent cases of canine pyoderma. Methods –  After bacterial identification and mecA testing, minimal inhibitory concentrations (MICs) of antiseptic agents were determined. Multidrug efflux pump genes, including qacA, qacB and smr, were identified. Results –  Of the 100 isolates, 57 were identified as meticillin‐resistant S. pseudintermedius. The MIC₉₀ of chlorhexidine acetate, chlorhexidine gluconate, acriflavine, ethidium bromide and benzalkonium chloride were 1, 1, 2, 0.5 and 2 μg/mL, respectively. Multidrug efflux pump genes qacA, qacB and smr were not detected in any of the isolates. Conclusions and clinical importance –  The MICs for chlorhexidine and other antiseptics remain low, and multidrug efflux pump genes were not found in the tested isolates.     

Nosological study of Borna disease virus infection in race horses.

To investigate the prevalence of diseases in the Borna disease virus (BDV) antibody positive race horses, we undertook seroepidemiological studies of BDV infection on 125 culled race horses in Hokkaido, Japan. The serological study disclosed the presence of antibodies only to BDV-p40 or -p24 in 19.2% (24/125) and 3.2% (4/125) of culled horses, respectively. Antibodies to both BDV-p40 and -p24 were found in 24.0% (30/125) of these horses. Of particular note was the finding that locomotorium disorders were detectable at a significantly higher rate in BDV antibody positive horses than that in the seronegative horses. These results imply that BDV infection may possibly contribute to an increase in the incidence rate of locomotorium disorders in race horses.     

Effects of midazolam-butorphanol, acepromazine-butorphanol and medetomidine on an induction dose of propofol and their compatibility in dogs

The effects of acepromazine-butorphanol (AB), midazolam-butorphanol (MB) and medetomidine (Med) on the induction dose of propofol and their compatibility with propofol were evaluated in client-owned dogs. All premedications induced good to excellent sedation and the induction dose of propofol was considerably reduced. Of the tested premedicants, Med induced the deepest sedation and the most potent dose-sparing effect. Induction of anesthesia was excellent to good in all dogs except for one dog premedicated with MB. Most dogs premedicated with AB or MB showed temporary apnea. Although other adverse effects such as bradycardia or hypotension may also occur, premedication with MB, AB or Med is a valuable technique for the induction of anesthesia with propofol in dogs in a clinical setting.     

Degenerative changes of the cranial cruciate ligament harvested from dogs with cranial cruciate ligament rupture

Degenerative cranial cruciate ligament (CCL) rupture is characterized histologically by degenerating extracellular matrix (ECM) and chondroid metaplasia. Here, we describe the progression of chondroid metaplasia and the changes in the expression of ECM components in canine CCL rupture (CCLR). CCLs from 26 stifle joints with CCLR (CCLR group) and normal CCLs from 12 young beagles (control group) were examined histologically and immunohistochemically for expression of type I (COLI), type II (COLII), type III collagen (COLIII) and Sry-type HMG box 9 (SOX9). Cell density and morphology of CCLs were quantified using hematoxylin–eosin staining. The percentage of round cells was higher in the CCLR group than in controls. COLI-positive areas were seen extensively in the connecting fibers, but weakly represented in the cytoplasm of normal CCLs. In the CCLR group, there were fewer COLI-positive areas, but many COLI-positive cells. The percentages of COLII-, COLIII- and SOX9-positive cells were higher in the CCLR group than in controls. The number of spindle cells with perinuclear halo was high in the CCLR group, and most of these cells were SOX9-positive. Deposition of COLI, the main ECM component of ligaments, decreased with increased COLIII expression in degenerated CCL tissue, which shows that the deposition of the ECM is changed in CCLR. On the contrary, expression of SOX9 increased, which may contribute to the synthesis of cartilage matrix. The expression of COLII and SOX9 in ligamentocytes showed that these cells tend to differentiate into chondrocytes.