Growth characteristics of canine distemper virus in a new cell line CCT cells originated from canine malignant histiocytosis

Canine distemper virus (CDV) growth and the morphological characterization were examined in a cell line established from a canine malignant histiocytosis (CCT cell line). The susceptibility of the CCT cells to 3 CDV strains, FXNO, YSA-TC and MD-77 was shown by detection of the antigen in the indirect fluorescent assay. After passaging 4 and 9 times through the CCT cells, only FXNO strain could produce the syncytia where demonstrated the antigens. Titers of 9 passaged viruses through the CCT cells showed slightly higher in the CCT cells than those in Vero cells. Morphological characterization of karyorrhexis and specific DNA ladder by extracted DNA electrophoresis indicated apoptosis in the CDV infected CCT cells.     

Cloning of swine desmoglein 1 and its direct proteolysis by Staphylococcus hyicus exfoliative toxins isolated from pigs with exudative epidermitis

Exudative epidermitis (EE) is an acute, often fatal skin disease of piglets caused by Staphylococcus hyicus. Clinical and histopathological manifestations of EE are similar to those of staphylococcal scalded skin syndrome (SSSS), a human blistering skin disease, in which exfoliative toxins produced by Staphylococcus aureus digest the extracellular domains of desmoglein (Dsg) 1 and cause loss of epidermal cell-cell adhesion. The aims of this study were to isolate and characterize cDNA for full length of swine Dsg1, and to determine whether the extracellular domains of swine Dsg1 produced by baculovirus (sDsg1-His) could be digested by four isoforms of exfoliative toxin produced by S. hyicus (ExhA, ExhB, ExhC and ExhD). Nucleotide sequencing revealed that swine Dsg1 cDNA consisted of an open reading frame of 3138 bp, encoding a precursor protein of 1045 amino acids. Deduced amino acid sequence of the swine Dsg1 precursor were highly homologous to corresponding bovine, canine, human and murine sequences. Immunoadsorption assay with a secreted form of sDsg1-His revealed that sDsg1-His specifically absorbs the immunoreactivity of 10 human pemphigus foliaceus sera against swine keratinocyte cell surfaces, suggesting its proper conformation. When sDsg1-His was incubated in vitro with Exhs, all four isoforms of Exh directly digested sDsg1-His into smaller peptides, whereas removal of calcium from sDsg1-His completely inhibited its proteolysis by these four Exhs. Recognition and digestion of calcium-stabilized structure on the extracellular domains of swine Dsg1 by Exhs indicated that EE shares similar molecular pathophysiological mechanisms of intra-epidermal splitting with SSSS in humans.     

Direct and indirect markers of cartilage metabolism in synovial fluid obtained from dogs with hip dysplasia and correlation with clinical and radiographic variables

OBJECTIVE: To compare activities of interleukin (IL)-1beta, IL-6, tumor necrosis factor (TNF)-alpha, and matrix metalloproteinase (MMP)-3 and contents of sulfated glycosaminoglycan (S-GAG) in joint fluid obtained from dogs with hip dysplasia (HD) and clinically normal dogs, evaluate correlations among these markers in joint fluid obtained from dogs with HD, and evaluate correlations between each marker and clinical and radiographic variables. Animals-26 dogs with HD (clinical group) and 43 clinically normal Beagles (control group). PROCEDURE: Joint fluid was aseptically collected from the hip joints of all dogs. For each dog in the clinical group, age, duration of lameness, radiographic osteoarthritis (OA) score, and Norberg angle in each affected joint were recorded. Activities of IL-1beta, IL-6, TNF-alpha, and MMP-3 and S-GAG contents were measured. Values were compared between groups by use of Mann-Whitney U tests, and the Spearman rank correlation test was used to evaluate correlations among markers and between each marker and clinical or radiographic variables. RESULTS: Values of all markers were significantly higher for the clinical group, compared with values for the control group. There was a moderate positive correlation between lameness duration and IL-6 activity and a strong negative correlation between the Norberg angle and IL-1beta activity. CONCLUSIONS AND CLINICAL RELEVANCE: Analysis of our results indicated that there was a significant increase in markers of OA in dogs with HD. Activities of IL-1beta and IL-6 in joint fluid of dogs with HD may be influenced by the severity of laxity in the hip joint and lameness duration, respectively.     

Melanocytic matricoma in a dog

We report a tumor that developed at the root of the tail in a male, 2-year-old Shih Tzu dog. Histologically, the tumor consisted of basaloid epithelial cells, with shadow cells and neoplastic melanocyte proliferation. Immunohistochemically, epithelial components were positive for keratin/cytokeratin but basaloid cells were negative. Mononuclear or multinuclear melanocytes were positive for vimentin, S-100 protein, and HMB-45. In conclusion, this dog was diagnosed with canine melanocytic matricoma on the basis of the similarity of the histologic and immunohistochemical features in humans, and it may be the first case in an animal.     

Antigenic analysis of avian Chlamydia psittaci using monoclonal antibodies to the major outer membrane protein.

Monoclonal antibodies to the major outer membrane protein (MOMP) of Chlamydia psittaci derived from a parrot were established for antigenic analysis of avian C. psittaci. With 17 monoclonal antibodies to MOMP, 17 reactivity patterns were identified on 112 strains of C. psittaci, C. pneumoniae and C. trachomatis, which were isolated from birds, mammals and humans in Japan, U.S.A., Canada and Taiwan, from 1938 to 1987. Immunological reactivity of budgerigar-derived strains to the monoclonal antibodies was different from that of pigeon-derived strains. Imported bird-derived strains were distinguishable from domestic bird-derived strains by the reactivity to the monoclonal antibodies. A close relationship between the subtypes and geographic origins was indicated on budgerigar-derived strains. On the contrary, various reactivity patterns were shown in pigeon-derived strains isolated in a narrow area. The monoclonal antibodies established in the present work may be useful probes for ecological study of avian C. psittaci.     

Distribution of TNF receptors and TNF receptor-associated intracellular signaling factors on equine tendinocytes in vitro

Although tumor necrosis factor (TNF) alpha is an important key factor in degeneration of equine superficial digital flexor tendon (SDFT), the dynamism of TNF receptors and associated factors on tendinocytes has not been elucidated. To reveal signaling events mediated by TNF-receptors (TNF-Rs) in tendinocytes, we focused on four signaling factors, TNF-R1, TNF-R2, TNF-R-associated factor 2 (TRAF2) and nuclear factor-kappa B (NF-kappaB), and investigated the distribution and production of these factors. Cultured tendinocytes were obtained from SDFTs of thoroughbred horses. The tendinocytes were treated with 10 ng/ml equine TNFalpha medium for 6 hours and then the four factors on tendinocytes were visualized by using an immunohistochemical method, and the amounts of the four factors were determined by Western blot analysis. Although TNF-R1 and TNF-R2 co-localized on the same tendinocyte, in untreated control cells (normal condition), immunoreactivity against TNF -R1 was very weak but TNF-R2 showed a strong reaction. However, TNF-R1 showed the same high level of reaction as TNF-R2 in TNFalpha-treated cells (inflamed condition). Intense TRAF2 and NF-kappaB were detected at inflamed condition, however both factors were also detected at normal condition. The distinct distributions of the four factors under different conditions (normal and inflamed condition) in vitro not only reflect the dynamism of the cytokines but may also provide important clues for a means to prevent from occurrence of tendonitis and progress of tendon degeneration.     

Fixation to the canine bone of artificial implant with new surface structure

Screw and laser (SL) column by making screw threads and forming small holes using laser irradiation on the base metal and conventional beads coating (BC) columns were embedded into the shaft of canine femurs, and compared the implant fixation to the host bone. The interfacial strength in SL columns was almost equivalent as BC columns, and bone-column contact rate was higher than BC columns significantly at twelve weeks after implantation. The newly devised SL surface had almost equivalent bone fixation strength comparable to the conventional BC surface. Also, this surface should provide a useful porous surface for use in artificial joints since there is no risk of surface structure detachment.     

Molecular epidemiological survey of benign Theileria parasites of cattle in Japan: detection of a new type of major piroplasm surface protein gene

Benign Theileria species of cattle are found in most parts of the world. The major piroplasm surface protein (MPSP), a conserved protein in all Theileria species, has been used as a maker for epidemiological and phylogenetical studies of benign Theileria species. Parasites with Ikeda- or Chitose-type MPSP genes are dominant in Japan, but we report here mixed infection cases of Theileria parasites with an additional MPSP type parasite infecting cattle in Abashiri District, Hokkaido. The MPSP gene sequence found in the additional type was closely related to MPSP genes of Theileria parasites found in Southeast Asian countries, including Thailand (Narathiwat) and Indonesia (Java). Theileria parasites from the blood sample were also distinguishable from the Ikeda or Chitose type parasites by the small subunit (SSU) rRNA gene sequence analysis, and they are grouped into the SSU rRNA types C/D found in Korea, North America, and Spain. The present finding of mixed infections of cattle with three different types of Theileria makes epidemiological feature of bovine theileriosis in Japan more complex. We have designed a set of primers specific to this MPSP type in order to conduct further epidemiological study.     

Use of monoclonal antibodies for analyses of Coxiella burnetii major antigens

Monoclonal antibodies (MAbs) to major antigens of Coxiella burnetii were produced. Some of the MAbs to a 62-kDa protein antigen, peptidoglycan protein complex and lipopolysaccharide (LPS) O-chains reacted with other bacteria whereas none of the MAbs to outer membrane proteins and LPS outer-core did. The LPS outer-core and OMPs may be useful antigens for specifically detecting antibodies to C. burnetii.