Isolation and characterization of immortalized porcine aortic endothelial cell lines

Primary porcine endothelial cells have a limited life span in culture. After four to five passages, they tend to de-differentiate and eventually reach senescence. The aim of this work was to establish immortalized porcine aortic endothelial cell lines (AOCs) to facilitate in vitro studies of different pathological process involving the endothelium. Primary porcine aortic endothelial cells (PAECs) were transfected with a plasmid containing the SV40 genome and selected on the basis of morphological and phenotypical features. Flow cytometry analysis demonstrated uptake of acetylated low density lipoproteins (Ac-LDL) and constitutive expression of SLA class I, CD29, CD31, CD41/61, CD80/86, CD46, SWC3, and LAMP-1 antigens by all analyzed lines and showed little differences to primary cells. The functional similarity between primary and immortalized endothelial cells was demonstrated in a cytotoxicity assay using a human natural killer cell line (NKL) as effector. The AOCs cell lines should be valuable tools for in vitro study of the human immune response against pig endothelial cells. In addition, they would be very useful to gain insight in the pathogenesis of some viral haemorrhagic diseases of pig such as African swine fever (ASF) or classical swine fever (CSF).     

Nasopharyngeal myxosarcoma in a Bengal tiger (Panthera tigris).

An 18- to 25-yr-old intact female Bengal tiger (Panthera tigris) was diagnosed antemortem and postmortem with nasopharyngeal myxosarcoma metastatic to the lung, chronic lymphoplasmacytic otitis media, and lymphoplasmacytic and eosinophilic stomatitis. Myxosarcomas are rare in domestic animals and seldom metastasize; this tumor has not been previously reported in an exotic felid. Computed tomography of the skull was used during the diagnoses.     

Facet joint geometry and intervertebral disk degeneration in the L5-S1 region of the vertebral column in German Shepherd dogs

OBJECTIVE: To evaluate the possible association between facet joint geometry and intervertebral disk degeneration in German Shepherd Dogs. ANIMALS: 25 German Shepherd Dogs and 11 control dogs of similar body weight and condition. PROCEDURE: Facet joint angles in the caudal portion of the lumbar region of the vertebral column (L5-S1) were measured by use of computed tomography, and the intervertebral discs were evaluated microscopically. The relationship between facet joint geometry and disk degeneration was evaluated by use of statistical methods. RESULTS: German Shepherd Dogs had significantly more facet joint tropism than control dogs, but an association with disk degeneration was not found. However, German Shepherd Dogs had a different facet joint conformation, with more sagittally oriented facet joints at L5-L6 and L6-L7 and a larger angle difference between the lumbar and lumbosacral facet joints, compared with control dogs. CONCLUSIONS AND CLINICAL RELEVANCE: A large difference between facet joint angles at L6-L7 and L7-S1 in German Shepherd Dogs may be associated with the frequent occurrence of lumbosacral disk degeneration in this breed.     

Anaplasma infection in free-ranging Iberian red deer in the region of Castilla-La Mancha, Spain

Organisms in the genus Anaplasma are obligate intracellular pathogens that multiply in both vertebrate and invertebrate hosts. The type species, Anaplasma marginale, causes bovine anaplasmosis and infects erythrocytes of the vertebrate host and undergoes a complex developmental cycle in ticks which serve as biological vectors. Infected cattle, wild ruminants and ticks can all serve as reservoirs of A. marginale. In this study, hunter killed Iberian red deer (Cervus elaphus hispanicus) from the region of Castilla-La Mancha in southwestern Spain were tested for Anaplasma infection. We found that 10% of the deer examined were seropositive for Anaplasma. Three A. marginale strains were subsequently obtained from salivary glands of Hyalomma marginatum that were removed from these deer, and the sequence of the major surface protein (msp)4 gene was determined for each strain and used for phylogenetic studies. Maximum parsimony analyses of msp4 sequences from H. marginatum ticks in comparison with New World cattle and bison isolates reported previously, suggested different origins for these Spanish A. marginale strains. The results of this study demonstrated that Iberian red deer are naturally infected with Anaplasma, and may therefore serve as a wildlife reservoir of the pathogen. Although the link between deer infection and the strains of A. marginale identified in ticks was not established, H. marginatum and Rhipicephalus bursa were identified as potential biological vectors for A. marginale in this region and may effect transmission of A. marginale between deer and cattle populations.     

Pasteurella multocida contains multiple immunogenic haemin- and haemoglobin-binding proteins

Iron-dependent outer membrane proteins (IROMPs) play an important role in bacterial pathogenesis and present several attributes of potential vaccine candidates. TBLASTN analysis of the Pasteurella multocida Pm70 genome using the same molecules of other bacterial pathogens as a query identified eight putative haemin and haemoglobin receptors for this organism. Quantitative binding assays have demonstrated that the proteins PM0040, PM0236, PM0741, PM1081, PM1428, PM0592 and HgbA bind both haemin and haemoglobin, whereas PM0576 and PM1282 ORFs only bind either haemoglobin or haemin, respectively. Furthermore, Western blot analysis showed that P. multocida-infected mice generate specific antibodies against PM0040, PM0236, PM0741, PM1081, PM1428, PM0592 and HgbA proteins. Nevertheless, inoculation of mice with any single one of these receptors alone did not protect against P. multocida infection.     

Rough vaccines in animal brucellosis: structural and genetic basis and present status

Brucellosis control and eradication requires serological tests and vaccines. Effective classical vaccines (S19 in cattle and Rev 1 in small ruminants), however, induce antibodies to the O-polysaccharide of the lipopolysaccharide which may be difficult to distinguish from those resulting from infection and may thus complicate diagnosis. Rough attenuated mutants lack the O-polysaccharide and would solve this problem if eliciting protective immunity; the empirically obtained rough mutants 45/20 and RB51 have been used as vaccines. Strain 45/20 is reportedly unstable and it is not presently used. RB51 is increasingly used instead of S19 in some countries but it is rifampicin resistant and its effectiveness is controversial. Some controlled experiments have found good or absolute protection in adult cattle vaccinated orally (full dose) or subcutaneously (reduced dose) and in one field experiment, RB51 was reported to afford absolute protection to calves and to perform better than S19. Controlled experiments in calves, however, have shown reduced doses of RB51 to be ineffective, full doses only partially effective, and RB51 less effective than S19 against severe challenges. Moreover, other observations suggest that RB51 is ineffective when prevalence is high. RB51 is not useful in sheep and evidence in goats is preliminary and contradictory. Rough mutants obtained by molecular biology methods on the knowledge of the genetics and structure of Brucella lipopolysaccharide may offer alternatives. The B. abortus manBcore (rfbK) mutant seems promising in cattle, and analyses in mice suggest that mutations affecting only the O-polysaccharide result in better vaccines than those affecting both core and O-polysaccharide. Possible uses of rough vaccines also include boosting immunity by revaccination but solid evidence on its effectiveness, safety and practicality is not available.     

The effects of a single injection of dexamethasone-21-isonicotinate on the lymphocyte functions of dairy cows at two weeks post partum

Dexamethasone is a potent therapeutic for treatment of the fatty liver syndrome or primary ketosis in post partum dairy cows. Reservations exist, however, among practitioners with respect to the risk of immunosuppression induced by corticosteroids. The aim of this study was to investigate the effect of a single injection of dexamethasone-21-isonicotinate on distinct immune functions of postpartum dairy cows because only scarce information is available on the effects of corticosteroid preparations when administered at a dosage and frequency for treatment of the fatty liver syndrome or primary ketosis. Sixteen Swedish red-pied dairy cows, between days 9 and 15 post partum, were allotted to either a control group (n = 8) or a treatment group (n = 8). The cows in the treatment group received a single intramuscular injection of a dexamethasone-21-isonicotinate suspension at a dosage of 0.02 mg/kg i.m. at the start of the experiment. White blood cell counts and selected lymphocyte functions (lymphocyte proliferation, expression of lymphocyte markers and the b2 and a4 chain of adhesion molecules belonging to the integrin family) and some parameters of the energy metabolism (glucose, insulin) were determined before the administration of corticosteroids (day 0) and subsequently at days 2, 4, 7 and 9 of the experiment. Changes in glucose and insulin were within the target range for treatment of the fatty liver syndrome or primary ketosis. Significant (P < 0.05) increases in the number of circulating white blood cells were observed in treated cows on the second day following treatment which was exclusively caused by an increase in the number of circulating neutrophils. Lymphocyte blastogenesis in response to ConA and the percentages of lymphocytes positive for CD2, CD4, CD8, CD49d and CD18 as well as the intensity of CD49d expression did not differ between the treatment and control groups. There was, however, a significant reduction (P < 0.01) in the intensity of CD18 expression on lymphocytes in the treated animals on the fourth day after treatment. In conclusion, a single administration of dexamethasone-21-isonicotinate in a dosage of 0.02 mg/kg i.m. at two weeks post partum in healthy cows had a significant but highly transient effect on CD18 expression on lymphocytes and the number of peripheral blood neutrophils, but did not affect lymphocyte blastogenesis or lymphocyte subpopulation patterns in peripheral blood.