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).     

Analysis of genetic diversity in Bolivian llama populations using microsatellites

South American camelids (SACs) have a major role in the maintenance and potential future of rural Andean human populations. More than 60% of the 3.7 million llamas living worldwide are found in Bolivia. Due to the lack of studies focusing on genetic diversity in Bolivian llamas, this analysis investigates both the genetic diversity and structure of 12 regional groups of llamas that span the greater part of the range of distribution for this species in Bolivia. The analysis of 42 microsatellite markers in the considered regional groups showed that, in general, there were high levels of polymorphism (a total of 506 detected alleles; average PIC across per marker: 0.66), which are comparable with those reported for other populations of domestic SACs. The estimated diversity parameters indicated that there was high intrapopulational genetic variation (average number of alleles and average expected heterozygosity per marker: 12.04 and 0.68, respectively) and weak genetic differentiation among populations (F_ST range: 0.003–0.052). In agreement with these estimates, Bolivian llamas showed a weak genetic structure and an intense gene flow between all the studied regional groups, which is due to the exchange of reproductive males between the different flocks. Interestingly, the groups for which the largest pairwise F_ST estimates were observed, Sud Lípez and Nor Lípez, showed a certain level of genetic differentiation that is probably due to the pattern of geographic isolation and limited communication infrastructures of these southern localities. Overall, the population parameters reported here may serve as a reference when establishing conservation policies that address Bolivian llama populations.     

Oestrogen,Progesterone and Oxytocin Receptors and COX‐2 Expression in Endometrial Biopsy Samples from the Induction of Ovulation to Luteolysis in Llamas (Lama glama)

Endometrial expression of oestrogen (ERα), progesterone (PR) and oxytocin receptor (OR) and cyclooxygenase‐2 (COX‐2) was evaluated from the induction of ovulation to luteolysis in llamas. Ovarian activity was daily assessed by ultrasonography in five females. Ovulation was induced immediately after the detection of an ovulatory follicle by a GnRH injection (Day 0). Endometrial samples were obtained by transcervical biopsies from the left and right horns on day 0 and days 4, 8, 10 and 12 post‐GnRH. Blood samples were collected daily for progesterone and estradiol‐17β determinations by RIA. An immunohistochemical technique was used to study receptors population and COX‐2 expression which were then evaluated by two independent observers. The expression of ERα and PR was highest on day 0 in the luminal epithelium and stroma in association with high plasma estradiol‐17β concentrations. Thereafter, a decrease in ERα population was registered on day 4 and a new increase of its expression was observed between days 8 and 12 in those cell types. Conversely, PR population was gradually down‐regulated until its lowest expression was reached on day 10 post‐GnRH in the luminal epithelium. Content of OR was similar throughout the study in all cell types. The expression of COX‐2 was highest from day 8 to 12 post‐GnRH in the luminal epithelium, in relation to the time of maximal PGF_2α release. Both steroid receptors populations and COX‐2 expression were similar between horns. Meanwhile, OR expression was higher in the right than in the left uterine horn. In summary, this study showed that the loss of endometrium sensitivity to progesterone by days 8–10 post‐induction of ovulation and the concomitant increase of COX‐2 expression could play a key role in the mechanism of luteolysis and somehow be related to the short corpus luteum lifespan of llamas.     

DOES MEASUREMENT OF SMALL INTESTINAL DIAMETER INCREASE DIAGNOSTIC ACCURACY OF RADIOGRAPHY IN DOGS WITH SUSPECTED INTESTINAL OBSTRUCTION?

The ratio between maximal small intestinal (SI) diameter and the height of the body of the fifth lumbar vertebra (L5) in radiographs has been reported as a diagnostic test in dogs with suspected intestinal obstruction. In order to assess the effect of the SI/L5 ratio on the accuracy of radiographic diagnosis of intestinal obstruction, lateral abdominal radiographs of 37 dogs with small intestinal obstruction and 48 nonobstructed dogs were mixed and examined independently by six observers who were unaware of the final diagnosis and who represented a range of experience. Observers first examined radiographs subjectively and stated the likelihood of obstruction (definitely not, probably not, equivocal, probably, definitely). Observers subsequently reexamined the radiographs, determined the SI/L5 ratio, and again stated the likelihood of obstruction. The most frequent cause of obstruction was foreign body (29/37, 78%). Dogs with SI obstruction had a significantly larger median SI/L5 ratio than nonobstructed dogs (P = 0.0002). Using an SI/L5 ratio of 1.7 for diagnosis of intestinal obstruction, sensitivity and specificity were 66%. Use of the SI/L5 ratio was not associated with increased accuracy of diagnosis for any observer, regardless of experience, hence this test may have no diagnostic impact.     

Pharmacokinetics of ciprofloxacin in sheep after single intravenous or intramuscular administration

Summary

The disposition and urinary excretion of ciprofloxacin (CIP) following intravenous (IV) or intramuscular (IM) administration of 7.5 mg/kg body weight in sheep (n = 5) was studied. The intravenous plasma concentration curve was best described pharmacokinetically by a two‐compartment open model, while the intramuscular administration data fitted better to a one‐compartment open model. Mean elimination half‐lives after IV and IM administration were 72 and 184 minutes, respectively. The absorption of intramuscularly administered CIP in sheep was fast: maximal plasma concentration (Cmax) was reached quickly (tmax 31.93 min) and attained values of 0.69 ± 0.27 mg/l. The bioavailability was 49%. The urinary data showed a significant decrease in the elimination rate constant of CIP when CIP was administered intramuscularly. The other parameters calculated did not display differences between the two routes of administration. The results obtained suggest that when CIP was administered by the IM route in the assayed dose, it was able to maintain serum concentrations above the MIC of most common pathogens over an 8‐hour period.     

Monoamine oxidase activity in some brain areas of sheep after hormonal stimulation

The influence of hormonal superovulatory preparations Folistiman (450 IU FSH, Spofa, Prague) and serum gonadotropin (1500 IU PMSG, Spofa, Prague) on monoamine oxidase (MAO), the degradative enzyme of catecholamines, was investigated in some areas of the brain regulating reproductive functions (area preoptica of the hypothalamus, pituitary gland, and pineal gland) in ewes with synchronized oestrus (20 mg chlorsuperlutin) during the oestrous period using a radiochemical method. After intramuscular administration of 1500 IU PMSG, marked increase of MAO activity was found in the area preoptica (p < 0.05) and in the pituitary gland (p < 0.01) in comparison with the control group. No change occurred in MAO activity after ovarian stimulation with FSH. Administration of the above superovulatory preparations failed to induce MAO activity in the pineal gland of sheep.     

Propofol or thiopentone as induction agents in romifidine-sedated and halothane-N2O-anesthetized dogs: a preliminary study.

The objective of this paper was to evaluate the use of romifidine as a premedicant in dogs before general anesthesia induced with propofol or thiopentone and maintained with halothane-N2O. Fifteen healthy dogs were anesthetized twice. Each dog received, as preanesthetic protocol, atropine (10 microg/kg, IM) and romifidine (40 microg/kg, IM); induction was delivered with propofol or thiopentone and anesthesia was maintained with halothane and N2O for 1 h. Some cardiovascular and respiratory variables and recovery times were recorded. Induction doses of propofol or thiopentone and the percentage of halothane necessary for maintaining anesthesia were also registered. Thiopentone as an induction agent is more respiratory depressive but is less hypotensive than propofol. Thiopentone reduces further the percentage of halothane necessary for maintaining the anesthesia. However, the quality of recovery is poorer, as the time to extubation is longer and the dogs occasionally had a violent recovery. The combination of romifidine, atropine, propofol, halothane, and N2O appears to be an effective combination for inducing and maintaining general anesthesia in healthy dogs.     

Antibacterial susceptibility patterns of Pseudomonas strains isolated from chronic canine otitis externa

Development of antibiotic resistance in bacteria is a problem of great concern. It is important to establish the convenience of antimicrobial susceptibility tests in animal infections. The aim of this study was to test the susceptibility to antibiotics of Pseudomonas strains isolated from chronic canine otitis externa. We tested 23 strains of Pseudomonas: 19 Ps. aeruginosa, three Ps. fluorescens and one Pseudomonas spp. The most effective antibiotics were tobramycin (100% susceptible), marbofloxacin (91.3%) and ceftazidime (91.3%). Ticarcillin and gentamicin, commonly used for the treatment of otitis externa also showed good results (susceptibility of strains was 86 and 65.2% respectively). Lower susceptibility was found using enrofloxacin (52.1%) probably due to its indiscriminate use. We emphasize the need for a rational policy of antibiotic prescribing in order to prevent the selection of resistant strains.     

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.     

Use of the scrotum as a transposition flap for closure of surgical wounds in three dogs

OBJECTIVE: To describe use of the scrotum as a pedicle flap to cover defects created by tumor excision in the perineum or caudal and medial aspect of the thigh. STUDY DESIGN: Clinical study. ANIMALS: Three dogs. METHODS: After tumor excision and prescrotal castration, an incision was made around the base of the scrotum, leaving a pedicle on the side opposite the skin defect. After removal of the tunica dartos and abdominal fascia, the scrotal skin was stretched and shaped to the defect, then sutured in position with a simple interrupted pattern. Bandages that were changed every 48 hours compressed the flaps, and healing was observed until sutures were removed. RESULTS: Two dogs had first intention healing in 15 days, whereas 1 dog developed necrosis of 10% of the flap, and this area healed by second intention. CONCLUSION: In male dogs, the scrotal skin can be used as a pedicle flap for reconstructive surgery of wounds in the perineum and the proximomedial and caudal aspect of the thigh. CLINICAL RELEVANCE: The perineal region remains a surgical challenge because of the lack of the available skin for reconstruction of surgical wounds. The scrotal skin should be considered for use as a transposition flap to cover skin defects in this region.