Effect of dietary Schizochytrium microalga oil and fish oil on plasma cholesterol level in rats

The purpose of the study was to test the hypothesis that the dietary oils with different content of n‐3 polyunsaturated fatty acids (PUFA) eicosapentaenoic acid (EPA) + docosahexaenoic acid (DHA) affect plasma lipid level in rats in a different degree. The diets with 6% of fish oil (FO) and Schizochytrium microalga oil (SchO; EPA+DHA content in the diets 9.5 + 12.3 and 2.6 + 29.5% of the sum of total fatty acids, respectively) were used; the diet with 6% of safflower oil (high content of n‐6 PUFA linoleic acid, 65.5%; EPA+DHA content 0.7 + 0.9%) was used as a control. The difference between FO and SchO was established only in the case of plasma triacylglycerol (TAG) level: plasma TAG of the FO‐fed rats did not differ from the control rats (p > 0.05), while SchO decreased (p < 0.05) plasma TAG to 46% of the control. On the other hand, FO and SchO decreased (p < 0.05) total plasma cholesterol (TC) in rats in the same extent, to 73% of the control. Regarding the underlying mechanisms for the TC decrease, both SchO and FO up‐regulated hepatic Insig‐1 gene (181 and 133% of the control; p < 0.05), which tended (p = 0.15 and p = 0.19 respectively) to decrease the amount of hepatic nSREBP‐2 protein (61 and 66% of the control). However, neither SchO nor FO influenced hepatic 3‐hydroxy‐3‐methyl‐glutaryl‐CoA reductase gene expression (p > 0.05); SchO (but not FO) increased (p < 0.05) low‐density lipoprotein receptor mRNA in the liver. It was concluded that the decrease of total plasma cholesterol might be caused by an increased cholesterol uptake from plasma into the cells (in the case of SchO), but also by other (in the present study not tested) mechanisms.     

In vivo induction of interferon gamma expression in grey horses with metastatic melanoma resulting from direct injection of plasmid DNA coding for equine interleukin 12

Whole blood pharmacokinetics of intratumourally injected naked plasmid DNA coding for equine Interleukin 12 (IL-12) was assessed as a means of in vivo gene transfer in the treatment of melanoma in grey horses. The expression of induced interferon gamma (IFN-g) was evaluated in order to determine the pharmacodynamic properties of in vivo gene transduction. Seven grey horses bearing melanoma were injected intratumourally with 250 μg naked plasmid DNA coding for IL-12. Peripheral blood and biopsies from the injection site were taken at 13 time points until day 14 post injection (p.i.). Samples were analysed using quantitative real-time PCR. Plasmid DNA was quantified in blood samples and mRNA expression for IFN-g in tissue samples. Plasmid DNA showed fast elimination kinetics with more than 99 % of the plasmid disappearing within 36 hours. IFN-g expression increased quickly after IL-12 plasmid injection, but varied between individual horses. Intratumoural injection of plasmid DNA is a feasible method for inducing transgene expression in vivo. Biological activity of the transgene IL-12 was confirmed by measuring expression of IFN-g.     

Laparoscopic cholecystectomy under field conditions in Asiatic black bears (Ursus thibetanus) rescued from illegal bile farming in Vietnam

Nine adult Asiatic black bears (Ursus thibetanus) previously rescued from illegal bile farming in Vietnam were examined via abdominal ultrasound and exploratory laparoscopy for liver and gall bladder pathology. Three bears demonstrated notable gall bladder pathology, and minimally invasive cholecystectomies were performed using an open laparoscopic access approach, standard 10 to 12 mmHg carbon dioxide pneumoperitoneum and a four-port technique. A single bear required insertion of an additional 5 mm port and use of a flexible liver retractor due to the presence of extensive adhesions between the gall bladder and quadrate and left and right medial liver lobes. The cystic duct was dissected free and this and the cystic artery were ligated by means of extracorporeal tied Meltzer knot sutures. The gall bladder was dissected free of the liver by blunt and sharp dissection, aided by 3.8 MHz monopolar radiosurgery. Bears that have had open abdominal cholecystectomies are reported as taking four to six weeks before a return to normal activity postoperatively. In contrast, these bears demonstrated rapid unremarkable healing, and were allowed unrestricted access to outside enclosures to climb trees, swim and interact normally with other bears within seven days of surgery.     

Influence of head positioning on the assessment of Chiari-like malformation in Cavalier King Charles spaniels

Chiari-like malformation (CM) is almost omnipresent in the Cavalier King Charles spaniels (CKCS), often leading to syringomyelia (SM). Morphometric studies have produced variable results concerning relationship between the brain parenchyma within the caudal cranial fossa (CCF) and SM. The present study assesses the effect of head position, one potential confounder. Magnetic resonance images of CKCS with CM were reviewed in extended and flexed head positions. Volumes were calculated from transverse T2-weighted brain images. Mid-sagittal images were used for measurement of cerebellar herniation and CSF space between cerebellum and brainstem. Fourteen CKCS were included into the study, seven dogs with CM and seven with CM/SM. There was no difference between the relative brain parenchyma within the CCF in extended position and flexed position, or the brain parenchyma within the rostral and middle cranial fossae proportion. Cerebellar herniation and CSF space between cerebellum and brainstem were significantly increased in the flexed position. Cerebellar herniation and CSF space differed significantly between CM and CM/SM in a flexed head position. Volumetric measurements did not vary with head position. Cerebellar herniation and CSF space between the cerebellum and the brainstem were larger in a flexed head position.     

Detection of Mycobacterium avium ssp. paratuberculosis in ileocaecal lymph nodes collected from elderly slaughter cows using a semi-nested IS900 polymerase chain reaction

The aim of this study was to investigate the occurrence of subclinical Mycobacterium avium spp. paratuberculosis (MAP) infections at slaughter by testing ileocaecal lymph nodes with a semi-nested IS900 PCR. Tissue samples were available within the framework of a parallel study investigating BSE-susceptibility factors in members of BSE-cohorts in the German Federal State of Lower Saxony. Ileocaecal lymph nodes were collected over a 2-year sampling period from 99 slaughter cattle of a mean age of 6.5 years (5.5-7.5 years). A recently developed IS900 semi-nested polymerase chain reaction (snPCR) assay offering a sensitivity of 1 genome equivalent was used for the detection of MAP-DNA. Based on this snPCR, 17 out of the 99 samples gave positive results, indicating a MAP occurrence of 17.17% in the random sample. All PCR products were sequenced for screening of polymorphisms. Nucleotide homologies of 98.5-100% were found with respect to the MAP K10 reference sequence IS900 (GenBank: AE16958). PCR analysis of ileocaecal lymph nodes collected from slaughter cattle proved to be a suitable technique to determine MAP occurrence in the local cattle population.     

The innate antiviral immune system of the cat: molecular tools for the measurement of its state of activation

The innate immune system plays a central role in host defence against viruses. While many studies portray mechanisms in early antiviral immune responses of humans and mice, much remains to be discovered about these mechanisms in the cat. With the objective of shedding light on early host-virus interactions in felids, we have developed 12 real-time TaqMan(?) qPCR systems for feline genes relevant to innate responses to viral infection, including those encoding for various IFNα and IFNω subtypes, IFNβ, intracellular antiviral factor Mx, NK cell stimulator IL-15 and effectors perforin and granzyme B, as well as Toll-like receptors (TLRs) 3 and 8. Using these newly developed assays and others previously described, we measured the relative expression of selected markers at early time points after viral infection in vitro and in vivo. Feline embryonic fibroblasts (FEA) inoculated with feline leukemia virus (FeLV) indicated peak levels of IFNα, IFNβ and Mx expression already 6h after infection. In contrast, Crandell-Rees feline kidney (CrFK) cells inoculated with feline herpes virus (FHV) responded to infection with high levels of IFNα and IFNβ only after 24h, and no induction of Mx could be detected. In feline PBMCs challenged in vitro with feline immunodeficiency virus (FIV), maximal expression levels of IFNα, β and ω subtype genes as well as IL-15 and TLRs 3, 7 and 8 were measured between 12 and 24h after infection, whereas expression levels of proinflammatory cytokine gene IL-6 were consistently downregulated until 48h post inoculation. A marginal upregulation of granzyme B was also observed within 3h after infection. In an in vivo experiment, cats challenged with FIV exhibited a 2.4-fold increase in IFNα expression in blood 1 week post infection. We furthermore demonstrate the possibility of stimulating feline immune cells in vitro with various immune response modifiers (IRMs) already known for their immunostimulatory properties in mice and humans, namely Poly IC, Resiquimod (R-848) and dSLIM?, a synthetic oligonucleotide containing several unmethylated CpG motifs. Stimulation of feline PBMCs with dSLIM? and R-848 effectively enhanced expression of IFNα within 12h by factors of 6 and 12, respectively, and Poly IC induced an increase in Mx mRNA expression of 28-fold. Altogether, we describe new molecular tools and their successful use for the characterization of innate immune responses against viruses in the cat and provide evidence that feline cells can be stimulated by synthetic molecules to enhance their antiviral defence mechanisms.     

Evaluation of miltefosine for the treatment of dogs naturally infected with L. infantum (=L. chagasi) in Brazil

Dogs naturally infected with Leishmania Infantum (=L. chagasi) were treated with miltefosine using different therapeutic regimens. The animals were evaluated for clinical evolution, biochemical parameters, parasite load (by real-time PCR), cytokine levels and humoral response. After treatment and during the following 24 months, there was progressive clinical improvement and complete recovery in 50% (7/14) of the treated animals. There was a decrease in the smear positivity of the bone marrow after treatment, and there was also a gradual and constant decrease in positive cultures at the end of the follow-up period. However, the PCR detection of parasite DNA remained positive. In general, all animals presented a significant increase in parasite load 6 months after treatment. The IFN-γ levels in all the groups tended to increase during follow-up period, regardless of the miltefosine dose administered. The IL-4 and IL-10 levels of the animals tended to decrease during follow-up, except after 300 days when only IL-10 increased. The serum antibodies identified antigens that ranged from 116 kDa to less than 29 kDa in the Western blot assay. Furthermore, 300 days after treatment, qualitative and quantitative differences in the antigen profiles were observed. Antigens of 97 and 46 kDa were the most intensely recognized. Higher levels of antigen-specific Leishmania IgG were detected before and 300 days after treatment in all groups. Taking together, the improvement in the clinical symptoms was not followed by parasitological clearance, suggesting that treatment with miltefosine is not recommended, especially in endemic areas like Brazil, where children are the major victims and dogs are involved in the maintenance of the parasite cycle.     

Evaluation of initial plasma lactate values as a predictor of gastric necrosis and initial and subsequent plasma lactate values as a predictor of survival in dogs with gastric dilatation‐volvulus: 84 dogs (2003–2007)

Objective – To test whether an initial plasma lactate≥6.0 mmol/L is associated with the presence of macroscopic gastric wall necrosis and overall survival in dogs presenting with gastric dilatation‐volvulus (GDV). Additionally, if no association was identified we sought to identify a different predictive initial plasma lactate concentration and to examine whether serial plasma lactate concentrations provide better prediction of survival. Design – Retrospective study over a 5‐year period (2003–2007). Setting – Urban private referral small animal teaching hospital. Animals – Eighty‐four client‐owned dogs with a diagnosis of GDV and plasma lactate measurements. Interventions – None. Measurements and Main Results – There was no statistically significant relationship found between survival and the presence of macroscopic gastric wall necrosis with the initial plasma lactate≥6 mmol/L. There was a significant relationship between the initial plasma lactate >2.9 mmol/L for predicting necrosis and <4.1 mmol/L for predicting survival to discharge. Forty dogs that had an increased initial plasma lactate (>2.5 mmol/L) also had a subsequent plasma lactate measured within 12 hours of presentation, with 37/40 dogs surviving and 70% of these surviving dogs having the subsequent lactate decrease by≥50% within 12 hours. The 3/40 that died failed to decrease their plasma lactate by≥50% from the initial blood lactate. Conclusion – The results of this study indicate that an initial presenting plasma lactate concentration≥6.0 mmol/L is not predictive of macroscopic gastric wall necrosis or survival in dogs presenting with GDV. A decrease in plasma lactate concentrations≥50% within 12 hours may be a good indicator for survival. Limitations to the study include its retrospective nature, the small number of patients, and the number of dogs that were euthanized rather than allowed to progress to a natural outcome.