New ruthenium(II)/phosphines/diimines complexes: Promising antitumor (human breast cancer) and Mycobacterium tuberculosis fighting agents

The synthesis and characterization of ruthenium compounds of the type [RuCl2(P)2(N-N)] [(P)2 = (PPh3) 2, dppb = 1,4-bis(diphenylphosphino)butano; dppp = 1,3-bis(diphenylphosphino)propane; N-N = 5,5′-dimethyl-2,2′dipyridyl (5,5′-mebipy) or 4,4′-dimethyl-2,2′dipyridyl (4,4′-mebipy)] are described. The complexes were characterized using elemental analysis, UV-Vis and infrared spectroscopies, cyclic voltammetry, and X-ray crystallography. In vitro evaluation of the complexes, using the MTT methodology, revealed their cytotoxic activities in a range of 5.4-15.7 μM against the MDA-MB-231 breast tumor cells and showed that, in this case, they are more active than the reference metallodrug cisplatin. The in vitro antimycobacterial activities of the complexes had their Minimum Inhibitory Concentration (MIC) for MTB cell growth measured, by the REMA method. The MICs for these complexes were found to be between 12.5 and 25.0 μg/mL. The results are comparable with the second line drug cycloserine (MIC = 12.5-50.0 μg/mL), commonly used in the treatment of TB. © 2013 Elsevier Ltd. All rights reserved.

Increase nitric oxide and oxidative stress in dogs experimentally infected by Ehrlichia canis: Effect on the pathogenesis of the disease

The aim of this study was to evaluate nitric oxide levels, lipid peroxidation, protein oxidation and glutathione reductase activity in serum of dogs experimentally infected by Ehrlichia canis. Banked serum samples of dogs divided into two groups were used: negative control (n=5) and infected by E. canis (n=5). The concentration of nitrite/nitrate (NOx), lipid peroxidation (TBARS), advanced oxidation protein products (AOPP), and glutathione reductase (GR) activity in sera were evaluated. Samples were collected on days 0, 3, 6, 18 and 30 post-infection (PI). NOx and TBARS levels were significantly (P<0.05) higher in the infected group at 18 and 30 days PI, as well as AOPP levels at 30 days PI when compared to samples from control group. The GR activity was significant (P<0.05) increased in serum of dogs infected by E. canis on days 18 and 30 PI. Based on the increased levels of NOx, TBARS, AOPP and GR activity we concluded that dogs experimentally infected by E. canis develop a state of redox imbalance and that these changes might be involved in the pathophysiology of the disease. © 2013 Elsevier B.V.

Oxidative stress, superoxide production, and apoptosis of neutrophils in dogs with chronic kidney disease

Oxidative stress is a key component in the immunosuppression of chronic kidney disease (CKD), and neutrophil function may be impaired by oxidative stress. To test the hypothesis that in uremic dogs with CKD, oxidative stress is increased and neutrophils become less viable and functional, 18 adult dogs with CKD were compared with 15 healthy adult dogs. Blood count and urinalysis were done, and the serum biochemical profile and plasma lipid peroxidation (measurement of thiobarbituric acid reactive substances) were determined with the use of commercial reagents. Plasma total antioxidant capacity (TAC) was measured with a spectrophotometer and commercial reagents, superoxide production with a hydroethidine probe, and the viability and apoptosis of neutrophils with capillary flow cytometry and the annexin V-PE system. The plasma concentrations of cholesterol (P = 0.0415), creatinine (P < 0.0001), and urea (P < 0.0001) were significantly greater in the uremic dogs than in the control dogs. The hematocrit (P = 0.0004), urine specific gravity (P = 0.015), and plasma lipid peroxidation (P < 0.0001) were significantly lower in the dogs that were in late stages of CKD than in the control group. Compared with those isolated from the control group, neutrophils isolated from the CKD group showed a higher rate of spontaneous (0.10 ± 0.05 versus 0.49 ± 0.09; P = 0.0033; median ± standard error of mean) and camptothecin-induced (18.53 ± 4.06 versus 44.67 ± 4.85; P = 0.0066) apoptosis and lower levels of superoxide production in the presence (1278.8 ± 372.8 versus 75.65 ± 86.6; P = 0.0022) and absence (135.29 ± 51.74 versus 41.29 ± 8.38; P = 0.0138) of phorbol-12-myristate-13-acetate stimulation. Thus, oxidative stress and acceleration of apoptosis occurs in dogs with CKD, the apoptosis diminishing the number of viable neutrophils and neutrophil superoxide production.

Deproteinized bovine bone mineral particles and osseointegration of implants without primary bone contact: An experimental study in dogs

Objectives: To evaluate the influence on osseointegration of Deproteinized bovine bone mineral (DBBM) particles used to fill defects of at least 1 mm around implants having no primary contact with bone. Material and methods: Premolars and first molars were extracted bilaterally from the mandible of six Labrador dogs. After 3 months of healing, mucoperiosteal full-thickness flaps were elevated, and one recipient site was prepared in the molar region of each hemi-mandible to place implants. These were installed with a deliberate circumferential and periapical space to the bone walls of 1.2 mm. All implants were stabilized with passive fixation plates to maintain the implants in situ and without any contact with the implant bed. The control sites were left to be filled with coagulum, while at the test sites, the residual gap was filled with DBBM. After 3 months of submerged healing, the animals were sacrificed. Ground sections were prepared and analyzed histomorphometrically. Results: Mineralized bone-to-implant contact was 4.0% and 3.9% for control and test sites, respectively. The width of the residual defects was 0.48 mm and 0.88 mm at the control and test sites, respectively. The percentage of implant surface covered by a layer of dense connective tissue of 0.12 mm of width on average was 84.9% and 88.5% at the control and test sites, respectively. Conclusion: A minor and not predictable degree of contact or distance osteogenesis was obtained on the implant surface when primary contact of the implant surface with the implant bed had deliberately been avoided. DBBM grafting of the artificial gap did not favor osseointegration. Neither did it enhance the ability to bridge the gap with newly formed bone in an artificial defect wider than 1 mm. © 2013 John Wiley & Sons A/S.

Enzyme effects on extruded diets for dogs with soybean meal as a substitute for poultry by-product meal

The effects of exogenous enzymes supplementation on kibble diets for dogs formulated with soybean meal (SBM) as a substitute for poultry by-product meal (PM) was investigated on nutrient digestibility, fermentation products formation, post-prandial urea response and selected faecal bacteria counts. Two kibble diets with similar compositions were used in two trials: PM-based diet (28.9% of PM; soybean hulls as a fibre source) and SBM-based diet (29.9% of SBM). In experiment 1, the SBM diet was divided into three diets: SBM-0, without enzyme addition; SBM-1, covered after extrusion with 7500U protease/kg and 45U cellulase/kg; and SBM-2, covered with 15000U protease/kg and 90U cellulase/kg. In experiment 2, the SBM diet was divided into three diets: SBM-0; SBM-1, covered with 140U protease/kg; 8U cellulase/kg, 800U pectinase/kg, 60U phytase/kg, 40U betaglucanase/kg and 20U xylanase/kg; and SMB-2, covered with 700U protease/kg, 40U cellulase/kg, 4000U pectinase/kg, 300U phytase/kg, 200U betaglucanase/kg and 100U xylanase/kg. Each experiment followed a block design with six dogs per diet. Data were submitted to analysis of variance and means compared by orthogonal and polynomial contrasts (p<0.05). In both experiments, nutrients and energy digestibility did not differ between diets (p>0.05). SBM consumption resulted in increased faecal moisture and production (p<0.05), without effect on faecal score. Higher concentration of propionate, acetate and lactate, and lower ammonia and pH were found in the faeces of dogs fed SBM (p<0.05). Higher post-prandial urea was verified in dogs fed SBM (p<0.05). In experiment 2, the addition of enzymes increased faecal concentration of propionate, acetate and total short-chain fatty acid (p<0.05) and tended to reduce post-prandial urea concentration (p=0.06). Although with similar digestibility, SBM shows a worse utilization of absorbed amino acids than the PM. Soybean oligosaccharides can beneficially change gut fermentation product formation. Enzymes can increase the gut fermentation activity and improve the SBM proteic value. © 2013 Blackwell Verlag GmbH.

Enzyme use in kibble diets formulated with wheat bran for dogs: Effects on processing and digestibility

Recently, there is an interest in technologies that favour the use of coproducts for animal nutrition. The effect of adding two enzyme mixtures in diets for dogs formulated with wheat bran (WB) was evaluated. Two foods with similar compositions were formulated: negative control (NC; without WB) and test diet (25% of WB). The test diet was divided into four treatments: without enzyme (positive control), enzyme mixture 1 (ENZ1; added before extrusion β-glucanase, xylanase, cellulase, glucoamylase, phytase); enzyme mixture 2 (ENZ2; added before extrusion the ENZ1 more α-amylase); enzyme mixture 2 added after the extrusion (ENZ2ex). ENZ1 and ENZ2 were used to evaluate the enzyme effect on extruder pre-conditioner (processing additive) and ENZ2ex to evaluate the effect of enzyme supplementation for the animal. Digestibility was measured through total collection of faeces and urine. The experiment followed a randomized block design with five treatments (diets) and six dogs per diet, totalling 30 dogs (7.0 ± 1.2 years old and 11.0 ± 2.2 kg of body weight). Data were submitted to analysis of variance and means compared by Tukey's test and orthogonal contrasts (p < 0.05). Reducing sugars showed an important reduction after extrusion, suggesting the formation of carbohydrate complexes. The apparent total tract digestibility (ATTD) of dry matter, organic matter, crude protein, acid-hydrolysed fat and energy was higher in NC than in diets with WB (p < 0.001), without effects of enzyme additions. WB diets resulted in higher faecal production and concentration of short-chain fatty acids (SCFA) and reduced pH and ammonia concentration (p < 0.01), with no effect of enzyme addition. The enzyme addition did not result in improved digestibility of a diet high in non-starch polysaccharides; however, only ATTD was measured and nutrient fermentation in the large intestine may have interfered with the results obtained. WB modified fermentation product formation in the colon of dogs. © 2013 Blackwell Verlag GmbH.

T and B lymphocytes in the brains of dogs with concomitant seropositivity to three pathogenic protozoans: Leishmania chagasi, Toxoplasma gondii and Neospora caninum

Background: Visceral leishmaniasis is a disease with great variability regarding the clinical manifestations in humans and dogs. Chronically infected dogs may develop neurological disorders, however, there are few reports that characterize the lesions and make clear the pathogenesis of the canine cerebral leishmaniasis. Concomitant with Leishmania chagasi, dogs may be infected by opportunistic pathogens, such as Toxoplasma gondii and Neospora caninum, which may contribute to the occurrence of lesions in the central nervous system. Hence, we aimed to compare the T and B lymphocytes population in the brains of infected dogs with seropositivity to L. chagasi, T. gondii and N. caninum concurrently (n = 24), seropositivity only to L. chagasi (n = 31), and seropositivity to T. gondii and N. caninum (n = 16). Uninfected dogs were used as control (n = 10). Results: Inflammatory lesions, characterised by mononuclear cell accumulation, composed mainly of CD3+ T lymphocytes predominated in several encephalic regions of the dogs from all the three infected groups, with no difference among them (P = 0.0004), whereas CD79α+ B lymphocytes were detected in very small intensity and presented no difference among groups (P = 0.5313). Furthermore, no association among diseases was detected at the serological enquire. Conclusions: We demonstrate that the peripheral infection by L. chagasi per se can promote the influx of lymphocytes within the nervous milieu as occurs during Toxoplasma and Neospora infections, and the concomitant seropositivity against these pathogens does not exacerbate the inflammatory brain lesions. Therefore, these findings give additional support that the brain should be included in the list of organs affected by visceral leishmaniasis and that even asymptomatic infected dogs may develop brain lesions. © 2013 Sakamoto et al.; licensee BioMed Central Ltd.

Effects of constant rate infusion of anesthetic or analgesic drugs on general anesthesia with isoflurane: A retrospective study in 200 dogs

Constant rate infusion (CRI) shows several advantages in balanced anesthesia, such as reduction of requirement for inhaled anesthetics and control of pain. The most commonly used drugs in these protocols are local anesthetics, dissociative, and opioids, which may be administered alone or in combinations. We evaluated the records of 200 dogs that underwent various surgical procedures with anesthetic or analgesic CRI in the perioperative period during 2011 and 2012 at the Veterinary Hospital of Franca University (Unifran), and identified possible complications during the transoperative period. Records evaluated included clinical state, laboratory tests, drugs used in premedication and induction, and CRI protocol. Acepromazine and morphine were the main drugs used in premedication. Propofol was used to induce anesthesia alone or in combination with other agents. We evaluated records of the 25 different CRI protocols. Fentanyl was the main drug employed in CRI, either alone or in combination. There were 128 episodes of anesthetic complications during CRI;the most common were hypotension, hypertension, and tachycardia, which occurred in 43 (32%), 35 (26.3%), and 19 (14.2%) dogs, respectively. Cardiac arrhythmia was reported in only 4 dogs. Signs of respiratory depression were present in dogs treated with 6 different CRI protocols. The consumption of isoflurane (vol %) reduced between 15.7% and 21.05% after 30minutes of the CRI in the fentanyl and fentanyl-lidocaine-ketamine CRI groups (p<0.05). In conclusion, CRI is a valid component of balanced anesthesia in dogs, safe, and has a low incidence of adverse effects. However, future studies are warranted to describe the results of the clinical use of CRI to better characterize and refine this technique.

Zymographic patterns of MMP-2 and MMP-9 in the CSF and cerebellum of dogs with subacute distemper leukoencephalitis

Distemper leukoencephalitis is a disease caused by the canine distemper virus (CDV) infection. It is a demyelinating disease affecting mainly the white matter of the cerebellum and areas adjacent to the fourth ventricle; the enzymes of the matrix metalloproteinases (MMPs) group, especially MMP-2 and MMP-9 have a key role in the myelin basic protein fragmentation and in demyelination, as well as in leukocyte traffic into the nervous milieu. To evaluate the involvement of MMPs during subacute distemper leukoencephalitis, we measured the levels of MMP-2 and MMP-9 by zymography in the cerebrospinal fluid (CSF) and in the cerebellum of 14 dogs naturally infected with CDV and 10 uninfected dogs. The infected dogs presented high levels of pro-MMP-2 in the CSF and elevated levels of pro-MMP-2 and pro-MMP-9 in the cerebellar tissue. Active MMP-2 was detected in the CSF of some infected dogs. As active MMP-2 and MMP-9 are required for cellular migration across the blood-brain barrier and any interference between MMPs and their inhibitors may result in an amplification of demyelination, this study gives additional support to the involvement of MMPs during subacute distemper leukoencephalitis and suggests that MMP-2 and MMP-9 may take part in the brain inflammatory changes of this disease. © 2013 Elsevier B.V.

Influence of presence or absence of keratinized mucosa on the alveolar bony crest level as it relates to different buccal marginal bone thicknesses. an experimental study in dogs

Objective: To investigate the influence of the presence or absence of keratinized mucosa on the alveolar bony crest level as it relates to different buccal marginal bone thicknesses. Material and methods: In six beagle dogs, the mandibular premolars and first molars were extracted bilaterally. In the right side of the mandible (test), flaps were elevated, and the buccal as well as part of the lingual masticatory mucosa was removed. The flap was released coronally to allow a primary wound closure. In the left side, the wounds were left unsutured with the keratinized mucosa remaining (control). After 3 months of healing, a complete absence of keratinized mucosa was found at the test sites. Two recipient sites were prepared at each side of the mandible, one in the premolar and one in the molar region. A buccal bony ridge width of approximately 1 and 2 mm was obtained at the premolar and molar region, respectively. Implants were installed with the shoulder flush with the buccal alveolar bony crest, and abutments were connected to allow a nonsubmerged healing. At least 2 mm of keratinized mucosa was surrounding the control sites, while at the test sites, the implants were bordered by alveolar mucosa. After 3 months, the animals were euthanized and ground sections obtained. Results: A higher vertical bony crest resorption was observed at the test compared with the control sites both at the premolar and molar regions, the differences being statistically significant. The top of the peri-implant mucosa was located more coronally at the control compared with the test sites. The horizontal resorption measured 1 mm below the implant shoulder was similar at the test and control sites. Only limited differences were found between premolar and molar sites, with the exclusion of the horizontal resorption that was higher at the test compared with the control sites. Conclusions: A higher alveolar buccal bony crest resorption and a more apical soft tissue marginal position should be expected, when implants are surrounded with thin alveolar mucosa at the time of installation, independently of the thickness of the buccal bony crest. © 2013 John Wiley & Sons A/S.

Improvement in clinical signs and cellular immunity of dogs with visceral leishmaniasis using the immunomodulator P-MAPA

This study investigated the immunotherapeutic potential of the protein aggregate magnesium-ammonium phospholinoleate-palmitoleate anhydride immuno-modulator (P-MAPA) on canine visceral leishmaniasis. Twenty mongrel dogs presenting clinical symptoms compatible with leishmaniasis and diagnosis confirmed by the detection of anti-leishmania antibodies were studied. Ten dogs received 15 doses of the immunomodulator (2.0mg/kg) intramuscularly, and 10 received saline as a placebo. Skin and peripheral blood samples were collected following administration of the immunomodulator. The groups were followed to observe for clinical signals of remission; parasite load in the skin biopsies using real-time PCR, the cytokines IL-2, IL-10 and IFN-γ in the supernatant of peripheral blood mononuclear cells stimulated in vitro with either total promastigote antigen or phytohemagglutinin measured by capture ELISA, and changes in CD4+ and CD8+ T cell subpopulations evaluated by flow cytometry. Comparison between the groups showed that treatment with the immunomodulator promoted improvement in clinical signs and a significant reduction in parasite load in the skin. In peripheral blood mononuclear cell cultures, supernatants showed a decrease in IL-10 levels and an increase in IL-2 and IFN-γ. An increase in CD8+ T cells was observed in peripheral blood. In addition, the in vitro leishmanicidal action of P-MAPA was investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and no leishmanicidal activity was detected. These findings suggest that P-MAPA has potential as an immunotherapeutic drug in canine visceral leishmaniasis, since it assists in reestablishing partial immunocompetence of infected dogs. © 2013 Elsevier B.V.

Normative study and evaluation of age influence on Brainstem auditory evoked potentials in mixed breed dogs]

The aim of this paper was to obtain normative data of auditory evoked potentials from 34 mixed breed dogs and evaluate the age influence. The animals were divided in two groups of different ages and auditory evoked potential was performed with a 85dB stimulus intensity. Group 1 included 16 dogs between 1 and 8 years of age, and group 2 included 18 dogs with over 8 years of age. The length and head diameter were measured and there was no statistical difference between the two groups. In group 1, mean latencies of waves I, III, and V were 1.13; 2.64, and 3.45ms, and the intervals I-III, III-V, and I-V were 1.51; 0.81, and 2.32 ms, respectively. In group 2, the mean latencies of waves I, III and V were 1.15, 2.62, and 3.55ms, and the intervals I-III, III-V, and I-V were 1.47, 0.93, and 2.40ms, respectively. The latencies observed in this study were similar to previous studies conducted by other authors. It was observed that significant differences were present for wave V and intervals III-V and I-V latencies when comparing groups with different ages, consequently this characteristic must be considered during BAEP result interpretation.

Correction of rectal sacculation through lateral resection in dogs with perineal hernia ? Technique description

The occurrence of perineal hernias in dogs during routine clinical surgery is frequent. The coexistence of rectal diseases that go undiagnosed or are not correctly treated can cause recurrence and postoperative complications. The objective of this report is to describe a surgical technique for treatment of rectal sacculation through lateral resection in dogs with perineal hernia, whereby restoring the rectal integrity.

Prevalence of neoplasia diagnosed in dogs at the veterinary hospital at eduardo mondlane university, mozambique

A retrospective study of tumors in dogs was carried out in the School Veterinary Hospital (HVE), Eduardo Mondlane University, Mozambique, between January 2004 and January 2009. Data from clinical-surgical records of tumor samples surgically removed, necropsy and cytological exams were used and sent for histopathology exams. During the study 210 tumors and about 27 different types of tumors were identified. Skin tumors (61.0%) were the most frequent, followed by mammary gland tumors (27.60%), reproductive system tumors (11.90%) and bone system tumors. The mammary gland tumors were observed only in females, with high prevalence in bitches which were not spayed or spayed after two years old. It was concluded that the neoplasia prevalence increase was correlated to the higher longevity of dogs, and breed also contributed for the development of determined tumors due to high incidence of mastocitomas in Boxer and German Shepherd breeds. The 100 % incidence of mammary gland neoplasia in females confirmed the higher predisposition for mammary gland neoplasia in females.