Standard and higher doses of atropine in a canine model of pulseless electrical activity

OBJECTIVE To determine whether standard or increased doses of atropine improve the return of spontaneous circulation (ROSC) rate in a canine model of pulseless electrical activity (PEA). METHODS A prospective, controlled, blinded laboratory investigation was performed using an asphyxial canine cardiac arrest model. After the production of asphyxial PEA, 75 dogs remained in untreated PEA for 10 minutes and then were randomized to receive placebo (group 1) or one of four doses of atropine (group 2, 0.04 mg/kg; group 3, 0.1 mg/kg; group 4, 0.2 mg/kg; group 5, 0.4 mg/kg). All the animals received mechanical external CPR and epinephrine (0.02 mg/kg every 3 minutes) throughout resuscitation. RESULTS The ROSC rates were not significantly different between the groups (group 1, 73%; group 2, 67%; group 3, 40%; group 4, 47%; group 5, 27%; p = 0.06). The heart rates and hemodynamics during resuscitation were not significantly different between the groups. CONCLUSION In this canine model of asphyxial PEA cardiac arrest, standard-dose atropine did not improve ROSC rates, compared with placebo. Increasing doses of atropine tended to decrease ROSC rates, compared with placebo and standard-dose atropine.

The effects of endothelin-1 on coronary perfusion pressure during cardiopulmonary resuscitation in a canine model

OBJECTIVE To study the hemodynamic effects of exogenously administered endothelin-1 (ET-1), a peptide produced by endothelial cells with potent non-adrenergically mediated vasoconstrictor properties. METHODS A prospective drug intervention study was carried out in a resuscitation research laboratory. Fifteen mixed-breed dogs were anesthetized and instrumented for hemodynamic monitoring. Asphyxia arrest was produced by clamping the endotracheal tube. Hemodynamic data were collected continuously. Following loss of aortic fluctuations monitored by thoracic aortic catheter, the animals remained in pulseless electrical activity (PEA) for 10 minutes. After 10 minutes of no-flow PEA, closed-chest CPR was begun and the animals were randomized to one of three treatment groups (EPI, 0.02 mg/kg epinephrine IV every 3 minutes; ENDO, 100 micrograms ET-1 IV at 0 minutes; and EPI/ENDO, a combination of the EPI and ENDO treatments). RESULTS ENDO and EPI alone produced similar coronary perfusion pressures (CPPs). The EPI/ENDO combination produced significantly improved CPP compared with that of either EPI or ENDO alone. In the EPI group, the best mean CPP was 16 +/- 14 mm Hg and occurred 7 minutes after drug administration. In the ENDO group, the best mean CPP was 28 +/- 7 mm Hg and occurred 13 minutes after drug administration. In the EPI/ENDO combination group, the best mean CPP was 61 +/- 37 mm Hg and occurred 7 minutes after drug administration (p < 0.05 compared with the EPI and ENDO groups alone). CONCLUSION ET-1 is a potent vasoconstrictor. The combination of EPI and ENDO significantly improved CPP compared with that for either agent alone. ET-1 should be investigated further as a vasoconstrictor in cardiac arrest.

Endogenous nitric oxide production in canine osteoarthritis: Detection in urine, serum, and synovial fluid specimens

OBJECTIVE To measure nitric oxide (NO) concentrations in serum, urine, and synovial fluid (SF) of dogs with naturally occurring cranial cruciate ligament (CCL) rupture and normal dogs, and to compare these with clinical and histologic changes of osteoarthritis (OA). STUDY DESIGN Prospective clinical study including 2 groups of animals selected from the hospital population. ANIMALS Forty-three dogs (CCL group) with OA secondary to CCL rupture; 30 healthy dogs (control group) without CCL rupture. METHODS Serum, urine, and SF were collected before and during surgery in the CCL group or immediately after euthanasia in the control group. Articular cartilage and synovial membrane tissue specimens were prepared for routine histologic examination. The stable end products of NO, total nitrite and nitrate (NOt) activity, were measured in body fluids and compared with macroscopic and histologic degrees of OA. Urinary NOt concentration was compared with urinary creatinine concentration and stated as urinary NOt:creatinine ratio (UNCR). RESULTS-SF NOt concentrations were not significantly different between the 2 groups. Serum NOt concentrations (45.6 vs 28.9 micromol/L; P =.042) and the UNCR (0.007 vs 0.004; P =.035) were significantly higher in dogs of the CCL group compared with the control population. An association between UNCR and histologic and macroscopical OA grades could be demonstrated. CONCLUSION UNCR might be a useful indicator of nitrite and nitrate production and, therefore, osteoarthritic changes in joints. CLINICAL RELEVANCE UNCR could be used as a tool to evaluate the NOt production by joint tissues over time and might therefore provide a method of evaluating the effects of drugs in the control of osteoarthritis.

Production of antibodies to canine IL-1beta and canine TNF to assess the role of proinflammatory cytokines

IL-1 and TNF are important proinflammatory cytokines implicated in both antimicrobial host defense and pathogenesis of diseases with an immune-mediated and/or inflammatory component. Respective studies in the dog have been hampered by the unavailability of reagents allowing the specific measurement of canine cytokine proteins and the effect of canine cytokine neutralization by Ab. Starting with recombinant canine (rcan) IL-1beta and rcanTNF, four polyclonal antisera and 22 mAb specific for rcanIL-1beta and rcanTNF were generated. Their usefulness in neutralization assays was determined. Using cytokine-containing supernatants of canine cells in bioassays, polyclonal antisera neutralized either canine IL-1beta or TNF. TNF was also neutralized by three antibodies developed in this study and one commercial mAb. The usefulness of monoclonal and polyclonal Ab in canine cytokine-specific Ab capture ELISA's was assessed. This resulted in the identification of a commercial mAb combination and one pair developed in this study allowing low levels of TNF to be detected by antibody capture ELISA. The detection limit was 141 pg/ml rcanTNF for both combinations. Using rcanIL-1beta as an antigen allowed the detection of lower concentrations of rcanIL-1beta (20 pg/ml, on the average) by a pair of polyclonal antisera than when monoclonals were used. By using such IL-1beta-specific and TNF-specific ELISA's, the respective cytokines were detected in supernatants of canine PBMC stimulated with LPS or heat-killed Listeria monocytogenes and interferon-gamma combined. Thus, monoclonal and polyclonal reagents were identified allowing the quantitation of canine IL-1beta and TNF production in vitro, and the neutralization of these cytokines.

Dominance of chemokine ligand 2 and matrix metalloproteinase-2 and -9 and suppression of pro-inflammatory cytokines in the epidural compartment after intervertebral disc extrusion in a canine model

BACKGROUND CONTEXT In canine intervertebral disc (IVD) disease, a useful animal model, only little is known about the inflammatory response in the epidural space. PURPOSE To determine messenger RNA (mRNA) expressions of selected cytokines, chemokines, and matrix metalloproteinases (MMPs) qualitatively and semiquantitatively over the course of the disease and to correlate results to neurologic status and outcome. STUDY DESIGN/SETTING Prospective study using extruded IVD material of dogs with thoracolumbar IVD extrusion. PATIENT SAMPLE Seventy affected and 13 control (24 samples) dogs. OUTCOME MEASURES Duration of neurologic signs, pretreatment, neurologic grade, severity of pain, and outcome were recorded. After diagnostic imaging, decompressive surgery was performed. METHODS Messenger RNA expressions of interleukin (IL)-1β, IL-2, IL-4, IL-6, IL-8, IL-10, tumor necrosis factor (TNF), interferon (IFN)γ, MMP-2, MMP-9, chemokine ligand (CCL)2, CCL3, and three housekeeping genes was determined in the collected epidural material by Panomics 2.0 QuantiGene Plex technology. Relative mRNA expression and fold changes were calculated. Relative mRNA expression was correlated statistically to clinical parameters. RESULTS Fold changes of TNF, IL-1β, IL-2, IL-4, IL-6, IL-10, IFNγ, and CCL3 were clearly downregulated in all stages of the disease. MMP-9 was downregulated in the acute stage and upregulated in the subacute and chronic phase. Interleukin-8 was upregulated in acute cases. MMP-2 showed mild and CCL2 strong upregulation over the whole course of the disease. In dogs with severe pain, CCL3 and IFNγ were significantly higher compared with dogs without pain (p=.017/.020). Dogs pretreated with nonsteroidal anti-inflammatory drugs revealed significantly lower mRNA expression of IL-8 (p=.017). CONCLUSIONS The high CCL2 levels and upregulated MMPs combined with downregulated T-cell cytokines and suppressed pro-inflammatory genes in extruded canine disc material indicate that the epidural reaction is dominated by infiltrating monocytes differentiating into macrophages with tissue remodeling functions. These results will help to understand the pathogenic processes representing the basis for novel therapeutic approaches. The canine IVD disease model will be rewarding in this process.

Cultivation and expansion of canine Schwann cells using reexplantation

Despite the numerous available possibilities for the surgical treatment of peripheral nerve lesions found in the dog, the success of these treatments is often unsatisfactory. It has been proven that Schwann cells (SC) have a positive influence on the regeneration of nerve stumps. Implanting a guidance channel seeded with autologous SC at the lesion site could be a new therapeutic approach. The aim of this research was to investigate the in vitro cultivation and expansion of canine SC as the main requirement for the treatment referred to above. Biopsies were carried out on 17 nerve samples originating from dogs of different breed, age, gender and condition. The reexplantation method was employed, followed by dissociation using hyaluronidase, collagenase and trypsin and further expansion. The samples were divided into six groups which were treated with a varying combination of mitogens (forskolin, bovine PEX, choleratoxin, heregulin). To obtain the quantities of SC, the specimens were immunostained by a p75-antibody. By employing a growing number of agents it was possible to obtain an increase in both the quantity of cells and purity of cultures. A maximum of 16x10(5) cells per millilitre of suspension was achieved. The largest SC purity measured 27.1%. The maximum SC quantity achieved was 43.3x10(4) SC per millilitre.

Surgical treatment of canine nasal aspergillosis by rhinotomy combined with enilconazole infusion and oral itraconazole.

OBJECTIVES To evaluate the effectiveness of rhinotomy and surgical debridement associated with topical administration of 2 per cent enilconazole and oral itraconazole in dogs with severe or recurrent sinonasal aspergillosis. METHODS A standard rhinotomy was performed on seven dogs. In the initial study, the bone flap was left attached cranially and replaced at the end of the procedure. In the main study group, the bone flap was discarded. Nasal passages were debrided and irrigated with enilconazole solution for one hour. Oral itraconazole was administered to four dogs for one month postoperatively. Follow-up rhinoscopy was performed in all dogs. RESULTS All three dogs in the initial study had recurrence of the disease and two dogs had a second surgery to remove the flap. The main study group included four dogs in which the flap was initially removed, and the two dogs from the initial study that required a second surgery. At follow-up rhinoscopy, five dogs were free of aspergillus but had bacterial or inflammatory rhinitis and one dog had a small aspergilloma but was subsequently asymptomatic. Telephone follow-up revealed that four dogs were asymptomatic, one dog had intermittent sneezing and serous nasal discharge, and one dog had intermittent epistaxis. CLINICAL SIGNIFICANCE Rhinotomy with removal of the flap combined with one-hour infusion of 2 per cent enilconazole and oral itraconazole resulted in satisfactory outcome in dogs with severe or recurrent aspergillosis.

Effect of synthetic agonists of toll-like receptor 9 on canine lymphocyte proliferation and cytokine production in vitro.

Synthetic agonists of TLR9 containing novel DNA structures and R'pG (wherein R=1-(2'-deoxy-beta-d-ribofuranosyl)-2-oxo-7-deaza-8-methyl-purine) motifs, referred to as immune modulatory oligonucleotides (IMOs), have been shown to stimulate T(H)-1-type-immune responses and potently reverse allergen-induced T(H)-2 responses to T(H)-1 responses in vitro and in vivo in mice. In order to investigate the immunomodulatory potential of IMOs in dogs, canine peripheral blood mononuclear cells (PBMC) from healthy dogs were stimulated with three different IMOs and a control IMO, alone or in combination with concanavalin A (ConA). Lipopolysaccharide (LPS) was used as a positive control for B lymphocyte activation. Carboxyfluorescein diacetate succinimidyl ester and phenotype staining was used to tag proliferating T and B lymphocytes (CD5(+) and CD21(+)) by flow cytometry. Real-time PCR and ELISA were processed to assay cytokine production of IFN-gamma, IL-10, TGF-beta, IL-6 and IL-10. Like LPS, IMOs alone induced neither proliferation of CD5(+) T cells nor CD21(+) B cells, but both LPS and IMO had the capacity to co-stimulate ConA and induced proliferation of B cells. In combination with ConA, one of the IMOs (IMO1) also induced proliferation of T cells. IMO1 also significantly enhanced the expression of IFN-gamma on the mRNA and protein level in canine PBMC, whereas expression of IL-10, TGF-beta and IL-4 mRNAs was not induced by any of the IMOs. These results indicate that in canine PBMC from healthy dogs, IMO1 was able to induce a T(H)-1 immune response including T- and B-cell proliferation.

Increasing incidence of canine leptospirosis in Switzerland.

A marked increase in canine leptospirosis was observed in Switzerland over 10 years with a peak incidence of 28.1 diagnosed cases/100,000 dogs/year in the most affected canton. With 95% affected dogs living at altitudes <800 m, the disease presented a seasonal pattern associated with temperature (r2 0.73) and rainfall (r2 0.39), >90% cases being diagnosed between May and October. The increasing yearly incidence however was only weakly correlated with climatic data including number of summer (r2 0.25) or rainy days (r2 0.38). Serovars Australis and Bratislava showed the highest seropositivity rates with 70.5% and 69.1%, respectively. Main clinical manifestations included renal (99.6%), pulmonary (76.7%), hepatic (26.0%), and hemorrhagic syndromes (18.2%), leading to a high mortality rate (43.3%). Similar to the human disease, liver involvement had the strongest association with negative outcome (OR 16.3). Based on these data, canine leptospirosis presents similar features and severity as the human infection for which it therefore can be considered a model. Its re-emergence in a temperate country with very high incidence rates in canines should thus be viewed as a warning and emphasize the need for increased awareness in other species.

The canine neuronal ceroid-lipofuscinosis: a review.

The present article gives a survey over the current scientific knowledge of the canine neuronal ceroid-lipofuscinosis (NCL). NCL is a heterogenous group of lysosomal storage diseases in humans and animals. In consequence of a gene mutation, there is an accumulation of ceroid-lipofuscin in neurons, cells of the retina and the skin and other cells. The stored ceroid-lipofuscin in neurons leads to an impaired cell function and subsequently to cell death. Recently, the underlying genetic defect was discovered in several dog breeds. Genetic testing permits an ante mortem diagnosis of the disease, which up to now was only possible with a positive biopsy result. Another advantage is the identification of carrier animals to eliminate the deleterious alleles.

Climatic conditions as a risk factor in canine gastric dilatation-volvulus

Canine acute gastric dilatation-volvulus (GDV) is a life-threatening condition of multifactorial origin. The risk of developing GDV is influenced by a variety of factors, including breed, age, gender, temperament, diet and management. A relationship between seasonal variations and the frequency of GDV has been previously documented although no association was found with any specific climatic event. Variables in weather conditions within a defined geographic region were investigated in a retrospective study of 287 client-owned dogs diagnosed with GDV between 1992 and 1999. Monthly incidences were evaluated and differences in atmospheric temperature, humidity and pressure between days in which GDV cases were observed and days in which no case was presented were examined. Although temperature was significantly associated with the occurrence of GDV, the difference in temperatures between days with and days without GDV cases was so small that it is unlikely to be of clinical relevance. Moreover, no significant association was found between GDV occurrence and atmospheric pressure or humidity, and a seasonal variation in GDV incidence was not observed.

Canine mast cell tumours: a review of the pathogenesis, clinical features, pathology and treatment

Mast cells (MCs) are well known for their neoplastic transformation in solitary and multiple cutaneous mast cell tumours (MCTs), as well as visceral and systemic mastocytosis. Dogs have a unique risk of developing cutaneous MCTs, and they account for 7% to 21% of all canine skin tumours. The aetiology of canine MCTs is unknown but is probably multifactorial. This article reviews up-to-date knowledge on the pathogenesis, the clinical presentation, the clinical prognostic factors, the diagnostic workup including clinical staging, cytological findings, histological findings and the various grading systems which have been evaluated based on morphology, the assessment of proliferation markers and other factors such as vessel density. Furthermore, detailed information about current treatment protocols for canine cutaneous MCTs is provided.

MRI findings in a case of canine tick born meningoencephalomyelitis

Tick borne encephalitis virus (TBE) is an endemic infectious agent in northeastern Switzerland causing mainly meningoencephalomyelitis in dogs. We report a canine case of tick born meningoencephalomyelitis resulting in flaccid tetraplegia and, subsequently, fatal respiratory failure. Magnetic resonance imaging (MRI) demonstrated intra-axial bilateral, symmetric, and hyperintense lesions in T2-weighted and Fluid Attenuated Inversion Recovery (FLAIR) sequences affecting thalamus, basal nuclei, cerebral white matter and ventral horns of the caudal cervical spine. These radiological findings overlap those described during flavivirus encephalitis affecting human beings. These lesions in MRI and diffusion weighted images correlated with areas of vasogenic edema detected histopathologically. In endemic regions, clinicians should be aware that bilateral, symmetrical hyperintense thalamic lesions in T2WI can be suggestive of flavivirus infection in dogs with encephalitis

Use of activity monitors for assessment of pruritus in an acute model of canine atopic dermatitis

BACKGROUND: We developed a canine model of acute atopic dermatitis to evaluate the potential of compounds to treat pruritus and skin lesions induced in Dermatophagoides farinae (Df)-sensitized dogs. HYPOTHESIS/OBJECTIVES: The aim was to investigate the effectiveness of long-term recording activity monitors to assess pruritus induced by allergen challenges. ANIMALS: Thirty-two Df-sensitized laboratory dogs. METHODS: In two blinded crossover studies, 28 Df-sensitized dogs were challenged on 3 days with a Df slurry applied to clipped abdominal skin. Dogs were treated with a positive control (prednisolone 1 mg/kg once daily for 5 days, starting 1 day before challenge) or left untreated; all were fitted with activity monitors. To confirm pruritus, a parallel study with four dogs was conducted, filming the dogs before and during challenge and assessing the film for pruritic behaviour. RESULTS: The activity of dogs treated with prednisolone was significantly lower between 00.00 and 03.00 h and between 03.00 and 06.00 h compared with untreated dogs (repeated-measures ANCOVA; P < 0.0001). To determine whether the recorded night-time activity corresponded to pruritic manifestations, we compared activity monitor and video recordings of four dogs for two periods (16.30-20.30 and 24.00-03.00 h) before and during a Df challenge. The correlation between night-time activity monitor activity and observed pruritic behaviour was highly significant (test of correlation coefficient versus zero: r = 0.57, P < 0.0001). CONCLUSIONS AND CLINICAL IMPORTANCE: Determination of night-time activity with activity monitors after allergen challenge appears to be an objective and practical way to assess pruritus in this experimental model of canine atopic dermatitis.

Comparative assessment of a canine-specific medium to support colony formation from canine hair follicular keratinocytes.

BACKGROUND Follicular stem cells and their progeny are responsible for the cyclical renewal of hair follicles and maintenance of the hair coat. The understanding of pathways involved in this process is essential to elucidate the pathogenetic mechanisms of primary alopecia. Stem cells and their direct descendants are located in the bulge region of the isthmus of hair follicles. Although these cells have been studied extensively in mice and humans, data for canine isthmic keratinocyte activation and proliferation are not available. HYPOTHESIS/OBJECTIVES The aim was to establish an accurate and reliable in vitro system to study the growth potential of canine isthmic keratinocytes. We assessed the colony-promoting capability of a commercially available canine-specific medium, CELLnTEC (CnT-09), compared with a well-established home-made medium, complete FAD (cFAD). The CnT-09 medium is specific for the growth of canine keratinocytes, while the cFAD medium can support growth and colony formation of keratinocytes from several species. ANIMALS Skin biopsies were obtained from 15 recently euthanized dogs of various breeds with no skin abnormalities. METHODS The isthmic region of compound hair follicles was isolated by microdissection and cell growth monitored using several parameters with colony-forming assays. RESULTS The CnT-09 and cFAD media provided similar growth as measured by the total number and size of colonies, as well as rate of cell differentiation. CONCLUSIONS The commercial canine-specific CnT-09 medium was comparable to the home-made cFAD medium in supporting the growth and proliferation of canine follicular keratinocytes in vitro. The CnT-09 medium should be a viable alternative growth medium for molecular studies of alopecic disorders in dogs.