The effect of 3 resuscitative fluid therapies on hemostasis as measured by rotational thromboelastometry in dogs.

BACKGROUND: Evidence suggests that administration of intravenous fluids impairs hemostasis. Thromboelastometry (TEM) may provide a more sensitive measure of the fluid effects on hemostasis than traditional coagulation tests. OBJECTIVE: The study aim was to determine if resuscitative fluid therapy affects hemostasis, as measured by TEM. MATERIALS AND METHODS: Using a randomized crossover design, 6 healthy dogs were administered intravenous colloid, crystalloid, and hypertonic saline at therapeutic doses. Whole blood samples were taken at baseline, 1, 4, and 8 h posttreatment for TEM analysis and measurement of PT, APTT, and fibrinogen. Univariate ANOVA on transformed data evaluated differences between groups and within groups. When significant differences were noted (P = .003), a Tukey test was performed. RESULTS: A statistically significant interaction between individual dogs and treatment was noted for most TEM variables. There was a significant decrease in clot firmness from baseline in the TEM assessment of the intrinsic pathway. Colloid treatment correlated with significantly higher clot firmness in the TEM extrinsic pathway assessment, which decreased over time, and PT was significantly shorter in colloid-treated dogs. Overall, PT was significantly prolonged at one hour; however, all PT values were within the RI. The fibrinogen concentration was significantly different between all treatments. CONCLUSIONS: Clinically relevant doses of resuscitative fluids resulted in a decreased clot firmness in the intrinsic pathway, as measured by TEM, which affected hemostatic variables in healthy dogs. There was also a significant individual response to treatment. The changes noted in this study are not expected to result in clinically apparent bleeding.

Evaluation of inhibition of F4ac positive Escherichia coli attachment with xanthine dehydrogenase, butyrophilin, lactadherin and fatty acid binding protein.

BACKGROUND: Neonatal and post-weaning colibacillosis caused by enterotoxigenic E. coli is responsible for substantial economic losses encountered by the pork industry. Intestinal colonization of young piglets by E. coli depends on the efficiency of bacterial attachment to host gastrointestinal epithelium that is mediated by fimbriae. We tested the effect of porcine individual milk fat globule membrane (MFGM) proteins on F4ac positive E. coli attachment to porcine enterocytes in vitro. RESULTS: Butyrophilin, lactadherin and fatty acid binding protein inhibited fimbriae-dependent adherence of E. coli to enterocytes in vitro, while xanthine dehydrogenase did not. The inhibiting activity was dose-dependent for all three proteins, but the inhibiting efficiency was different. CONCLUSIONS: The results indicate that MFGM proteins may interfere with attachment of E. coli to porcine neonatal intestinal mucosa.

A comparative proteomic study of plasma in feline pancreatitis and pancreatic carcinoma using 2-dimensional gel electrophoresis to identify diagnostic biomarkers: A pilot study.

While pancreatitis is now recognized as a common ailment in cats, the diagnosis remains challenging due to discordant results and suboptimal sensitivity of ultrasound and specific feline pancreatic lipase (Spec fPL) assay. Pancreatitis also shares similar clinical features with pancreatic carcinoma, a rare but aggressive disease with a grave prognosis. The objective of this pilot study was to compare the plasma proteomes of normal healthy cats (n = 6), cats with pancreatitis (n = 6), and cats with pancreatic carcinoma (n = 6) in order to identify potential new biomarkers of feline pancreatic disease. After plasma protein separation by 2-dimensional gel electrophoresis, protein spots were detected by Coomassie Brilliant Blue G-250 staining and identified by mass spectrometry. Alpha-1-acid glycoprotein (AGP), apolipoprotein-A1 (Apo-A1), and apolipoprotein-A1 precursor (Pre Apo-A1) appeared to be differentially expressed, which suggests the presence of a systemic acute-phase response and alteration of lipid metabolism in cats with pancreatic disease. Future studies involving greater case numbers are needed in order to assess the utility of these proteins as potential biomarkers. More sensitive proteomic techniques may also be helpful in detecting significant but low-abundance proteins.

Bien que la pancréatite soit maintenant reconnue comme un problème peu fréquent chez les chats, le diagnostic demeure un défi étant donné les résultats discordants et la sensibilité sous-optimale de l'échographie et de l'épreuve spécifique de la lipase pancréatique féline (Spec fPL). La pancréatite partage également des similarités cliniques avec le carcinome pancréatique, une maladie rare mais agressive ayant un pronostic grave. L'objectif de cette étude pilote était de comparer les protéomes plasmatiques de chats normaux en santé (n = 6), de chats avec une pancréatite (n = 6), et de chats avec un carcinome pancréatique (n = 6) afin d'identifier de nouveaux biomarqueurs potentiels de maladie pancréatique féline. Après séparation des protéines plasmatiques par électrophorèse en gel en deux dimensions, les taches de protéines furent détectées par coloration avec du bleu brillant de Coomassie G-250 et identifiées par spectrométrie de masse. La glycoprotéine acide alpha-1 (AGP), l'apolipoprotéine A1 (Apo-A1), et le précurseur de l'apolipoprotéine A1 (Pre Apo-A1) apparaissent comme étant exprimées de manière différentielle, ce qui suggère la présence d'une réponse de phase-aiguë systémique et une altération du métabolisme des lipides chez les chats avec une maladie pancréatique. Des études additionnelles regroupant un plus grand nombre de cas sont nécessaires afin d'évaluer l'utilité de ces protéines comme biomarqueurs potentiels. Des techniques plus sensibles de protéomique pourraient également être utiles pour détecter des protéines significatives mais de faible abondance.(Traduit par Docteur Serge Messier).

Identification of Escherichia coli F4ac-binding proteins in porcine milk fat globule membrane.

F4ac-positive enterotoxigenic Escherichia coli (ETEC) must attach to the intestinal mucosa to cause diarrhea in piglets. Prevention of bacterial attachment to the intestinal mucosa is the most effective defense against ETEC-induced diarrhea. Porcine milk fat globule membranes (MFGM) were shown to be able to inhibit attachment of ETEC to the intestinal brush border; however, the specific components of porcine MFGM that inhibited attachment of ETEC to enterocytes were not identified. Accordingly, the purpose of this study was to identify F4ac-binding MFGM proteins by overlay Western blot and affinity chromatography. The proteome of porcine MFGM was characterized and the following F4ac-binding proteins were detected by overlay Western blot and affinity chromatography: lactadherin, butyrophilin, adipophilin, acyl-CoA synthetase 3, and fatty acid-binding protein 3. The biological function of these proteins was not investigated but it is possible that their interaction with F4ac fimbria interferes with bacterial attachment and colonization.

Les Escherichia coli entérotoxinogénique (ETEC) positif pour F4ac doivent s'attacher à la muqueuse intestinale pour causer la diarrhée chez les porcelets. L'empêchement de l'attachement bactérien à la muqueuse intestinale est le moyen de défense le plus efficace contre la diarrhée induite par les ETEC. Les membranes de globules de gras de lait porcin (MFGM) ont été montré comme étant capable d'inhiber l'attachement des ETEC à la bordure en brosse intestinale; toutefois, les composantes spécifiques des MFGM porcines qui inhibaient l'attachement des ETEC aux entérocytes ne furent pas identifiées. Ainsi, le but de la présente étude était d'identifier les protéines des MFGM liant F4ac par immunobuvardage par superposition et chromatographie d'affinité. Le protéome des MFGM porcine fut caractérisé et les protéines liant F4as suivantes furent détectées par immunobuvardage par superposition et chromatographie d'affinité : lactadhérine, butyrophiline, adipophiline, acyl-CoA synthétase 3, et la protéine 3 liant les acides gras. La fonction biologique de ces protéines ne fut pas étudiée mais il est possible que leur interaction avec les fimbriae F4ac interfère avec l'attachement bactérien et la colonisation.(Traduit par Docteur Serge Messier).

Evaluation of lymph node aspirates at diagnosis and relapse in dogs with high-grade multicentric lymphoma and comparison with survival time.

BACKGROUND: Canine high-grade multicentric lymphoma, a common disease with variable response to chemotherapy, is often diagnosed using cytology. OBJECTIVES: The purpose of the study was to compare cytologic features of canine peripheral lymph node aspirates collected at diagnosis and at relapse, and evaluate their usefulness in predicting survival. METHODS: Cytologic scoring based on a rubric and nuclear morphometry analyses were performed on cytologic smears collected at diagnosis and at relapse. Scores at diagnosis and relapse were compared by paired t-test and evaluated in relation to time from diagnosis to remission, remission to relapse, relapse to death, and total survival time, using the Cox proportional-hazards regression model. RESULTS: Number of mitoses and total cytologic score were significantly higher at relapse compared to diagnosis (P < .05). None of the nuclear morphometry measures were significantly different between diagnosis and relapse. The presence of binucleated or multinucleated cells at diagnosis was associated with a shorter remission and decreased total survival (P < .05). Increased mean nucleoli at relapse was associated with longer remission and total survival (P < .05). Increased minimum nuclear radius and diameter at diagnosis were associated with a decreased time from relapse to death (P < .05). Several nuclear morphometry measures at relapse were associated with a shorter time from diagnosis to remission (P < .05). CONCLUSIONS: Number of mitoses and total score were higher at relapse than at diagnosis in canine lymphoma. The presence of binucleated or multinucleated cells at diagnosis may be useful as indicator of a poor prognosis. Further studies including a larger number of cases are required to reinforce the prognostic values of these cytologic features.

Reticulocytosis in nonanemic dogs: increasing prevalence and potential etiologies.

BACKGROUND: An increasing prevalence of reticulocytosis in the absence of anemia (RAA) in dogs has been suspected in recent years. OBJECTIVES: The objectives were to determine whether prevalence of RAA in our canine population has been increasing over the last years, and to identify potential predisposing factors. METHODS: The annual prevalence of RAA in adult dogs was determined between 2000 and 2012. Clinical histories and CBC data were analyzed for all dogs, as well as owner response to a questionnaire including information on nutrition and supplements was conducted for dogs with RAA identified between 2011 and 2012. In addition, serum iron concentration (Fe), total iron-binding capacity (TIBC), and percent transferrin saturation (%TS) were determined in 14 dogs with RAA and compared with 8 healthy control dogs. RESULTS: Reticulocytosis in the absence of anemia was identified in 1035 dogs, with the prevalence increasing since 2006. Dogs with RAA evaluated after 2006 (n = 853) had significantly lower MCV and were more likely to have microcytosis than those prior to 2006 (n = 182). Increased incidence of osteoarthritis was observed in dogs evaluated after 2006, including the dogs studied between 2011 and 2012 (n = 31), and administration of nonsteroidal anti-inflammatory drugs, omega-3 fatty acids, and glucosamine was more common in the latter. Significantly lower mean Fe and %TS, and higher TIBC were found in dogs with RAA compared to unaffected dogs. CONCLUSIONS: Prevalence of RAA has increased in recent years in our canine population. More ubiquitous use of anti-inflammatory medications and nutraceuticals, associated with increased diagnosis of osteoarthritis should be considered as contributing factors.

Utility of nuclear morphometry in the cytologic evaluation of canine cutaneous soft tissue sarcomas.

Cytopathologists lack reliable criteria to distinguish neoplastic from reactive spindle cells; however, with computer-based nuclear morphometry, it is now possible to more objectively and precisely quantify differences between selected populations of cells. Forty-four cutaneous soft tissue sarcomas and 5 cases of reactive spindle cell proliferations in the dog were morphometrically analyzed with regard to median and standard deviation (SD) of nuclear area, diameter (max, min, mean), radius (max, min), perimeter, and roundness. Overall, nuclei from reactive spindle cells were larger, with greater variation in nuclear size and shape. Significant differences (P < 0.05) were found for several nuclear parameters, including the median and SD of maximum diameter and radius, as well as the SD of roundness. No significant differences were found in nuclear parameters between soft tissue sarcomas divided by histologic grade, mitotic index, or tumor necrosis score. Analysis of the sources of variation indicated near-perfect intraobserver and substantial interobserver agreement. The largest source of variation was due to selection of different measurement fields, reflecting the inherent biological variation in nuclear size within the tumor cell population. The results indicate that nuclear morphometry on cytologic preparations is a reproducible method that may be able to differentiate cutaneous soft tissue sarcomas from reactive mesenchymal lesions in the dog. Further studies, including a larger number of cases, are warranted to assess repeatability of results.

Localization of Bovine papillomavirus in equine sarcoids and inflammatory skin conditions of horses using laser microdissection and two forms of DNA amplification.

Equine sarcoids are the most common tumor of horses. Bovine papillomavirus (BPV) has been suggested as the cause of sarcoids. Studies have shown that BPV is present in swabs or biopsies from nonsarcoid-bearing equine skin. Skin biopsies from a variety of different conditions and normal skin from horses with no reported history of sarcoids were examined by polymerase chain reaction (PCR) for the presence of BPV, which was found in all different types of skin conditions as well as normal skin. Forty-one out of 86 skin biopsies from horses without sarcoids were found to contain BPV DNA. Laser microdissection, followed by DNA amplification through both PCR and isothermal loop-mediated amplification, was performed on these 41 biopsies and on 70 additional BPV-positive sarcoid biopsies to localize the virus. Location of BPV DNA was different between sarcoid and nonsarcoid groups. Nonsarcoid skin biopsies were more likely to have BPV within intact or inflamed epidermis than sarcoids (P = 0.016 and P = 0.007, respectively). Areas of inflammation within the dermis and epidermis were more likely to contain BPV than in noninflamed areas (P = 0.008 and P = 0.009, respectively). Bovine papillomavirus was also found in the epidermis of all types of sarcoids examined, more frequently in occult sarcoids than in fibroblastic and nodular types (P = 0.03 and P = 0.01, respectively). Results suggest that BPV is commonly found in normal and inflamed equine skin, and it is likely an important predisposing factor in the development of sarcoids.

Serum C-reactive protein concentrations in healthy Miniature Schnauzer dogs.

BACKGROUND: C-reactive protein (CRP) is a sensitive marker for inflammation in people and dogs. In people, an association between CRP concentration and atherosclerosis has been reported. Atherosclerosis is rare in dogs, but the Miniature Schnauzer breed may be at increased risk for developing this vascular disease. It is not known if CRP concentrations in Miniature Schnauzer dogs differ from those in other dog breeds. OBJECTIVES: Our objectives were to validate an automated human CRP assay for measuring CRP in dogs and compare CRP concentrations in healthy Miniature Schnauzer dogs with those in non-Miniature Schnauzer breeds. METHODS: Sera from 37 non-Miniature Schnauzer dogs with inflammatory disease were pooled and used to validate a human CRP immunoturbidimetric assay for measuring canine CRP. Blood was collected from 20 healthy Miniature Schnauzer dogs and 41 healthy dogs of other breeds. Median serum CRP concentration of healthy Miniature Schnauzer dogs was compared with that of healthy non-Miniature Schnauzer dogs. RESULTS: The human CRP assay measured CRP reliably with linearity between 0 and 20 mg/L. CRP concentration for healthy Miniature Schnauzer dogs (median 4.0 mg/L, minimum-maximum 0-18.2 mg/L) was significantly higher than for the healthy non-Miniature Schnauzer dogs (median 0.1 mg/L, minimum-maximum 0-10.7 mg/L); 17 of the 20 Miniature Schnauzer dogs had values that overlapped with those of the non-Miniature Schnauzer dogs. CONCLUSIONS: Median CRP concentration of Miniature Schnauzer dogs was slightly higher than that of other breeds of dogs. A relationship between higher CRP concentration in Miniature Schnauzer dogs and idiopathic hyperlipidemia, pancreatitis, and possible increased risk for atherosclerosis remains to be determined.

C-reactive protein as a prognostic indicator in dogs with acute abdomen syndrome.

The objective of the present study was to measure serum C-reactive protein (CRP) concentrations in 32 dogs with acute abdomen syndrome at presentation and after 48-72 hr. Data were evaluated to determine if there was an association between CRP concentration and outcome, and if CRP concentration correlated with the white blood cell (WBC) count at both time points. An immunoturbidimetric assay for human CRP, previously validated for use in dogs, was used for serum CRP analysis. Increased serum CRP concentrations were found in 21 dogs at presentation. Fifteen of these dogs had declining serum CRP concentrations by 48-72 hr, but 3 of the 15 dogs were later euthanized. Serum CRP concentrations increased by 48-72 hr in 4 dogs. Of the 32 dogs, 4 were dead or were euthanized prior to the 48-72 hr time point. No significant difference between initial CRP concentration and outcome was found (P = 0.054). Initial and 48-72 hr CRP values taken together were significantly different between outcome groups (P < 0.001). Serum CRP concentrations that were elevated at both time points were associated with a poor prognosis. No correlation was found between CRP concentrations and WBC counts at presentation (P = 0.83); however, a significant correlation was noted at 48-72 hr (P = 0.03). Evaluation of sequential CRP concentrations in dogs with acute abdomen syndrome may be helpful in assessing clinical response to treatment and predicting outcome. Also, serum CRP may be better in detecting tissue injury and/or inflammation at presentation than WBC counts in select cases.

Clonality and phenotyping of canine lymphomas before chemotherapy and during remission using polymerase chain reaction (PCR) on lymph node cytologic smears and peripheral blood.

Polymerase chain reaction (PCR) assays for the immunoglobulin and T-cell receptor genes were utilized to determine phenotype and clonality from lymph node cytologic smears and peripheral blood lymphocytes from 10 dogs with lymphoma, before chemotherapy and during remission. Results were compared with those from 13 dogs with a cytologic diagnosis of lymph node hyperplasia. Clonality was identified in 7 of the lymphomas on the basis of either lymph node cytology or peripheral blood lymphocytes before treatment. No lymph node hyperplasia samples were clonal. In 6 of the dogs with lymphoma, clonality was demonstrated during clinical remission. Detection of PCR clonality during clinical remission is an effective means of identifying minimal residual disease in canine lymphoma and thus additional work is warranted to determine if molecular remission is prognostic or predictive for outcome in well-controlled and well-defined lymphoma subtypes.

Epidemiology of equine sarcoids in horses in western Canada.

Sarcoids are the most common tumor of the equine skin but only 1 study describing the epidemiology of sarcoids in Canadian horses has been published. The records of 5 veterinary diagnostic laboratories in western Canada were searched to identify submissions of sarcoids from horses. The submission records and diagnostic reports of 802 separate submissions of equine sarcoids were reviewed for age, breed, and gender of the horse and the number, location, and clinical type of sarcoid. From these records, the 307 submissions to laboratories in Saskatchewan were compared to a reference group to test for breed and gender predisposition. Based on clinical history and lesion descriptions, 5 clinical types of sarcoids were identified. Horses of various ages and 23 equine breeds were affected; donkeys were over-represented. Polymerase chain reaction (PCR) for the bovine papillomavirus (BPV) DNA was performed on formalin-fixed paraffin-embedded tissues from a stratified subset of 96 of the different clinical types; BPV2 was present in 60 of 74 (81%) for which a PCR product was obtained. Unlike other areas in the world, in western Canada, equine sarcoids are most commonly associated with BPV type 2.

Immunohistochemical expression of vascular endothelial growth factor and vascular endothelial growth factor receptor-2 in canine simple mammary gland adenocarcinomas.

The expression of 5 markers associated with angiogenesis, proliferation, and apoptosis was studied in 26 canine simple mammary gland adenocarcinomas (SMGAs). The adenocarcinomas were graded histologically, and tissue sections were immunohistochemically stained for the expression of vascular endothelial growth factor (VEGF), VEGF receptor-2 (VEGFR-2), intra-tumor microvessel density, and tumor proliferation (PI) using antibodies against VEGF, VEGFR-2, von Willebrand factor, and Ki-67 antigen, respectively. Apoptotic indices (AI) were determined by an apoptosis assay. Markers VEGF and VEGFR-2 were detected in 96% and 100% of SMGAs, respectively. A high correlation between histologic grade and PI (r = 0.73), a moderate correlation between VEGF and histologic grade (r = 0.33), and between VEGF and PI (r = 0.42) were found. There was a significant difference in median PI among the 3 histologic grade groups (r < 0.05). Vascular endothelial growth factor may stimulate tumor cell proliferation through an autocrine loop, since VEGF and VEGFR-2 were expressed in most tumors.

Guidelines for resident training in veterinary clinical pathology. III: cytopathology and surgical pathology.

The Education Committee of the American Society for Veterinary Clinical Pathology has identified a need for improved structure and guidance of training residents in clinical pathology. This article is the third in a series of articles that address this need. The goals of this article are to describe learning objectives and competencies in knowledge, abilities, and skills in cytopathology and surgical pathology (CSP); provide options and ideas for training activities; and identify resources in veterinary CSP for faculty, training program coordinators, and residents. Guidelines were developed in consultation with Education Committee members and peer experts and with evaluation of the literature. The primary objectives of training in CSP are: (1) to develop a thorough, extensive, and relevant knowledge base of biomedical and clinical sciences applicable to the practice of CSP in domestic animals, laboratory animals, and other nondomestic animal species; (2) to be able to reason, think critically, investigate, use scientific evidence, and communicate effectively when making diagnoses and consulting and to improve and advance the practice of pathology; and (3) to acquire selected technical skills used in CSP and pathology laboratory management. These guidelines define expected competencies that will help ensure proficiency, leadership, and the advancement of knowledge in veterinary CSP and will provide a useful framework for didactic and clinical activities in resident-training programs.

Sarcoids in two captive tapirs (Tapirus bairdii): clinical, pathological and molecular study.

This case report describes for the first time sarcoids in tapirs (Tapirus bairdii), namely, a 2-year-old male and a 3.6-year-old female born and housed at the same facility. The male presented with a 3-cm nodular, red, pedunculated, hairless, ulcerated mass on the inner surface of the left pinna. No recurrence or additional growths were present during the 3 years following surgical excision of the mass. The female presented with a similar 2-cm mass on the inner surface of the right pinna, which recurred 2 months following surgical excision, but was subsequently successfully treated locally with liquid nitrogen with no further recurrence during a 2-year follow-up period. Histologically, these two masses closely resembled equine sarcoids. Similarly, an association with bovine papillomavirus 1 was demonstrated using polymerase chain reaction and in situ hybridization.

Alkaline phosphatase: beyond the liver.

The alkaline phosphatases comprise a heterogeneous group of enzymes that are widely distributed in mammalian cells. They often are associated with cell membranes, but their exact physiologic function is unknown. Despite this, alkaline phosphatase activity is a very useful serum biochemical indicator of liver disease, particularly cholestatic disease. However, increases in the activity of alkaline phosphatase in serum and other body fluids may reflect physiologic or pathologic changes beyond those of hepatic origin. For example, nonhepatic increases in serum alkaline phosphatase activity are found in young animals, in pregnant and lactating females, and in association with high fat diets. Bone disease, endocrine disease, neoplasia, and other disorders can result in increased alkaline phosphatase activity. In addition, alkaline phosphatase activity may be increased due to induction by certain drugs such as glucocorticoids and anticonvulsants. In this article, we will review the physiologic and pathologic factors influencing the activity of alkaline phosphatase in serum and other body fluids, with an emphasis on disorders beyond liver disease.

Serial blood lactate concentrations in systemically ill dogs.

BACKGROUND: Lactate concentration often is quantified in systemically ill dogs and interpreted based on human data. To our knowledge, there are no published clinical studies evaluating serial lactate concentrations as a prognostic indicator in ill dogs. OBJECTIVES: Our objective was to perform a prospective study, using multivariate analysis, to determine whether serial lactate concentrations were associated with outcome in ill dogs requiring intravenous fluids. METHODS: Eighty sick dogs had lactate concentrations evaluated, using an analyzer that measures lactate in the plasma fraction of heparinized whole blood, at 0 hours and 6 hours after initiation of treatment. Severity of illness and outcome (survivor, nonsurvivor) were determined by reviewing the patient's record 2 weeks after admission. Lactate concentrations, age, body weight, gender, and severity of illness were evaluated using multivariate analysis to determine their effects on outcome. RESULTS: Dogs with lactate concentrations greater than the reference interval at 6 hours were 16 times (95% confidence interval = 2.32-112.71 times, P <.01) more likely not to survive compared to dogs with lactate concentrations within the reference interval. Lactate concentrations above the reference interval at 0 hours were not significantly related to outcome. However, hyperlactatemia that did not improve by > or = 50% within 6 hours was significantly associated with mortality (P = .024). CONCLUSION: Dogs with a lactate concentration higher than the reference interval at 6 hours were more likely not to survive. These results indicate an association between lactate concentration and outcome and emphasize the importance of serial lactate concentrations in evaluating prognosis.