C-reactive protein measurement in canine saliva.

An established time-resolved immunofluorometric assay designed for measurement of C-reactive protein (CRP) in canine blood was evaluated and validated for use in canine saliva. C-reactive protein was measured in saliva specimens from 5 healthy dogs before and after the injection of casein and in 37 dogs with different disease conditions. The analytical and functional limits of detection were 0.000053 microg/ml and 0.0091 microg/ml, respectively, and intra- and interassay coefficients of variation ranged between 6.7-9.9% and 8.5-16.5%, respectively. A recovery experiment showed no significant disagreement between detected values and expected ones, and saliva CRP concentration was measured in a linear and proportional manner. A positive correlation was found between CRP levels obtained in saliva and serum samples in the experimental (R2 = 0.76) and clinical studies (R2 = 0.70). The assay was able to detect significant differences between salivary CRP levels in healthy dogs and dogs with inflammatory processes. These results suggest that saliva can be used for CRP measurement in dogs. The use of saliva presents the advantage of an easier and less stressful sampling method for the animals, which might be performed outside of hospital environments.     

Allogeneic tooth transplantation in the dog

Allogeneic tooth transplantation was evaluated as a functional and aesthetic treatment for dental fracture in the dog. Of 7 dogs that received tooth transplants, 5 were research animals and 2 were clinical patients. Canine teeth were transplanted immediately after extraction in the research dogs. Endodontic therapy had been performed on the affected canine tooth of one clinical patient. The other clinical patient had bilateral maxillary canine fractures 2 months earlier. One of the research dog transplants failed at 3 weeks as a result of improper surgical technique. Four of the research dogs had a solid implant for 18 months, after which time the dogs were euthanatized serially. All transplanted teeth were anchored firmly into the alveoli, but were nonviable. Root resorption, with bone replacement, was first noticed at 24 months. The transplanted tooth in the first clinical patient remained functional for 3 months, after which time the tooth was fractured. The right canine transplant in the second clinical patient failed by 3 months, probably because of preexisting periapical inflammation. The left transplanted tooth remains stable at 38 months. It was concluded that allogeneic tooth transplantation may have merit as a rapid and inexpensive method for replacement of fractured teeth in the dog. Function is compromised gradually as a result of root resorption and ankylosis, with tooth fracture likely to occur after 2 years.     

Reattachment of tooth fragment: an in vitro study

Canine tooth fracture is common in dogs. Application of an esthetic and durable restoration may be challenging in veterinary dental practice. This study used traditional human dental laboratory methods to evaluate fracture strength of intact dog canine teeth and fractured teeth that had been restored by reattachment of the tooth fragment. The results showed that the teeth restored by reattachment of the tooth fragment supported a test load equal to 45.4 % of the load necessary to fracture intact canine teeth.     

Transmission of multiple antimicrobial-resistant Staphylococcus intermedius between dogs affected by deep pyoderma and their owners

The occurrence of antimicrobial-resistant Staphylococcus intermedius strains was investigated in 13 dogs affected by deep pyoderma, their owners and 13 individuals without daily contact with dogs (control group). A total of 90 canine and 33 human S. intermedius isolates were typed by pulsed field gel electrophoresis (PFGE) to determine their possible identity. The occurrence of S. intermedius in dog-owners was significantly higher compared with the control group (Fisher's exact test, P=0.03), with S. intermedius being detected in seven dog-owners and in one individual not exposed to dogs. The results of the PFGE analysis showed that six out of 13 (46%) owners carried strains identical to those isolated from their dogs. Strains detected in both dogs and humans were resistant up to five different antimicrobial classes, including penicillins, fusidic acid, macrolides/lincosamides, tetracycline and chloramphenicol. Based on the results of this study, owners of dogs affected by deep pyoderma often carry multiple antimicrobial-resistant strains of S. intermedius occurring in their dogs. Independent of the direction and modalities of transmission, this finding raises questions concerning the possible transfer of resistance genes from canine S. intermedius to human pathogenic staphylococci.     

Wnt5a expression in canine osteosarcoma

Osteosarcoma is the most common primary malignancy of bone in dogs and is associated with poor long‐term outcomes due to its highly metastatic nature. A better understanding of the signalling pathways and proteins involved with osteosarcoma pathogenesis may aid in improved outcomes through the use of targeted therapies. The Wnt5a protein, a ligand for the non‐canonical Wnt signalling pathway, is implicated in mediating the aggressiveness of cancer cell lines, including those of human osteosarcoma origin. Given the close relationship between human and canine osteosarcoma, the primary goal of this study was to characterize Wnt5a expression in canine osteosarcoma. Second, if Wnt5a expression was present in canine osteosarcoma, the study aimed to determine any potential association with clinical outcome and clinical variables in similarly treated osteosarcoma‐bearing dogs. Wnt5a expression was present in 26 of the 48 (54%) cases of canine osteosarcoma. Wnt5a expression was not associated with progression‐free survival (P = 0.4) or overall survival (P = 0.1).     

CLINICAL AND ULTRASONOGRAPHIC CHARACTERISTICS OF SALIVARY MUCOCELES IN 13 DOGS

Salivary mucocele is one of the causes of submandibular swelling in dogs and is due to a collection of mucoid saliva that has leaked from a damaged salivary gland. The purpose of this case series report was to describe the clinical and ultrasonographic characteristics of confirmed salivary mucoceles in 13 dogs admitted to the Faculty of Veterinary Medicine at Cairo University. The final diagnosis of salivary mucocele was based on aspirate cytology for all dogs and additional surgical excision for seven dogs. For dogs admitted from 2 weeks to 1 month from the onset of clinical signs, the cervical mucocele appeared as a round echogenic structure with a large volume of central anechoic content. The wall was a clearly identified hyperechoic structure surrounding the gland. For dogs admitted between 1 to 2 months from the onset of clinical signs, the volume of anechoic material appeared less than that seen in the acute cases. The overall appearance of the salivary mucocele was heterogenous. For dogs admitted after 2 months from the onset of clinical signs, the salivary mucocele appeared grainy or mottled, with a heterogenous appearance and a further decrease in anechoic content. For one dog that presented after 3 months from the onset of clinical signs, the salivary mucocele was hard on palpation and appeared hyperechoic with distal acoustic shadowing. Findings from this study indicated that ultrasonographic characteristics of salivary mucoceles in dogs vary depending on the chronological stage of the disease.     

CD20 expression in normal canine B cells and in canine non-Hodgkin lymphoma

We examined the expression of CD20 in normal canine peripheral blood mononuclear cells, normal canine spleen, and canine non-Hodgkin lymphoma (NHL) to determine the feasibility of using this antigen as a diagnostic aid and as a possible target for therapy. An antibody generated against a C-terminal (intracytoplasmic) epitope of human CD20 recognized proteins of 32-36 kd in normal and malignant canine lymphocytes. This antibody showed restricted membrane binding in a subset of lymphocytes in peripheral blood, in the B-cell regions from a normal canine spleen and lymph node, and in malignant cells from 19 dogs with B-cell NHL, but not from 15 dogs with T-cell NHL. The patterns of CD20 reactivity in these samples overlapped those seen using an antibody that recognizes canine CD79a. This anti-CD20 antibody is therefore suitable as an aid to phenotype canine NHL. In contrast, normal canine B cells were not recognized by any of 28 antibodies directed against the extracellular domains of human CD20 (including the chimeric mouse-human antibody Rituximab) or by any of 12 antibodies directed against the extracellular domains of mouse CD20. Thus, the use of CD20 as a therapeutic target will require the generation of specific antibodies against the extracellular domains of canine CD20.     

Expression of class II beta-tubulin by proliferative myoepithelial cells in canine mammary mixed tumors

Benign mammary mixed tumors in dogs resemble human salivary pleomorphic adenomas with regard to their histogenesis, including the occurrence of cartilaginous or bony metaplasia as well as the expression pattern of cytoskeletal proteins in proliferative myoepithelial cells. Recently, a monoclonal antibody specific for class II beta-tubulin has been developed. The epitope it recognizes was determined to be the heptapeptide Glu-Glu-Glu-Glu-Gly-Glu-Asp, which is the common sequence found among the canine, rat, mouse, and human class II beta-tubulin-specific regions. We carried out immunohistochemical studies on mammary mixed tumors obtained from three female dogs using this the monoclonal antibody. The antibody to class II beta-tubulin reacted intensely with proliferative myoepithelial cells in canine mammary mixed tumors, whereas staining was barely detectable in normal myoepithelial cells surrounding alveoli and alveolar ducts within the tumor and adjacent normal tissue. Proliferative myoepithelial cells also expressed vimentin, but alpha-smooth muscle actin (alphaSMA) staining was barely detectable. Immunoblot analysis showed that class II beta-tubulin and vimentin were expressed in myoepithelial cell lines prepared from the three mammary mixed tumors. On the other hand, only one cell line, which was negative for alphaSMA, produced cartilage-specific type II collagen. These results suggest that class II beta-tubulin could be a new molecular marker of proliferating myoepithelial cells in canine mammary mixed tumors and that differential expression of cytoskeletal components is associated with cartilaginous metaplasia of proliferative myoepithelial cells in mixed mammary tumors.     

Evaluation of a radioimmunoassay for type III procollagen peptide in the dog

Type III procollagen peptide (P-3-P) is a serum marker for hepatic fibrosis in humans. The utility of a commercially available radioimmunoassay for P-3-P was evaluated in the dog. The specificity of the assay was assessed by polyacrylamide gel electrophoresis (PAGE) of canine serum and purified bovine P-3-P, followed by Western immunoblotting with rabbit aniti-P-3-P serum. The sensitivity was assessed by performing the radioimmunoassay on dilutions of sera from 22 dogs. Polyacrylamide gel electrophoresis of purified bovine P-3-P and sera from two dogs suspected of having elevated P-3-P concentrations revealed no homologous bands of staining. Western immunoblotting showed marked cross-reactivity of the high antisera concentrations with several components of the serum proteins, but none corresponding to the purified P-3-P. All tested sera from dogs had minimal competitive binding with radiolabeled P-3-P in the radioimmunoassay. Dilution curves of dog sera did not parallel either the standard curve or the dilution curve of a known test human serum. There were no statistically different P-3-P concentrations in any of the groups of dogs studied. It was concluded that currently available radioimmunoassay kits for the measurement of P-3-P in the human are not applicable in the dog. Seemingly, the structure or metabolism of canine P-3-P may vary significantly from that of the bovine or human, limiting the sensitivity and specificity of this assay in the dog.     

Detection of potential markers for systemic disease in saliva of pigs by proteomics: A pilot study

Animals with different health status have been studied in order to extend the knowledge about protein composition of porcine saliva samples and to discover potential salivary markers for systemic disease in porcine production. Clinical examination of animals was performed at farm level where 10 healthy pigs and 10 animals with evident clinical signs of disease were randomly selected. Saliva and blood samples were obtained and afterwards animals were humanely sacrificed to perform a complete necropsy. Levels of two acute phase proteins, haptoglobin and C-reactive protein, were used to identify possible active infections of the animals. Moreover, serological analysis, to the main porcine infectious diseases in the area, was performed. Salivary proteins were separated by two-dimensional gel electrophoresis followed by mass spectrometry for the identification of specific proteins. A total of 58 spots out of 75 were successfully identified by MS, which correspond to 20 unique proteins. Two different approaches were used to perform a statistical comparison of saliva protein patterns from healthy and diseased animals using the relative spot volume (% spot volume/total volume of all spot in the gel, approach “A”) or taking also into account the total protein content of each saliva sample (μg of spot/mL of saliva, approach “B”). Both analyses showed three proteins in common that are differentially regulated between states. However, approach B was selected for biomarker searching since it gave an estimation of protein concentration and showed differential expression of proteins between both health states in a total of 10 proteins, which were up-regulated in disease. Mass spectrometric analysis identified those proteins as salivary lipocalin, lipocalin 1, double headed protease inhibitor protein, adenosine deaminase, haptoglobin, albumin fragments, S100-A8, S100-A9, S100-A12 and pancreatic alpha amylase. These proteins could be considered as potential salivary markers of disease.     

Diseased dogs isoamylases in clinically normal and diseased dogs

Isoamylases in normal canine sera were separated on cellulose acetate membranes using a discontinuous buffer system without EDTA. Four peaks of amylase activity were present in 17 of 24 sera. Normal values were established. The majority of activity was present in Peak 4 (cathodal isoamylase). Tissue extracts of pancreas, duodenum, kidney, lung, testis, spleen and uterus-ovaries contained Peak 4 isoamylase. Liver and salivary gland lacked all isoamylase activity. Pancreas contained Peak 3 in addition to Peak 4 isoamylase. A tissue origin for Peaks 1 and 2 was not identified. An overall lack of resolution resulted from the inclusion of EDTA in the electrophoresis buffer system. This may account for previous findings suggesting that pancreatic amylase is not present in normal canine serum. An increase in the Peak 3 isoamylase was present in dogs with pancreatitis while dogs with pancreatic atrophy had a decrease in all isoamylases. Total amylase activity was significantly (p < 0.05) decreased in dogs with pancreatic atrophy.     

Salivary Cortisol Concentrations in Healthy Dogs and Dogs with Hypercortisolism

Background: Measurement of salivary cortisol is a useful diagnostic test for hypercortisolism (HC) in humans. Objectives: To determine whether measurement of salivary cortisol concentration is a practical alternative to plasma cortisol to diagnose HC, to validate the use of salivary cortisol, and to examine the effect of time of day and sampling location on salivary cortisol. Animals: Thirty healthy dogs and 6 dogs with HC. Methods: Prospective, observational clinical trial including healthy volunteer dogs and dogs newly diagnosed with HC. Salivary and plasma cortisol concentrations were measured with an immunoassay analyzer. Intra‐ and interassay variability, linearity, and correlation between salivary and plasma cortisol concentrations were determined. Results: The required 300 μL of saliva could not be obtained in 88/326 samples from healthy dogs and in 15/30 samples from dogs with HC. The intra‐assay variability for measurement of salivary cortisol was 5–17.7%, the interassay variability 8.5 and 17.3%, and the observed to expected ratio 89–125%. The correlation (r) between salivary and plasma cortisol was 0.98. The time of day and location of collection did not affect salivary cortisol concentrations. Dogs with HC had significantly higher salivary cortisol values than healthy dogs (10.2 ± 7.3 nmol/L versus 1.54 ± 0.97 nmol/L; P < .001). Conclusions and Clinical Importance: The ROCHE Elecsys immunoassay analyzer correctly measured salivary cortisol in dogs. However, a broad clinical application of the method seems limited, because of the large sample volume required.     

Thoracic actinomycosis (arcanobacteriosis) or nocardiosis causing thoracic pyogranuloma formation in three dogs

OBJECTIVE: To describe three cases of canine thoracic actinomycosis (arcanobacteriosis) or nocardiosis in which the primary pathological lesion was a pyogranulomatous abscess in the mediastinum. Clinical signs, difficulties in diagnosis, treatment and prognosis are examined. Comparisons are made between human and veterinary literature to assist in formulating a rational treatment plan. DESIGN: Retrospective clinical study. PROCEDURE: Review of case records from 1984 to 1998. RESULTS: Three dogs presented with large intrathoracic pyogranulomas producing variable clinical signs, not necessarily associated with the respiratory tract. Ages ranged from 2 to 5 years old. Two dogs responded to surgical opening and passive drainage of the abscess, or surgical excision of the granuloma with associated structures, and medical therapy. One dog died intra-operatively. CONCLUSION: A combination of surgical and antimicrobial therapy may carry a fair-to-good prognosis for thoracic granuloma caused by actinomycosis (arcanobacteriosis) or nocardiosis. The extent of surgery should be based on assessment of individual cases and must include surgical biopsy for histology and culture to enable a specific diagnosis to be made. Complete surgical excision is not necessarily required. Prolonged antimicrobial therapy is indicated.     

Serum isoamylase values in normal dogs and dogs with exocrine pancreatic insufficiency

Estimations were made of the serum isoamylase values of normal dogs and of dogs with confirmed exocrine pancreatic insufficiency. A statistically significant difference was demonstrated between the two groups in respect of the values of one of the isoamylase fractions measured. Further study has confirmed that canine salivary tissue lacks amylase activity and that the source of the isoamylase fractions was the pancreas.This knowledge has potential value in the diagnosis of canine exocrine pancreatic insufficiency.     

The expression of calretinin and cytokeratins in canine acanthomatous ameloblastoma and oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) and canine acanthomatous ameloblastoma (CAA) represent two epithelium‐derived neoplasms that affect the oral cavity of dogs. The expression of cytokeratins (CKs) and calretinin has been previously established in the canine tooth bud and odontogenic tumours. The aim of this study was to characterize the CK and calretinin expression profile of OSCC in comparison to CAA and canine tooth bud tissues. Samples from 15 OSCC and 15 CAA cases, as well as 6 tooth buds and 2 normal gingival tissues were examined. OSCC CK expression was consistent with the CK expression profile of CAA and canine tooth bud tissue. Calretinin was positively expressed in 10 of 15 OSCC cases, with 5 cases demonstrating high staining intensity. Only 2 of 15 CAA cases demonstrated mild‐moderate staining intensity. The statistically significant difference in staining pattern and intensity of calretinin in OSCC and CAA can help distinguish between these two tumour types.     

Gnotobiotics and immunopathology: the use of the gnotobiotic environment to study acquired and inherited immunodeficiency diseases

Gnotobiotic animals are highly valued for the study of infectious diseases wherein the clinical signs and lesions of disease can be directly related to host-pathogen interactions and not to the additive effects of environmental influences and other confounding factors. Gnotobiotic dogs have been used to study the pathogenesis of acquired immunodeficiencies associated with canine distemper virus (CDV). In recent years, the laboratory at OSU, in conjunction with University of Pennsylvania personnel have begun a series of long-term studies of dogs affected with the canine X chromosome-linked severe combined immunodeficiency (XSCID) syndrome. This fatal inherited defect is caused by mutation in the common gamma chain (IL2RG) gene and renders affected animals profoundly immunodeficient. XSCIDs dogs, raised within a gnotobiotic environment for up to 3 years remain clinically healthy and are, in every respect normal except for the persistent T-cell defect and the failure to develop lymph nodes. Bone marrow transplantation (unfractionated or enriched for CD34+ stem cells) is the treatment of choice for both the XSCID dogs and male human infants affected with this syndrome. In preliminary studies, we have shown that human CD34+ stem cells colonized XSCIDs-affected gnotobiotic dogs, migrated to the thymus and demonstrated post-thymic activation (CD45RA+ phenotype) in peripheral blood. While many issues are unresolved, these data suggest that, through the use of the gnotobiotic environment, xenotransplantation (human-to-dog) may yield a stable and immunologically functional human-dog chimera.     

Diagnostic value of the use of lateral and occlusal radiographic views in comparison with periodontal probing for the assessment of periodontal attachment of the canine teeth in dogs

OBJECTIVE: To determine the diagnostic value of 2 intraoral bisecting angle radiographic views in comparison with periodontal probing for the assessment of periodontal attachment of the canine teeth in dogs. STUDY POPULATION: 466 canine teeth from 117 dogs. PROCEDURE: Periodontal probing measurements were recorded, and clinical attachment levels (CAL) were calculated at the mesial, buccal, distal, and lingual (or palatal) surfaces on each canine tooth. Occlusal and lateral radiographs of the canine teeth were obtained. Alveolar margin height (AMH) was measured at the same 4 surfaces. Values for AMH and CAL were compared on the basis of tooth surface, dental arch, and radiographic view. RESULTS: The AMH at the mesial and distal surfaces of the mandibular canine teeth was measurable on the lateral view and was significantly correlated with CAL. Similar results were found for the mesial and distal surfaces of the maxillary canine teeth. Buccal and lingual AMH were measured on the mandibular occlusal radiographic view, and values were significantly correlated with CAL, but only the buccal AMH could be assessed on the occlusal radiographic view of the maxilla with values that correlated significantly with CAL. CONCLUSIONS AND CLINICAL RELEVANCE: The lateral radiographic view is suitable for evaluating periodontal attachment at the mesial and distal surfaces of the canine teeth in dogs. The occlusal radiographic view is suitable for assessing buccal surfaces as well as the lingual surface of mandibular canine teeth but not the palatal surface of maxillary canine teeth in dogs.