peer reviewed | OBJECTIVE: To evaluate the usefulness of multisite quantitative ultrasonography for noninvasive assessment of bone in horses. SAMPLE POPULATION: 12 healthy horses and both forelimbs from 8 clinically normal horses. PROCEDURE: For in vivo measurements, various regions of interest (ROI) were examined on the third metacarpal bone, radius, and tibia. Precision error for speed of sound (SOS) measurements was obtained by measuring each ROI of 4 horses 10 times with probe repositioning. Additionally, 3 operators measured each aspect of the third metacarpal bone of 6 horses 5 times each. For ex vivo measurements, third metacarpal bones were examined at 9 ROI, and SOS measurements were performed before and after soft tissue removal. One ROI of a single forelimb was subjected to 96 ex vivo measurements with 3 different contact media. RESULTS: The lateral aspect of the third metacarpal bone had significantly higher SOS values than the dorsal and medial aspect of the third metacarpal bone. No difference was obtained between SOS values of the lateral and medial aspect of the radius. The tibia had significantly higher SOS values than the lateral aspect of the radius and the dorsal and medial aspect of the third metacarpal bone. Intraoperator coefficients of variation ranged from 0.62 to 3.15%, and interoperator coefficients of variation ranged from 0.78 to 2.70%. Values of SOS were highest when silicone oil was used as the contact medium. CONCLUSIONS AND CLINICAL RELEVANCE: Speed of sound measurements obtained by quantitative ultrasonography in axial transmission mode can be used to precisely measure superficial cortical bone properties of third metacarpal bone, radius, and tibia in horses.

Objective-To clone segments of the canine liver alkaline phosphatase (LALP) and corticosteroidinduced alkaline phosphatase (CIALP) genes and use those clones to determine the tissue source of CIALP, the kinetics of LALP and CIALP mRNA expression for glucocorticoid-treated dogs, and the correlation between LALP and CIALP transcript concentrations and isoenzyme activities. Sample Population-Tissues obtained from 7 dogs treated with prednisone (1 mg/kg, SC, q 24 h) for up to 32 days and 1 untreated (control) dog. Procedure-Gene segments of LALP and CIALP were obtained by reverse transcription-polymerase chain reaction (RT-PCR) assay. The tissue source of CIALP and IALP mRNA was determined by northern blot analysis of tissues from 1 of the glucocorticoidtreated dogs. Hepatic tissues and serum samples were obtained from the 6 remaining glucocorticoidtreated dogs on days 0, 2, 5, 10, and 32 of prednisone treatment, and relative expression of LALP and CIALP mRNA was correlated with LALP and CIALP activity. Results-A 2,246-base pair (bp) segment of canine LALP and a 1,338-bp segment of CIALP were cloned. Northern blot analysis revealed CIALP mRNA expression in hepatic tissues only after glucocorticoid treatment. Kinetics of LALP and CIALP mRNA expression in the liver of glucocorticoid-treated dogs paralleled liver and serum activities of LALP and CIALP. Conclusions and Clinical Relevance-The liver is the most likely source for CIALP in dogs. Analysis of kinetics of serum and hepatic LALP and CIALP mRNA suggests that after glucocorticoid treatment, both are regulated by modification of mRNA transcript concentrations, possibly through differing mechanisms.

Objective-To correlate tissue distribution with development of lesions after experimental infection with a virulent strain of noncytopathic bovine viral diarrhea virus (BVDV) type 2 in calves. Animals-Ten 14-day-old and two 2-month-old colostrum-deprived calves. Procedure-Calves were intranasally inoculated with BVDV type-2 strain 1373 from an outbreak of clinically severe bovine viral diarrhea (BVD). Two 14-day-old calves served as noninfected controls. Two calves each were euthanatized on postinoculation days 3, 6, and 12, and 1 each on days 8, 9, 13, and 14. Tissues were collected for immunohistologic and histologic examination. Results-Inoculated calves developed nonspecific clinical signs characterized by high fever and decreased numbers of leukocytes and thrombocytes. Viral antigen was detected focally in lymphoid tissues on day 3. On days 6, 8, 9, 12, and 14, viral antigen became increasingly widespread throughout organs and tissues. Viral antigen in lymphoid tissues was associated with severe depletion of all compartments. Lesions in other tissues were not well correlated with distribution of viral antigen. Depletion of lymphoid tissues was observed in a calf on day 13, but viral antigen had been cleared from most tissues and was detected in vascular walls only. Conclusions and Clinical Relevance-Infection with a virulent BVDV strain resulted in wide dissemination of viral antigen in host tissues. Severe lymphoid depletion developed in lymphoid tissues, whereas viral antigen was generally not associated with lesions in other tissues. Findings suggest that development of lesions in acute BVD is not solely a function of viral replication and is also attributable to host reaction to infection.

Objective-To characterize structural changes in pulmonary vessels of dogs with dirofilariosis. Animals-8 dogs with dirofilariosis and 2 unaffected control dogs. Procedure-Pulmonary artery pressure was measured in affected dogs, and dogs then were euthanatized. Scanning electron microscopy was used to examine vascular corrosion casts of pulmonary vasculature. Tissue sections of pulmonary vasculature were evaluated by use of histologic examination. Results-Pulmonary artery pressure was higher in dogs with severely affected pulmonary vessels. In tissue sections, dilatation, as well as lesions in the tunica intima and proliferative lesions resulting in constriction or obstruction, were frequently observed in branches of the pulmonary artery. Numerous dilated bronchial arteries were observed around affected pulmonary arteries. Hyperplastic venous sphincters were observed in small pulmonary veins and venules. In corrosion casts, affected pulmonary lobar arteries had dilatation, pruning, abnormal tapering, constriction, and obstruction. In small arteries and arterioles, surface structures representing aneurisms and edema were seen. Bronchial arteries were well developed and extremely dilated, and they formed numerous anastomoses with pulmonary arteries at all levels, from the pulmonary trunk to peripheral vessels. Capillaries in the lungs were dilated with little structural change. Small pulmonary veins and venules had irregular annular constrictions that were caused by hyperplastic smooth muscle cells of venous sphincters. Conclusions and Clinical Relevance-Scanning electron microscopy of microvascular casts delineated links between the bronchial and pulmonary circulations in dogs with dirofilariosis. Results of scanning electron microscopy provided a structural explanation for the development of pulmonary circulatory disturbances and pulmonary hypertension in dogs affected by dirofilariosis.

Objective-To investigate accumulation of extracellular adenosine (ADO) by equine articular chondrocytes and to compare effects of adenosine kinase inhibition and adenosine deaminase inhibition on the amount of nitric oxide (NO) produced by lipopolysaccharide (LPS)-stimulated chondrocytes. Sample Population-Articular cartilage from metacarpophalangeal and metatarsophalangeal joints of 14 horses. Procedure-Chondrocytes were cultured as monolayers, and cells were incubated with LPS, the adenosine kinase inhibitor 5'-iodotubercidin (ITU), or the adenosine deaminase inhibitor erythro-9-(2-hydroxy-3- nonyl)adenine hydrochloride (EHNA). Concentrations of ADO in cell supernatants were measured by use of reverse-phase high-performance liquid chromatography. Effect of inhibition of enzymatic metabolism of ADO on induced NO production was evaluated by exposing cells to a combination of LPS and ITU or LPS and EHNA. Results-Articular chondrocytes accumulated extracellular ADO when exposed to LPS or ITU. Chondrocytes exposed to ITU accumulated ADO in a time-dependent manner. Unstimulated chondrocytes did not accumulate ADO. Similarly, EHNA alone did not produce detectable ADO concentrations; however, addition of EHNA and ITU resulted in a synergistic effect on accumulation of ADO. Lipopolysaccharideinduced NO production was more effectively suppressed by exposure to ITU than to EHNA Conclusions and Clinical Relevance-Equine articular chondrocytes release ADO in response to the proinflammatory stimulus of bacterial LPS. Inhibition of the metabolism of ADO increases accumulation of extracellular ADO. Autocrine release of ADO from chondrocytes may play a role in the cellular response to tissue damage in arthritic conditions, and pharmacologic modulation of these pathways in joints of arthritic horses could be a potential method of therapy.

Objective-To determine immunoreactivity of matrix metalloproteinase (MMP)-1, -3, and -13 in cartilaginous tumors of dogs, correlate expression of MMP with histologic grade of tumors and clinical outcome of dogs, and compare MMP immunoreactivity between chondrosarcomas and chondromas. Sample Population-Formalin-fixed, paraffin-embedded tissues obtained from samples of naturally occurring chondrosarcomas (n = 31) and chondromas (8) of dogs that were submitted to our veterinary medical diagnostic laboratory. Procedure-Histologic sections from each sample were stained with H&E and monoclonal antibody to MMP-1, -3, and -13 by use of an avidin-peroxidase immunohistochemical technique. For each section, histologic grade (I, II, or III) and immunohistochemical expression (0, 1, 2, or 3) were evaluated. Clinical outcome was obtained from medical records or interviews with referring veterinarians and scored as a good outcome, moderate outcome, or poor outcome. Correlations among variables and differences between chondrosarcomas and chondromas were analyzed. Results-Samples from chondrosarcomas had significantly higher immunoreactivity of MMP-1 and -13, compared with immunoreactivity in samples from chondromas. In chondrosarcomas, a significant positive correlation (r, 0.386) was found between MMP-1 and -13 immunoreactivities, and a significant negative correlation (r, -0.390) was detected between MMP-3 and -13 immunoreactivities. Conclusion and Clinical Relevance-A significant increase in expression of collagenases (MMP-1 and - 13) in chondrosarcomas, compared with expression in chondromas, suggests that collagenases may play an important role in tumor progression, and possibly metastasis, in chondrosarcomas of dogs.

Objective-To evaluate molecular abnormalities in the c-kit gene of canine mast cell tumors (MCT) with different grades of cellular differentiation. Sample Population-31 normal tissue specimens from dogs and 45 canine MCT classified according to grade of cell differentiation. Procedure-Genomic DNA extractions were made from canine MCT and normal tissues. Parts of exon 11, intron 11, and exon 12 of the c-kit gene were amplified by use of polymerase chain reaction. These regions were cloned, sequenced, and compared with GenBank sequences of the National Center for Biotechnology International. A statistical analysis was used to compare sequences from canine MCT and normal tissues. Results-A significantly higher percentage of homozygous intron 11 deletion was found in canine MCT (49%) than in normal tissues (13%). This percentage was also higher in moderately and poorly differentiated MCT, compared with well-differentiated MCT. Although no mutations were detected in any of the specimens, a polymorphism at amino acid position 606 of the canine c-kit sequence was found in all the studied sequences. Conclusion and Clinical Relevance-Results indicated a relationship between intron 11 deletion and MCT, and the grade of MCT differentiation. We suggest that intron 11 deletion may be implicated in the pathogenesis of MCT and could be used as a marker for diagnosis and prognosis of canine MCT.

Objective-To compare preoperative administration of meloxicam and butorphanol to perioperative administration of butorphanol alone for control of postoperative signs of pain in dogs. Animals-40 client-owned dogs scheduled for surgical repair of a cranial cruciate ligament rupture. Procedure-Group-1 dogs received butorphanol (0.2 mg/kg, IV) and meloxicam (0.2 mg/kg, IV) just prior to surgery. Group-2 dogs received butorphanol just prior to surgery (0.2 mg/kg, IV) and at incision closure (0.1 mg/kg, IV). Pain assessment began 1 to 2 hours before surgery and from extubation until 24 hours after surgery by obtaining the following measurements: the visual analog scale (VAS) score, cumulative pain score (CPS), adjusted cumulative pain score, modified cumulative pain score, and the adjusted modified cumulative pain score (AMCPS). Serum cortisol concentration was measured between 12 to 24 and between 1 to 2 hours prior to surgery, and at 30 minutes, and 1, 2, 4, 8, 18, and 24 hours after extubation. Results-No significant differences between treatment groups were observed in CPS or VAS score. At 8, 9, 10, and 11 hours after extubation, meloxicambutorphanol- treated dogs had a significantly lower AMCPS, compared with butorphanol-alone-treated dogs. Total serum cortisol concentration (area under the curve) during the measurement period was significantly lower in meloxicam-butorphanol-treated dogs, compared with butorphanol-alone treated dogs. Conclusions and Clinical Relevance-Preoperative single dose administration of meloxicam-butorphanol is equivalent to or slightly better than the administration of 2 perioperative doses of butorphanol for the control of postoperative signs of pain in dogs.

Objective-To evaluate the usefulness of multisite quantitative ultrasonography for noninvasive assessment of bone in horses. Sample Population-12 healthy horses and both forelimbs from 8 clinically normal horses. Procedure-For in vivo measurements, various regions of interest (ROI) were examined on the third metacarpal bone, radius, and tibia. Precision error for speed of sound (SOS) measurements was obtained by measuring each ROI of 4 horses 10 times with probe repositioning. Additionally, 3 operators measured each aspect of the third metacarpal bone of 6 horses 5 times each. For ex vivo measurements, third metacarpal bones were examined at 9 ROI, and SOS measurements were performed before and after soft tissue removal. One ROI of a single forelimb was subjected to 96 ex vivo measurements with 3 different contact media. Results-The lateral aspect of the third metacarpal bone had significantly higher SOS values than the dorsal and medial aspect of the third metacarpal bone. No difference was obtained between SOS values of the lateral and medial aspect of the radius. The tibia had significantly higher SOS values than the lateral aspect of the radius and the dorsal and medial aspect of the third metacarpal bone. Intraoperator coefficients of variation ranged from 0.62 to 3.15%, and interoperator coefficients of variation ranged from 0.78 to 2.70%. Values of SOS were highest when silicone oil was used as the contact medium. Conclusions and Clinical Relevance-Speed of sound measurements obtained by quantitative ultrasonography in axial transmission mode can be used to precisely measure superficial cortical bone properties of third metacarpal bone, radius, and tibia in horses.

Objective-To determine tracheal mucociliary clearance rate (TMCCR) by use of a standard protocol in healthy anesthetized cats and to determine the effect of theophylline on TMCCR in healthy anesthetized cats. Animals-6 healthy cats. Procedure-Cats were anesthetized with propofol, and a droplet of the radiopharmaceutical technetium Tc 99m macroaggregated albumin was placed endoscopically at the carina. Dynamic acquisition scintigraphic imaging was performed, using the larynx as the end point. The TMCCR was determined by measuring the distance the droplet traveled by frame rate. Each cat was imaged 6 times as follows: 3 times following placebo administration and 3 times following the administration of sustained release theophylline (25 mg/kg, PO). Serum theophylline concentrations were assessed during imaging to ensure therapeutic concentrations. Results-The TMCCR in healthy adult cats anesthetized with propofol was 22.2 +/- 2.8 mm/min. Tracheal mucociliary clearance rate in cats receiving theophylline was 21.8 +/- 3.5 mm/min. Theophylline administration did not significantly alter TMCCR. Conclusion and Clinical Relevance-Theophylline has been shown to increase TMCCR in humans and dogs. In our study, we determined TMCCR in healthy anesthetized cats and found that it was not accelerated by the administration of theophylline.