Objective-To evaluate the origin and degree of activity of nitric oxide (NO) and matrix metalloproteinase (MMP) in explants of cranial cruciate ligaments (CCLs) obtained from dogs and cultured with and without inflammatory activators. Sample Population-Tissue specimens obtained from 7 healthy adult Beagles that were (mean +/- SD) 4.5 +/- 0.5 years old and weighed 12.5 +/- 0.8 kg. Procedure-The CCLs were harvested immediately after dogs were euthanatized, and specimens were submitted for explant culture. Cultures were stimulated by incubation with a combination of interluekin-1, tumor necrosis factor-α, and lipopolysaccharide, or they were not stimulated. Culture supernatants were examined for production of NO nitrite-nitrate metabolites (NOts) and activity of MMP. Cultured specimens were evaluated by use of immunohistochemical analysis to detect activity of inducible NO synthase (iNOS). Results-All ligament explants produced measurable amounts of NOts. Stimulated cultures produced significantly more NOts after incubation for 24 and 48 hours, compared with nonstimulated cultures. Production of MMP in supernatants after incubation for 48 hours was significantly higher in stimulated cultures than in nonstimulated cultures. Cells with positive staining for iNOS were detected on all slides. Positively stained cells were predominantly chondroid metaplastic. There was a significant difference in intensity of cell staining between stimulated and nonstimulated cultures. Conclusion and Clinical Relevance—Explant cultures of intact CCLs obtained from dogs produce iNOS-induced NO. Stimulation of chondroid metaplastic cells in CCL of dogs by use of inflammatory activators can increase production of iNOS, NOts, and MMP.

Objective-To determine whether the hyoepiglotticus muscle has respiratory-related electromyographic activity and whether electrical stimulation of this muscle changes the position and conformation of the epiglottis, thereby altering dimensions of the aditus laryngis. Animals-6 Standardbred horses. Procedure-Horses were anesthetized, and a bipolar fine-wire electrode was placed in the hyoepiglotticus muscle of each horse. Endoscopic images of the nasopharynx and larynx were recorded during electrical stimulation of the hyoepiglotticus muscle in standing, unsedated horses. Dorsoventral length and area of the aditus laryngis were measured on images obtained before and during electrical stimulation. Electromyographic activity of the hyoepiglotticus muscle and nasopharyngeal pressures were measured while horses exercised on a treadmill at 50, 75, 90, and 100% of the speed that produced maximum heart rate. Results-Electrical stimulation of the hyoepiglotticus muscle changed the shape of the epiglottis, displaced it ventrally, and significantly increased the dorsoventral length and area of the aditus laryngis. The hyoepiglotticus muscle had inspiratory activity that increased significantly with treadmill speed as a result of an increase in phasic and tonic activity. Expiratory activity of the hyoepiglotticus muscle did not change with treadmill speed in 4 of 6 horses. Conclusions and Clinical Relevance-Findings reported here suggest that contraction of the hyoepiglotticus muscle increases dimensions of the airway in horses by depressing the epiglottis ventrally during intense breathing efforts. The hyoepiglotticus muscle may be an important muscle for dilating the airway in horses, and contraction of the hyoepiglotticus muscle may induce conformational changes in the epiglottis.

Objective-To determine the prevalence of activating internal tandem duplications (ITDs) in exons 11 and 12 of c-kit in mast cell tumors (MCTs) of dogs and to correlate these mutations with prognosis. Sample Population-157 formalin-fixed, paraffinembedded MCTs from dogs in the pathology database of the Veterinary Medical Teaching Hospital at the University of California, Davis. Procedure-Genomic DNA was isolated from tumor specimens and a polymerase chain reaction procedure was performed to determine whether there were ITDs in exons 11 and 12. Results-We identified ITDs in 1 of 12 (8%) grade-I, 42 of 119 (35%) grade-II, and 9 of 26 (35%) grade-III tumors (overall prevalence, 52 of 157 [33%]). Logistic regression analysis revealed that the odds of grade-II and -III tumors possessing an ITD were approximately 5 times greater than that for grade-I tumors, although these odds did not differ significantly. Although MCTs possessing an ITD were twice as likely to recur after excision and twice as likely to result in metastasis as those without an ITD, these values also did not differ significantly. Conclusions and Clinical Relevance-These results provide evidence that ITDs in c-kit occur frequently in MCTs of dogs. The high prevalence of c-kit activating mutations in MCTs of dogs combined with the relative abundance of mast cell disease in dogs provide an ideal naturally developing tumor in which to test the safety and efficacy of novel small-molecule kinase inhibitors such as imatinib mesylate.

Objective-To examine the role of bovine viral diarrhea virus (BVDV) biotype on the establishment of fetal infection in cattle. Animals-30 mixed-breed pregnant cows. Procedure-Pregnant cows were inoculated oronasally with either i-VVNADL, originating from an infectious BVDV cDNA clone of the National Animal Disease Laboratory (NADL) isolate, or the parental virus stock, termed NADL-A. Results-All cows developed neutralizing antibodies to BVDV, and virus was commonly isolated from peripheral blood mononuclear cells or nasal swab specimens of NADL-A inoculated cows; however, virus was rarely isolated from specimens of i-VVNADL inoculated cows. i-VVNADL did not cause fetal infection, whereas all fetuses harvested from NADL-A inoculated cows at 6 weeks after inoculation had evidence of infection. Immunoblot analysis of fetal virus isolates revealed the absence of NS3, confirming a noncytopathic (NCP) biotype BVDV in the NADL-A stock. The sequence of the NCP contaminant (termed NADL-1102) and the i-VVNADL genome were virtually identical, with the exception of a 270 nucleotide-long insert in the i-VVNADL genome. Phylogenetic analyses revealed that NADL-1102 forms a monophyletic group with 6 other NADL genomes. Conclusions and Clinical Relevance-These data suggest that the contaminating NCP virus in the NADL-A stock was the ancestral NADL virus, which originally infected a bovine fetus and recombined to produce a cytopathic (CP) variant. Following oronasal infection of pregnant cows, viremia and transplacental transmission of CP BVDV to the fetus is rare, compared with the high occurrence of maternal viremia and fetal infection observed with NCP BVDV.

Objective-To develop a laparoscopic-assisted technique for cystopexy in dogs. Animals-8 healthy male dogs, 7 healthy female dogs, and 3 client-owned dogs with retroflexion of the urinary bladder secondary to perineal herniation. Procedure-Dogs were anesthetized, and positive pressure ventilation was provided. In the healthy male dogs, the serosal surface of the bladder was sutured to the abdominal wall. In the healthy female dogs, the serosa and muscular layer of the bladder were incised and sutured to the aponeurosis of the external and internal abdominal oblique muscles. Dogs were monitored daily for 30 days after surgery. Results-All dogs recovered rapidly after surgery and voided normally. In the female dogs, results of urodynamic (leak point pressure and urethral pressure profilometry) and contrast radiographic studies performed 30 days after surgery were similar to results obtained before surgery. Cystopexy was successful in all 3 client-owned dogs, but 1 of these dogs was subsequently euthanatized because of leakage from a colopexy performed at the same time as the cystopexy. Conclusion and Clinical Relevance-The laparoscopic-assisted cystopexy technique was quick, easy to perform, and not associated with urinary tract infection or abnormalities of urination.

Objective-To determine whether Haemobartonella canis and Mycoplasma haemofelis (formerly known as H felis [large form]) can be differentiated by use of comparative analysis of gene sequences. Sample Population-Blood samples obtained from 3 dogs infected with H canis and 2 cats infected with M haemofelis. Procedure-The partial 16S rDNA and ribonuclease P RNA (RNase P) genes were amplified, cloned, and sequenced in blood samples obtained from H canis-infected dogs and M haemofelis-infected cats. The DNA sequences were subjected to comparative analysis. Results-The 16S rDNA sequences of H canis and M haemofelis were nearly identical (homology of 99.3 to 99.7%). In contrast, RNase P gene sequences had a lower degree of sequence homology between the 2 organisms (94.3 to 95.5%). Conclusions and Clinical Relevance-Haemobartonella canis and M haemofelis are not identical organisms. Molecular differentiation of H canis and M haemofelis is more clearly evident by use of comparative analysis of RNase P gene sequences than by comparative analysis of 16S rDNA gene sequences.

Objective-To determine the comparative virulence of 5 isolates of bovine viral diarrhea virus (BVDV) type II by inoculating 6- to 9-month-old beef calves with isolates originating from the tissues of cattle affected with naturally occurring, transient, acute, nonfatal infections or naturally occurring, peracute, fatal infections. Animals-22 calves that were 6 to 9 months old. Procedure-The study used BVDV isolates 17011, 713, and 5521 that originated from fetuses aborted from cows with transient, nonfatal, acute BVDV infections and isolates 23025 and 17583 that originated from the tissues of cattle with peracute, fatal BVDV infections. Calves were allotted to 6 groups (1, mockinfected control calves [n = 2]; 2, inoculated with BVDV 17011 [4]; 3, inoculated with BVDV 713 [4]; 4, inoculated with BVDV 5521 [4]; 5, inoculated with BVDV 23025 [4]; and 6, inoculated with BVDV 17583 [4]). Rectal temperatures and clinical signs of disease were recorded daily. Total and differential WBC and platelet counts were performed. Histologic examination and immunohistochemical analysis were conducted to detect lesions and distribution of viral antigens, respectively. Results-Calves inoculated with BVDV 23025 or 17583 developed more severe clinical signs of disease (fever and diarrhea), more severe lymphopenia, and more severe lesions (alimentary epithelial necrosis, lymphoid depletion, and BVDV antigen deposition in lymphatic tissues), compared with calves inoculated with BVDV 713, 5521, or 17011. Conclusions and Clinical Relevance-Relative severity of experimentally induced infections corresponded to severity of clinical signs of naturally occurring infections with respective BVDV isolates.

Objective-To develop a model for measuring rotary stability of the canine elbow joint and to evaluate the relative contribution of the anconeal process (AN), lateral collateral ligament (LCL), and medial collateral ligament (MCL). Sample Population-18 forelimbs from 12 canine cadavers. Procedure-Forelimbs were allocated to 3 experimental groups (6 forelimbs/group). Each intact forelimb was placed in extension at an angle of 135° and cycled 50 times from -16° (pronation) to +28° (supination) in a continuous manner at 2.0 Hz. Cycling was repeated following sectioning of the structure of interest (group 1, AN; group 2, LCL; and group 3, MCL). Torque at -12° (pronation) and +18° (supination) was measured for each intact and experimentally sectioned limb. A Student t test was performed to compare torque values obtained from intact verses experimentally sectioned limbs and for comparison with established criteria for differentiation of primary (≥ 33%), secondary (10 to 33%), and tertiary rotational stabilizers (< 10%). Results-In pronation, the AN was the only primary stabilizer (65%). For supination, the LCL was a primary stabilizer (48%), AN was a secondary stabilizer (24%), and MCL was a tertiary stabilizer (7%). Conclusions and Clinical Relevance-With the elbow joint in extension at an angle of 135°, the AN is a primary rotational stabilizer in pronation, and the LCL is a primary stabilizer in supination. Disruption of the AN or LCL may affect rotary range of motion or compromise stability of the elbow joint in dogs.

Objective-To assess the heritability of pancreatic acinar atrophy (PAA) in German Shepherd Dogs (GSDs) in the United States. Animals-135 GSDs belonging to 2 multigenerational pedigrees. Procedure-Two multigenerational pedigrees of GSDs with family members with PAA were identified. The clinical history of each GSD enrolled in the study was recorded, and serum samples for canine trypsinlike immunoreactivity (cTLI) analysis were collected from 102 dogs. Dogs with a serum cTLI concentration ≤ 2.0 µg/L were considered to have exocrine pancreatic insufficiency (EPI) and were assumed to have PAA. Results-Pedigree I consisted of 59 dogs and pedigree II of 76 dogs. Serum cTLI concentrations were measured in 48 dogs from pedigree I and 54 dogs from pedigree II. A total of 19 dogs (14.1%) were determined to have EPI, 9 in pedigree I (15.3%) and 10 in pedigree II (13.6%). Of the 19 dogs with EPI, 8 were male and 11 were female. Conclusion and Clinical Relevance-Evaluation of data by complex segregation analysis is strongly suggestive of an autosomal recessive mode of inheritance for EPI in GSDs in the United States.

Objective-To describe neuroendocrine responses that develop in dogs subjected to prolonged periods of ventricular pacing. Animals-14 adult male hound-type dogs. Procedure-Samples were obtained and neuroendocrine responses measured before (baseline) and after 3 periods of ventricular pacing. A pacemaker was used to induce heart rates of 180, 200, and 220 beats/min (BPM). Each heart rate was maintained for 3 weeks before increasing to the next rate. Atrial natriuretic peptide, antidiuretic hormone, aldosterone, norepinephrine, epinephrine, and dopamine concentrations and plasma renin activity were measured. Severity of left ventricular compromise was estimated. Results-Shortening fraction decreased significantly with increasing heart rates (mean +/- SE, 35.5 +/- 1.4, 25.0 +/- 1.4, 19.5 +/- 1.9, and 12.2 +/- 2.3 for baseline, 180 BPM, 200 BPM, and 220 BPM, respectively). Atrial natriuretic peptide concentrations increased significantly at 180 BPM (44.1 +/- 3.0 pg/mL) and 200 BPM (54.8 +/- 5.5 pg/mL), compared with baseline concentration (36.8 +/- 2.6 pg/mL). Dopamine concentration increased significantly at 200 BPM (70.4 +/- 10.4 pg/mL), compared with baseline concentration (44.2 +/- 7.3 pg/mL). Norepinephrine concentrations increased significantly from baseline concentration (451 +/- 46.2 pg/mL) to 678 +/- 69.8, 856 +/- 99.6, and 1,003 +/- 267.6 pg/mL at 180, 200, and 220 BPM, respectively. Conclusions and Clinical Relevance-Dogs subjected to ventricular pacing for 9 weeks developed neuroendocrine responses similar to those that develop in humans with more chronic heart failure and, except for epinephrine concentrations, similar to those for dogs subjected to ventricular pacing for < 6 weeks.