The present study investigated whether the 14-3-3 η and γ proteins, which are potent matrix metalloprotease (MMP) stimulators, are detectable in the synovial fluid of dogs with cranial cruciate ligament rupture (CCLR). Synovial fluid samples from 7 dogs with unilateral CCLR and control samples from 4 dogs without a history of any joint inflammation or any other abnormalities underwent Western blot analysis for the 14-3-3 η, γ, and σ proteins as well as MMP-1 and MMP-3. Craniocaudal and lateral radiographic projections of the stifle joint were evaluated for the presence and severity of 13 specific radiographic markers of osteoarthritis and graded numerically. The Spearman method was used to detect any correlation between the 14-3-3-η level in the synovial fluid and the radiograph-based grade. The η isoform was present only in the samples from the dogs with CCLR. The levels of 14-3-3-γ, MMP-1, and MMP-3 were significantly higher in the samples from the dogs with CCLR than in the control samples (P < 0.05). However, there was no significant difference between the CCLR and control samples in the level of the σ isoform. The Spearman method showed a significant correlation between the 14-3-3-η level in the synovial fluid and the presence of either patellar osteophytes or lateral or medial (or both) condylar periarticular osteophytes (P < 0.05). The MMP stimulatory effect of the 14-3-3 η and γ isoforms may be the reason for the high levels of MMP-1 and MMP-3 observed. Thus, 14-3-3 proteins, especially the η isoform, may be important markers of osteoarthritis caused by CCLR.

The objective of this study was to experimentally infect calves with bovine viral diarrhea virus (BVDV) isolated from free-ranging white-tailed deer. Twelve colostrum-deprived male Holstein calves were used. Eight were inoculated intranasally with a BVDV type 1a isolated from free-ranging white-tailed deer, and the other four were inoculated with the cell culture medium only and served as a control group. Whole blood, saliva, and nasal and rectal secretions were collected on days 0, 3, 7, 10, 14, 17, and 21 after inoculation for virus isolation and real-time reverse-transcriptase polymerase chain reaction (RT-PCR). On days 14 and 21, 4 calves in the infected group and 2 in the control group were euthanized; multiple tissue samples were collected for histopathologic study. Histopathologic changes included thymic atrophy and lymphoid depletion of the Peyer's patches in all 8 infected calves. The RT-PCR gave positive results with the buffy coat of all 8 infected calves, the nasal samples of 7, and the saliva samples of 2. Virus neutralization testing of the serum gave positive results for 4 of the 8 infected calves, and enzyme-linked immunosorbent assay of the serum gave positive results for 3. All of the samples from the control calves yielded negative results.

Tissues unsuitable for standard immunohistochemical and histopathological examinations for chronic wasting disease (CWD) in cervids and for scrapie in sheep are frequently submitted for testing. This study investigated the effects of experimental autolysis on the detection of abnormal prion protein (PrPsc) in lymphoid and central nervous system (CNS) tissues from elk and sheep. The PrPsc was detected using a Western blotting (WB) test following PrPsc enrichment using sodium phosphotungstic acid (PTA) precipitation (PTA-WB). A commercial enzyme-linked immunosorbent assay (ELISA) was used as a reference test for quantitative measurement. This study showed that the amount of PrPsc in lymphoid and CNS tssues from elk and sheep decreased gradually as a result of autolysis, but PrPsc was still detectable after 5 and 15 d incubation at 37°C by PTA-WB for all lymphoid and CNS samples. The results of the ELISA supported those of PTA-WB, particularly for CNS tissues. In conclusion, autolysis at 37°C for 15 d would not significantly affect the detection of PrPsc in lymphoid and CNS tissues by WB and ELISA and, particularly, PTA-WB is a valuable and alternative confirmatory test to detect PrPsc in autolyzed lymphoid and CNS samples.

Objective—To determine whether trilostane or ketotrilostane is more potent in dogs and determine the trilostane and ketotrilostane concentrations that inhibit adrenal gland cortisol, corticosterone, and aldosterone secretion by 50%. Sample—24 adrenal glands from 18 mixed-breed dogs. Procedures—Adrenal gland tissues were sliced, placed in tissue culture, and stimulated with 100 pg of ACTH/mL alone or with 5 concentrations of trilostane or ketotrilostane. Trials were performed independently 4 times. In each trial, 6 samples (1 for each time point) were collected for each of the 5 concentrations of trilostane and ketotrilostane tested as well as a single negative control samples. At the end of 0, 1, 2, 3, 5, and 7 hours, tubes were harvested and media and tissue slices were assayed for cortisol, corticosterone, aldosterone, and potassium concentrations. Data were analyzed via pharmacodynamic modeling. One adrenal slice exposed to each concentration of trilostane or ketotrilostane was submitted for histologic examination to assess tissue viability. Results—Ketotrilostane was 4.9 and 2.4 times as potent in inhibiting cortisol and corticosterone secretion, respectively, as its parent compound trilostane. For trilostane and ketotrilostane, the concentrations that inhibited secretion of cortisol or corticosterone secretion by 50% were 480 and 98.4 ng/mL, respectively, and 95.0 and 39.6 ng/mL, respectively. Conclusions and Clinical Relevance—Ketotrilostane was more potent than trilostane with respect to inhibition of cortisol and corticosterone secretion. The data should be useful in developing future studies to evaluate in vivo serum concentrations of trilostane and ketotrilostane for efficacy in the treatment of hyperadrenocorticism.

Objective—To develop an antibody-based flow cytometric assay to detect coated platelets in dogs and to characterize the interaction of recombinant human coagulation factor VIIa with activated platelets from dogs with hemophilia A. Sample—Platelets from 4 dogs with hemophilia A, 4 dogs with hemophilia B, 4 dogs with von Willebrand disease, and 6 hemostatically normal dogs. Procedures—Freshly isolated platelets were activated with thrombin, convulxin, or a thrombin-convulxin combination. Resulting platelet phenotypes were resolved on the basis of P-selectin and fibrinogen expression, and binding of recombinant human coagulation factor VIIa to these distinct platelet subpopulations was measured by use of a flow cytometric assay. Results—Coated platelets were identified on the basis of expression of α-granule fibrinogen and were generated in response to stimulation with the thrombin-convulxin combination but not to stimulation with either agonist alone. Approximately 70% of the platelets from dogs with hemophilia A, hemophilia B, and von Willebrand disease and from the control dogs had the coated platelet phenotype. Recombinant human coagulation factor VIIa bound preferentially to coated platelets with a mean ± SD binding equilibrium constant of 2.6 ± 0.5μM. Conclusions and Clinical Relevance—Formation of coated platelets in dogs was similar to that in humans. Recombinant human coagulation factor VIIa bound preferentially to coated platelets from dogs. Impact for Human Medicine—A similar mechanism of action for recombinant human coagulation factor VIIa may exist in dogs and humans. The potential for use of dogs in the study of bleeding disorders in humans was strengthened.

Objective—To estimate the risk of subclinical Mycobacterium avium subsp paratuberculosis (MAP) infection in cows that ingested MAP DNA–positive raw colostrum as calves, compared with risk in cows that ingested MAP DNA–negative raw colostrum as calves. Animals—205 calves born in 12 commercial dairy herds. Procedures—Each calf was separated from its dam within 30 to 60 minutes after birth and fed raw colostrum. For each calf, samples of the colostrum fed were collected and tested for the presence of MAP DNA by use of a nested PCR assay for the target gene ISMAP02. Calves fed colostrum positive or negative for MAP DNA were classified into exposed (n = 69) and unexposed (136) groups, respectively. Each calf was tested for MAP infection at 30, 42, and 54 months of age by use of a serum ELISA and bacterial culture of feces. Weibull hazard regression models were used to evaluate the association between exposure to MAP DNA–positive colostrum and time to testing positive for MAP infection. Results—Hazard of MAP infection was not different between groups (exposed vs unexposed) when serum ELISA, bacterial culture of feces, or both diagnostic tests (parallel interpretation) were positive. Conclusions and Clinical Relevance—Heifer calves fed MAP DNA–positive colostrum were at no greater risk of MAP infection, compared with heifer calves fed MAP DNA–negative colostrum. This result contradicts findings from other studies and should be interpreted with caution.

Objective: To determine repeatability of a wireless, inertial sensor–based lameness evaluation system in horses. Animals: 236 horses. Procedures: Horses were from 2 to 29 years of age and of various breeds and lameness disposition. All horses were instrumented with a wireless, inertial sensor-based motion analysis system on the head (accelerometer), pelvis (midline croup region [accelerometer]), and right forelimb (gyroscope) before evaluation in 2 consecutive trials, approximately 5 minutes apart, as the horse was trotted in a straight line. Signal-processing algorithms generated overall trial asymmetry measures for vertical head and pelvic movement and stride-by-stride differences in head and pelvic maximum and minimum positions between right and left sides of each stride. Repeatability was determined, and trial difference was determined for groups of horses with various numbers of strides for which data were collected per trial. Results: Inertial sensor–based measures of torso movement asymmetry were repeatable. Repeatability for measures of torso asymmetry for determination of hind limb lameness was slightly greater than that for forelimb lameness. Collecting large numbers of strides degraded stride-to-stride repeatability but did not degrade intertrial repeatability. Conclusions and Clinical Relevance: The inertial sensor system used to measure asymmetry of head and pelvic movement as an aid in the detection and evaluation of lameness in horses trotting in a straight line was sufficiently repeatable to investigate for clinical use.

Objective—To evaluate the diagnostic value of plasma N-terminal pro-B-type natriuretic peptide (NT-proBNP) concentrations in Doberman Pinschers in various stages of dilated cardiomyopathy (DCM). Animals—328 Doberman Pinschers. Procedures—Staging of DCM was determined via analysis of results of physical examinations, 24-hour ambulatory ECG (Holter) recordings, and echocardiographic evaluations. Plasma samples for NT-proBNP assays were obtained at each examination. Concentrations of NT-proBNP were measured in 337 samples obtained from 196 healthy Doberman Pinschers (control dogs) and in 195 samples obtained from 132 Doberman Pinschers in various stages of DCM. These included dogs that had ventricular premature contractions (VPCs; 79 samples), echocardiographic changes (23 samples), or both (51 samples); 16 samples were from dogs with overt DCM, and 26 were from dogs that were considered normal during initial examination but developed DCM within 1.5 years after this assessment. Receiver operating characteristic curves were analyzed to determine sensitivity and specificity of NT-proBNP concentrations for detection of DCM. Results—NT-proBNP concentrations in dogs that had or developed DCM were significantly higher than those of control dogs. Sensitivity and specificity of NT-proBNP concentrations (cutoff value, > 400 pmol/L) to detect all stages of DCM were 81.1 % and 75.0%, respectively; sensitivity was 90.0% and specificity was 75.0% to predict echocardiographic changes. Specificity to detect echocardiographic changes was 90.4% at a cutoff value of 550 pmol/L. Conclusions and Clinical Relevance—Plasma concentrations of NT-proBNP were increased in dogs with DCM and in apparently healthy dogs that developed DCM within 1.5 years after samples were obtained, compared with concentrations in control dogs.

Objective—To determine effects of duration and type of anesthetic on tear production in dogs. Animals—8 female Beagles. Procedures—Each dog was randomly allocated into 1 of 4 groups according to a Latin square design to receive anesthesia as follows: 1 hour with isoflurane, 1 hour with desflurane, 4 hours with isoflurane, and 4 hours with desflurane. Each dog was anesthetized with the selected inhalant 4 times during a 4-week period, with at least 5 days separating anesthetic episodes. Aqueous tear production was measured via the Schirmer I tear test at baseline and 10 minutes, 30 minutes, and 1 hour after induction of anesthesia as well as 2, 3, and 4 hours after induction for the 4-hour groups. Tear production was also measured after the dogs were standing after recovery from anesthesia and 2, 10, and 22 hours after recovery from anesthesia. Results—Aqueous tear production was significantly reduced in dogs during anesthesia and returned to baseline values immediately after recovery and until 10 hours after anesthesia in all treatment groups. Inhalant type and duration had no significant effect. Neither lateral recumbency nor left versus right eyes had a significant effect. Conclusions and Clinical Relevance—Results suggested that inhalant anesthetics did not reduce tear production after anesthesia and that longer-duration anesthesia did not cause decreased tear production, compared with shorter-duration anesthesia.

Objective—To determine the mode of inheritance for cerebellar abiotrophy (CA), a neurologic disease in Arabians. Animals—804 Arabians, including 29 horses (15 males and 14 females) with CA. Procedures—Most horses (n = 755) belonged to 1 of 4 paternal families. Among the 29 CA-affected horses, all had clinical signs consistent with the disease; the disease was confirmed histologically following euthanasia in 8 horses. From the pedigree information, inbreeding coefficients were calculated for 16 affected horses and compared with coefficients for a subgroup of 16 unaffected horses. Complex segregation analysis was used to determine the effect of a putative Mendelian locus on the development of the disease and the probable mode of inheritance of CA. Results—The mean inbreeding coefficient was 0.0871 for CA-affected and unaffected horses, suggesting that all of the Arabians were inbred to the same degree and that affected horses were not more inbred than were unaffected horses. Results of the complex segregation analysis were consistent with a single Mendelian autosomal recessive mode of inheritance. Conclusions and Clinical Relevance—Knowledge of the mode of inheritance of CA should help breeders to make informed decisions regarding the selection of animals for mating when closely related horses have developed CA or produced CA-affected foals.