Objective—To determine immunomodulatory effects of synbiotics administered in ovo on immune-related gene expression in adult chickens. Animals—30 Green-legged Partridgelike chickens. Procedures—On incubation day 12, eggs were injected with 3 synbiotics (Lactococcus lactis subsp lactis IBB SL1 with raffinose family oligosaccharides [RFOs; S1], Lactococcus lactis subsp cremoris IBB SC1 with RFOs [S2], and Lactobacillus acidophilus and Streptococcus faecium with lactose [S3]). Control eggs were injected with RFOs prebiotic or saline (0.9% NaCl) solution. Gene expression of 6 cytokines (interleukin [IL]-4, IL-6, IL-12p40, IL-18, interferon [IFN]-β, and IFN-γ) and 1 chemokine (IL-8) was analyzed in the cecal tonsils and spleen of 6-week-old chickens by means of reverse transcription quantitative PCR assays. Results—Gene expression for IL-4, IL-6, IFN-β, and IL-18 was significantly upregulated in the spleen of chickens in groups S2 and S3. In contrast, IL-12 expression was downregulated in group S2 and IFN-γ expression was downregulated in group S3. Expression of IL-8 did not change in chickens treated with synbiotics in ovo. Gene expression of all cytokines, except for IL-18, was downregulated in cecal tonsils. Conclusions and Clinical Relevance—In ovo administration of synbiotics activated the immune system in adult chickens. The intestinal immune system (cecal tonsils) had downregulation of expression for the cytokines evaluated, which indicated an increase in oral tolerance, whereas in the peripheral part of the immune system (spleen), expression of IL-4 and IL-6 was upregulated. Evaluation of immune-related gene expression patterns may be useful when monitoring the effectiveness of synbiotic selection with respect to immunobiotic properties.

Up to 60% of cases of equine colitis have no known cause. To improve understanding of the causes of acute colitis in horses, we hypothesized that Clostridium perfringens producing enterotoxin (CPE) and/or beta2 toxin (CPB2) are common and important causes of severe colitis in horses and/or that C. perfringens producing an as-yet-undescribed cytotoxin may also cause colitis in horses. Fecal samples from 55 horses (43 adults, 12 foals) with clinical evidence of colitis were evaluated by culture for the presence of Clostridium difficile, C. perfringens, and Salmonella. Feces were also examined by enzyme-linked immunosorbent assay (ELISA) for C. difficile A/B toxins and C. perfringens alpha toxin (CPA), beta2 toxin (CPB2), and enterotoxin (CPE). Five C. perfringens isolates per sample were genotyped for the following genes: cpa, cpb, cpb2 consensus, cpb2 atypical, cpe (enterotoxin), etx (epsilon toxin), itx (iota toxin), netB (necrotic enteritis toxin B), and tpeL (large C. perfringens cytotoxin). The supernatants of these isolates were also evaluated for toxicity for an equine cell line. All fecal samples were negative for Salmonella. Clostridium perfringens and C. difficile were isolated from 40% and 5.4% of samples, respectively. All fecal samples were negative for CPE. Clostridium perfringens CPA and CPB2 toxins were detected in 14.5% and 7.2% of fecal samples, respectively, all of which were culture-positive for C. perfringens. No isolates were cpe, etx, netB, or tpeL gene-positive. Atypical cpb2 and consensus cpb2 genes were identified in 15 (13.6%) and 4 (3.6%) of 110 isolates, respectively. All equine C. perfringens isolates showed far milder cytotoxicity effects than a CPB-producing positive control, although cpb2-positive isolates were slightly but significantly more cytotoxic than negative isolates. Based on this studied population, we were unable to confirm our hypothesis that CPE and CPB2-producing C. perfringens are common in horses with colitis in Ontario and we failed to identify cytotoxic activity in vitro in the type A isolates recovered.

The purpose of this study was to manually measure corneal thickness in canine eyes using a spectral-domain optical coherence tomography (SD-OCT) device and to assess intra- and inter-observer reliability of this technique. Twenty healthy dogs with a mean age of 4.7 y were examined. A 6-mm corneal pachymetry protocol was carried out by 1 operator using 1 SD-OCT device in both eyes of each animal. Measurements were obtained manually and in duplicate by 2 independent investigators (> 24 h apart), using the built-in caliper function. Measurements included epithelial thickness (ET), non-epithelial thickness (NET), and central corneal thickness (CCT). The overall mean ET, NET, and CCT for all eyes examined were 72.3 ± 4.6 μm, 538.9 ± 42.5 μm, and 611.2 ± 40.3 μm, respectively. There was no significant difference in ET, NET, or CCT based on the eye examined [oculus dexter (OD) versus oculus sinister (OS)], age, or gender of the animal. There was no significant difference in replicate measurements of ET, NET, or CCT done by the same operator, although a small but significant difference was noted between operators for ET measurements only. The mean difference in ET between operators was 0.6 μm (P = 0.03). The coefficient of variation ranged from 0.5% to 9.27% and intraclass correlation coefficient ranged from 0.35 to 0.97. Based on these results, manual measurements of corneal thickness in canine eyes using a portable SD-OCT device provided ET, NET, and CCT measurements with clinically acceptable intra- and inter-observer reliability.

Objective—To ultrasonographically measure the thickness of the individual wall layers of the duodenum, jejunum, and colon of dogs. Animals—85 dogs with no clinical signs or ultrasonographic evidence of gastrointestinal tract disease. Procedures—Total wall thickness and thickness of the mucosa, submucosa, muscularis, and serosa were measured ultrasonographically in the duodenum, jejunum, and colon of each dog. Results—The mucosal layer was the thickest layer of the duodenum and jejunum. There was a significant difference in thickness of the mucosal layer between small and large dogs. Mean ± SD thickness of the mucosal layer of the duodenum for small, medium, and large dogs was 2.4 ± 0.5 mm, 2.6 ± 0.6 mm, and 2.8 ± 0.5 mm, respectively. Mean ± SD thickness of the mucosal layer of the jejunum for small, medium, and large dogs was 1.8 ± 0.4 mm, 2.0 ± 0.4 mm, and 2.2 ± 0.5 mm, respectively. The remaining wall layers of the duodenum and jejunum were similar in thickness, and there were no significant differences among small, medium, and large dogs. All layers contributed equally to the total colonic wall thickness. Mean ± SD thickness of the colonic wall for small, medium, and large dogs was 1.5 ± 0.3 mm, 1.4 ± 0.5 mm, and 1.6 ± 0.4 mm, respectively. Conclusions and Clinical Relevance—Values for thickness of the wall layers of the duodenum, jejunum, and colon of dogs reported here may be useful for assessing gastrointestinal tract diseases primarily targeting a specific wall layer.

Objective- To describe the optimal thoracoscopic approach to the bovine pleural cavity and evaluate the short-term effects of thoracoscopy on cardiovascular and pulmonary function of healthy cattle. Sample- 6 healthy adult Holstein cows (12 hemithoraxes). Procedures- For each cow, thoracoscopy was performed in both the left and right hemithoraxes with a 24-hour interval between procedures. Cows were sedated and restrained in a standing position for each thoracoscopic examination. Examination of each hemithorax lasted for 30 minutes. Arterial blood gas variables, heart rate, and respiratory rate were assessed at predetermined times before, during, and after the procedures to monitor cardiovascular and pulmonary function. Thoracic ultrasonography was performed immediately and at 24 hours and 1 week after each thorascopic examination to evaluate the extent of residual pneumothorax. Results- Insertion of the laparoscope into the pleural cavity at the ninth intercostal space 15 cm ventral to the transverse processes of the thoracic vertebrae provided optimal visibility of structures in both the left and right hemithoraxes. Most structures of the pleural cavity were equally visible from both sides except the esophagus and the dorsal branch of the vagus nerve, which were best observed in the left hemithorax, and the pericardium, which was best observed in the right hemithorax. Mild increases in heart and respiratory rates and moderate decreases in arterial oxygen saturation and Pao2 were detected during the procedures. Conclusions and Clinical Relevance-Standing thoracoscopy was well tolerated in healthy adult dairy cattle and needs to be evaluated in cattle with pulmonary disease.

Objective-To evaluate the effects of sequential anesthesia of the individual compartments of the equine stifle joint on lameness induced by intra-articular deposition of interleukin (IL)-1β. Animals-6 horses. Procedures-For each horse, baseline hind limb lameness was first evaluated. A randomly selected compartment of 1 stifle joint was then injected with IL-1β to induce synovitis and lameness; subsequently, the same compartment was anesthetized with 2% mepivacaine hydrochloride, and lameness was reevaluated. Two weeks later, baseline lameness was evaluated, and lameness was similarly induced; thereafter, the 2 synovial compartments of the stifle joint not injected with IL-1β were anesthetized sequentially in random order (ie, first and second blocks); lameness was evaluated after each block. Finally, the IL-1β–treated compartment was anesthetized (third block); lameness was again evaluated. This second experiment was repeated for the contralateral stifle joint 2 weeks later. Throughout the study, lameness was quantified objectively by assessing vertical pelvic movement asymmetry with a wireless, inertial sensor-based system. Results-Intra-articular deposition of IL-1β induced lameness in all injected limbs. In the first experiment, anesthesia of the compartment injected with IL-1β resulted in a significant decrease in lameness, with vertical pelvic movement asymmetry approaching baseline. In the second experiment, lameness improved significantly after the second and third blocks and was almost completely abolished after all 3 synovial compartments were anesthetized. Conclusions and Clinical Relevance-In horses, lameness caused by a lesion in 1 compartment of a stifle joint can be improved more by instillation of local anesthetic solution into that compartment than by anesthesia of the other compartments.

Objective- To determine whether a flexible vaccination regimen provides protection against challenge exposure with a virulent Leptospira borgpetersenii serovar Hardjo isolate. Animals- Fifty-five 4-week-old calves seronegative for antibodies against L borgpetersenii serovar Hardjo. Procedures- Calves were assigned to 3 groups and administered 2 doses of adjuvant (control calves; n = 11), 1 dose of serovar Hardjo bacterin and 1 dose of adjuvant (22), or 2 doses of the serovar Hardjo bacterin (22); there was a 16-week interval between dose administrations. Three weeks after the second dose, all calves were challenge exposed by use of conjunctival instillation of a heterologous strain of L borgpetersenii serovar Hardjo for 3 consecutive days. Urine samples for leptospiral culture were collected for 5 weeks after challenge exposure; at that time, all calves were euthanized and kidney samples collected for leptospiral culture. Results- Antibody titers increased in both leptospiral-vaccinated groups of calves. A significant increase in antibody titers against L borgpetersenii serovar Hardjo was detected after administration of the second dose of L borgpetersenii serovar Hardjo bacterin and challenge exposure. In 10 of 11 adjuvant-treated control calves, serovar Hardjo was isolated from both urine and kidney samples. Leptospira borgpetersenii serovar Hardjo was not isolated from the urine or kidney samples obtained from any of the 21 remaining calves that received 1 dose of bacterin or the 20 remaining calves that received 2 doses of bacterin. Conclusions and Clinical Relevance- Protection in young calves was induced by vaccination with 1 or 2 doses of a serovar Hardjo bacterin.

Objective-To determine the safety and short-term efficacy of intrabursal administration of botulinum toxin type B (BTXB) to alleviate lameness in horses with degenerative injury to the podotrochlear apparatus (PA). Animals-10 Quarter Horses with degenerative injury to the PA. Procedures-Degenerative injury to the PA was confirmed with diagnostic analgesia and imaging. Then, BTXB (3.8 to 4.5 U/kg) was injected into the podotrochlear (navicular) bursa of each horse. Three horses were used in a safety evaluation. Subsequently, video recordings of lameness evaluations were obtained for 7 client-owned horses 5 days before (baseline) and 7 and 14 days after BTXB treatment and used to determine the effect of BTXB injection on lameness; 1 horse was removed from the study 8 days after BTXB treatment. Three investigators who were unaware of the treated forelimbs or time points separately reviewed the recordings and graded the lameness of both forelimbs of the horses. Results-Improvement in lameness of the treated forelimbs was detected at 1 or both time points after BTXB administration in all horses. However, all horses had some degree of lameness at the end of the study. Two horses developed transient increases in lameness 48 to 72 hours after treatment; lameness resolved uneventfully. Conclusions and Clinical Relevance-Intrabursal injection of BTXB temporarily alleviated chronic lameness in horses with degenerative injury to the PA, without causing serious short-term adverse effects. Further investigation into the potential use of BTXB in horses affected by degenerative injury to the PA is warranted.

Objective-To determine effects of the glycoprotein IIb/IIIa receptor antagonists abciximab and eptifibatide on in vitro inhibition of cat platelets. Sample-Venous blood samples from 10 healthy cats. Procedures-Blood samples were anticoagulated with hirudin. Aliquots of whole blood from each cat were allocated to 5 treatments (baseline, 50 μg of abciximab/mL, abciximab volumetric control treatment, 4μM eptifibatide, and eptifibatide volumetric control treatment). Impedance platelet aggregometry was performed with 6.5μM ADP or 32μM thrombin receptor activator peptide (TRAP). Magnitude of platelet aggregation was determined by measuring the area under the curve 15 minutes after addition of ADP or TRAP. Results-Eptifibatide caused a significant reduction in platelet aggregation, compared with baseline values, for aggregometry with both ADP (median, 50.0; range, 8 to 122 [baseline median, 306.0; baseline range, 130 to 664]) and TRAP (median, 75.5; range, 3 to 148 [baseline median, 219.0; baseline range, 97 to 578]). There was no significant difference in platelet aggregation with abciximab, the abciximab volumetric control treatment, or the eptifibatide volumetric control treatment for aggregometry with ADP or TRAP. Conclusions and Clinical Relevance-Eptifibatide caused a significant reduction in platelet aggregation in vitro, but there was no identifiable antiplatelet effect for abciximab. Eptifibatide and abciximab have different binding and inhibitory actions; therefore, it can be hypothesized that abciximab would be ineffective in cats because of a lack of receptor binding, reduced binding kinetics, or lack of downstream signaling. Eptifibatide may be useful in identifying hyperreactive platelets in cats in an in vitro platelet inhibitory assay.

Objective—To determine the differentiation of canine adipose tissue–derived mesenchymal stem cells (ASCs) into endothelial progenitor cells (EPCs). Animals—3 healthy adult Beagles. Procedures—Canine ASCs were isolated and cultured from adipose tissue, and endothelial differentiation was induced by culturing ASCs in differentiation medium. Morphological and immunophenotypic changes were monitored. Expression of endothelial-specific markers was analyzed by conventional and real-time RT-PCR assay. The in vitro and in vivo functional characteristics of the endothelial-like cells induced from canine ASCs were evaluated by use of an in vitro solubilized basement membrane tube assay, low-density lipoprotein uptake assay, and in vivo solubilized basement membrane plug assay. Results—After differentiation culture, the cells developed morphological changes, expressed EPC markers such as CD34 and vascular endothelial growth factor receptor 2, and revealed functional characteristics in vitro. Furthermore, the induced cells allowed vessel formation in solubilized basement membrane plugs in vivo. Conclusion and Clinical Relevance—Results indicated that canine ASCs developed EPC characteristics when stimulated by differentiation medium with growth factors. Thus, differentiated canine ASC-EPCs may have the potential to provide vascularization for constructs used in regenerative medicine strategies.