Is the caudal auricular axial pattern flap robust? A multi-centre cohort study of 16 dogs and 12 cats (2005 to 2016).

OBJECTIVE: To determine the frequency and type of healing complications arising after the use of the caudal auricular axial pattern flap to close defects on the head in dogs and cats. MATERIAL AND METHODS: Multi-centre retrospective cohort study. Centres were recruited by the Association for Veterinary Soft Tissue Surgery Research Cooperative. Medical records of 11 centres were reviewed, and data from all dogs and cats treated with a caudal auricular axial pattern flap were retrieved. The following data were recorded: signalment, reason for reconstruction, flap dimensions, anatomic landmarks used, histological diagnosis, flap healing and whether revision surgery was required. RESULTS: Twenty-eight cases were included: 16 dogs and 12 cats. Flap length: width ratio was approximately 3:1 and flap length extended to the scapular spine in most cases. Optimal wound healing occurred in five of 16 (31%) dogs and six of 12 (50%) cats. Wound dehiscence without flap necrosis occurred in one of 16 (6%) dogs and one of 12 (8%) cats. Wound dehiscence with flap necrosis occurred in 10 of 16 (63%) dogs and five of 12 (42%) cats. Revision surgery was performed in eight of 16 (50%) dogs and three of 12 (25%) cats. CLINICAL SIGNIFICANCE: The caudal auricular axial pattern flap can provide full thickness skin coverage for large defects on the head in dogs and cats. Partial flap necrosis is a common complication, and revision surgery may be required in order to achieve final wound closure.

The incidence of surgical site dehiscence following full-thickness gastrointestinal biopsy in dogs and cats and associated risk factors.

OBJECTIVES: The objectives of this study were to: (1) document the incidence of surgical site dehiscence after full-thickness gastrointestinal biopsy in dogs and cats and (2) identify potential risk factors. METHODS: Data relating to dogs and cats undergoing full-thickness gastrointestinal biopsy were reviewed retrospectively following submission of a completed questionnaire by 12 referral institutions. Outcome measures were definite dehiscence, possible dehiscence (clinical records suggestive of dehiscence but not confirmed), suspected dehiscence (definite and possible combined) and death within 14 days. Logistic regression was planned for analysis of association of dehiscence with low preoperative serum albumin, biopsy through neoplastic tissue, biopsy alongside another major abdominal surgical procedure and biopsy of the colon. RESULTS: Of 172 cats, two (1·2%) had definite dehiscence, and four (2·3%) had possible dehiscence. Low preoperative serum albumin was significantly associated with definite dehiscence in univariable analysis and with suspected dehiscence and death within 14 days in univariable analysis, but all odds ratios had wide 95% confidence intervals. A histopathological diagnosis of neoplasia was significantly associated with death within 14 days in univariable analysis. Of 195 dogs, two (1·0%) had definite dehiscence, and three (1·5%) had possible dehiscence. In dogs, there was no association between any outcome measure and the putative risk factors. CLINICAL SIGNIFICANCE: Incidence of dehiscence following full-thickness gastrointestinal biopsy was low in this study. When determining the appropriateness of biopsy in individual cases, this information should be balanced against the potentially life-threatening consequences of dehiscence.

Feline talocrural luxation: a cadaveric study of repair using ligament prostheses.

Currently recommended surgical techniques to treat severe biaxial feline talocrural soft-tissue injuries commonly lead to unsatisfactory outcome. Data relating to canine talocrural stabilisation may not be useful in cats due to major differences in tarsal anatomy between the species. This experimental biomechanical cadaveric study used specimens (n = 10) prepared from the distal pelvic limbs of five adult cats. The aim was to design a technique for treating talocrural luxation using suture prostheses and bone tunnels, and to investigate its suitability for use in clinical cases. Four prosthetic ligaments were placed through a series of five 1.5 mm bone tunnels. Two prostheses, the caudoproximal pair, were taut in talocrural flexion and two prostheses, the craniodistal pair, were taut in extension. The intact specimens had their range-of-motion (ROM) and stability tested, after which they were transected at the talocrural joint (simulated luxation) and repaired using the technique described. The ROM and stability of the repaired specimens were tested and compared to the intact specimens. The repaired specimens had comparable stability to the intact specimens, although the ROM was different (p <0.05) in six of 16 positions (p <0.003125). These corresponded to the positions where the lateral prostheses were taut. The repair technique described may be useful in the treatment of talocrural luxation, as it is low-profile in an area of limited soft-tissue cover, allows anatomic reduction, restores normal talocrural joint stability and near-normal tarsal ROM.

Complications associated with temporary tracheostomy tubes in 42 dogs (1998 to 2007).

OBJECTIVE: To identify the type and frequency of tracheostomy tube complications, and to determine factors associated with these complications, and with poor outcome. METHODS: A database search for dogs undergoing temporary tube tracheostomy was performed. The medical records were scrutinised. The signalment, respiratory disease history, diagnosis, surgery, type and frequency of tube care procedures, type of complications and outcome were recorded. RESULTS: Forty-two records were found. Complications occurred in 36 of 42 (86%) cases. Bulldogs were more likely to dislodge the tube than other breeds (P=0·0376), and cases with three or more complication types underwent more routine care procedures than those with fewer complications (P=0·0370). Thirty-four of 42 (81%) dogs had a successfully managed tracheostomy tube and survived until tube removal, or elective euthanasia without significant tube complications. Bulldogs were also significantly (P=0·0376) more likely to have an unsuccessful tube outcome, as were dogs experiencing severe bradycardia during treatment (P=0·0176). Dogs with unsuccessful tube outcome were significantly (P=0·0331) younger than dogs with successful tube outcome. CLINICAL SIGNIFICANCE: Tracheostomy tubes in dogs have a high complication rate but a good outcome in most dogs. Careful management may improve the outcome of dogs with tracheostomy tubes, especially bulldogs and dogs exhibiting bradycardia during treatment.

Rapid screening for the detection of HLA-B57 and HLA-B58 in prevention of drug hypersensitivity.

HLA-B57 and HLA-B58 are major histocompatibility class (MHC)-I allotypes that are potentially predictive of important clinical immune phenotypes. HLA-B*5701 is strongly associated with hypersensitivity to the HIV drug abacavir, liver toxicity from the antibiotic flucloxacillin and is a marker for slow progression of HIV AIDS. HLA-B*5801 is associated with hypersensitivity to allopurinol used to treat hyperuricaemia and recurrent gout. Here we describe a monoclonal antibody (mAb) specific for HLA-B57 and HLA-B58 that provides an inexpensive and sensitive screen for these MHC-I allotypes. The usefulness of HLA-B57 screening for prediction of abacavir hypersensitivity was shown in three independent laboratories, including confirmation of the mAb sensitivity and specificity in a cohort of patients enrolled in the PREDICT-1 trial. Our data show that patients who test negative by mAb screening comprise 90%-95% of all individuals in most human populations and require no further human leukocyte antigen (HLA) typing. Patients who test positive by mAb screening should proceed to high-resolution typing to ascertain the presence of HLA-B*5701 or HLA-B*5801. Hence, mAb screening provides a low-cost alternative to high-resolution typing of all patients and lends itself to point-of-care diagnostics and rapid ascertainment of low-risk patients who can begin immediate therapy with abacavir, flucloxacillin or allopurinol.

Treatment of caudal mandibular fracture and temporomandibular joint fracture-luxation using a bi-gnathic encircling and retaining device.

Fractures of the caudal portion of the mandible and temporomandibular joint (TMJ) fracture-luxation can be challenging to treat with direct fixation methods. This paper describes a simple technique for the indirect treatment of caudal mandibular fracture and TMJ fracture-luxation using a subcutaneous loop of nylon leader line tunnelled around the maxilla, incisive and nasal bones, and under the mandible, placed just caudal to the canine teeth, and crimped ventral to the mandibular skin: a bignathic encircling and retaining device (BEARD). A BEARD was used to treat two immature dogs with simple, unilateral caudal mandibular fractures, six cats with unilateral injury (two with TMJ luxation, three with TMJ fracture-luxation, one with caudal mandibular fracture), and two cats with bilateral injury (comminuted caudal mandibular fracture with contralateral TMJ luxation; bilateral condylar neck fracture). The BEARD treatment failed short-term due to poor tolerance in one cat, and concurrent injuries and poor initial reduction in another cat. One cat was lost to long-term follow-up. Rostral dental occlusion was normal in six out of seven cases, and reported jaw function was normal in seven out of seven cases. The case with poor occlusion had imperfect initial reduction. Complications included dorsal nasal skin swelling or discharge, oesophagostomy tube dislodgement or blockage, BEARD loosening, and regurgitation. Treatment of uni- or bilateral caudal mandibular trauma using a BEARD can lead to clinical union, and normal rostral occlusion, provided that case selection is appropriate and immediate-post-surgical occlusion has been corrected.

Computed tomography as an aid to management of chronic oropharyngeal stick injury in the dog.

OBJECTIVE: To describe the use of computed tomography scanning in the management of dogs with chronic signs after oropharyngeal stick injury. METHODS: Dogs with a final diagnosis of chronic oropharyngeal stick injury that underwent a computed tomography scan during their investigation were selected retrospectively from case files at the Royal Veterinary College, London. RESULTS: The six dogs were young (median age 3.1 years) and medium to large breed (19.0 to 42.0 kg). By the time of referral the most common clinical sign was cervical swelling (five dogs). Stick foreign bodies were apparent on the plain computed tomography images in all cases and appeared as well-demarcated, linear abnormalities. A ventral mid-line approach was used for foreign body retrieval, and the computed tomography findings corresponded well with the surgical findings, with stick foreign body length ranging from 1 to 7 cm. Closed suction drainage was used in five dogs, for two to four days. Clinical signs fully resolved postoperatively in all cases, although cervical swelling recurred three weeks after surgery in one case. This dog had the smallest foreign body, the greatest number of surgical interventions before referral (three) and the longest disease course before referral (eight months). CLINICAL SIGNIFICANCE: Computed tomography scanning is accurate in identifying the presence and location of chronic stick foreign bodies. Recurrence of disease is possible despite successful retrieval of the wood fragments found by computed tomography scan.

Toll-like receptors.

Toll-like receptors are a family of transmembrane receptors responsible for recognition and initiation of a response to invading microbes by the immune system. As part of the innate immune system, Toll-like receptors recognise pathogen-associated molecular patterns, highly conserved components that are essential to microbial function. Some of ten toll-like receptors identified in humans are able to recognise several pathogen-associated molecular patterns.

Telomerase reverse transcriptase (TERT) expression and proliferation in canine brain tumours.

Telomerase is a ribonucleoprotein enzyme complex that synthesizes telomere DNA. It is detected in 85-90% of malignant tumours in humans, but not in most somatic cells. Because telomerase plays a critical role in cell immortality, it represents an important target for anticancer therapies. We have previously shown that the dog is a potentially useful model for evaluating telomerase-based therapeutics. In this present study we analysed 93 canine brain tumours for telomerase reverse transcriptase (TERT) expression by immunohistochemistry. TERT immunoreactivity was detected in 16 of 50 grade 1 (32%) and 29 of 43 grade 2 tumours (67.4%), demonstrating a statistically significant association with histological grade (P = 0.00012). A subset of 51 tumours was also assessed for MIB-1 expression. The MIB-1 labelling index (LI) was found to correlate significantly with tumour grade, with a mean MIB-1 LI of 1.5% for grade 1 tumours, as compared with a mean MIB-1 LI of 21.7% for grade 2 tumours (P << 0.001). The MIB-1 LI was also significantly associated with TERT expression in all brain tumours (P << 0.001). These data further support the dog as a model for the preclinical development of telomerase-based therapeutics in brain tumours.

Conservation of leukocyte cell surface proteins: implications for the generation of monoclonal antibodies against newly identified leukocyte cell surface proteins.

The availability of mouse monoclonal antibodies has been integral to the classification of human leukocyte cell surface proteins under the "Cluster of Differentiation" or "CD" nomenclature system. The sequencing of the human genome has identified many more proteins that have characteristics similar to the known leukocyte cell surface proteins, but which have not so far been identified using monoclonal antibodies. One factor that may have limited the generation of monoclonal antibodies to some of these proteins is the high level of sequence conservation between the mouse and human proteins, in particular in the extracellular regions that are recognized by most of the widely used antibodies. An alternative approach is to use a more distant species, such as chickens, for the generation of antibody reagents. Here we compare the extent of amino acid differences in the protein CD molecules expressed by human leukocytes and their mouse and chicken homologs. The analysis confirms that the human proteins are more similar to the mouse homologs than the chicken homologs. The results indicate that chicken antibodies have the potential to be used as an alternative to mouse reagents where human-mouse sequence conservation is high.

Improved antigen binding by a CD20-specific single-chain antibody fragment with a mutation in CDRH1.

We have prepared single-chain immunoglobulin Fv fragments from the CD20-specific hybridoma HB13d. One scFv clone demonstrated strong binding to a CD20-derived peptide by ELISA and to CD20-positive cells by flow cytometry, a second had reduced binding, and a third clone did not bind the target antigen. Sequence analysis showed that all three constructs contained shared and unique amino acid changes when compared to the nearest germline match. Molecular modelling of the scFv variants revealed that several of the mutations are located in regions predicted to contact antigen, including a mutation in the heavy chain CDR1 of the strongest binding scFv construct. No similar mutation is present in the highly conserved protein sequences of a number of CD20-specific monoclonal antibodies. BIACORE analysis demonstrated that the mutated scFv had approximately three-fold greater antigen-binding activity than another clone. Competition studies showed that the scFv is able to compete with intact CD20 monoclonal antibody for binding to the target antigen. The improved antigen binding of this scFv will permit the construction of novel CD20-specific reagents for the therapy of lymphomas.

In silico evaluation of two mass spectrometry-based approaches for the identification of novel human leukocyte cell-surface proteins.

The identification and quantitation of cell-surface proteins expressed by leukocytes currently use the wide availability of monoclonal antibodies (mAb) in immunohistochemical and flow cytometric assays. Presently, approximately 400 such proteins have been characterized; however, analysis of the completed human genome sequence indicates that it may contain several thousand as-yet unidentified molecules, which may be expressed on the leukocyte cell surface. Recent advances in protein isolation and analysis using mass spectrometry illustrate that it is now feasible to identify the protein composition of a complex sample such as a plasma membrane extract. Such an approach may be useful for the identification of the cell-surface proteins that have not been identified using mAb techniques. Here, we detail the results of an in silico evaluation of the peptides isolated using two methods used to label plasma membrane proteins to determine whether these methods are suitable for the identification of known leukocyte cell-surface proteins by mass spectrometry. The labeling of cell-surface proteins before isolation and characterization is a valuable means of differentiating between plasma membrane and internal membrane proteins The results indicate that although the majority of cell-surface proteins can be identified using either of the approaches, others known to be important diagnostically and/or therapeutically would not be identified using either approach. The implication of this for the use of these techniques in the discovery of new leukocyte cell-surface proteins is discussed.

Rapid simultaneous measurement of multiple cytokines using 100 microl sample volumes--association with neonatal sepsis.

Early diagnosis of neonatal infection has proved problematic due to the inadequacy of currently available laboratory tests. Neonatal sepsis is associated with an increase in plasma-derived cytokine levels, but an increase of a single cytokine cannot identify neonatal sepsis specifically and multiple cytokine levels are required. The time constraints and relatively large volume of plasma required to measure multiple cytokines from newborn infants by conventional enzyme-linked immunosorbent assay (ELISA) techniques is prohibitive. We therefore applied cytometric bead array (CBA) technology for simultaneous measurement of multiple cytokines from a group of 18 term neonates with infection confirmed by culture and a control group. 'Normal' ranges were established for each cytokine from 1-7-, 8-14- and 15-21-day-old newborns. There was no significant change in the levels of cytokines from infants in different control age groups, suggesting that basal cytokine levels are unchanged in the first 3 weeks of life. In the patient groups, however, there was a significant difference in several cytokines between the different age groups. Interleukin (IL)-6, IL-10 and IL-12 were increased significantly in the 1-7-day-old patient group compared to either the 8-14 and 15-21 age group, suggesting that infection in utero is associated with increased levels of these cytokines compared to infection acquired following birth. When individual patient cytokine levels were compared to normal control reference ranges, two patients failed to show significant elevation of any cytokine tested. All other patients showed elevated levels of between one and nine cytokines tested (mean of 4.6). There was no correlation between elevated cytokine levels and types of infective organism or patient age. In conclusion, neonatal sepsis is associated with the elevation of multiple plasma cytokines. The use of CBA kits is a rapid, easy, low sample volume and sensitive method to measure multiple plasma cytokines.

Chimeric receptors with 4-1BB signaling capacity provoke potent cytotoxicity against acute lymphoblastic leukemia.

To develop a therapy for drug-resistant B-lineage acute lymphoblastic leukemia (ALL), we transduced T lymphocytes with anti-CD19 chimeric receptors, consisting of an anti-CD19 single-chain variable domain (reactive with most ALL cases), the hinge and transmembrane domains of CD8alpha, and the signaling domain of CD3zeta. We compared the antileukemic activity mediated by a novel receptor ('anti-CD19-BB-zeta') containing the signaling domain of 4-1BB (CD137; a crucial molecule for T-cell antitumor activity) to that of a receptor lacking costimulatory molecules. Retroviral transduction produced efficient and durable receptor expression in human T cells. Lymphocytes expressing anti-CD19-BB-zeta receptors exerted powerful and specific cytotoxicity against ALL cells, which was superior to that of lymphocytes with receptors lacking 4-1BB. Anti-CD19-BB-zeta lymphocytes were remarkably effective in cocultures with bone marrow mesenchymal cells, and against leukemic cells from patients with drug-resistant ALL: as few as 1% anti-CD19-BB-zeta-transduced T cells eliminated most ALL cells within 5 days. These cells also expanded and produced interleukin-2 in response to ALL cells at much higher rates than those of lymphocytes expressing equivalent receptors lacking 4-1BB. We conclude that anti-CD19 chimeric receptors containing 4-1BB are a powerful new tool for T-cell therapy of B-lineage ALL and other CD19+ B-lymphoid malignancies.

T*1e and T*2e maps derived in vivo from the rat using longitudinally detected electron spin resonance phase imaging: application to abdominal oxygen mapping.

A novel imaging modality is introduced which uses radiofrequency longitudinally detected electron spin resonance (RF-LODESR). It is capable of providing qualitative and semiquantitative information on a variety of parameters reflecting physiological function, the most significant being tissue oxygenation. Effective spin-lattice (T*1e) and spin-spin (T*2e) electronic relaxation time maps of the abdomen of living 200-g rats were generated after intravenous administration of a triarylmethyl free radical (TAM). These maps were used to evaluate oxygen distribution. Differences between the liver, kidneys, and bladder were noted. Conclusions were made regarding the distribution, perfusion, and excretion rate of the contrast medium. Ligature-induced anoxia in the kidney was also visualized. LODESR involves transverse ESR irradiation with a modulated excitation, and observing oscillations in the spin magnetization parallel to the main magnetic field. The T*1e and T*2e maps were calculated from a set of LODESR signal phase images collected at different detection frequencies. Each phase image also provides qualitative information on tissue oxygen levels without any further processing. This method presents an alternative to the conventional transverse ESR linewidth-based oximetry methods, particularly for animal whole-body imaging applications.

Clostridial antibody response from injection-site lesions in beef cattle, long-term response to single or multiple doses, and response in newborn beef calves.

Experiments were conducted to compare clostridial antibody response of beef heifers that do and do not develop injection-site lesions, evaluate long-term antibody response of a single- and multiple-dose toxoid, and evaluate the ability of a clostridial toxoid to elicit an active antibody response in newborn calves. In Exp. 1, 37 weaned heifers were vaccinated (d 0) with a clostridial vaccine (Alpha-7, 2 mL, s.c.). Serum samples were collected on d 0, 28, 56, 84, and 112 to determine clostridial antibody titers. On d 28, heifers were visually inspected and palpated for injection-site lesions. The percentage of heifers that developed lesions was 64.9%. Lesioned heifers had elevated antibody titers for Clostridium chauvoei (CC) on d 28 (P < 0.08) and 84 (P < 0.07) compared with non-lesioned heifers. Clostridium sordellii (CS) and perfringens type D (CPD) antibody titers were greater in lesioned heifers than in non-lesioned heifers on d 28 and 56. In Exp. 2, long-term antibody response of Alpha-7 (A7) and Ultrabac 7 (UB7) was investigated in stocker heifers. The A7 heifers (n = 15) received one 2-mL vaccination (d 0), and the UB7 heifers (n = 15) received a 5-mL vaccination on d 0 and 28. Blood samples were collected on d 0, 28, 56, 84, 112, 140, and 180. Clostridium chauvoei, CPD, and Cl. novyi (CN) antibody titers from the A7 heifers were greater than those from the UB7 heifers on d 28. Due to the second UB7 injection, CC, CS, CN, and Cl. perfringens type C (CPC) antibody titers were greater in UB7 heifers than in A7 heifers on d 56. By d 112, titers were not different, and by d 140 all antibody titers were below detectable levels. In Exp. 3, 58 pregnant, mature, crossbred cows were vaccinated with A7 before calving. At birth, calves were carefully observed to ensure consumption of colostrum. Calves were blocked according to parturition date, and calves in each block were randomly allocated to receive A7 (s.c. at 3 +/- 3 d of age) or remain unvaccinated controls. Calves were bled at the time of vaccination (d 0) and on d 28, 56, 84, and 112. Antibody titers for CC, CPC, and CPD were elevated on d 0 and decreased throughout the experimental period (P < 0.01), but no titer differences (P > 0.10) were detected between treatment groups on any of the days sampled. These data indicated that antibody titers against clostridial diseases are enhanced when injection-site lesions develop. One injection of Alpha-7 seemed to provide the same length of protection as two injections of Ultrabac 7.

Radiofrequency lesions produced by handheld temperature controlled probes for use in atrial fibrillation surgery.

OBJECTIVES: Detailed analysis of the size and shape of lesions produced by handheld radiofrequency ablation devices at open heart surgery has not been reported previously. METHODS: Radiofrequency lesions were made from the epicardial surface of the cardiac ventricles in open-chested dogs. The effects of electrode size, electrode temperature and duration of ablation were studied. In a second group of experiments simultaneous multielectrode ablation was performed on the ventricular epicardium after cold cardioplegia. RESULTS: Using a single 12 x 2.5 mm electrode and a target temperature of 80 degrees C the lesion depth increased from 3.8+/-0.9 mm at 15 s, to 6.1+/-0.9 mm at 120 s (P=0.01). Increasing the target temperature from 70 to 90 degrees C (for 60 s) increased lesion depth from 5.0+/-1.2 to 5.6+/-1.7 mm (P=0.2). There was no difference in depth of lesions with the two electrode widths (4.0+/-0.5 mm (large) vs. 3.9+/-1.0 mm (small)). Lesions produced using the multielectrode probe (80 degrees C, 60 s) were 30-35 mm long with even penetration into the tissue. The mean depth of these lesions on microscopic sections was 3.9 mm. The mean width was 7.1 mm. CONCLUSIONS: Handheld probes can be used to make deep linear lesions in the myocardium. Lesions expand rapidly and are wider than they are deep. A multielectrode ablation device allows rapid formation of linear lesions.