Dr Charlotte Maile

A glutamic acid to lysine (E40K) residue substitution in superoxide dismutase 1 (SOD1) is associated with canine degenerative myelopathy: the only naturally occurring large animal model of amyotrophic lateral sclerosis (ALS). The E40 residue is highly conserved across mammals, except the horse, which naturally carries the (dog mutant) K40 residue. Here we hypothesized that in vitro expression of mutant dog SOD1 would recapitulate features of human ALS (ie, SOD1 protein aggregation, reduced cell viability, perturbations in mitochondrial morphology and membrane potential, reduced ATP production, and increased superoxide ion levels); further, we hypothesized that an equivalent equine SOD1 variant would share similar perturbations in vitro, thereby explain horses' susceptibility to certain neurodegenerative diseases. As in human ALS, expression of mutant dog SOD1 was associated with statistically significant increased aggregate formation, raised superoxide levels (ROS), and altered mitochondrial morphology (increased branching (form factor)), when compared to wild-type dog SOD1-expressing cells. Similar deficits were not detected in cells expressing the equivalent horse SOD1 variant. Our data helps explain the ALS-associated cellular phenotype of dogs expressing the mutant SOD1 protein and reveals that species-specific sequence conservation does not necessarily predict pathogenicity. The work improves understanding of the etiopathogenesis of canine degenerative myelopathy.

Reasons for performing study: Muscle biopsy is increasingly used in equine veterinary practice for investigating exertional, inflammatory or immune mediated myopathies and unexplained muscle atrophy. Although formalin-fixed samples are often used, for complete evaluation, fresh-frozen tissue is required. Freezing muscle in veterinary practice is impractical: samples sent to specialist laboratories for processing are therefore susceptible to delays, potentially leading to artefact and compromising histological interpretation. Hypothesis: Altered temperature, duration and hydration status influence the severity of storage-induced artefact in equine muscle. Methods: Skeletal muscle obtained immediately post euthanasia was divided into 6 independent samples from each of 8 horses. One sample per horse was frozen immediately in isopentane precooled in liquid nitrogen. Additional samples were stored in conditions designed to mimic possible situations encountered in practice, including increased storage times, temperature and hydration status. Following storage, stored samples were frozen as before. Cryosections were stained using haematoxylin and eosin and ranked for artefact on 2 occasions by 2 blinded observers. The best samples were processed subsequently with a panel of routine stains and immunolabelled for collagen V to enable the measurement of minimum fibre diameters. Results: Both prolonged storage and increased hydration resulted in more storagc-associated artefact. Samples stored for 24 h chilled on dry gauze were ranked higher than those stored on damp gauze; however, a panel of routinely-used histochemical staining techniques was unaffected by chilled 24 h storage. There was no significant effect of storage on mean fibre diameter; however, both chilled dry and damp storage for 24 h caused a significant increase in fibre-size variability. Conclusion and potential relevance: Caution should be exercised when interpreting fibre size profiles in shipped samples. Equine muscle biopsy samples are optimally shipped in dry gauze, sealed in plastic containers and shipped on ice packs to be processed within 24 h and can thus be interpreted by the receiving laboratory with minimal artefact.

Background: Equine type 1 polysaccharide storage myopathy (PSSM1) is associated with a missense mutation (R309H) in the glycogen synthase (GYS1) gene, enhanced glycogen synthase (GS) activity and excessive glycogen and amylopectate inclusions in muscle. Methods: Equine muscle biochemical and recombinant enzyme kinetic assays in vitro and homology modelling in silico, were used to investigate the hypothesis that higher GS activity in affected horse muscle is caused by higher GS expression, dysregulation, or constitutive activation via a conformational change. Results: PSSM1-affected horse muscle had significantly higher glycogen content than control horse muscle despite no difference in GS expression. GS activity was significantly higher in muscle from homozygous mutants than from heterozygote and control horses, in the absence and presence of the allosteric regulator, glucose 6 phosphate (G6P). Muscle from homozygous mutant horses also had significantly increased GS phosphorylation at sites 2+2a and significantly higher AMPK alpha 1 (an upstream kinase) expression than controls, likely reflecting a physiological attempt to reduce GS enzyme activity. Recombinant mutant GS was highly active with a considerably lower K-m for UDP-glucose, in the presence and absence of G6P, when compared to wild type GS, and despite its phosphorylation. Conclusions: Elevated activity of the mutant enzyme is associated with ineffective regulation via phosphorylation rendering it constitutively active. Modelling suggested that the mutation disrupts a salt bridge that normally stabilises the basal state, shifting the equilibrium to the enzyme's active state. General significance: This study explains the gain of function pathogenesis in this highly prevalent polyglucosan myopathy. (C) 2016 Elsevier B.V. All rights reserved.

The use of clinical skills models is now commonplace in veterinary education with the aim of improving proficiency and competency when subsequently performing clinical procedures on patients. However, it is important to evaluate the construct and content validity of the models and protocols being used to replace live animal teaching. Performing in-depth validation studies takes considerable time and resources which may not be readily available in an educational setting. This study describes a fast and effective method using expert feedback to evaluate the validity of clinical skills models and their associated protocols used in veterinary teaching. Thirty skills used in the teaching of undergraduate veterinary students at the University of Surrey were evaluated, ten from each of the core species (companion animal, equine and production animal). Qualified veterinary surgeons with experience of performing each skill were invited to read through the protocol and perform the skills. They were then asked to provide anonymous ratings, using a 5-point Likert scale, regarding: the realism of the model, the suitability of the protocol and the suitability of the model and protocol to prepare students to perform the skill in clinical practice. Results showed that 80% of respondents agreed that the performing the skill was realistic compared to the live animal for 63.3% of skills, that the written protocol is appropriate for performing this skill for 96.7% of skills and or that the model and protocol are suitable to prepare students to perform the skill in clinical practice for 76.7% of skills. This study presents an innovative approach to high throughput clinical skills teaching validation.

The equine veterinary profession is frequently considered to be inaccessible to veterinary students that have had limited exposure to the equine industry. One of the perceived reasons for this inaccessibility is the use of what is often coined ‘horsey’ language within the industry. If a student has limited experience of the equine industry, there is a potential language barrier to teaching which may result in poor engagement with equine teaching and placements. Students that otherwise might consider a career in equine veterinary medicine are discouraged by an apparent barrier in communication between horse owners, equine vets and themselves. This project aimed to develop a new teaching resource to increase student engagement with the equine content of the veterinary programme at the University of Surrey.An online anonymous survey was shared via social media and was used to generate a list of terms that equine veterinary surgeons, horse owners and paraprofessionals believed essential for a new graduate veterinary surgeon to understand in the equine industry. The survey design was based on the ‘Twenty Statements Test’ where in this case participants were asked for up to ten equine specific jargon terms. University of Surrey equine academics provided definitions of each of the terms. Definitions were derived from a variety of sources including textbooks and equestrian websites. Additional jargon terms were added to the resource when the equine academics identified incomplete topics to ensure students received a comprehensive list of terms. The list of terms and their definitions were then developed into an e-book glossary resource for students to be able to download and access offline to ensure access when on placement in remote locations. The eBook was presented to students during a communication session for fourth year veterinary students at the University of Surrey. Students were surveyed on the usefulness of the resource and whether the resource improved their confidence in using and understanding equine industry specific language.The data collected in this study provides evidence that language is a barrier to learning and that with the development of an easily navigable eBook resource, this barrier can be reduced. By increasing student engagement and enabling students to feel less like outsiders during clinical placements, this teaching resource could have a direct effect on the proportion of graduates considering a career in equine practice and thereby go towards helping with the current recruitment and retention crisis within the field of equine veterinary medicine.