Dystrophin-deficient muscular dystrophy in an old English sheepdog

MUSCULAR dystrophies are a heterogeneous group of inherited, degenerative, mostly non-inflammatory disorders characterised by progressive muscle weakness and wasting (Braund 1997, Shelton and Engvall 2002). In companion animals, dystrophies associated with an absence of or abnormality in dystrophin, dystrophin-associated proteins and laminin α2 have been recognised (Cooper and others 1988, O’Brien and others 2001, Shelton and others 2001). Dystrophin-deficient muscular dystrophy, resulting from mutations in the dystrophin gene, is the most common and best studied form in dogs, and is known to be X-linked (Shelton and Engvall 2002). It has been reported in several breeds, including the golden retriever (Kornegay and others 1988, Sharp and others 1992), rottweiler (Winand and others 1994), German shorthaired pointer (Schatzberg and others 1999), Irish terrier (Wentink and others 1972), Belgian Groenendaeler shepherd dog (Van Ham and others 1993), samoyed (Presthus and Nordstoga 1993), miniature schnauzer (Paola and others 1993), Brittany spaniel (Van Ham and others 1995), rat terrier (Wetterman and others 2000), Pembroke Welsh corgi (Woods and others 1998), labrador retriever (Bergman and others 2002) and Japanese spitz (Jones and others 2004). Specific mutations have been identified for the golden retriever (Sharp and others 1992), rottweiler (Winand and others 1994) and German shorthaired pointer (Schatzberg and others 1999). This short communication describes, to the authors’ knowledge, the first case of dystrophin-deficient muscular deficiency in an old English sheepdog. A three-month-old, male old English sheepdog was presented to the neurology/neurosurgery department of the Animal Health Trust, for evaluation of a three-week history of difficulty in eating, excessive salivation, mild exercise intolerance and impaired growth. The littermates and parents were apparently unaffected. The animal weighed 8·2 kg; physical examination confirmed a poor body condition, with atrophy of the temporalis, paraspinal and distal limb muscles. Marked swelling of the tongue and pharyngeal region was evident, accompanied by excessive salivation. Neurological findings included a stiff gait with an increased muscle tone of both pelvic limbs, decreased to absent withdrawal reflexes on all four limbs, decreased tongue movement and absent gag reflex. A generalised myopathic disorder was suspected, with an inflammatory, infectious, dystrophic or other congenital myopathy as the primary differential diagnoses; the possibility of a neuropathy was also considered. The results of a complete blood count and serum biochemistry revealed a marked elevation of serum creatine kinase (CK) (53,396 iu/l, reference range 21 to 56 iu/l), suggesting muscle disease. Serum antibody titres for Toxoplasma gondii and Neospora caninum were negative. Electrodiagnostic testing was performed under general inhalational anaesthesia. Concentric needle electromyography of the major muscle groups of the foreand hindlimbs and paraspinal, tongue and laryngeal muscles, as well as the temporalis and masseter muscles, revealed fibrillation potentials, positive sharp waves and complex repetitive discharges. Cardiac ultrasound was normal, as were thoracic and abdominal radiographs. A barium swallow revealed poor laryngeal motility. Muscle biopsies were taken from the right cranial tibial, right temporalis and left myohyoideus muscles, and unfixed and fixed (immersed in 10 per cent buffered formalin) biopsies from each site were submitted under refrigeration to the Comparative Neuromuscular Laboratory, University of California, San Diego, USA, by a courier service. Upon receipt, the fixed muscle specimens were embedded in paraffin, and the unfixed, refrigerated specimens were flash frozen in isopentane precooled in liquid nitrogen and stored at –80°C before further processing. Cross-sections 8 μm thick were cut using a cryostat, and stained and reacted with a standard panel of histological and histochemical stains and enzyme reactions, including the general stains haematoxylin and eosin and modified Gomori trichrome for morphological abnormalities, periodic acid-Schiff for glycogen, myofibrillar ATPases at pH 9·8 and 4·3 for fibre typing, esterase for localisation of motor endplates and macrophage activity, nicotinamide adenine dinucleotide dehydrogenase for oxidative activity, acid phosphatase for macrophages, oil red O for neutral triglycerides, and the alizarin and von Kossa stains for calcium (Dubowitz 1985, Dickinson and LeCouteur 2002). Degeneration and regeneration were the predominant pathological changes, including variability in myofibre size, with large groups of necrotic fibres, and clusters of small basophilic regenerating fibres (Figs 1a, b, c). Multifocal areas of myofibre mineralisation were present with both the alizarin and von Kossa stains for calcium (Fig 1d). Cellular infiltrates were limited to macrophages within necrotic fibres, without overt lymphocytic infiltration. Fibrosis was not a feature. A non-inflammatory myopathy with a distinctly dystrophic phenotype was diagnosed based on the pathological changes. Immunohistochemical staining was performed on frozen muscle specimens from the affected dog and a control dog (a young adult dog that had been euthanased because of trauma, Veterinary Record (2006) 158, 270-273