Evaluation of the dystrophin±glycoprotein complex, a-actinin, dysferlin and calpain 3 in an autosomal recessive muscular dystrophy in Labrador retrievers Natasha J. Olby a, * , Nick J.H. Sharp a , Louise V.B. Anderson b , Louis M. Kunkel c , Carsten G. Bo Ènnemann c,1 a Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606, USA b Neurobiology Department, University Medical School, Newcastle upon Tyne NE2 4HH, UK c Division of Genetics and Department of Neurology, The Children's Hospital, Enders 570, Boston, MA 02115, USA Received 26 November 1999; received in revised form 12 May 2000; accepted 2 June 2000 Abstract Labrador retrievers suffer from an autosomal recessive muscular dystrophy of unknown aetiology. Dogs affected with this disease develop generalized weakness associated with severe, generalized skeletal muscle atrophy and mild elevations in creatine kinase in the ®rst few months of life. The severity of signs tends to progress over the ®rst year of life but can vary from mild exercise intolerance to non-ambulatory tetraparesis. Beyond 1 year of age, the signs usually stabilize and although muscle mass does not increase, affected dogs' strength may improve slightly. The pathological changes present on muscle biopsy include marked variation in muscle ®bre size with hypertrophied and round atrophied ®bres present. There is an increased number of ®bres with central nuclei and split ®bres can be seen. It has been suggested that the disorder is a model for limb-girdle muscular dystrophy. In recent years, mutations in genes encoding the proteolytic enzyme, calpain 3, a novel protein named dysferlin, and components of the dystrophin-glycoprotein complex have been identi®ed as causes of autosomal recessive limb-girdle muscular dystrophy. We have evaluated these proteins in normal dogs and in three Labrador retrievers with autosomal recessive muscular dystrophy using immunohistochemistry and Western blot analysis on frozen skeletal muscle. The results demonstrate that dystrophin, the sarcoglycans, a-actinin, dysferlin and calpain 3 are present in the normal and affected dogs. We conclude that this autosomal recessive muscular dystrophy is not due to a de®ciency of a-actinin, or any of the known autosomal recessive limb-girdle muscular dystrophy proteins, although we cannot rule out a malfunction of any of these proteins. q 2001 Elsevier Science B.V. All rights reserved. Keywords: Autosomal recessive muscular dystrophy in Labrador retrievers; Limb-girdle muscular dystrophy; Congenital muscular dystrophy; Sarcoglycan; Merosin; Calpain 3; Dysferlin 1. Introduction Autosomal recessive muscular dystrophy in Labrador retrievers is a clinically and histopathologically variable disorder that affects skeletal muscle, causing exercise intol- erance in young Labrador retrievers. Cardiac muscle is not affected [1]. It is inherited by a monogenic autosomal reces- sive trait [2] and has been reported world wide [3,4], but is particularly prevalent in certain strains of working Labrador in the United Kingdom [1,4]. In the veterinary literature it has been given various names including type 2 muscle ®bre de®ciency, muscular dystrophy and more recently Labrador retriever hereditary myopathy [2]. The clinical [1] and histo- pathological [4,5] features of the disease have been described in detail but as yet the underlying defect is unknown. Neuromuscular Disorders 11 (2001) 41±49 0960-8966/01/$ - see front matter q 2001 Elsevier Science B.V. All rights reserved. PII: S0960-8966(00)00166-8 www.elsevier.com/locate/nmd Table 1 Known mutations associated with autosomal recessive limb-girdle muscu- lar dystrophy (LGMD) and congenital muscular dystrophy (CMD) MD Gene location Protein LGMD type 2A [14] 15q Calpain 3 LGMD type 2B [15] 2p Dysferlin LGMD type 2C [16] 13q12 g-Sarcoglycan LGMD type 2D [17] 17q21 a-Sarcoglycan LGMD type 2E [18,19] 4q12 b-Sarcoglycan LGMD type 2F [20] 5q33 d-Sarcoglycan LGMD type 2G [12] 17q11±12 Unknown LGMD type 2H [13] 9q31±34.1 Unknown CMD: merosin de®cient [21,22] 6q2 Laminin a-2 chain * Corresponding author. 1 Present address: Abt. Neuropa Èdiatrie, Georg-August-Universita Èt Go Èttingen, 37075 Go Èttingen, Germany.