P.P.5 11 Autosomal dominant congenital fibre type disproportion. Study of a family

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to mild weakness in adulthood. The severity of the phenotype may be related to a skewed pattern of X inactivation. Here we report on a 77year-old woman belonging to a pedigree without miscarriages or early deaths in males. She had a progressive ptosis since childhood, proximal limb weakness, hemidiaphragmatic paresis and a severe restrictive respiratory dysfunction leading to death at age 84 years. The muscle biopsy showed a large predominance of type I fibres, centrally nucleated fibres and some mitochondrial abnormalities. A novel stop mutation Leu498X was identified in MTM1 gene. Two heteroplasmic deletions were detected by Southern blot in the muscle mitochondrial DNA. There was no skewed pattern of X-inactivation in her blood DNA. The present study illustrates three aspects of importance: (1) the investigation of the MTM1 gene is indicated in females, even if very old and isolated cases, when histopathological findings show muscle fibers that retain an immature phenotype; (2) survival is possible until the 80s in a female carrier, despite the deleterious functional effect of the MTM1 mutation; (3) a random pattern of X-inactivation is present in females carriers with an intermediate phenotype. doi:10.1016/j.nmd.2006.05.153

P.P.5 09 A new combined phenotype of congenital myopathy, hypohydrotic ectodermal dysplasia and immunodeficiency in two brothers J. Kirschner 1,*, U. Osmers 2,3, A. Rao 2,3, S. Ehl 4, S. Feske 2,5 1 University of Freiburg, Dept. of Neuropediatrics and Muscle Disorders, Freiburg, Germany; 2 CBR Institute of Biomedical Research, Boston, MA, USA; 3 Harvard Medical School, Dept. of Pathology, Boston, MA, USA; 4 University of Freiburg, Dept. of Pediatrics and Adolescent Medicine, Freiburg, Germany; 5 Harvard Medical School, Department of Pediatrics, Boston, MA, USA Hypohydrotic ectodermal dysplasia (HED) is a congenital disorder of teeth, hair, and eccrine sweat glands, which can be caused by a variety of gene defects. Here we report to brothers born to consanguineous parents with a novel disease syndrome of HED, Severe Combined Immunodeficiency (SCID), and congenital myopathy. The SCID is characterized by abnormal T cell activation and a defect in store-operated Ca2+ influx in lymphocytes and fibroblasts from both patients. The ectodermal dysplasia manifested with abnormal sweat production and severely dysplastic dental enamel. The older brother died of septicemia due to SCID at the age of 11 months. The younger brother received a bone marrow transplant at the age of 6 months and appears to be immunologically fully competent since then. Both patients showed congenital generalized muscle weakness. The surviving brother reached independent ambulation at the age of 20 months. At the age of 11 years the boy showed generalized weakness (3+ to 4/5 according to MRC-scale) including facial muscles. He was not able to climb stairs independently and his maximal walking distance was limited to about one kilometer. His speech is hypernasal due to velopharyngeal weakness. A muscle biopsy at the age of 5 years revealed variation in fiber size but no dystrophic changes. Fiber typing showed a predominance of type I and atrophic type II fibers. NADH staining did not reveal any cores or other structural abnormalities. Although overlap syndromes between ectodermal dysplasia and immunodeficieny have been described (mutations in IKKg/NEMO and IkBa) to our knowledge this is the first description of patients with additional clinical and morphologic features of a congenital myopathy. The identification of the underlying genetic defect in the presented patients will contribute to the understanding of pathogenetic mechanisms in congenital myopathies. doi:10.1016/j.nmd.2006.05.154

P.P.5 10 A sarcoplasmic body myopathy ˚ hlberg, M. Engvall, B. Hedberg *, H. Stibler, F. Xiang, G. A T. Ansved, L. Edstro¨m Karolinska Institute, Clinical Neuroscience, Stockholm, Sweden In 1980 a new hereditary myopathy with adult onset was described in a Swedish family. The most characteristic feature of this myopathy was the sarcoplasmic inclusion bodies (SB), seen in the muscle biopsies. Distal muscle weakness seemed to be the most prominent early symptom. Now, we have further analysed the phenotype to distinguish the different stages of myopathy from a subclinical stage to fully developed myopathy with patients requiring wheelchair for ambulation. We have found that even the subclinical cases have these specific sarcoplasmic bodies in their muscle biopsies together with elevated creatine kinase (CK) levels. The sarcoplasmic bodies from the symptom-free carriers have now been thoroughly studied by transmission electron microscopy on epoxy embedded material, supplemented with ultrastructural immuno cytochemistry on muscle tissue from one patient. The SB’s were found to be of different shapes, from homogenous electrondense structures, with distinct borders, to a more mottled appearance. It was earlier suggested that mutant, abnormal stable desmin might be the cause of the sarcoplasmic bodies due to deficient break-down of desmin. A mutation of the desmin gene has now been excluded as a possible cause of this myopathy, by direct sequencing. It seems, however to be a dramatic upregulation of desmin and also of vimentin in the highly affected patients. The increase of vimentin might be explained by upregulation in immature regenerating muscle cells. However, the cause of elevated desmin levels is more difficult to explain. The presence of sarcoplasmic bodies, as a presymptomatic phenomenon, mostly combined with increase in CK, is helpful in establishing possible carriership of the mutant gene in symptomfree family members. doi:10.1016/j.nmd.2006.05.155

P.P.5 11 Autosomal dominant congenital fibre type disproportion. Study of a family A. Nucci 1,*, C.A. Chagas 3, A.V. Faria 2, E.M.B. Pacheco 2, L.S. Queiroz 3 1 Campinas State University (UNICAMP), Department of Neurology, Campinas, Brazil; 2 Campinas State University (UNICAMP), Department of Radiology, Campinas, Brazil; 3 Campinas State University (UNICAMP), Department of Pathology, Campinas, Brazil Congenital fibre type disproportion (CFTD) is a rare myopathy, with heterogeneous clinical phenotype and inheritance. Muscle biopsy is characteristic, showing predominance and smallness of type 1 fibres. Few cases were studied by magnetic resonance imaging of muscles (mMR). To present mMR data in three cases of CFTD in two successive generations and to analyse topography of muscle involvement and degree of affection. Patients were studied clinically, by muscle biopsy, CK, mMR and neurophysiological exams. MR was performed in a 2Tesla system, with two sets of T1-weighted axial slices, 6–8 mm thick, of thighs and legs. Each muscle was scored zero (normal) to four (end stage) blindly of clinical data. For each muscle, on both limbs, scores obtained in the three patients were summated and compared between muscles and compartments. The father was 44 years old and his siblings, a girl of 11 and a boy of 9. They had congenital myopathy with hypotonia and delayed motor milestones. All were thin and hypotonic, with arched palate and normal facial and external eye movements. Axial muscle involvement caused scoliosis, severe in the father, moderate and mild in the children. There was diffuse motor deficit with predom-

Abstracts / Neuromuscular Disorders 16 (2006) 644–726 inance in lower limbs, especially foot and toe dorsiflexors. CK was normal to mildly increased. The father had typical fibre type disproportion in biceps brachii. Two cases had a myopathic pattern in EMG. Total mMR scores were: extensor hallucis longus (18); extensor digitorum longus (18), tibialis anterior (16,5), sartorius (15), adductor brevis (14), adductor magnus (14), adductor longus (13), vastus medialis (12), soleus (12), tibialis posterior (11), peroneus (10), semitendinosus (10), semimembranosus (10), gracilis ( 9), biceps femoris (8), rectus femoris (8), vastus intermedius (8), vastus lateralis (8), gastrocnemius lateralis (7), gastrocnemius medialis (4) Posterior muscles of thighs were relatively spared compared to anterior and medial compartments. Anterior compartments of legs were most affected, followed by deep posterior, superficial posterior and lateral compartments. Scores had correlation with age and clinical severity of disease. doi:10.1016/j.nmd.2006.05.156

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G.P.5 01 Inflammatory muscle disease in dogs is associated with Leishmania infantum O. Paciello 1,*, G. Oliva 2, L. Gradoni 3, L. Manna 2, S. Papparella 1 1 Department of Pathology and Animal Health, University of Naples Federico II, Naples, Italy; 2 Department of Veterinary Clinical Science, University of Naples Federico II, Naples, Italy; 3 Istituto Superiore di Sanita`, Rome, Italy Leishmaniases, caused by Leishmania spp., are associated with a complex of wide-ranging diseases having various clinical and epidemiological features. Polymyositis (PM) is an autoimmune inflammatory muscle disease of unknown causes. In dogs PM might be associated with infectious diseases caused by Toxoplasma gondii, Neospora caninum, Ehrlichia canis or Hepatozoon canis; however, its association with Leishmania infection has not been well documented. To evaluate the possibility of Leishmania involvement in PM, we studied muscle biopsies of 15 dogs with a parasitologically and serologically established diagnosis of leishmaniosis and clinical evidence of muscle disease. Biopsy samples from the biceps femoris muscle were examined with routine stains and immunohistochemical analyses using: specific antibody against Leishmania spp., monoclonal antibodies against canine leucocytes antigens, MHC class I and MHC class II. The presence and the amount of Leishmania DNA in the muscle samples was evaluated by a real time PCR and correlated with a degree of muscle inflammation. With laser micro-dissector single muscle fibers were isolated and tested for the presence of Leishmania DNA inside the fibers. Characteristic histological features of PM were observed in all studied cases with various degrees of lymphocytes infiltration. The inflammatory cells were CD3+, CD4+, CD18+, CD45+ positive and expressed MHC class I and MHC class II. In addition, several muscle fibers had MHC class I and II positivity on the sarcolemma. By light microscopy, Leishmania immunoreactivity was observed within macrophages and in the connective tissue adjacent muscle fibers. Although no Leishmania DNA was amplified from the isolated muscle fibers, the degree of muscle damage and inflammatory cells infiltrations were proportional to the total amount of Leishmania DNA. Our studies provide evidence that: (1) Leishmania might be considered as possible cause of PM in dogs; (2) Leishmania is not present within muscle fibers and that (3) the muscle damage might be related to immunological alteration, always associated to Leishmania infectious. We suggest that Leishmania has to be considered in the differential diagnosis of PM in dogs and that PM could occur in human leishmaniosis – if such new treatment approach can be utilized. (We thank Dr. Mercedes Domı´ n-

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guez, Servicio Inmunologia, Instituto de Salud Carlos III, Spain, for kindly providing the anti-leishmania antibody.) doi:10.1016/j.nmd.2006.05.157

G.P.5 02 MRL/MpJ wound-healing phenotype increases the myofiber size in mdx mouse skeletal muscle Y. Ohsawa *, H. Hagiwara, N. Naoe, Y. Kuroda, T. Kawase, T. Murakami, Y. Sunada Kawasaki Medical School, Neurology, Kurashiki, Japan MRL/MpJ mice have been highlighted as a mouse model of accelerated wound-healing and tissue regeneration. Genetic background of this phenotype has not been determined. In order to determine the effect of MRL/MpJ wound- healing phenotype on the dystrophin-deficient muscular dystrophy, we crossed mdx mice with MRL/MpJ mice to generate and characterize F2 offspring. By crossing with F1 offspring, we obtained F2 offspring with four distinct genotypes: wildtype, mdx, mdx/MRL/MpJ, and MRL/MpJ. The mdx genotype was determined by single-nucleotide mismatch PCR amplification of mutant dystrophin as described previously (Do AA, Muscle Nerve 1996;19). MRL/MpJ genotype was determined by the closure of earpunched hole as described previously (Clark LD, Clin Immunol Immunopathol 1998;88). The single-myofiber area (n = 5, 250 myofibers for each mice) in quadriceps femoris muscle was measured from the florescence images of anti-laminin alpla-2 immunostaining using two color technique with Lumina Vision software. Compared to the wild-type mice, the mdx mice showed a significant reduction in single-myofiber area (4445.8 ± 72.2 microm2 vs. 8326.6 ± 67.1 microm2, P < 0.05). In contrast, the MRL/MpJ mice exhibited a significant increase in single- myofiber area (10917.8 ± 68.8 microm2, P < 0.05). In the mdx/ MRL/MpJ mice, single-myofiber area was significantly increased (8681.9 ± 61.5 microm2, P < 0.05), compared to the mdx mice. MRL/MpJ wound-healing phenotype could accelerate myofiber regeneration in mdx mice. Thus, the MRL/MpJ wound healing mechanism may be applicable for the treatment of muscular dystrophy. doi:10.1016/j.nmd.2006.05.158

G.P.5 03 Laser microdissection-based expression analysis of key muscle regeneration genes in degenerative–regenerative groups of mdx mice M. Marotta 1,*, Y. Sarria 1, C. Ruiz 1, M. Roig-Quilis 1,2 1 Institut de Recerca, Hospital Universitari Vall d’Hebron, Barcelona, Spain; 2 Hospital Materno-Infantil Vall d’Hebron, Barcelona, Spain Study the gene expression throughout the complete muscle degenerative–regenerative process by the application of laser capture microdissection (LCM) to the obtaining of tissue from mdx-mice skeletal muscle degenerative–regenerative groups (DRGs) representing all the stages of muscle regeneration process. The obtaining of microdissected tissue from these DRGs allows to the accurate gene expression analysis in the exactly site of injury and in a precise point in time of skeletal muscle regeneration process. Tissue from the different DRGs were obtained from 20-lm thick serial frozen transversal sections from mdx-mice gastrocnemius muscle group (GMG), mounted on pre-treated poly-L-lysine RNase-free membrane-slides and immediately processed for hematoxylin-eosin staining and dehydration with RNasefree graded ethanol solutions and xylene. LCM was performed using a PALMÒ Robot Microbeam (PALM technologies). Total RNA obtained from each DRG was extracted by the RNeasy Micro Kit