Novel splicing dysferlin mutation causing myopathy with intra-familial heterogeneity.

Dysferlinopathies belong to the heterogeneous group of autosomal recessive muscular disorders, caused by mutations in the dysferlin gene and characterized by a high degree of clinical variability even though within the same family. This study aims to describe three cases, belonging to a consanguineous Tunisian family, sharing a new splicing mutation in the dysferlin gene and presenting intra-familial variability of dysferlinopathies: Proximal-distal weakness and distal myopathy with anterior tibial onset. We performed the next generation sequencing for mutation screening and reverse transcriptase-PCR for gene expression analysis. Routine muscle histology was used for muscle biopsy processing. The clinical presentation demonstrated heterogeneous phenotypes between the three cases: Two presented intermediate phenotypes of dysferlinopathy with proximal-distal weakness and the third had a distal myopathy with anterior tibial onset. Genetic analysis yielded a homozygous splicing mutation (c.4597-2A>G) in the dysferlin gene, giving rise to the suppression of 28 bp of the exon 43. The splicing mutation found in our family (c.4597-2A>G) is responsible for the suppression of 28 bp of the exon 43 and a wide clinical intra-familial variability.

Dysferlinopathy in Tunisia: clinical spectrum, genetic background and prognostic profile.

Dysferlinopathy is a rare group of hereditary muscular dystrophy with an autosomal recessive mode of inheritance caused by a mutation in the DYSF gene. It encodes for the dysferlin protein, which has a crucial role in multiple cellular processes, including muscle fiber membrane repair. This deficit has heterogeneous clinical presentations. In this study, we collected 20 Tunisian patients with a sex ratio of 1 and a median age of 50.5 years old (Interquartile range (IQR) = [36,5-54,75]). They were followed for periods ranging from 5 to 48 years. The median age at onset was 17 years old (IQR = [16,8-28,4]). Five major phenotypes were identified: Limb-girdle muscular dystrophy (LGMDR2) (35%), a proximodistal phenotype (35%), Miyoshi myopathy (10%),  Distal myopathy with anterior tibial onset (DMAT) (10%), and asymptomatic HyperCKemia (10%). At the last evaluation, more than half of patients (55%) were on wheelchair. Loss of ambulation occurred generally during the fourth decade. After 20 years of disease progression, two patients with a proximodistal phenotype (10%) developed dilated cardiomyopathy and mitral valve regurgitation. Restrictive respiratory syndrome was observed in three patients (DMAT: 1 patient, proximodistal phenotype: 1 patient, LGMDR2: 1 patient). Genetic study disclosed five mutations. We observed clinical heterogeneity between families and even within the same family. Disease progression was mainly slow to intermediate regardless of the phenotype.

Novel Missense CAPN3 Mutation Responsible for Adult-Onset Limb Girdle Muscular Dystrophy with Calves Hypertrophy.

CAPN3 gene encodes for calpain-3; this protein is a calcium-dependent intracellular protease. Deficiency of this enzyme leads to weakness of the proximal limb muscles and pelvic and shoulder girdles, the so-called limb-girdle muscular dystrophy type 2A (LGMD2A). Here, we reported the case of a Tunisian patient with LGMD2A associated with a novel missense mutation (c.T1681C/p.Y561H). A 61-year-old man, with consanguineous parents, was referred for gait difficulties and slowly progressive proximal weakness of the four limbs associated with moderate hypertrophy of the calves but his facial muscles were unaffected. Electromyography showed that the profile was myopathic pattern and creatine kinase (CK) level was high. Muscle biopsy processing included routine histological, immunohistochemical, and Western Blot reactions, using a panel of antibodies directed against dystrophin, dysferlin, calpain-3, sarcoglycan α, ß, γ, and δ. For mutation analysis, we designed an NGS-based screening. Immunological analyses demonstrated a total deficiency in calpain-3 and δ-sarcoglycan, and a reduced expression of dysferlin. The genetic study yielded a homozygous missense mutation (c.T1681C) of the 13th exon of the CAPN3 gene. The mutation found in our patient (c.T1681C/p.Y561H) has not been previously reported. It is responsible for complete calpain-3 and δ-sarcoglycan deficiency and reduced dysferlin expression. The genetic study is mandatory in such cases with multiple-protein deficiency and ambiguous results of immune-histology and Western Blot studies.