engleski

Clinical and non-invasive genetic study of first six Croatian patients with dysferlinopathy

Mutations in the human dysferlin (DYSF) gene cause autosomal recessive muscular dystrophies characterized by degeneration and weakness of proximal and/or distal muscles: limb girdle muscular dystrophy 2B and Miyoshi myopathy (MM). Current diagnostic methods require muscle biopsy for immunodiagnosis, followed by direct gene analysis. Aim. To present an alternative, non-invasive approach to diagnosis based on linkage analysis and a non-invasive blood diagnostic assay. Patients and Methods. Six patients from five unrelated, informative, Croatian families affected by presumed MM/LGMD2B were selected on clinical and laboratory features. Haplotype analysis was done by microsatellites flanking the dysferlin gene: D2S292, D2S2113, D2S291 and D2S2111. Non-invasive diagnostics of dysferlin from peripheral blood used the new method (Ho M et al. Ann Neurol 2001 ; 51:129-133) consisted in: Isolation of peripheral blood mononuclear cells (PBMC) by Ficoll and CD14MicroBeads followed by SDS-PAGE and immunoblotting using the NCL-Hamlet anti-dysferlin monoclonal antibody. Immunoreactive bands were detected with chemiluminiscence system (Amersham). Results. All six patients showed linkage with studied markers. Moreover, three MM patients from two families showed two different homozygous haplotypes suggesting homozygosity of one or two mutations of independent origin. Immunoblotting of dysferlin on PBMC and skeletal muscle was detected in controls, but was absent in four and weak in two patients. These preliminary results need direct detection of gene mutation. Conclusion. Since we lack sensitive and specific biopsy screening methods for detecting patients with dysferlinopathy, linkage analysis in informative families followed by Western blot analysis of dysferlin on PBMC could avoid the need of invasive procedure for muscle specimens.

dysferlinopathy; LGMD2B; clinics; linkage analysis; Western blot; monocytes

doi:10.1016/j.nmd.2005.06.006