Ultrastructural changes in dysferlinopathy support defective membrane repair mechanism

Abstract

Background: The dysferlin gene has recently been shown to be involved in limb girdle muscular dystrophy type 2B and its allelic disease, Miyoshi myopathy, both of which are characterised by an active muscle degeneration and regeneration process. Dysferlin is known to play an essential role in skeletal muscle fibre repair, but the process underlying the pathogenetic mechanism of dysferlinopathy is not completely understood.

Aims: To define both specific alterations of muscle fibres and a possible sequential mechanism of myopathy development.

Methods: A histological, immunohistochemical, and ultrastructural analysis of 10 muscle biopsies from patients with molecularly diagnosed dysferlinopathy.

Results: An inflammatory response was seen in most of the muscle biopsies. The immunohistochemical pattern demonstrated active regeneration and inflammation. Non-necrotic fibres showed alterations at different submicroscopic levels, namely: the sarcolemma and basal lamina, subsarcolemmal region, and sarcoplasmic compartment. In the subsarcolemmal region there were prominent aggregations of small vesicles, probably derived from the Golgi apparatus, which consisted of empty, swollen cisternae. In the sarcolemma there were many gaps and microvilli-like projections, whereas the basal lamina was multilayered.

Conclusions: The histopathological, immunohistochemical, and ultrastructural data show that dysferlinopathy is characterised by a very active inflammatory/degenerative process, possibly associated with an inefficient repair and regenerative system. The presence of many crowded vesicles just beneath the sarcolemma provides submicroscopical proof of a defective resealing mechanism, which fails to repair the sarcolemma.

Figure 3

Electron microscopy of skeletal muscle from a patient with dysferlinopathy. (A) Subsarcolemmal accumulation of vesicles (asterisk) derived from the Golgi apparatus with swollen cisternae seen in (B) (asterisk); (C) vacuoles (asterisk on the left) with degenerating myelin figures and microvilli-like projections (asterisk on the right); (D) severe loss of myofilaments, dilated rough endoplasmic reticulum, and non-specific degenerative changes of mitochondria; large subsarcolemmal vacuoles with loss of microfilaments are also seen (asterisk). Original magnification, ×9100 (A–C) and ×5600 (D).

Figure 1

Skeletal muscle morphology and immunohistochemistry in dysferlinopathy. A set of serial muscle biopsy sections (patient CR) shows muscle morphology and immunolabelling with different antibodies. (A) Haematoxylin and eosin (H&E) staining. Immunostaining with antibodies to (B) the Golgi zone, (C) caveolin 3, and (D) fetal myosin. The asterisks indicate the same fibres in different serial sections. Two basophilic muscle fibres (A) are positive for fetal myosin (D) and show greatly reduced caveolin 3 labelling (C) and an absence of Golgi labelling (B). Additional atrophic fibres are negative for fetal myosin and the Golgi zone, but they are strongly positive for caveolin 3. Original magnification, ×200.

Figure 2

Electron microscopy of skeletal muscle from a patient with dysferlinopathy. (A) Elongated microvilli-like projections oriented parallel to the cell surface and close to sarcolemmal gap (asterisk); (B) basal lamina reduplication (asterisk); (C) many electrondense globular aggregates (asterisk); (D) a lymphocyte localised in the virtual space between the sarcolemma and the basal lamina. Original magnification, ×9100.