Mutations of the selenoprotein N gene, which is implicated in rigid spine muscular dystrophy, cause the classical phenotype of multiminicore disease: reassessing the nosology of early-onset myopathies

Abstract

Multiminicore disease (MmD) is an autosomal recessive congenital myopathy characterized by the presence of multiple, short core lesions (known as "minicores") in most muscle fibers. MmD is a clinically heterogeneous condition, in which four subgroups have been distinguished. Homozygous RYR1 mutations have been recently identified in the moderate form of MmD with hand involvement. The genes responsible for the three other forms (including the most prevalent phenotype, termed the "classical" phenotype) remained, so far, unknown. To further characterize the genetic basis of MmD, we analyzed a series of 62 patients through a combined positional/candidate-gene approach. On the basis of clinical and morphological data, we suspected a relationship between classical MmD and the selenoprotein N gene (SEPN1), which is located on chromosome 1p36 (RSMD1 locus) and is responsible for the congenital muscular dystrophy with rigid spine syndrome (RSMD). A genomewide screening, followed by the analysis of 1p36 microsatellite markers in 27 informative families with MmD, demonstrated linkage to RSMD1 in eight families. All showed an axial myopathy with scoliosis and respiratory failure, consistent with the most severe end of the classical MmD spectrum; spinal rigidity was evident in some, but not all, patients. We excluded linkage to RSMD1 in 19 families with MmD, including 9 with classical MmD. Screening of SEPN1 in the 8 families that showed linkage and in 14 patients with classical sporadic disease disclosed 9 mutations affecting 17 patients (12 families); 6 were novel mutations, and 3 had been described in patients with RSMD. Analysis of three deltoid biopsy specimens from patients with typical RSMD revealed a wide myopathological variability, ranging from a dystrophic to a congenital myopathy pattern. A variable proportion of minicores was found in all the samples. The present study represents the first identification of a gene responsible for classical MmD, demonstrates its genetic heterogeneity, and reassesses the nosological boundaries between MmD and RSMD.

Figure 1

Histochemical and ultrastructural features of MmD. a–d, Serial transverse cryostat sections. a and b, Patient 16 at 5 years of age, left deltoid, stained with hematoxylin-eosin (HE) (a) and myosin adenosine triphosphatase, pH 9.4 (b). Note variability of fiber size and predominance of type 1 (lighter) fibers. Although all the type 2 fibers show large diameters, both large and small type 1 fibers are observed, which produces a smaller mean diameter (“type 1 relative hypotrophy”). c and d, Patient 2 at 14 years of age, right deltoid; nicotinamide adenine dinucleotide–tetrazolium reductase (NADH-TR) (c) and succinate dehydrogenase (d). Multiple focal areas lacking oxidative activity are present in most type 1 (darker) and type 2 fibers. As shown here in two different biopsy specimens, there was no significant increase in endomysial connective tissue. e–g, Longitudinal electron micrograph sections from patients 8 (e, f) and 16 (g); left quadriceps and left deltoid at 23 and 5 years of age, respectively. e and f, Mild, early-stage minicore lesions; the sarcomeres appear out of register, Z-line streaming is limited to one sarcomere in length and 3–4 myofibrils in width, and the longitudinal myofilament array is still recognizable. g, Electron-dense material of Z-line origin, forming irregular zones, with severe focal disorganization of the myofibrillar structure. Bar=40 μm in a and b, 25 μm in c, 12.5 μm in d, and 2.5 μm in e–g.

Figure 2

Clinical features. a, Patient 12 at age 5.5 years. Slender neck with absent sternocleidomastoideus bulk, flat thorax, and moderate deltoid atrophy are present from early stages of the disease, as is an unusual appearance of the lower limbs, characterized by amyotrophic inner thighs, straight calves, and flat feet. Scoliosis also follows a particular pattern, with associated dorsal lordosis (“hollow back”) and a lateral trunk deviation and frequently requires an extensive arthrodesis (b and c). b, Patient 7 at age 15 years, immediately before surgical correction. c, Stabilization of the spinal deformity in patient 6 at age 17 years, 4 years after arthrodesis.

Figure 3

Histochemical and ultrastructural analysis of deltoid muscle biopsies taken from patients with RSMD at the ages of 11 (RSMD1 and RSMD2) and 14 years (RSMD3). a–f, Transverse cryostat sections; g–i, longitudinal electron micrograph sections. All samples show fiber size variability (a–c, HE) and focal oxidative-negative areas (arrows; d–f, NADH-TR) corresponding, at the ultrastructural level, to short regions of Z-line streaming and sarcomere disarray (g–i). The myopathological pattern in RSMD1 is equivalent to that in patients with typical MmD; absence of necrosis, regeneration, and endomysial fibrosis coexists with the presence of short core lesions in most fibers. In contrast, in RSMD3, there is a predominance of dystrophic features, such as regenerative fibers (asterisks) and marked increase of endomysial connective tissue, while small zones of Z-line streaming were present in less than half of the fibers. Sample RSMD2 presented an intermediate pattern, with some rare regenerative fibers (asterisk), no significant fibrosis, and multiple minicores in approximately two-thirds of the fibers. Bar = 40 μm in a–c, 25 μm in d–f, and 2.5 μm in g–i.