Muscle histopathology in nebulin-related nemaline myopathy: ultrastrastructural findings correlated to disease severity and genotype

Abstract

Nemaline myopathy (NM) is a rare congenital myopathy characterised by hypotonia, muscle weakness, and often skeletal muscle deformities with the presence of nemaline bodies (rods) in the muscle biopsy. The nebulin (NEB) gene is the most commonly mutated and is thought to account for approximately 50% of genetically diagnosed cases of NM. We undertook a detailed muscle morphological analysis of 14 NEB-mutated NM patients with different clinical forms to define muscle pathological patterns and correlate them with clinical course and genotype. Three groups were identified according to clinical severity. Group 1 (n = 5) comprises severe/lethal NM and biopsy in the first days of life. Group 2 (n = 4) includes intermediate NM and biopsy in infancy. Group 3 (n = 5) comprises typical/mild NM and biopsy in childhood or early adult life. Biopsies underwent histoenzymological, immunohistochemical and ultrastructural analysis. Fibre type distribution patterns, rod characteristics, distribution and localization were investigated. Contractile performance was studied in muscle fibre preparations isolated from seven muscle biopsies from each of the three groups. G1 showed significant myofibrillar dissociation and smallness with scattered globular rods in one third of fibres; there was no type 1 predominance. G2 presented milder sarcomeric dissociation, dispersed or clustered nemaline bodies, and type 1 predominance/uniformity. In contrast, G3 had well-delimited clusters of subsarcolemmal elongated rods and type 1 uniformity without sarcomeric alterations. In accordance with the clinical and morphological data, functional studies revealed markedly low forces in muscle bundles from G1 and a better contractile performance in muscle bundles from biopsies of patients from G2, and G3.In conclusion NEB-mutated NM patients present a wide spectrum of morphological features. It is difficult to establish firm genotype phenotype correlation. Interestingly, there was a correlation between clinical severity on the one hand and the degree of sarcomeric dissociation and contractility efficiency on the other. By contrast the percentage of fibres occupied by rods, as well as the quantity and the sub sarcolemmal position of rods, appears to inversely correlate with severity. Based on our observations, we propose myofibrillar dissociation and changes in contractility as an important cause of muscle weakness in NEB-mutated NM patients.

Target resequencing of neuromuscular disease-related genes using next-generation sequencing for patients with undiagnosed early-onset neuromuscular disorders

“…Notably, in CM-2, muscle biopsy was not performed, but NM was identified because previously reported pathogenic mutations (NEB c.24580-1G4A and c.8980delA) were identified and the patient's clinical features clearly corresponded to the phenotypes of the reported cases. 25,26 In the case of CM-5, who had childhood-onset slowly progressive muscle weakness, no specific results were obtained except for muscle biopsy findings of myogenic changes and type II fiber predominance (Figure 2l). However, a previously reported mutation (MYH7 p.His1901Leu) was detected and the patient's clinical features were clearly consistent with those of definite cases; 27 thus, MSM was definitively diagnosed.…”

Severe myopathy in mice lacking the MEF2/SRF-dependent gene leiomodin-3

“…Skeletal muscle from Lmod3-KO mice showed a marked disruption of sarcomeric organization by conventional histology, whereas sarcomeric disorganization in Klhl40-KO skeletal muscle was comparatively mild at the histological level and only detectable by electron microscopy. Skeletal muscle from Lmod3-KO mice also displays abnormal glycogen accumulation and nemaline rods, which appear as electron-dense inclusions attributed to the accumulation of disarrayed Z-line proteins, a hallmark of nemaline myopathy (2,33).…”

Mutation Update: The Spectra of Nebulin Variants and Associated Myopathies

“…Reference sequence: c.NM_001271208.1 (contains all of the 183 exons including the triplicated region). cons, consanguineous; HOZ, homozygous mutation, del ex55, Ashk., Ashkenazi Jewish founder mutation; Finnish founder mutations: FIN1, a missense mutation Ser > Ile in exon 122; FIN2, a missense mutation Thr > Pro in exon 151; FIN3, a frameshift mutation in exon 122; ALT1, mutation in the alternatively spliced exons 63–66 (always expressed together); ALT2, mutation in the alternatively spliced exons 143 or 144; ALT3, mutation in the alternatively spliced exons 167–177 (expressed independently), TRI, mutation in the triplicate region. Published mutations in Table 2: A, Pelin et al (1999); B, Pelin et al (2002); C, Anderson et al (2004); D, Lehtokari et al (2006); E, Wallgren-Pettersson et al (2007); F, Lehtokari and Greenleaf et al (2009); G, Romero and Lehtokari et al (2009); H, Lawlor et al (2011); I, Ochala et al (2011); J, Lehtokari et al (2011); K, Kapoor et al (2013); L, Yonath et al (2012); M, Böhm et al (2013); N, Scoto et al (2013); O, Gajda et al (2013); P, Kiiski et al (2013); Q, Malfatti et al (2014); R, Malfatti et al (in preparation). …”

“…Published mutations in Table 2: A, Pelin et al (1999); B, Pelin et al (2002); C, Anderson et al (2004); D, Lehtokari et al (2006); E, Wallgren-Pettersson et al (2007); F, Lehtokari and Greenleaf et al (2009); G, Romero and Lehtokari et al (2009); H, Lawlor et al (2011); I, Ochala et al (2011); J, Lehtokari et al (2011); K, Kapoor et al (2013); L, Yonath et al (2012); M, Böhm et al (2013); N, Scoto et al (2013); O, Gajda et al (2013); P, Kiiski et al (2013); Q, Malfatti et al (2014); R, Malfatti et al (in preparation).…”

“…All pathogenic variants hitherto known in NEB have been recessive, mostly compound heterozygous, and the most common cause of autosomal-recessive nemaline myopathy (NM; MIM #256030) [Pelin et al, 1999; Pelin et al, 2002; Anderson et al, 2004; Lehtokari et al, 2006; Lawlor et al, 2011; Ochala et al, 2011; Kapoor et al, 2013; Malfatti et al, 2014]. Other, less common entities include early-onset distal myopathy without nemaline bodies [Wallgren-Pettersson et al, 2007], a distal form of NM [Lehtokari et al, 2011], core-rod myopathy with generalized muscle weakness [Romero et al, 2009], and a childhood-onset distal myopathy with rods and cores [Scoto et al, 2013].…”