Myofibrillar Myopathies: A Clinical and Myopathological Guide

Abstract

Myofibrillar myopathies (MFMs) are histopathologically characterized by desmin‐positive protein aggregates and myofibrillar degeneration. Because of the marked phenotypic and pathomorphological variability, establishing the diagnosis of MFM can be a challenging task. While MFMs are partly caused by mutations in genes encoding for extramyofibrillar proteins (desmin, αB‐crystallin, plectin) or myofibrillar proteins (myotilin, Z‐band alternatively spliced PDZ‐containing protein, filamin C, Bcl‐2‐associated athanogene‐3, four‐and‐a‐half LIM domain 1), a large number of these diseases are caused by still unresolved gene defects. Although recent years have brought new insight into the pathogenesis of MFMs, the precise molecular pathways and sequential steps that lead from an individual gene defect to progressive muscle damage are still unclear. This review focuses on the clinical and myopathological aspects of genetically defined MFMs, and shall provide a diagnostic guide for this numerically significant group of protein aggregate myopathies.

INTRODUCTION

Protein aggregation is the pathomorphological hallmark of major neurodegenerative diseases such as Alzheimer's and Parkinson's diseases. However, pathological protein aggregation is also the characteristic feature in the expanding group of myofibrillar myopathies (MFMs). The paradigm of the latter is a disease caused by mutations in the desmin gene that result in a skeletal myopathy, frequently associated with cardiac abnormalities 17, 18, 19, 34, 38. Striated muscles from patients with mutations in the desmin gene characteristically display cytoplasmic accumulation of desmin‐immunoreactive material and myofibrillar alterations 12, 19, 34, 38. Desminopathy shares its structural myofibrillar and intermyofibrillar abnormalities with the large group of MFMs (synonyms: desmin‐related myopathies, secondary desminopathies) 11, 38. These disorders comprise sporadic and familial neuromuscular conditions of considerable clinical and genetic heterogeneity. While these MFMs are partly caused by αB‐crystallin, myotilin, Z‐band alternatively spliced PDZ‐containing protein (ZASP), filamin C, four‐and‐a‐half LIM domain 1 (FHL1), Bcl‐2‐associated athanogene‐3 (BAG3) or plectin mutations 15, 20, 23, 28, 29, 31, 35, 36, 37, 38, 40, 43, 44, a large number of these diseases are caused by so far unidentified gene defects. MFMs are clinically characterized by a progressive course leading to severe disability. To date, no causative or even ameliorating therapies exist for this numerically significant group of hereditary myopathies (17).

The knowledge of the structure and function of wild‐type desmin is a prerequisite for the understanding of human MFM. Desmin is the classical type III intermediate filament (IF) protein of striated and smooth muscle cells. It has a tripartite structure comprising a central α‐helical coiled‐coil rod domain flanked by non‐α‐helical head and tail domains. The central rod domain, formed by four α‐helical segments (1A, 1B, 2A, 2B) which are separated by three short polypeptide linkers (L1, L12, L2), plays a critical role in desmin filament assembly and the formation of the extrasarcomeric cytoskeleton (2). This filamentous structure forms a three‐dimensional scaffold around myofibrillar Z‐disks, thereby interlinking neighboring myofibrils and connecting the myofibrillar apparatus to nuclei, mitochondria and sarcolemma (Figure 1). However, the ordered formation, spacing and anchorage of the IF system are dependent on the proper interaction of desmin with its binding partners, that is, plectin and αB‐crystallin 24, 43. Insights into the molecular pathogenesis of desmin mutations have been obtained from transfection experiments and in vitro assembly studies 1, 2, 3, 4, 32, which indicate that the majority of desmin rod mutants is either: (i) incapable of forming a de novo desmin IF network; or (ii) forming abnormal IF structures, inducing the collapse of a preexisting IF network and leading to desmin‐positive protein aggregates. These results imply that mutated desmin compromises the filament formation competence and that filament–filament interactions should be key events in the molecular pathogenesis of desminopathy. However, if desmin mutants have such a toxic effect on the desmin filament system in vitro, why does it take such a long time until the clinical symptoms of progressive muscular damage become apparent in humans? The previous “simple” mechanistic explanation is further challenged by the recent observation that certain desmin mutants exhibit assembly defects in vitro when analyzed on their own, but facilitate proper filament formation when studied in one‐to‐one mixtures of mutant proteins with wild‐type desmin (5). Thus, it is unlikely that the complex human pathology is solely related to direct effects of desmin mutants on the assembly of desmin IFs. As an alternative explanation, desmin mutants may interfere with the interaction with binding partners, thereby influencing the structural and functional organization of the extrasarcomeric cytoskeleton as well as intracellular signaling cascades. This view is further substantiated by the observation that mutations in genes encoding cytoskeletal linker proteins in muscle cells, namely plectin, filamin C, ZASP, FHL1 and myotilin, lead to similar desmin‐positive cytoplasmic protein aggregates 28, 31, 36, 37, 44. Other contributing factors relate to metabolic abnormalities. Several studies demonstrated that the filamentous desmin–plectin network plays an essential role in the subcellular positioning and function of mitochondria (24) (Figure 1). In addition, several studies demonstrated mitochondrial pathology in MFM 23, 27, 31, 32. Further evidence for the disease‐contributing mitochondrial pathology are the generation of reactive oxygen species and associated oxidation and nitration of desmin and other cytoskeletal proteins (22). Hence, focal disturbances in the production of ATP may have a negative influence both on the structural integrity of myofibrils and other highly ATP‐dependent cellular compartments and processes. Other central factors are primary or secondary disturbances of protein quality control mechanisms related to, in particular, the ubiquitin–proteasome system (UPS) and the autophagic–lysosomal pathway (types 1 and 2 of programmed cell death) 26, 30 (Figure 1). A direct link between protein quality control mechanisms and desmin protein aggregate pathology is substantiated by a recent study showing that desmin mutants impair the proteolytic function of the UPS (6). Conversely, mutations in the genes encoding the molecular chaperones αB‐crystallin and valosin‐containing protein (VCP) also share key features of the desmin‐positive aggregate pathology 21, 43. In particular, mutant VCP has been attributed to impair UPS function (46). As UPS and autophagic buildup are inversely related, skeletal muscle from patients with VCP mutations exhibits signs of increased autophagy (21). This observation implies a direct molecular cross‐talk between the UPS and the autophagic system. However, rimmed and non‐rimmed vacuoles, which are the morphological hallmark of autophagic processes, are also characteristic features in muscle biopsies from patients with mutations in desmin, filamin C, plectin, FHL1, myotilin and ZASP 8, 15, 23, 30, 31, 47. For a more detailed synopsis on the molecular aspects of MFM pathology, please refer to the following excellent review (13).

Figure 1.

Schematic drawing of a muscle fiber with regard to myofibrillar myopathies (MFMs) causing mutations in genes coding for extramyofibrillar (desmin, αB‐crystallin, plectin) and myofibrillar (filamin C, myotilin, Z‐band alternatively spliced PDZ‐containing protein, four‐and‐a‐half LIM domain 1, Bcl‐2‐associated athanogene‐3) proteins. Note that the extramyofibrillar cytoskeleton forms a three‐dimensional scaffold around myofibrillar Z‐disks, thereby interlinking neighboring myofibrils and connecting the myofibrillar apparatus to myonuclei, sarcolemma and mitochondria. Mutations in these known MFM genes lead to misfolded proteins and desmin‐positive protein aggregates in conjunction with Z‐disk alterations and mitochondrial abnormalities. The ubiquitin–proteasome and the autophagic–lysosomal system are essential components of the cellular protein quality control framework, which is responsible for the degradation of misfolded proteins and protein aggregates.

This review focuses on the clinical (Table 1) and myopathological (Table 2) aspects of MFM, and shall provide a diagnostic guide for this numerically significant and often under‐recognized group of protein aggregate myopathies.

Table 1.

Myofibrillar myopathies: clinical presentation. Abbreviations: CB = conduction block; CK = creatine kinase; DCM = dilative cardiomyopathy; RSS = rigid spine syndrome; SPS = scapuloperoneal syndrome; EBS = epidermolysis bullosa simplex.

Disease	Muscle weakness	Heart	Respiratory system	CK levels	Extramuscular
Desminopathy	Distal > proximal, SPS	DCM, CB	Insufficiency	n − 5x
aBCopathy	Proximal > distal	DCM,CB		n − 7x	Cataracts
Filaminopathy	Proximal > distal	DCM, CB	Insufficiency	n − 8x	Neuropathy
Myotilinopathy	Distal > proximal	DCM	Insufficiency	n − 5x	Neuropathy, contractures
BAG3opathy	Proximal	DCM	Insufficiency	n − 15x	RSS, scoliosis, contractures neuropathy
FHL1opathy	Distal = proximal, hypertrophy, SPS	CB	Insufficiency	n − 10x	RSS, scoliosis contractures
ZASPopathy	Distal > proximal hand muscle atrophy	DCM, CB		n − 6x	Neuropathy
Plectinopathy	Distal > proximal			n − 5x	EBS, nail dystrophy

Table 2.

Myofibrillar myopathies: histopathological features. Abbreviations: AV = autophagic vacuoles; CPB = cytoplasmic bodies; GFM = granulofilamentous material; G‐Tri = modified Gomori trichrome; PA = protein accumulation; RB = reducing bodies; RV = rimmed vacuoles; SB = spheroid bodies.

Protein	H&E	G‐Tri	COX	EM
Desmin	Inclusions, AV	PA	Rubbed out	GFM
Abc	Inclusions, AV	PA	Rubbed out	GFM
Filamin‐C	Inclusions, AV	PA	Rubbed out	GFM
Myotilin	Inclusions, AV SB, CPB	PA	Rubbed out	GFM, SB, CPB
BAG3	Inclusions, AV bodies	PA	Rubbed out	GFM, apoptotic
FHL1	Inclusions, AV	PA		GFM, CPB, RB
ZASP	Inclusions, AV	PA	Rubbed out	GFM, CPB
Plectin	Inclusions	PA	Rubbed out	GFM

MFM: CLINICAL CLUES TO THE DIAGNOSIS

Disease onset

The vast majority of MFM patients have an adult onset of their progressive muscle symptoms. Exceptions from this rule are MFM caused by mutations in genes encoding for BAG3, FHL1, plectin and desmin, which may manifest in childhood, adolescence and adulthood 17, 39, 40, 41. While MFMs caused by desmin and αB‐crystallin tend to manifest in early and middle adulthood, disease onset beyond the fourth decade of life points toward the diagnosis of myotilin‐, ZASP‐ and filamin C‐related MFMs.

Patterns of muscle weakness and atrophy

MFMs are associated with marked clinical variability. Skeletal muscle weakness in the lower extremities is the most frequent initial clinical symptom. MFM may present with the phenotypic presentation of: (i) distal myopathy; (ii) limb girdle muscular dystrophy; (iii) scapuloperoneal syndrome; (iv) isolated muscle group involvement; and (v) generalized myopathy. Generalized muscle weakness and wasting involving trunk and neck muscles are frequent features in late disease stages. As a rough rule of the thumb, predominant distal muscle involvement is seen in desminopathy and myotilinopathy, whereas involvement of proximal muscle groups is the leading symptom in other MFM forms. Scapuloperoneal phenotypes have been described in relation to desmin and FHL1 mutations. Late‐stage manifestations of facial muscle weakness, swallowing difficulties or dysarthria seem to be restricted to desmin‐, αB‐crystallin‐ and myotilin‐related MFMs.

Beyond skeletal muscle: clinical signs for specific MFM forms

Careful history and clinical examination often provide essential clues to the specific diagnosis. Cardiac involvement comprising multiple forms of arrhythmias, that is, AV‐nodal conduction blocks; supraventricular and ventricular ectopic beats and tachycardia; or true dilated, hypertrophic or restrictive cardiomyopathy is a classical feature in desminopathy, which may precede, coincide with or succeed skeletal muscle weakness (25). Cardiomyopathy manifesting in childhood is also a typical clinical finding in BAG3opathy and in a subset of patients with ZASP and αB‐crystallin mutations. Respiratory failure has been reported to be a frequent feature in patients with filamin C, FHL1 and BAG3 mutations, and in a subgroup of MFM patients with desmin, αB‐crystallin and myotilin mutations. A blistering skin disease since birth in conjunction with skeletal muscle myopathy is the pathognomonic sign of plectin‐related epidermolysis bullosa simplex with muscular dystrophy (EBS‐MD) (41). While lower back pain seems to be a common initial symptom in filamin C‐ and FHL1‐related MFMs, myalgia has been reported in myotilinopathy and FHL1opathy. Joint contractures are common in BAG3‐, FHL1‐ and myotilin‐related MFMs, but are absent in desminopathy, αB‐crystallinopathy, filaminopathy and ZASPopathy. Rigid spine and scoliosis are classical features of childhood MFMs caused by BAG3 and FHL1 mutations. Clinical and electrophysiological signs of polyneuropathy are numerous in MFM patients with ZASP and filamin C mutations. Cataracts have solely been reported in a subset of patients with αB‐crystallin mutations (43).

Family history

With the exception of autosomal recessive EBS‐MD and X‐linked FHL1opathy, all known MFM‐associated gene defects follow an autosomal dominant mode of inheritance. Proven recessive mutational patterns have only been reported in very few desminopathy patients. However, a significant number of MFMs show sporadic disease manifestation.

Creatine kinase (CK) levels

CK levels in MFMs are either normal or only slightly (<8‐fold) elevated. Exceptions from this rule are BAG3‐ and myotilin‐related MFMs, in which more marked increases of CK levels (up to 15‐fold) have been reported.

Electromyography

Needle electromyography in MFM patients usually shows a myopathic pattern with short‐duration polyphasic motor unit potentials with decreased amplitudes. In addition, fibrillation potentials, positive sharp waves and myotonic and pseudomyotonic phenomena are frequent electromyographic features. Some MFM patients display a mixture of myopathic and neurogenic findings.

Interpretation of clinical data

MFMs are still an underdiagnosed group of hereditary myopathies. Thus, we would like to stress the point that the differential diagnosis of MFMs should be taken into consideration in basically all patients suffering from a myopathy of unknown aetiology. In particular, in patients with clinical and morphological features of sporadic inclusion body myositis, MFM should be considered as a top differential diagnosis. Certain clinical constellations are highly indicative for special subtypes of MFM. In patients presenting with a distal myopathy phenotype, a disease onset in the third or fourth decade of life and additional evidence of cardiac pathology, a desminopathy should be suspected. Painless proximal muscle weakness in association with cataracts and cardiomyopathy starting in the third or fourth decade should raise the suspicion of an αB‐crystallinopathy. An epidermolytic skin disorder since birth in conjunction with a myopathy is pathognomonic for plectin‐related EBS‐MD. A disease onset beyond the fourth decade of life should alert the clinician to putative filamin C‐, ZASP‐ or myotilin‐related MFM. Here, distal myopathy phenotypes with facial and bulbar symptoms, myalgia and joint contractures point toward a myotilinopathy, whereas LGMD phenotypes with cardiac involvement and polyneuropathy are in keeping with the diagnosis of ZASP‐ and filamin‐C related MFMs. In all non‐facioscapulohumeral muscular dystrophy‐related scapuloperoneal syndromes, desmin and FHL1 gene mutations should be considered.

MFM: MYOPATHOLOGICAL CLUES TO THE DIAGNOSIS

Routine muscle biopsy workup

Formalin fixation of muscle tissue is the early death of a good myopathalogical analysis of MFM muscle specimens. Proper handling of biopsy specimens (beware of freezing artifacts!) cryopreservation and separate storage of muscle in 4% glutaraldehyde for electron microscopy (EM) are mandatory. A standard light microscopic workup should at least include the following stains: hematoxylin & eosin (H&E), modified Gomori trichrome (G‐Tri), reduced nicotinamide adenine dinucleotide tetrazolium reductase (NADH‐TR), ATPase at different pHs (4.2, 4.6 and 9.4), periodic acid Schiff (PAS), oil red O, succinic dehydrogenase (SDH) and cytochrome‐C oxidase (COX).

A broad spectrum of light microscopic changes ranging from mild to severe degenerative muscle alterations has been reported in MFM biopsy specimens. In addition to myopathic features (eg, rounding of muscle fibers, pathological fiber size variation, internally located myonuclei, fiber splitting), subsarcolemmal and/or sarcoplasmic protein aggregates (basophilic or eosinophilic in H&E stains; dark blue or pink in G‐Tri), cytoplasmic bodies, rimmed and non‐rimmed vacuoles, rubbed‐out fibers and core‐like lesions are the classical myopathological findings in MFM 8, 17, 35 (Figure 2). Spheroid bodies, which appear as coiled aggregates arranged in linear packets of greenish material in G‐Tri stains, are highly indicative of myotilinopathy (Figure 3). Sarcoplasmic reducing bodies appearing as dark green or reddish inclusions in G‐Tri stains and showing a positive reaction with menadione–NBT clearly point toward X‐linked FHL1opathy (Figure 4). A recent study on genetically confirmed MFM cases pointed out that rubbed‐out fibers tend to be more frequent in desminopathy and αB‐crystallinopathy, whereas rimmed and non‐rimmed vacuoles are more prominent in myotilinopathy and ZASPopathy (8). Furthermore, oxidative enzyme abnormalities comprising fibers with focal areas of attenuated or absent NADH‐TR, SDH and COX staining, or areas with increased enzymatic staining as well as a low amount of COX‐negative fibers are frequent in MFM 1, 8, 23, 31 (2, 5). In contrast, ragged red fibers and an abundance of necrotic muscle fibers do not belong to the classical MFM spectrum. While the outlined features are fairly characteristic, certainly not all MFM muscle biopsies meet these criteria. In cases with only subtle muscle pathology, the diagnostic findings are easily overlooked, and unspecific or even normal muscle biopsy analyses have been described in genetically confirmed MFM cases 42, 45. On the other side of the spectrum, a genetically confirmed desminopathy with the myopathological presentation of a PAS‐positive vacuolar myopathy has recently been reported (10). Thus, immunostaining and often EM are essential diagnostic tools for establishing the definite diagnosis of MFM.

Figure 2.

Histopathological findings in myofibrillar myopathies (MFMs). Hematoxylin & eosin (H&E) (A) and Gomori trichrome (G‐Tri) (B) staining in a genetically proven desminopathy. Arrows indicate the presence of isolated sarcoplasmic and subsarcolemmal protein aggregates (bars = 50 µm). H&E (C) and G‐Tri (D) staining in a genetically proven myotilinopathy and in a patient with MFM of unknown aetiology, respectively. Note the vacuolar changes (arrows) in myotilinopathy (bar = 75 µm) and the polymorphic protein aggregates in MFM of unknown aetiology (bar = 50 µm). Succinic dehydrogenase (E) and cytochrome‐C oxidase (F) staining in a genetically proven desminopathy. Note the presence of rubbed‐out fibers (*) and multiple core‐like lesions (bars = 40 µm). Bar in (A), (B), (D) = 50 µm. Bar in (C) = 75 µm. Bar in (E), (F) = 40 µm.

Figure 3.

Spheroid bodies (arrows) appear as coiled aggregates arranged in linear packets of greenish material in Gomori trichrome stain (A). At the ultrastructural level (B), a spheroid body is a well‐circumscribed and demarcated body (asterisk). This body is devoid of mitochondria or other cytoplasmic organelles. A spheroid body is highly indicative of myotilinopathy. Bar in (A) = 30 µm.

Figure 4.

Reducing bodies are characterized by the presence of intracytoplasmic inclusion bodies strongly dark blue stained in the menadione‐linked α‐glycerophosphate dehydrogenase preparation without substrate, α‐glycerophosphate. Here (A), an Azan stain nicely reveals numerous reducing bodies in many muscle fibers (arrows). Electron microscopy (B) shows reducing bodies (asterisk) composed of dense osmiophilic material consisting of closely packed granulofibrillar particles. Myofibrils with reducing bodies have disorganized cross‐striations. This condition is also commonly associated with rimmed vacuoles and cytoplasmic bodies. A reducing body is highly indicative of FHL1opathy. Bar in (A) = 40 µm.

Figure 5.

Myopathological findings in an epidermolysis bullosa simplex with muscular dystrophy patient with a homozygous plectin mutation (31). Hematoxylin & eosin (A) and cytochrome‐C oxidase (COX) (B) stains demonstrate severe myopathic and mitochondrial alternations. * denotes a COX‐negative fiber. (C) Note the subsarcolemmal and sarcoplasmic accumulation of desmin‐positive material. (D) Desmin immunogold electron microscopy shows highly unordered desmin filaments labeled by multiple gold particles. * denotes cross‐sectioned myofibrils. Bar in (A) = 50 µm.

Immunostaining

Plethoras of accruing proteins have been described in MFM (for details, please refer to 11, 34). With respect to their diagnostic value and commercial availability, antibodies directed against desmin, αB‐crystallin and myotilin have demonstrated to be sensitive diagnostic tools to depict pathological protein aggregation in MFM. Indeed, the extent of pathological protein aggregation can be highly variable both in terms of quantity and quality in MFM. While some MFMs show an abundance of labeled aggregates, others only display scarce protein aggregation pathology. Special emphasis should be put on the identification of desmin‐, αB‐crystallin‐ or myotilin‐positive band‐like structure in the subsarcolemmal region and polymorphic sarcoplasmatic aggregates (Figure 6). To avoid diagnostic confusion, the interpretation of muscle biopsies displaying structures with increased desmin, αB‐crystallin or myotilin labeling should take into account that central cores, minicores, as well as neurogenic target fibers also show increased immunoreactivity with these antibodies 14, 33. Other indirect signs hinting at the diagnosis of MFM are the labeling of membranous structures within muscle fibers with antibodies directed against sarcolemmal proteins, such as dystrophin, sarcoglycans and caveolin‐3. Because of an overlap of myopathological findings between MFM and VCP‐related inclusion body myopathy associated with Paget's disease of bone and frontotemporal dementia (IBMPFD), and cases of inclusion body myopathy/myositis, differentiating these disorders from another can be a challenging diagnostic task (21). As nuclear inclusions with the exception in myotilinopathy and sarcolemmal expression of MHC‐1 are absent in MFM, antibodies directed against MHC‐1 and ubiquitin, preferentially in conjunction with 4,6‐diamidino‐2‐phenylindole, are useful diagnostic markers to delineate these disorders from each other. Although automated immunoperoxidase techniques are widely in use and have several practical advantages, indirect immunofluorescence is clearly our preferred immunolabeling method in the assessment of protein aggregate pathology in MFM.

Figure 6.

Indirect immunofluorescence findings in myofibrillar myopathies. Desmin (A–C) staining in genetically proven desminopathies. Note the high variability of protein aggregate formation. Arrows in (A) denote sarcoplasmic and arrowheads subsarcolemmal aggregates. In (B), desmin‐positive aggregates are restriced to the subsarcolemmal region. An extensive vacuolar presentation of a desminopathy is demonstrated in (C). Desmin‐ and αB‐crystallin‐positive inclusions in genetically proven myotilinopathy and desminopathy are highlighted in (D) and (E), respectively. (F) Caveolin‐3‐positive membranous structures within muscle fibers of a genetically proven desminopathy. Bar in (A) = 60 µm. Bar in (C) = 100 µm. Bar in (B), (D), (E), (F) = 50 µm.

EM

As EM has a central role in the diagnostic workup of MFM, it is rather unfortunate that the appropriate expertise in ultrastructural pathology is slowly but unmistakably fading in many neuromuscular and histopathology laboratories. In suspected MFM/protein aggregate myopathy cases, EM analysis should particularly focus on the identification of pathological protein aggregation and signs of myofibrillar degeneration. The following baseline of pathological ultrastructural findings may serve as a useful diagnostic MFM checklist. With regard to pathological protein aggregation, specifically focus on the identification of: (i) sarcoplasmic granulofilamentous material in the subsarcolemmal and intermyofibrillar region (Figure 7A,B); (ii) sarcoplasmic and/or nuclear filamentous inclusions (Figure 7C); (iii) cytoplasmic bodies; (iv) spheroid bodies (Figure 3B); (v) reducing bodies (Figure 4); (vi) tubulofilamentous inclusions (Figure 7D); (vii) autophagic vacuoles; and (viii) membranous/myelin‐like whorls. To address the aspect of myofibrillar degeneration, specifically look for: (i) Z‐line alterations (streaming, irregularities, Z‐line loss, rods); (ii) myofibrillar remnants; and (iii) cores and core‐like lesions. Furthermore, the additional features should be addressed: (i) apoptotic myonuclei; and (ii) areas with depletion or accumulation of mitochondria. The interpretation of findings should consider several important aspects. Sarcoplasmic granulofilamentous material in conjunction with signs of myofibrillar degeneration is the classical ultrastructural hallmark of MFM. A recent study pointed out that the extent of granulofilamentous material is more prominent in desminopathy and αB‐crystallinopathy, whereas patients with ZASP, myotilin and filamin C mutation tend to display more filamentous cytoplasmic inclusions (7). However, as desmin‐positive granulofilamentous material has recently been reported in a genetically confirmed case of VCP‐related IBMPFD (21), this feature is not entirely specific for MFM. The presence of spheroid bodies or reducing bodies is highly indicative of myotilin and FHL1 mutations, respectively 16, 47. With the exception of myotilinopathy (7), MFM muscle biopsy specimens do not contain nuclear inclusions, which allows a distinction from sporadic and hereditary inclusion body myopathy/myositis. Here, it is important to note that cytoplasmic tubulofilamentous inclusions as well as autophagic vacuoles are shared features of MFM and inclusion body myopathy/myositis pathology.

Figure 7.

Electron microscopy findings in myofibrillar myopathy. (A,B) Granulofilamentous material (*) in genetically proven desminopathy. Note the clustering of mitochondria (arrow) in close relation to granulofilamentous material and myofibrillar structures. (C,D) Filamentous sarcoplasmic inclusions (*) in genetically proven filaminopathy. Arrows denote Z‐disk remnants.

Western blotting

As the vast majority of MFMs, caused by desmin, αB‐crystallin, ZASP, myotilin, filamin C and BAG3 mutations, have heterozygous amino acid substitutions or small in‐frame deletions, diagnostic differences in the expression of wild‐type and mutant proteins cannot be addressed by standard immunodetection techniques. Furthermore, the total desmin, αB‐crystallin and myotilin protein contents in MFM muscle seem to be unchanged when compared to normal controls. With the exception of some rare mutations in desminopathy and αB‐crystallinopathy, in which the individual mutation type led to a truncation of the mutant protein 10, 35, standard Western blotting does not play a diagnostic role in desmin‐, αB‐crystallin‐, ZASP‐, myotilin‐ and filamin C‐related MFMs. However, X‐linked FHL1 and homozygous plectin mutations lead to a reduction (FHL1, plectin) or even absence (plectin) of protein expression, and thus, can be helpful in terms of diagnostic evaluation 31, 47. Furthermore, heat shock protein 27 expression pattern analysis by two‐dimensional gel electrophoresis has been reported as a diagnostic tool to differentiate desminopathy from other MFM forms (9).

Genetic analysis

As outlined in this review, the careful interpretation of clinical and myopathological findings may provide essential clues for establishing the diagnosis of a distinct MFM disease entity. Nevertheless, the identification of a pathogenic mutation is nowadays the gold standard in making the diagnosis of a hereditary neuromuscular disorder. However, this approach is often hampered by the lack of specialized national laboratories, which can offer an appropriate and affordable genetic service.