Desminopathies: pathology and mechanisms

Since detailed epidemiological studies on MFM are not available, the incidence and prevalence of desmin myopathy and/or cardiomyopathy are currently unclear. Interpretation of the thus far published data allows the assumption that desminopathies fulfill the definition of a rare disease with no more than 5 affected individuals in 10,000. In a study on the prevalence of desmin mutations in a cohort of 116 families and 309 additional patients with pure dilatative cardiomyopathy, desmin mutations accounted for up to 2 % of disease manifestation [178]. Desmin mutations also seem to be one of the more frequently encountered gene defects in the MFM group. In a cohort of 53 patients from 35 Spanish MFM families, myotilin mutations were the predominant cause affecting 18 families followed by desmin mutations in 11 families [130]. In earlier studies reporting on the Mayo MFM cohort of 63 patients, 6 carried myotilin and 4 carried desmin mutations [165, 166]. Desminopathies have been reported in diverse ethnic groups and affect both female and male patients. Gender effects have been reported in two studies, in which male heterozygous mutation carriers were more prone to cardiac disease manifestations [5, 183]. The disease manifestation is highly variable with an age of onset ranging from the 1st to the 8th decade of life. In rare recessive forms the disease manifests in the 1st decade of life [31, 58, 121, 140]. In the more frequently encountered familial and sporadic cases due to heterozygous desmin mutations, a disease onset ranging from the 2nd to the 4th decade of life has been reported in the majority of patients [160, 183].

Initially, desminopathies have been associated with a progressive distal myopathy phenotype starting in the lower legs. However, subsequent studies reported the association between desmin mutations and limb girdle, scapuloperoneal, and generalized myopathy phenotypes [9, 35, 191]. A meta-analysis based on the interpretation of published data from 159 patients carrying 40 different heterozygous desmin mutations provided highly valuable insights into this complex issue [183]. Signs of combined distal and proximal muscular weakness were found in two thirds (67 %; 71/106) of mutation carriers, whereas true distal and proximal myopathy phenotypes were only present in 27 % (29/106) and 6 % (6/106), respectively. In this study, 74 % (110/148) of carriers had signs of skeletal muscle disease. Isolated skeletal muscle disease was reported in 22 % (31/141) of carriers. A combination of signs of skeletal muscle and cardiac pathology was found in 49 % (67/137).

Creatine kinase (CK) levels in desmin mutation carriers are of limited diagnostic value; 57 % (62/109) of mutation carriers had elevated CK levels (91 % displayed a ≤4-fold increase). Remarkably, 30 % (25/83) of patients with manifest skeletal muscle disease were reported to have normal CK levels [183]. Needle electromyography (EMG) typically reveals a myopathic pattern with short duration, polyphasic, and low amplitude motor unit potentials. In addition, positive sharp waves, fibrillation potentials, and pseudomyotonic/myotonic discharges have frequently been documented in desminopathy patients [160]. Sensory and motor nerve conduction studies usually give normal results [59]. Muscle MRI studies pointed out that signal alterations in the gluteus maximus, semitendinosus, sartorius, gracilis, and peroneal muscles are early and predominant signs of desminopathies [48, 156].

Cardiac disease manifestations in desminopathies, which may precede, coincide with, or succeed skeletal muscle weakness, comprise true cardiomyopathy as well as various forms of cardiac conduction defects (CCD) and arrhythmias [92, 173, 178, 183, 185]. The above-cited meta-analysis revealed the presence of cardiological signs in 74 % (105/141) of desmin mutation carriers [183]. Isolated cardiological signs were reported in 22 % (34/152) of carriers. Out of 67 patients with verified cardiomyopathy (49 %; 67/138), 23 were classified as dilatative, 18 as unspecified, 16 as restrictive, 8 as hypertrophic, and 2 as arrhythmogenic right ventricular cardiomyopathy. Desmin mutations frequently lead to clinically symptomatic and asymptomatic ECG abnormalities (62 %; 83/133). In this group, CCD seems far more frequent (39 %; 52/133) than isolated arrhythmias (5 %; 6/133). A combination of both was found in 19 % (25/133) of carriers. A more detailed characterization of 77 patients with CCD revealed that atrioventricular block (47) and right bundle branch block (14) were the most prevalent manifestations. In 31 patients with arrhythmias, atrial fibrillation (9), ventricular premature beats (8), and ventricular tachycardia (7) were the most frequently detected ECG abnormalities. Thus, a basic electrophysiological workup should include a 12-lead surface ECG and a 24-h Holter ECG. The cardiac function is routinely assessed by transthoracic echocardiography. However, cardiac MRI has been proposed as a more sensitive diagnostic tool to detect focal cardiac pathology in early and clinically asymptomatic stages of desminopathy [173].

Patients with desmin mutations are at risk to develop respiratory problems. In the desminopathy meta-analysis 26 % (29/110) of carriers were reported to suffer from respiratory insufficiency [183]. Thus, blood gas analysis and spirometry are mandatory in patients with desminopathies. Cataracts, swallowing difficulties, intestinal pseudo-obstruction, and repetitive episodes of diarrhea and constipation have been reported as miscellaneous or putative disease-related symptoms [6, 58, 129, 183].

Desmin myopathies and cardiomyopathies are characterized by a progressive course and may change their initial clinical presentation. In a case report, the transformation of an initially hypertrophic cardiomyopathy into a restrictive and finally dilated cardiomyopathy has been documented [65]. A further study reported on a 10-year follow-up of 28 patients from 19 families with desmin mutations [190]. In 11 patients, primary skeletal muscle involvement was followed by cardiac disease after 6.3 ± 3.5 years, whereas in 16 patients with a primary cardiac disease manifestation skeletal muscle problems occurred after 6.8 ± 4.1 years. Out of the latter group, six patients presented with major cardiac complications. A progression from mild conduction defects to high degree conduction blocks requiring permanent pacing was observed in 8 out of 19 patients. In this cohort of 28 patients, 5 died at a mean age of 58.0 ± 6.5 years, accounting for a mortality rate of 17.8 % [190]. In the desminopathy meta-analysis, however, a mortality rate of 26 % (27/104) of desmin mutation carriers with a mean age 49 ± 9.3 years was reported [183]. Documented causes of death in both studies were sudden cardiac death, heart failure, respiratory insufficiency, chest infection, and iatrogenic complications of cardiac treatment. Both studies conclusively underline that the cardiac disease manifestation is the major cause of premature death in desminopathies.

Diagnostic skeletal muscle biopsies from patients with desminopathies usually show mild to severe signs of a degenerative myopathy with rounding of muscle fibers, fiber splitting, internalization of myonuclei, and increased connective and fat tissue. Pathological protein aggregates, the hallmark of MFM, generally emerge as subsarcolemmal and/or sarcoplasmic inclusions. In addition, sarcoplasmic bodies as well as rimmed and non-rimmed vacuoles may be present. The typical pathology of MFM is best visualized in H&E and modified Gomori trichrome (GT) stains (Fig. 1a, b). Enzymatic stains (NADH-TR, SDH, and COX) may show further characteristic oxidative enzyme and mitochondrial abnormalities comprising rubbed-out fibers and core-like lesions (Fig. 1c). The stage of disease progression in individual muscles often mirrors the severity of the observed myopathological changes. However, one should keep in mind that the myopathological picture of desminopathies is highly variable. The myopathological findings in genetically proven desminopathies range from no overt pathology over subtle myopathic changes with or without pathological protein aggregates to the picture of a vacuolar myopathy [35, 160, 161].

Desmin immunostaining is mandatory to depict desmin-positive pathological protein aggregates in the subsarcolemmal and/or sarcoplasmic region. In addition to the characteristic immunoreactivity to desmin (Fig. 2a), the pathological protein aggregates are positively stained by a wide variety of antibodies directed against cytoskeletal proteins (e.g., dystrophin, F-actin, filamin C, myotilin, plectin, synemin), heat shock proteins (e.g., αB-crystallin, Hsp27), ubiquitin, Alzheimer-related proteins (e.g., β-APP), and cyclin-dependent kinases (e.g., CDK2, p21). Out of this long list, stains for αB-crystallin (Fig. 2b), filamin-C, and myotilin are sensitive diagnostic tools to depict pathological protein aggregation in desminopathies and other forms of MFM [161, 166].

Granulofilamentous material in the subsarcolemmal region and/or between neighboring myofibrils is the classical ultrastructural hallmark of desminopathies (Fig. 3). However, granulofilamentous material is not specific for this disease entity as it has been described in other forms of MFM, in particular in αB crystallinopathies. Other features of pathological protein aggregation are cytoplasmic bodies and autophagic vacuoles. These changes are found in conjunction with signs of myofibrillar degeneration comprising Z-disc alterations (streaming, irregularities, loss, and rods), myofibrillar remnants, and core and core-like lesions. Typical signs of concomitant mitochondrial pathology are areas with accumulation or depletion of mitochondria with normal or abnormal morphology.

The human desmin gene (DES) on chromosome 2q35 is a single copy gene that spans over a length of approximately 8.3 kb and comprises nine exons coding for a 470-amino acid protein with a molecular weight of 53.5 kDa [106]. DES belongs to a gene cluster further comprising APEG1 (synonym SPEG or striated muscle preferentially expressed protein kinase) and CHPF. This gene cluster is most likely regulated by the DES 5′ locus control region (LCR), which has been identified 9–18 kb upstream of DES [177]. The currently available information on the epigenetic regulation of DES is still limited. The DES promoter contains a CpG-island also covering the DES transcription start site and exon 1. It has been described that the DES promoter is non-methylated regardless of its expression status. The expression of desmin in muscle cells seems to be activated by acetylation of histones H3 and H4 as well as methylation of histone H3 at lysine residue 4 (H3K4me2 and me3) around the transcription start site of DES. In non-muscle cells, the DES promoter is silenced by methylation of histone H3 lysine residue 27 (H3K27me3) [108].

Disease-causing mutations spread over the entire DES gene. They significantly cluster in exon 6, which encodes the C-terminal half of the coil 2 domain (Fig. 4). Mutations in this domain have primarily been associated with a skeletal muscle phenotype, whereas mutations residing in the head and tail domains of the desmin protein seem more frequently associated with a cardiac phenotype [183]. As of June 2012, 67 disease-causing mutations of the DES gene have been published. These mutations may lead to the expression of 61 different mutant forms of desmin (Table 1).

The vast majority of familial desminopathies follow an autosomal dominant mode of inheritance. The most frequent DES mutations are missense mutations leading to single amino acid substitutions. Splice site mutations causing the loss of exon 3 (p.Asp214_Glu245del), small in-frame deletions of one, three or seven codons, and frame shift mutations, which may lead to the expression of truncated desmin protein species, have been reported only in a small number of patients (Table 1). For the p.Arg350Pro mutation, the most frequently encountered pathogenic desmin missense mutation in Germany, a founder allele has been established [191].

Five families with an autosomal recessive mode of inheritance have been published thus far. In one family, the homozygous Arg16Cys missense mutation was observed [4]. In two families, a small in-frame deletion has been identified leading to p.Arg173_Glu179del [121, 140]. In another family, a 22 base pair deletion in exon 6 causing a premature stop codon associated with a virtually complete lack of desmin protein has been reported [31]. In the fifth family, the disease manifestation has been attributed to a compound heterozygote DES mutation (p.Ala360Pro/p.Asn393Ile [58]) (Table 1).

In addition to these familial cases, an increasing number of sporadic desminopathies have been published. These sporadic disease manifestations are due to missense, splice site, or frame-shift mutations (Table 1).

Human desmin is a 470-amino acid protein ([79, 153]; UniProtKB/Swiss-Prot database entry P17661) with a calculated molecular weight of 53.5 kDa. Like all IF proteins, it is fibrous in nature, exhibiting a tripartite structure with a central, mostly α-helical domain of conserved size, i.e., ~45 nm. This so-called “rod” is flanked by non-α-helical amino-terminal (“head”) and carboxy-terminal (“tail”) domains of highly varying amino acid number [73] (Fig. 4a). The “rod” domain comprises two continuous α-helical segments, coil 1 and coil 2, which are connected by a “linker” (L12) of unknown structure. The current structural model of desmin is derived from recent structural analyses of the closely related class III IF protein vimentin [33] (Fig. 4b). Coil 1 consists of two α-helical subdomains, i.e., coil 1A and coil 1B. They are separated by linker L1, which could also accommodate α-helical conformation. While coil 1B forms a stable dimeric coiled coil, the coil 1A segment has only a weak coiled-coil forming capacity. In addition, the current model omits the previously used separation into coil 2A and 2B domains, but demonstrates it to be a continuous α-helix [123].

Desmin is a highly insoluble protein and hence it can be kept in solution as a monomer and purified only under highly denaturing conditions as provided by buffers containing 8 M urea. For assembly, it is usually first re-natured by dialysis into buffers of low salt such as 2 mM sodium phosphate (pH 7.5). The assembly process starts actually during the course of re-naturation, and two molecules dimerize by coiled-coil formation of the central α-helical rod domains of two desmin monomers in parallel orientation [77]. A so-called heptad repeat pattern, which is characteristic for a coiled-coil structure [26], drives the supercoiling of the two helices, yielding the dimeric, elongated structure. Two of these coiled-coil dimers further associate in a half-staggered, anti-parallel fashion to a tetramer of ~60 nm length [77, 107] (Fig. 5a). These tetramers are able to spontaneously assemble into highly ordered long filaments upon increase of the ionic strength to physiological values. The assembly process can be described to occur in three phases: (1) The lateral parallel assembly of tetramers yielding full-width, 60-nm-long filaments that have been termed unit-length filaments or ULFs [71, 77] (Fig. 5b). (2) The formation of extended intermediate filaments by serial longitudinal annealing of ULFs and further longitudinal annealing of short filaments [71] (Fig. 5c1,c2,c3,d). (3) After a few minutes of assembly, filaments undergo a final maturation step characterized by a radial compaction process [76, 79].

Desmin is the most abundant IF protein in striated and smooth muscle cells. Immunoblotting after one-dimensional SDS-PAGE with desmin-specific antibodies shows a single band corresponding to an apparent molecular weight of 58 kDa. In addition to muscle, desmin expression has been described in a wide variety of normal and diseased cells, for example, pericytes [24], hepatic stellate cells [124], myoid stromal cells of placenta [170], Sertoli cells [148], decidual cells [67], injured glomerular podocytes [70], and mesotelioma cells [85].

Desmin is one of the earliest markers of muscle development [27], and this is true for all vertebrates including amphibian and fish, pointing to an important evolutionary conservation [75, 153]. Its expression precedes all other muscle-specific structural proteins and even—with the exception of myf-5—the expression of the myogenic transcription factors myoD, myogenin, and mrf4 (myf-6) [99, 103, 112]. During early muscle development desmin and vimentin are co-expressed. Vimentin is the most abundant IF protein of immature myoblasts. Upon further differentiation into mature muscle cells, desmin is strongly upregulated, while the expression of vimentin is completely ceased [43, 152].

Post-translational modifications do play an important role in regulating the functions of intermediate filaments. In vitro translation of chicken skeletal and smooth muscle desmin using a cell-free rabbit reticulocyte system resulted in predominantly non-phosphorylated desmin, suggesting a low amount of phosphorylated desmin under basal conditions [125]. On the other hand, desmin has been described as one of the major acceptors of ³²P in differentiating chicken myotubes. Two-dimensional tryptic analysis of desmin revealed multiple sites of phosphorylation [52]. Another study showed that desmin is a substrate of the p21-activated kinase (PAK) mainly leading to phosphorylation of serine residues within the desmin “head” domain. PAK-mediated phosphorylation of desmin strongly inhibited its filament-forming ability [127]. Assembly is also inhibited by ADP ribosylation of desmin via an arginine-specific mono-ADP-ribosyltransferase of muscle, which primarily targets arginine 48 and to a lesser extent arginine 68 within the desmin “head” domain [201]. A subsequent study provided evidence that ADP-ribosylated desmin neither co-assembles with nor affects the filament formation of non-modified desmin. In contrast, the ADP-ribosylation of pre-formed desmin IFs resulted in their disassembly. Furthermore, this process is dependent on the phosphorylation of additional amino acid residues [198].

Desmin immunostaining of cross-sectioned striated muscle revealed sarcoplasmic and subsarcolemmal localizations. In longitudinal sections, desmin immunostains revealed a cross-striated pattern [162]. In addition, desmin is enriched at the level of myotendinous and neuromuscular junctions of skeletal muscle as well as of intercalated discs in cardiac muscle [30, 180]. Immunogold electron microscopy demonstrated the presence of desmin at costameres, filamentous structures spanning between myofibrils and the overlaying sacolemma, filamentous inter-Z-disc structures of neighboring myofibrils and mitochondria, and at the desmosomal plaque of intercalated discs, where desmin IFs bind to desmoplakin, a member of the plakin family of crossbridging proteins [90, 111, 146, 162].

This three-dimensional filamentous extra-sarcomeric desmin cytoskeleton interlinks neighboring myofibrils (Fig. 6) and connects the myofibrillar apparatus with myonuclei, other cell organelles, and the extracellular matrix via the subsarcolemmal cytoskeleton [27, 160]. Beyond a sole mechanical integration, this complex interaction is thought to be the basis for a mechano-chemical signaling between various compartments. IFs in general and desmin in particular have also been hypothesized to directly bind to genomic DNA and to exert a role in transcription regulation and DNA organization [182]. Roles of desmin in the morphology and homeostasis of myonuclei, mitochondria, and lysosomes are discussed in the respective paragraphs on muscle pathology, desmin binding partners, and desmin mouse models. Biomechanical aspects of desmin at the level of single IFs, myoblasts, myofibers, and whole muscle are summarized in the respective paragraph on biomechanics.

Desmin exerts its multiple functions through direct and indirect binding to various other cellular molecules. In the past 4 decades of desmin research, a growing number of desmin binding partners has been described (see also a previous review on desmin interactions [38]). The following description of desmin interactions is confined to binding partners that are present in muscle cells, i.e., IF proteins, IF-associated proteins, sarcomeric proteins, membrane-associated proteins, small heat shock proteins, apoptosis-related proteins, and nucleic acids.

Analysis of single skeletal muscle fibers from a patient with the heterozygous p.Arg350Pro desmin mutation showed the presence of pathological desmin-positive protein aggregates in conjunction with the normal cross-striated desmin staining pattern [158, 159] (Fig. 7). This finding raises the question whether mutant desmin forms mixed desmin IFs or mixed aggregates with wild-type desmin or if the mutant protein segregates from the wild-type protein. This issue cannot be solved using commercially available desmin antibodies as they do not distinguish between the wild-type and the point-mutated desmin protein species.

Studies on desmin protein expression in desminopathies due to deletion or frame-shift mutations would allow at least the determination of the expression level of the mutant desmin species. However, only two studies reported such data yet. In one study immunoblotting revealed two desmin bands in cardiac and skeletal muscle with the lower band corresponding to a truncated desmin protein species showing an intensity of approximately 30 % of the upper wild-type band. However, no desmin mutation analysis was reported [3]. In another study on a desminopathy due to a heterozygous c.735G>C mutation, desmin immunoblotting revealed the presence of two bands at the positions of 53 kDa and 50 kDa. However, RT-PCR analysis in this particular family identified three distinct desmin mRNA species coding for wild-type and p.Glu245Asp desmin, which both are visible at 53 kDa, as well as p.Asp214_Glu245del corresponding to the truncated 50-kDa protein. Here, the amount of the truncated desmin protein varied between 5 and 43 % [35]. Two-dimensional desmin gel electrophoresis in desminopathies revealed non-uniform results. In a desminopathy due to the heterozygous p.Lys240del mutation, an aberrant and more acidic spectrum of desmin spots was visible [158, 159], whereas the p.Asp214_Glu245del desmin mutant species presented with a more alkaline pI [35].

Desmin has been found to be oxidated and nitrated in muscle fibers containing pathological protein aggregates. These modifications may have a direct cytotoxic effect and may impair the degradation via the ubiquitin-proteasom system [87]. Beyond specific desmin modifications, skeletal muscle samples from desminopathies exhibited increased levels of glycoxidated, lipoxidated, and nitrated proteins [88].

A study demonstrated an increased immunoreactivity of the 20S proteasome core as well as its 19S and 11S (synonyms: PA28alpha/beta, PSME1/PSME2) regulators co-localizing with pathological protein aggregates in desminopathies. Further components of the immunoproteasome have also been reported to colocalize with the protein aggregates [46]. Another study demonstrated an enrichment and co-localization of the mutant ubiquitin UBB+1 as well as the autophagy-related p62 with desmin-positive protein aggregates in desminopathies [131]. Desmin-positive protein aggregates in desminopathies also show a positive immunoreactivity with antibodies directed against HspB1 (synonyms: Hsp25, Hsp27) and HspB5 (synonym: αB-crystallin) [160]. With regard to HspB1, two-dimensional gel electrophoresis demonstrated a shift of the main HspB1 spot to a more alkaline pI [36]. Further relationships between the expression of mutant desmin and the ubiquitin–proteasome system, autophagy, and heat shock proteins are provided in the respective paragraphs on desmin mouse models.

Desmin transfection studies have been performed in a variety of muscle and non-muscle cell types: BHK21 (ATCC CCL-10) hamster kidney fibroblasts express desmin and vimentin intermediate filaments [80]. HL-1 cardiac [34], C2C12 (ATCC CRL-1772), and inducible C2.7 [141] skeletal muscle-derived mouse myoblasts, human coronary artery smooth muscle (hcasmc), and neonatal rat cardiac ventricular myocytes (nrcm) [178] as well as C3H/10T1/2 (ATCC CCL-226) mouse fibroblasts ectopically expressing Myf5 or MyoD all contain desmin and vimentin IFs. 3T3 (ATCC CRL-1658) mouse fibroblasts contain vimentin. Vimentin-free mouse embryonic fibroblasts (MEFs) isolated from vimentin knockout mice and spontaneously immortalized do not express cytoplasmic IF proteins [37]. Moreover, both MCF7 (ATCC HTB-22) human epithelial cells and bovine mammary gland epithelium cells grown in the presence of hormones BMGE+H [154] are vimentin-free and express only keratin IF proteins. SW13 (ATCC CCL-105) human epithelial cells do not express any IF protein.

The overall data derived from studies using these cell types indicated that the majority of desmin mutants are incapable of forming a de novo desmin IF network, but instead form non-IF structures and desmin-positive protein aggregates. In addition, most of them induce the collapse of a pre-existing IF network such as in 3T3 cells, although several mutant desmin proteins integrate well, just like the wild-type protein, into the vimentin network [9, 11, 16, 167]. As an example, we depict here the transfection of the p.Arg406Trp desmin rod mutant into various cell types. In the vimentin-free SW13 and BMGE+H cells, it forms dot-like and short rod-like structures (Fig. 8a, b); in addition, in 3T3 cells it completely segregates from the endogenous vimentin filaments and causes the reorganization of vimentin IFs around the nucleus (Fig. 8c). Similarly, the p.Leu345Pro desmin mutant completely avoids integrating into the vimentin system (Fig. 8d–f); however, some affinity for vimentin is observed in extended vimentin IFs below the cell nucleus and outside of the area of the collapsed vimentin IFs (Fig. 8f, arrowheads denote green desmin dots aligning with the red vimentin IFs). In the collapsed area, the green and the red signals superimpose to yield a yellow signal; however, the signals differ entirely in shape, dots versus fibers, indicating that they are not contained within the same structure but are only located close to each other and thereby generate a yellow signal.

The first disease mutant desmin to be analyzed at the in vitro level for its assembly properties was the rather drastic p.Arg173_Glu179del desmin “rod” mutation, which misses seven amino acids in a row in coil 1B. In both cell culture after cDNA-transfection and after forced assembly of recombinant proteins, only non-IF structures were obtained [121]. Later on, several other desmin disease mutants were studied, and their ability to form IF networks in vivo was analyzed by cellular cDNA transfection [11]. In order to reveal how desmin mutations influence basic filament assembly properties, a consecutive study investigated their assembly properties at the protein level [12]. Four major classes of pathological effects were observed (Fig. 9): (1) after 10 s of assembly, filaments form by lateral association of tetramers and subsequent longitudinal annealing of ULFs (see also Fig. 5); however, upon further incubation these IF-like filaments show an abnormal lateral annealing leading to the formation of large “sheets” (Fig. 9, p.Asn342Asp); (2) a second class forms IFs similar to those generated from desmin WT, although the filaments are less regular (Fig. 9, p.Ala360Pro); (3) other mutants exhibit extended filaments at 10 s of assembly, but thereafter immediately decay into ball-like aggregates [13] (Fig. 9, p.Leu370Pro); (4) filament assembly starts with apparently normal ULFs; however, they fail to longitudinally anneal as observed with wild-type desmin and instead stay at the ULF-state or loosely associate longitudinally such that the sub-filament structure remains visible (Fig. 9, p.Arg406Trp). Notably, all desmin disease mutants that were not able to form stable filaments on their own in vitro also did not form IFs after transfection into IF-free cells. Instead, they did segregate from the endogenous IF system when transfected into vimentin- or desmin-containing cells [13]. While the finding that truncated desmin mutants did not properly assemble is not that unexpected, the observed high pathogenicity of missense mutations suggests that desmin filament assembly is a delicate process that can be easily distorted.

A characteristic biophysical property of IFs is their elasticity. In response to mechanical stretch, desmin filaments have been shown to extend in length in conjunction with a thinning of their diameter. Experiments demonstrated that upon a 3.4-fold extension, desmin filaments reduced their diameter from 12.6 to 3.5 nm [98]. Moreover, the phenomenon that IFs become more viscoelastic when exposed to an external mechanical force is referred to as “strain stiffening” [86, 155]. In striated muscle, the elasticity of desmin filaments has been attributed to play a cell-protective role against mechanical stress. Thus, desmin filaments may dissipate mechanical energy during muscle contraction.

In this context, mutant desmin may exert a pathogenic effect on the viscoelastic properties of the desmin filament system, which consecutively may lead to progressive muscle fiber damage and muscle weakness. The viscoelastic properties of several disease-associated mutant desmin protein species have been analyzed. Here, the focus was on desmin mutants that are able to form apparently normal filament networks in vitro and in vivo. Some of these mutants showed nanomechanical properties similar to that of wild-type desmin, while others displayed changes in their tensile properties with reduced strain stiffening. Such an increase in the resistance of desmin filaments to an external pulling force may inflict changes in the adaptability of muscle cells to mechanosensing and mechanotransduction [14, 97]. Analysis of primary cultured myoblasts derived from a patient with a heterozygous p.Arg350Pro desmin mutation revealed an aberrant response to mechanical stress. These cells displayed increased cell stiffness and a higher rate of cell death and substrate detachment [22]. A number of biomechanical studies on mice lacking desmin have been reported. One study provided evidence that the lack of desmin is associated with a reduction of the overall passive elasticity of intact isolated soleus muscle [2]. Another study focusing on the diaphragm reported a reduced stiffness and viscoelasticity of this muscle together with an increased tetanic force production [23]. A further study showed a reduced passive tensile strength after eccentric contraction associated with disrupted Z-discs in individual myofibrils. In addition, the maximal isometric force production in the extensor digitorum longus muscle was reported to be decreased, which points to a possible involvement of desmin in excitation–contraction coupling [149]. This assumption was further underlined by an additional study reporting a defective excitation–contraction coupling in intact extensor digitorum longus and soleus muscles of desmin knockout mice. Here, the increased fatigue in extensor digitorum longus muscle was assessed by continuous high frequency field stimulation and could be overcome by the addition of caffeine. This combination of results argues against a defect in the sarcoplasmic reticulum calcium storage capacity [197]. However, this interpretation was challenged by yet another study, which used a more physiological high frequency stimulation protocol with titanic trains. In this setting, a reduced fatigue in intact soleus muscle without any change in the calcium sensitivity of the contractile apparatus was observed [7]. Finally, a study described that the absence of desmin resulted in an increased number of branched myofibers in the flexor digitorum brevis muscle. This work reported neither differences in the resting sarcoplasmic calcium concentration nor in the sarcoplasmic calcium release amplitude in desmin knockout myofibers [61].