Research - what's new? 2004

A sample of MAJOR articles from the medical literature.

Primary respiratory failure in inclusion body myositis.
Voermans NC, Vaneker M, Hengstman GJ, ter Laak HJ, Zimmerman C, Schelhaas HJ, Zwarts MJ. Neuromuscular Centre Nijmegen, Department of Neurology, University Medical Centre Nijmegen, The Netherlands.
Neurology. 2004 Dec 14;63(11):2191-2.
Case Report
The idiopathic inflammatory myopathies are a group of disorders characterized by acquired muscle weakness and presence of inflammatory infiltrates in skeletal muscle. The three most common diseases within this group are dermatomyositis (DM), polymyositis (PM), and inclusion body myositis (IBM). Respiratory muscle weakness with respiratory failure is a well-recognized complication in PM and DM but has only rarely been reported in IBM. Symptomatic respiratory failure in IBM is considered to be secondary to coincidental pulmonary disease. We report a patient with IBM who developed subacute respiratory failure caused by primary respiratory muscle weakness.

Tau aggregates are abnormally phosphorylated in inclusion body myositis and have an immunoelectrophoretic profile distinct from other tauopathies.
Maurage CA, Bussiere T, Sergeant N, Ghesteem A, Figarella-Branger D, Ruchoux MM, Pellissier JF, Delacourte A. INSERM U422, Faculte de Medecine, 1 place de Verdun, Lille cedex, France. ca-maurage@chru-lille.fr
Neuropathol Appl Neurobiol. 2004 Dec;30(6):624-34.
Sporadic inclusion body myositis (s-IBM) is the most frequent progressive acquired inflammatory myopathy in people older than 50 years. Abnormal aggregates of 'Alzheimer's proteins', including tau proteins, have been previously demonstrated in s-IBM. In the present study, we have investigated by immunohistochemistry and immunoblotting analysis the presence of tau proteins in muscle biopsy samples from patients with s-IBM and other myopathies with rimmed vacuoles, using newly developed antibodies raised against tau protein epitopes found in Alzheimer's disease brain. Tau immunoreactivity was shown in rimmed vacuoles or inclusions, preferentially with antibodies directed against phosphorylated carboxy-terminal epitopes of tau proteins. Cytoplasmic reactivity was also demonstrated in atrophic, nonvacuolated fibres, as well as in non-necrotic fibres invaded by inflammatory cells. Abnormally phosphorylated tau aggregates were also found in other compartments of the muscle fibre in s-IBM and other myopathies. Tau immunoblotting showed an electrophorectic profile of a doublet within the range of 60-62 kDa isovariants, which was different from tauopathies of the central nervous system. Finally, the unique pattern of immunoreactivity of s-IBM samples towards anti-tau antibodies is a new clue to a possible distinct subclass of peripheral tauopathy, different from the tauopathies of the central nervous system.

Associations with autoimmune disorders and HLA class I and II antigens in inclusion body myositis.
Badrising UA, Schreuder GM, Giphart MJ, Geleijns K, Verschuuren JJ, Wintzen AR, Maat-Schieman ML, van Doorn P, van Engelen BG, Faber CG, Hoogendijk JE, de Jager
AE, Koehler PJ, de Visser M, van Duinen SG; Dutch IBM Study Group.
Department of Neurology, K5Q, Leiden University Medical Center, PO Box 9600, 2300 RC Leiden, The Netherlands. ubadrising@lumc.nl
Neurology. 2004 Dec 28;63(12):2396-8.
Whether autoimmune mechanisms play a role in the pathogenesis of inclusion body myositis (IBM) is unknown. Human leukocyte antigen (HLA) analysis in 52 patients, including 17 with autoimmune disorders (AIDs), showed that patients were more likely to have antigens from the autoimmune-prone HLA-B8-DR3 ancestral haplotype than healthy control subjects, irrespective of the presence of AIDs.
Patients lacked the apparently protective HLA-DR53 antigen. The results provide further support for an autoimmune basis in IBM. I have included a pdf file of the whole ARTICLE HERE

The prion protein in human neuromuscular diseases.
Kovacs GG, Kalev O, Gelpi E, Haberler C, Wanschitz J, Strohschneider M, Molnar MJ, Laszlo L, Budka H. National Institute of Psychiatry and Neurology, Budapest, Hungary.
J Pathol. 2004 Nov;204(3):241-7.
The basis of human prion diseases affecting the nervous system is accumulation of a disease-associated conformer (PrPSc) of the normal cellular prion protein (PrPC). Earlier studies demonstrated increased expression of PrPC in inclusion body myositis (IBM), dermato-, and polymyositis, as well as neurogenic muscle atrophy. To define the spectrum and reliability of PrPC immunoreactivity, its expression was examined systematically in a series of pathologically characterized muscular disorders by means of immunohistochemistry, confocal laser microscopy, and immunogold electron microscopy. Anti-PrPC immunolabelling of rimmed vacuoles was observed in IBM, inclusions of myofibrillary myopathy, targets, regenerating, and atrophic fibres, mononuclear cells, in addition to ragged red fibres in mitochondrial myopathies, and focal sarcolemmal immunostaining in non-diseased controls. Quantitative analysis demonstrated that, in neurogenic muscle lesions, anti-PrPC staining detects a significantly broader spectrum of fibres than anti-vimentin or anti-NCAM. In dystrophic muscle, PrPC expression was mainly restricted to regenerating fibres. In IBM, PrPC expression was not confined to rimmed vacuoles or vacuolated fibres and only a small percentage (7.1%) of rimmed vacuoles were PrPC positive. Ultrastructurally, PrPC was observed in the cytoplasm of lymphocytes, in the myofibrillar network of targets, and in rimmed vacuoles. Knowledge of disease circumstances with altered expression of PrPC is important in the setting of a potentially increased chance for extraneural PrPC-PrPSc conversion. In addition, our observations suggest that PrPC may have a general stress-response effect in various neuromuscular disorders.
Introduction (p. 241)
Human prion diseases, or transmissible spongiform encephalopathies (TSEs), occur as sporadic, acquired, and genetic disorders [1]. Change in the conformation of the normal cellular form of prion protein (PrPC) produces a pathological, disease-associated PrP (PrPd or PrPSc) which accumulates predominantly in the brain. Recent studies indicate that PrPd may be present in skeletal muscle, spleen, vessel wall, and peripheral nerves in sporadic Creutzfeldt-Jakob disease (CJD) [2-5] in addition to widespread presence in the lymphoid tissue of variant CJD [6]. Yet, prion disease predominantly affecting extraneural sites has never been described.
Discussion: (p. 245-47)
In the present study we demonstrate a fairly uniform morphological pattern of increased PrPC expression in distinct muscular disease entities. PrPC appears in lymphocytes in inflammatory infiltrates, targets in neurogenic lesions, rimmed vacuoles in IBM, and regenerating and atrophic fibres in various diseases. In addition to these, PrPC expression is found in the sarcoplasm of normal-appearing fibres in IBM, neurogenic lesions, and muscular dystrophies. This is in contrast to nondiseased controls where PrPC expression is restricted to the sarcolemma. Occasional nerve branches and muscle spindles also show PrPC immunoreactivity which may be relevant to the accumulation of PrPd in skeletal muscle [22].
      In fact, quantification demonstrates in neurogenic lesions that anti- PrPC immunostaining detects a significantly broader spectrum of fibres than antivimentin or anti-NCAM, which are thought to identify regenerating and denervated fibres, respectively [23]. This could suggest a neurotrophic role for PrPC. However, this has not been confirmed in other studies, since neural regulation of the expression of acetylcholine receptors at neuromuscular synapses in skeletal muscle is not related to the PrP gene [24].
     In IBM, PrPC immunoreactivity is infrequent in rimmed vacuoles. The percentage of PrPC-positive fibres without rimmed vacuoles in Gömöri-stained sections is significantly higher than that of PrPC-positive vacuolated fibres. Thus, PrPC-positive vacuolated and non-vacuolated fibres, as well as inflammatory infiltrates, contribute to the increased expression of PrPC detected in western blotting [20]. Accumulation of PrPC in targets, and in tubulofilamentous-like structures in rimmed vacuoles in IBM, seems to represent a stereotyped response to myofibrillar breakdown or occasional co-aggregation [25]. This is supported by our observation that the majority of rimmed vacuoles in IBM do not contain PrPC. Another explanation could be that PrPC, as a potential stress-response protein, may be partially involved in a cytoprotective process similar to the ubiquitin-proteosome system. The latter is part of an intracellular surveillance complex aiming to repair altered proteins (eg by chaperones) or to isolate and eliminate these from the cell (eg by the autophagic-lysosomal system) [21]. This is supported by our and previous observations that ubiquitin is present in autophagic rimmed vacuoles (Figure 2f) [19]. We have seen sarcoplasmic ubiquitin immunoreactivity overlapping with that of PrPC, suggesting a common response of these proteins to disease-associated alterations. Interestingly, in myofibrillar myopathy, PrP-immunoreactive deposits do not overlap exclusively with specific desmin immunopositivity [26].
     As apoptosis and oxidative stress are common pathogenetic steps in the aforementioned disease states [27], and PrPC has a suggested antioxidant or antiapoptotic capacity, this may be one reason for its
higher expression [15,16].
     Recently, similar to our findings with PrPC, enhanced alpha-B-crystallin expression was reported in structurally intact fibres (so-called X fibres) in IBM and other diseases including polymyositis and myofibrillar myopathy [28]. In that study, in contrast to our findings with PrPC, alpha-B-crystallin-immunopositive X fibres were much more frequent in IBM than in other disorders. Thus, PrPC seems to have rather a more general responder effect to various stress actions irrespective of the type of neuromuscular disease, in contrast to the pathogenetic stressor action of alpha- B-crystallin mostly limited to IBM.
     Overexpression of wild-type hamster or sheep PrPC transgenes in transgenic mice produces a myopathy characterized by degenerating muscle fibres with some phagocytosis, variation in fibre size, and central nuclei [29]. It cannot be excluded that this may play a part in degeneration of muscle fibres in certain neuromuscular disease states with uncontrolled upregulation of PrPC expression. Whether PrPC is helpful or harmful in these circumstances has not yet been characterized. However, our observations suggest that PrPC may play a general stress response effect in various neuromuscular disorders.
     PrPC is known to have a role in T-lymphocyte activation, and macrophages also express PrP [30,31]. Here we show that, in inflammatory muscle disease, PrPC is more abundant in lymphocytes than in
     Knowledge of disease states with altered expression of PrPC is important when a potentially increased chance for extraneural PrPC-PrPSc conversion is considered. It may well be that patients with prion disorders who suffer from any kind of additional muscle disease with inflammation, neurogenic lesions, or widespread regeneration are more prone to accumulation of pathological PrPd in muscle [3].

* Prion, as defined by MedlinePlus at http://medlineplus.gov/, is "a protein particle that lacks nucleic acid and has been implicated as the cause of various neurodegenerative diseases (as scrapie, Creutzfeldt-Jakob disease, and bovine spongiform encephalopathy)."

Two major histocompatibility complex haplotypes influence susceptibility to sporadic inclusion body myositis: critical evaluation of an association with HLA-DR3.
Authors: P. Price; L. Santoso; F. Mastaglia; M. Garlepp; C.C. Kok; R. Allcock; N. Laing
Tissue Antigens, November 2004, vol. 64, no. 5, pp. 575-580(6)
Abstract: Previous studies of sporadic inclusion body myositis (sIBM) have shown a strong association with HLA-DR3 and other components of the 8.1 ancestral haplotype (AH) (HLA-A1, B8, DR3), where the susceptibility locus has been mapped to the central major histocompatibility complex (MHC) region between HLA-DR and C4. Here, the association with HLA-DR3 and other genes in the central MHC and class II region was further investigated in a group of 42 sIBM patients and in an ethnically similar control group (n = 214), using single-nucleotide polymorphisms and microsatellite screening. HLA-DR3 (marking DRB1*0301 in Caucasians) was associated with sIBM (Fisher's test). However, among HLA-DR3-positive patients and controls, carriage of HLA-DR3 without microsatellite and single-nucleotide polymorphism alleles of the 8.1AH (HLA-A1, B8, DRB3*0101, DRB1*0301, DQB1*0201) was marginally less common in patients. Patients showed no increase in carriage of the 18.2AH (HLA-A30, B18, DRB3*0202, DRB1*0301, DQB1*0201) or HLA-DR3 without the central MHC of the 8.1AH, further arguing against HLA-DRB1 as the direct cause of susceptibility. Genes between HLA-DRB1 and HOX12 require further investigation. BTL-II lies in this region and is expressed in muscle. Carriage of allele 2 (exon 6) was more common in patients. BTL-II(E6)*2 is characteristic of the 35.2AH (HLA-A3, B35, DRB1*01) in Caucasians and HLA-DR1, BTL-II(E6)*2, HOX12*2, RAGE*2 was carried by several patients. The 8.1AH and 35.2AH may confer susceptibility to sIBM independently or share a critical allele.
This study established that sIBM could be promoted by alleles characteristic of the 8.1AH or the 35.2AH. HLA-DR3 (8, 9) [or DRB1*03 (7)] has previously been associated with sIBM. However, when HLA-DR3-positive controls and sIBM patients were compared, carriage of HLA-DR3 without other components of the 8.1AH was less common in patients, suggesting HLA-DR3 is not the direct cause
of disease. This is supported by the lack of any association between sIBM and the 18.2AH (HLA-A30, B18, DRB1*0301) and represents a fundamental break with the traditional interpretation of HLA-DR3 associations. Hence, it is worth reviewing previous studies of the cohort and comparing the 8.1 and 18.2AH.
With a subset of the present cohort, Kok et al. (9) used novel markers of the 8.1AH in the HSP70 gene cluster to define a region of the 8.1AH between HLA-DR3 and C4A that was carried by all DR3-positive patients. The authors suggested that the RAGE gene might mediate the genetic differences, as RAGE is a receptor for the bA4 fragment of amyloid precursor protein present in the rimmed vacuoles of skeletal muscle in sIBM and senile plaques of the brain in Alzheimer's disease (16). However, the HLA-DR typing was serological and hence reflected the DRB1 gene. MHC class II genes telomeric of DRB1 were not considered. Here, carriage of DR3 with no other markers of the 8.1AH was less common in patients, but the effect was lost rapidly as we traversed the central MHC so the critical gene is likely to be close to DRB1.
The 18.2AH and the 8.1AH share HLA-DRB1*0301, DQA1*0501, and DQB1*0201 but differ in carriage of DRB3*0202 (18.2AH) rather than DRB3*0101 (8.1AH) and at all loci telomeric of this gene. DRB3 alleles have been associated with susceptibility to Graves' disease in African Americans (17) and Jamaicans (18). These studies compared the class II alleles of the 8.1 and 18.2AH (although this terminology was not used). Central MHC and other class II genes were not addressed and require further consideration, as the haplotypes diverge immediately telomeric of DRB1. This provides a precedent for consideration of DRB3 as a candidate gene. Other candidates (in chromosomal order away from DRB3) are DRA, BTL-II (13), TSBP (testis-specific basic protein), NOTCH4 (a transmembrane receptor which regulates cell fate decisions) (19), and G18 (a predicted gene with no known function) (20).
BTL-II lies between TSBP and DRA and is expressed in skeletal muscle. Carriage of allele 2 of exon 6 was more common in the patients. As BTL-II(E6)*2 is characteristic of the 35.2AH (HLA-A3, B35, DR1) in Caucasians, carriage of other haplotypic markers was investigated. A region of the 35.2AH defined by HLA-DR1, BTLI( E6)*2, HOX12*T, RAGE*T was more common among patients than controls. As several patients carried these alleles without any component of the 8.1AH, the two haplotypes may confer susceptibility to sIBM independently or may share a critical allele. Studies of further sIBM patients, including the rare cases from other ethnic groups, may help identify MHC genes responsible for susceptibility to sIBM.

Myositis: an update on pathogenesis
Lisa Christopher-Stine and Paul H. Plotz
Curr Opin Rheumatol 16:700-706. [November 2004]
Excerpt from article (pages 703-705)
Inclusion body myositis
It is debated whether sIBM is primarily immunemediated; some experts, particularly neurologists, prefer the name inclusion body myopathy. A recent case describing the concomitant findings of anti-PM-SCl antibodies in a patient with IBM supports a role for autoimmunity pathogenesis of IBM [37]. In addition, IBM has been described in concert with autoimmune rheumatic diseases, albeit rarely. Most recently, the combination of SLE, Sjogren syndrome, and IBM was reported for the first time [38]. The patient had SLE for over a decade when she presented with muscle weakness. Electron microscopy demonstrated intranuclear and intracytoplasmic tubulofilamentous inclusions; the patient improved with high-dose methotrexate therapy.
Muntzing et al. [39] demonstrated the presence of clonal restriction of TCR expression in muscle-infiltrating lymphocytes in IBM. Identical T-cell clones predominate in different muscles and exist for many years, suggesting an antigen-driven inflammatory reaction in IBM.
Protein folding and trafficking
The "unfolded protein response" (UPR) is a functional mechanism whereby cells protect themselves from endoplasmic reticulum (ER) stress by assuring proper folding and preventing buildup of unfolded proteins in the ER. This is accomplished with the help of molecular chaperones such as calnexin, calreticulin, GRP94, BiP/GRP78, and ERp72. Expression and immunolocalization of these proteins was studied in patients with sIBM and controls, and in amyloid-ß-precursor protein (AßPP) overexpressing cultured human muscle fibers [40]. All five ER chaperones physically associated with AßPP in sIBM muscle, implying that they may play a role in AßPP folding and processing.
The lysosomal system of muscle fibers may also play a critical role in rimmed vacuole formation [41]. Work by Kumamoto et al. [42] showed that the transport of newly synthesized lysosomal enzymes via the Golgi apparatus and autophagic vacuole formation in the lysosome system is activated in sIBM. This was demonstrated by the observation of clathrin and M6PR, which facilitate receptor- mediated intracellular transport inside rimmed vacuoles and in the sarcoplasm of vacuolated or nonvacuolated fibers, but not in control specimens. PrPSc, a hallmark of prion diseases such as spongiform encephalopathies, was demonstrated to be a prominent constituent in sporadic IBM muscle tissue of a patient with concomitant Creutzfeldt-Jakob Disease (CJD)[43]. The existence of muscle disease in subtypes of CJDmay deserve further systematic investigation, as distinct glycotyopes of PrSc may be present in muscle and brain. The muscle glycotype in this case report resembled that found in vCJD brain.
Although it is attractive to connect vacuolar trafficking abnormalities of glycoproteins to IBM because of the known genetic lesion in one type of hereditary IBM (see below), for the moment, it seems safest to consider these observations as evidence of yet another group of proteins being trapped in IBM inclusions.
UDP-N-acetylglucosamine-2-epimerase/N-acetylmannosamine kinase (GNE) mutation
The genetic absence of GNE activity leads to muscle weakness in hereditary inclusion body myositis (HIBM), perhaps by interfering with the trafficking of glycoproteins. Huizing et al. [44] examined the glycosylation status of [alpha]-dystroglycan (a central protein of skeletal muscle dystrophin) in muscle biopsies of four HIBM patients and found absent or markedly reduced [alpha]-dystroglycans using antibodies specific for ß-dystoglycan and laminin [alpha]-2. They suggest that HIBM may therefore be a "dystroglycanopathy," similar to the process in congenital muscular dystrophies-perhaps providing one mechanism for the muscle weakness of HIBM patients.
In an important series of observations, inflammation has now been clearly recognized in hereditary IBM. Yabe et al. [45] reported two cases of distal myopathy with rimmed vacuoles (DMRV) in a Japanese family in which inflammation (an unusual feature of DMRV) was present as well as a compound heterozygous mutation in their GNE gene. Mutations associated with DMRV in this study were localized to the kinase domain. An additional novel homozygous mutation was discovered in a nonJewish Iranian population with quadriceps-sparing myopathy consistent with adult onset hIBM: G-to-A mutation in exon 7 changing valine to isoleucine in the epimerase domain of GNE [46]. Muscle inflammation was present in this case as well.
These important observations blur the boundary between purely hereditary and sporadic IBM, in particular by raising the possibility that the inflammation seen in apparently sporadic IBM might be a downstream secondary event following damage induced by as-yet unrecognized genetic mutations.
The Middle East cluster of hIBM is the result of a founder mutation with incomplete penetrance and is not limited only to the Jewish population. One hundred twenty-nine patients in 55 families with a known history of hIBM were homozygous for the M712T mutation, initially described as the "Persian Jewish Mutation" [47]. Argov et al. [47] have found that the phenotypic spectrum is wider than initially thought. In a study by Del Bo et al., genetic analysis of the GNE gene in an Italian family with autosomal recessive h-IBM demonstrated two novel mutations: a heterozygous deletion involving exons 1-9 and the missense R162C mutation [48]. The quadriceps weakness was apparently distinct from that found in the quadriceps sparing nonPersian Jewish families with a different GNE mutation.
Hereditary IBM with early-onset Paget disease of bone and frontotemporal dementia (IBMPFD) is a rare form of hereditary myopathy. Its clinical attributes distinguish it from the other forms of hIBM linked to chromosome 9. Watts et al. [49] found no relation of GNE mutations with IBMPFD, confirming genetic heterogeneity with IBM2.
Other genetic evidence
The muscle morphology in X-EDMD is not pathognomonic; rather, there are nonspecific myopathic changes including endomysial connective tissue proliferation, fiber splitting, type II fiber predominance, and type I fiber atrophy. A recent paper reported the co-existence of x-linked Emery-Dreifuss muscular dystrophy (EDMD) and IBM [50]. Thus typical IBM morphologic features may be found in other neuromuscular disorders.
Although the pathogenesis of the inflammatory myopathies remains obscure, great strides over the past year have placed us closer to understanding the etiologies of these diverse disease entities. We have deepened our understanding that although these diseases may share some components of the clinical phenotype as well as some serologic similarities, they differ on a molecular and cellular level. The inflammatory myopathies result from a combination of interactions between environmental and genetic risk factors, and the need for definitive, safer therapeutic options in inflammatory myopathies makes the search for defining detailed pathogenesis of inflammation and muscle fiber damage at the molecular level essential.

Have recent immunogenetic investigations increased our understanding of disease mechanisms in the idiopathic inflammatory myopathies?
Hector Chinoy, William E.R. Ollier and Robert G. Cooper
Current Opinion in Rheumatology 2004, 16:707-713 [Nov 2004]
Rheumatic Diseases Centre, Hope Hospital, Salford M6 8HD, UK.
PURPOSE OF REVIEW: The idiopathic inflammatory myopathies (IIM) continue to provide a challenge given the variable effectiveness of the available treatments, and immunogenetic studies are ongoing to further elucidate IIM disease mechanisms. This review examines how recent research has improved our understanding of the mechanisms that lead to IIM.
RECENT FINDINGS: HLA-DRB1 studies in a large homogenous cohort of UK Caucasian patients have confirmed that polymyositis (PM) and dermatomyositis (DM) are not genetically identical diseases while other studies have shown that tumor necrosis factor alpha is genetically implicated in disease susceptibility. Some remarkable results from an international collaboration, correlating gene-environment interactions, clearly suggest that ultraviolet light is capable of modulating both clinical and immunologic features of IIMs. Studies on microchimerism are unraveling interesting associations in juvenile DM patients, and bolstering the hypothesis that myositis may be an 'allo-immune' disease. mRNA gene expression profiling is helping to increase our understanding of myositis pathogenesis, whilst animal models have provided new information on the roles of Th1 responses and nitric oxide synthase in muscle disease. New candidate genes have been examined in inclusion body myositis (IBM), and a novel gene transfer experiment has been conducted, which led to significant changes in expression of the IBM phenotype.
SUMMARY: Improving the understanding of the immunogenetics and immunopathogenesis of the IIMs may in the future provide novel therapeutic targets, and thus improve outcomes in these difficult diseases.
Excerpt from article (710-711)
Inclusion body myositis
As IBM usually presents with distal, rather than proximal, muscle weakness, and thus mimicking peripheral neuropathy, it is rarely seen by rheumatologists. However, due to presence of inflammation in muscle biopsies, sporadic IBM is considered one of the IIMs and is therefore included in this update. Several HLA and non-HLA loci are already known risk factors for sporadic IBM [3]. HLA class I and II associations have recently been examined in 47 sporadic IBM patients and 29,670 controls [38]. HLA-A*03 (OR 2.6, 95% CI 1.6-4.1), B*08 (OR 2.8, 95% CI 1.6-4.6), DRB1*03 (OR 3.5, 95% CI 2.1-5.6), and DQB1*05 (2.0, 95% CI 1.2-3.3) alleles were all significantly increased compared with controls, even after adjustment for multiple comparisons. The ethnicity of the cases in this study was not stated and these results should thus be interpreted with caution. There are currently no clinical or biochemical parameters that predict the outcome of IBM or response to treatment, and HLA typing may help subgroup IBM patients and predict such parameters.
Sporadic IBM muscle biopsies possess structural abnormalities similar to those in brain tissue plaques from Alzheimer's disease patients, including amyloid- precursor protein (AßPP), amyloid- ß(Aß), and apolipoprotein E (apoE) [39]. Using an adenovirus vector, A gene transfer into normal cultured human muscle fibers induced phenotypic abnormalities similar to those found in IBM, suggesting the likelihood of a key role of AßPP/A in IBM pathogenesis [40]. The same gene was transferred into muscle fibers from a patient with known sporadic IBM and associated cardiac amyloidosis in whom a Val122Ile transthyretin (TTR) mutation was also expressed [41]. The resulting overexpression of the A gene amplified the abnormalities found in this patient's cultured muscle fibers, including accelerated degeneration, inclusions, and vacuole formation, which were over and above those seen in the normal muscle. The TTR mutation could either be a genetic risk factor, or perpetuate the existing IBM. Polymorphisms of two further intracellular amyloid deposits, ApoE and [alpha](1)-antichymotrypsin, have been tested in the peripheral blood of 35 sporadic IBM patients [42], but no significant associations or correlations with age of onset were found. An important gene expression profiling study has also found increased expression of amyloid-ß and ApoE in IBM, but significantly elevated levels of the same genes were also demonstrated in PM and DM, suggesting that accumulation of such proteins in IBM may be due to posttranscriptional events [43].
T cell receptors
Immunohistochemistry studies of biopsies from patients with PM have demonstrated a predominance of CD8+ T lymphocytes invading non-necrotic muscle fibers that express HLA class I on their cell surface [44]. Previous studies have demonstrated clonally expanded T cell receptor (TCR) families in muscle fibers of patients with PM. To characterize the role of these T cells further, a process of CDR3 spectratyping was combined with laser microdissection and single-cell polymerase chain reaction (PCR), to select individual pathogenically relevant T cells from PM muscle biopsies [45]. After examining repeat biopsies in one patient, it was shown that T cell clones could persist for many years. In another patient, several T cells had minor nucleotide changes in the CDR3 region of the T cell receptor, which did not alter the amino acid sequences, thus suggesting the presence of different T cell clones driving a common antigendriven response. Oligoclonal CDR3 spectratype peaks disappeared during immunosuppressive treatment. Recent TCR expression work has also been performed in IBM, again suggesting only a limited number of antigens drive the inflammatory reaction [46]. These pathogenically relevant T cells may represent future epitopes for targeted immunotherapy.
Animal models
Myositis is thought to be predominantly Th1 driven, initiated by an (unknown) antigen and MHC interaction, leading to T cell expansion, maturation, and subsequent cytokine proliferation (eg, IFN-[gamma] , IL-1, IL-18). Theoretically, dampening down of the Th1 cell response, and switching to a Th2-driven system, producing alternative cytokines (eg, IL-4, IL-6, IL-10), could dampen down the pathogenicity of autoreactive T cells [47]. A strain of nonobese diabetic (NOD) mice has been developed, rendered Th1-deficient by a CD2 promoter driven IFN-[gamma] receptor ß-chain transgene [47]. An unexpected consequence was the development of an early lethal myopathy due to a CD8+ T cell-dependent myositis syndrome, characterized by a massive leukocyte inflammatory response in the muscle fibers. By interacting with other genetic components in the NODmodel, the inhibition of Th1 responses may conversely have exacerbated certain autoimmune responses.
Another way of influencing the inflammatory response in muscle is through nitric oxide (NO), which can be pro- or anti-inflammatory depending on its concentration and locality. A further transgenic mouse model has been developed with muscle-specific overexpression of neuronal nitric oxide synthase (nNOS), driven by a human skeletal muscle actin promoter [48]. Using a rodent hindlimb unloading/reloading model, overexpression of nNOS inhibited neutrophil production, and prevented any increases in muscle membrane damage. Muscle-derived NO evidently functions as an anti-inflammatory molecule, and may provide a future potential therapeutic target.
New and future approaches
The work done by Okada et al. [16] and ourselves demonstrates the advantages, and indeed necessity, of undertaking national/international genetic collaborations. The results of these larger studies illustrate the importance during genetic testing of treating IIM subtypes as discrete, rather than grouped, diagnoses. Due to the rarity of IIMs, only further collaboration is likely to elucidate the complex interactions between genetic and environmental factors. Recently doubt has been cast on the diagnostic category PM, as some patients have been shown to have IBM or muscular dystrophy [49]. mRNA microarray gene expression profiling studies may assist in a more robust molecular reclassification of the IIMs [43], whilst also providing novel molecular insights into the role of genes in IIM subtypes (reviewed in [50]). It is relatively easy to obtain muscle biopsies, allowing ready analysis of target tissue in IIM patients. Measurements of mRNA levels allow the demonstration of which genes are upregulated or downregulated (albeit without establishing cause and effect), and clarification of which genes to analyze in further candidate gene studies. A recent landmark study examined the molecular profiles of patients with IIMs, showing distinct changes from normal muscle and differing between the IIMs [43]. An important distinguishing feature of DM, compared with PM and IBM, was the increased expression of interferoninducible genes, and this finding was reproduced in a JDM study [20], raising the hypothesis that DM has an antigen-driven pathogenesis, and again supporting the idea that PM and DM are genetically different. In the future, improved techniques and reduced costs may allow the use of powerful high-density SNP arrays to conduct whole genome scans by association [50-52].
Research continues worldwide into our understanding of the underlying pathogenesis of the IIMs. A basic science approach led to the elucidation of the key role of TNF alpha in RA, and the subsequent use of anti-TNFalphatherapy. By analogy, ongoing collaborative genetic work may help identify key hierarchical molecules implicated in IIM pathology.

Transglutaminase catalyzes differential crosslinking of small heat shock proteins and amyloid-beta.
Boros S, Kamps B, Wunderink L, de Bruijn W, de Jong WW, Boelens WC. Department of Biochemistry 161, Nijmegen Center for Molecular Life Sciences, University of Nijmegen, P.O. Box 9101, 6500HB Nijmegen, The Netherlands.
FEBS Lett. 2004 Oct 8;576(1-2):57-62.
Crosslinking of proteins by tissue transglutaminase (tTG) is enhanced in amyloid (Abeta) deposits characteristic of Alzheimer's disease and sporadic inclusion body myositis. Small heat shock proteins (sHsps) also occur in amyloid deposits. We here report the substrate characteristics for tTG of six sHsps. Hsp27, Hsp20 and HspB8 are both lysine- and glutamine-donors, alphaB-crystallin only is a lysine-donor, HspB2 a glutamine-donor, and HspB3 no substrate at all. Close interaction of proteins stimulates crosslinking efficiency as crosslinking between different sHsps only takes place within the same heteromeric complex. We also observed that alphaB-crystallin, Hsp27 and Hsp20 associate with Abeta in vitro, and can be readily crosslinked by tTG.

Direct evidence for a chronic CD8+-T-cell-mediated immune reaction to tax within the muscle of a human T-cell leukemia/lymphoma virus type 1-infected patient with sporadic inclusion body myositis.
Ozden S, Cochet M, Mikol J, Teixeira A, Gessain A, Pique C. Unite d'Epidemiologie et Physiopathologie des Virus Oncogenes, Paris, France.
J Virol. 2004 Oct;78(19):10320-7.
Human T-cell leukemia/lymphoma virus type 1 (HTLV-1) infection can lead to the development of HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), concomitantly with or without other inflammatory disorders such as myositis. These pathologies are considered immune-mediated diseases, and it is assumed that migration within tissues of both HTLV-1-infected CD4(+) T cells and anti-HTLV-1 cytotoxic T cells represents a pivotal event. However, although HTLV-1-infected T cells were found in inflamed lesions, the antigenic specificity of coinfiltrated CD8(+) T cells remains to be determined. In this study, we performed both ex vivo and in situ analyses using muscle biopsies obtained from an HTLV-1-infected patient with HAM/TSP and sporadic inclusion body myositis. We found that both HTLV-1-infected CD4(+) T cells and CD8(+) T cells directed to the dominant Tax antigen can be amplified from muscle cell cultures. Moreover, we were able to detect in two successive muscle biopsies both tax mRNA-positive mononuclear cells and T cells recognized by the Tax11-19/HLA-A*02 tetramer and positive for perforin. These findings provide the first direct demonstration that anti-Tax cytotoxic T cells are chronically recruited within inflamed tissues of an HTLV-1 infected patient, which validates the cytotoxic immune reaction model for the pathogenesis of HTLV-1-associated inflammatory disease.

Mutant ubiquitin UBB+1 is accumulated in sporadic inclusion-body myositis muscle fibers.
NEUROLOGY 2004;63:1114-1117
Fratta P, Engel WK, Van Leeuwen FW, Hol EM, Vattemi G, Askanas V.
USC Neuromuscular Center, Department of Neurology, University of Southern California Keck School of Medicine, Good Samaritan Hospital, Los Angeles, CA 90017-1912, USA.
Abstract - Mutant ubiquitin (UBB+1), a product of "molecular misreading," is toxic to cells because its ubiquitinated form inhibits the proteasome, contributing to accumulation of misfolded proteins and their ensuing toxicity. The authors demonstrate in 10 sporadic inclusion body myositis (s-IBM) muscle biopsies that UBB+1 is accumulated in aggregates containing amyloid-beta and phosphorylated-tau. In s-IBM, UBB+1 may be pathogenic by inhibiting proteasome, thereby promoting accumulation of cytotoxic misfolded amyloid-beta and phosphorylated-tau.

Inflammatory disorders of muscle: progress in polymyositis, dermatomyositis and inclusion body myositis.
Dalakas MC.
Current Opinion in Neurology. 2004 Oct;17(5):561-567.
Neuromuscular Diseases Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland, USA.
PURPOSE OF REVIEW: To provide an update on the major advances in inflammatory myopathies.
RECENT FINDINGS: Polymyositis is an uncommon disorder that can be misdiagnosed when the old, and never validated, criteria of Bohan and Peter are used. New diagnostic criteria were recently introduced, in which the MHC/CD8 complex is considered a specific immunopathological marker because it distinguishes the antigen-driven inflammatory cells that characterize polymyositis and sporadic inclusion-body myositis from the non-specific, secondary inflammation seen in other disorders, such as dystrophies. In sporadic inclusion-body myositis the inflammatory cells invade non-vacuolated fibers, whereas the vacuolated fibers are not invaded by T cells, implying two independent processes, a primary immune process with antigen-driven T cells identical to polymyositis, and a degenerative process in which beta-amyloid and amyloid-related proteins participate in vacuolar degeneration. In polymyositis and sporadic inclusion-body myositis, antigen-specific and clonally expanded autoinvasive T cells persist for years, even in different muscles, as reconfirmed by proof-of-principle techniques involving CDR3 spectratyping combined with laser microdissected single-cell polymerase chain reaction of the T-cell receptor genes. The formation of immunological synapse between autoinvasive T cells and muscle fibers was recently strengthened by the upregulation of co-stimulatory molecules ICOS/ICOS-L and PD-L1. A new, distinct myopathy characterized by T-cell-triggered macrophage hyperactivation has now been recognized in patients with dermatomyositis-like disease.
SUMMARY: Despite recent progress, the antigen(s) responsible for T-cell activation in polymyositis and sporadic inclusion-body myositis and the cause of vacuolar degeneration in sporadic inclusion-body myositis remain unclear. Newer, more aggressive immunotherapies may be encouraging, but control trials are needed to prove efficacy.

Journey into muscular dystrophies caused by abnormal glycosylation.
Muntoni F. Dubowitz Neuromuscular Centre, Department of Paediatrics, Imperial College of Medicine, Hammersmith Hospital, London, UK. f.muntoni@imperial.ac.uk
Acta Myol. 2004 Sep;23(2):79-84.
An increasing number of genes encoding for putative or demonstrated glycosyltransferases are being associated with muscular dystrophies of variable severity, ranging from severe congenital onset and associated structural eye and brain changes, to relatively mild forms with onset into adulthood. Five of these genes (POMT1; POMGnT1; FXRP; Fukutin; LARGE) encode for proteins involved in the glycosylation of alpha-dystroglycan and, indeed, abnormal glycosylation of this molecule is a common finding in all the respective conditions (Walker Warburg syndrome; Muscle-Eye-Brain disease; congenital muscular dystrophy type 1C and Limb girdle muscular dystrophy type 21; Fukuyama muscular dystrophy; congenital muscular dystrophy type 1D). A 6th gene, GNE, responsible for the hereditary form of inclusion body myositis, encodes for a glycosyltransferase the substrate(s) of which is, however, still unclear. This article provides an overview of the clinical, biochemical and genetic features of this group of disorders.

Randomized pilot trial of high-dose betaINF-1a in patients with inclusion body myositis.
Muscle Study Group. Neuromuscular Disease Center, 601 Elmwood Avenue, Box 673, Rochester, NY 14642-8673. Rabi_Tawil@URMC.Rochester.edu
Neurology. 2004 Aug 24;63(4):718-20.
Beta-interferon-1a (betaINF-1a) is well tolerated at low dose (30 microg IM/week) in inclusion body myositis (IBM). The authors investigated the safety and tolerability of high-dose (60 microg IM/week) betaINF-1a in a randomized, placebo-controlled trial in IBM. Twenty-seven of the 30 subjects enrolled completed the study. The adverse event profile was similar for the placebo and betaINF-1a groups. betaINF-1a, at a dose of 60 microg IM/week, is well tolerated in IBM, but no differences in muscle strength or mass were observed between the placebo and betaINF-1a groups at 6 months in this pilot study.

Interferon beta-responsive inclusion body myositis in a hepatitis C virus carrier.
Yakushiji Y, Satoh J, Yukitake M, Yamaguchi K, Nakamura I, Nishino I, Kuroda Y.
The Division of Neurology, Department of Internal Medicine, Saga University School of Medicine, Saga, Japan. yakushij@hsp.ncvc.go.jp
Neurology. 2004 Aug 10;63(3):587-8.
Case Report

Sporadic inclusion body myositis: morphology, regeneration, and cytoskeletal structure of muscle fibres.
Arnardottir S, Borg K, Ansved T. Department of Clinical Neuroscience, Division of Neurology, Karolinska Hospital, Stockholm, Sweden. snjolaug.arnardottir@ks.se
J Neurol Neurosurg Psychiatry. 2004 Jun;75(6):917-20.
OBJECTIVE: To characterise morphological abnormalities in relation to muscle fibre type in sporadic inclusion body myositis (s-IBM). METHODS: 14 muscle biopsies from 11 patients with s-IBM were characterised for morphological abnormalities and fibre type composition as well as muscle fibre regeneration and cytoskeletal structure, using histochemical and immunohistochemical techniques. RESULTS: Morphological abnormalities included inflammatory infiltrates and "rimmed vacuoles," and pronounced variation in fibre size. There were no significant differences in fibre type composition between s-IBM patients and controls based on the myofibrillar ATPase staining. A differential effect on muscle fibre sizes was noted, type II fibres being smaller in the s-IBM patients than in the controls. Conversely, the mean type I muscle fibre diameter in the s-IBM patients was larger than in the controls, though this difference was not significant. An ongoing intense regeneration process was present in s-IBM muscle, as indicated by the expression of neonatal myosin heavy chain, vimentin, and CD56 (Leu-19) in most of the muscle fibres. The cytoskeletal proteins dystrophin and desmin were normally expressed in s-IBM muscle fibres that were not undergoing degeneration or regeneration. CONCLUSIONS: There are extensive morphological and morphometric alterations in s-IBM, affecting different muscle fibre types in different ways. The cytoskeletal structure of type I and II muscle fibres remains unaffected in different stages of the disease.

Insulin-like growth factor I in inclusion-body myositis and human muscle cultures.
Broccolini A, Ricci E, Pescatori M, Papacci M, Gliubizzi C, D'Amico A, Servidei S, Tonali P, Mirabella M. Department of Neuroscience, Catholic University, Rome, Italy. a.broccolini@rm.unicatt.it
J Neuropathol Exp Neurol. 2004 Jun;63(6):650-9.
Possible pathogenic mechanisms of sporadic inclusion-body myositis (sIBM) include abnormal production and accumulation of amyloid beta (A beta), muscle aging, and increased oxidative stress. Insulin-like growth factor I (IGF-I), an endocrine and autocrine/paracrine trophic factor, provides resistance against A beta toxicity and oxidative stress in vitro and promotes cell survival. In this study we analyzed the IGF-I signaling pathway in sIBM muscle and found that 16.2% +/- 2.5% of nonregenerating fibers showed increased expression of IGF-I, phosphatidylinositide 3'OH-kinase, and Akt. In the majority of sIBM abnormal muscle fibers, increased IGF-I mRNA and protein correlated with the presence of A beta cytoplasmic inclusions. To investigate a possible relationship between A beta toxicity and IGF-I upregulation, normal primary muscle cultures were stimulated for 24 hours with the A beta(25-35) peptide corresponding to the biologically active domain of A beta. This induced an increase of IGF-I mRNA and protein in myotubes at 6 hours, followed by a gradual reduction thereafter. The level of phosphorylated Akt showed similar changes. We suggest that in sIBM. IGF-I overexpression represents a reactive response to A beta toxicity, possibly providing trophic support to vulnerable fibers. Understanding the signaling pathways activated by IGF-I in sIBM may lead to novel therapeutic strategies for the disease.

Apolipoprotein E and alpha-1-antichymotrypsin polymorphisms in sporadic inclusion body myositis.
Gossrau G, Gestrich B, Koch R, Wunderlich C, Schroder JM, Schroeder S, Reichmann H, Lampe JB.
Department of Neurology, Medical Clinic II, Technical University Dresden, Dresden, Germany. ggossrau@uni-bonn.de
Eur Neurol. 2004;51(4):215-20. Epub 2004 May 17.
Sporadic inclusion body myositis (s-IBM) is a progressive muscle disease of unknown aetiology. Characteristically, intracellular amyloid deposits are detectable, including beta-amyloid precursor protein, phosphorylated tau, alpha1-antichymotrypsin (alpha1-ACT) and apolipoprotein E (ApoE). Polymorphisms and mutations of the encoding genes have been identified in a variety of neurodegenerative diseases including Alzheimer's disease (AD). Beside other factors, polymorphisms may lead to protein accumulation in both diseases. In particular, polymorphisms within the ApoE and alpha1-ACT gene have been implicated in the aetiology of AD and s-IBM. We analysed ApoE and alpha1-ACT gene polymorphisms in 35 s-IBM patients. We could not identify any statistical significant correlation between distinct ApoE and alpha1-ACT genotypes and the risk of developing s-IBM. Additionally, ApoE and alpha1-ACT genotypes seem not to influence the onset age of s-IBM. A combination of different alpha1-ACT and ApoE genotypes appears not to enhance the risk of developing s-IBM. Therefore, allelic variations of alpha1-ACT and ApoE are unlikely to be genetic key factors in the aetiology of s-IBM.

Proteasomal expression, induction of immunoproteasome subunits, and local MHC class I presentation in myofibrillar myopathy and inclusion body myositis.
Ferrer I, Martin B, Castano JG, Lucas JJ, Moreno D, Olive M. Instituto de Neuropatologia, Servicio de Anatomia Patologica, Hospital Universitario de Bellvitge, Hospitalet de Llobregat, Spain. 8082ifa@comb.es
J Neuropathol Exp Neurol. 2004 May;63(5):484-98.
Inclusion body myositis (IBM) and myofibrillar myopathy (MM) are diseases characterized by the abnormal accumulation of proteins in muscle fibers, including desmin, alphaB-crystallin, gelsolin, actin, kinases, and phospho-tau, along with ubiquitin in muscle fibers, suggesting abnormal protein degradation as a possible cause of the surplus myopathy. Since the ubiquitin-proteasome system plays a crucial role in non-lysosomal protein degradation, the present study has examined by immunohistochemistry the expression of components of the catalytic core of 20S proteasomes and its regulators: 19S and PA28alpha/beta, and the expression of immunoproteasome subunits LMP2, LMP7, and MECL1 in 8 patients with MM and 10 patients with IBM. The patients with MM were from 6 unrelated families, 2 sporadic cases, I with autosomal recessive and 5 with autosomal dominant inheritance. One sporadic patient had a de novo R406W mutation in the desmin gene, and 1 patient with autosomal dominant MM had a single amino acid deletion at position 366 in the desmin gene. Increased immunoreactivity to 20S, 19S, and PA28alpha/beta colocalizing abnormal protein deposits, as revealed in consecutive serial sections, was seen in all cases with MM and IBM. In all cases, the subunits of the immunoproteasome LMP2, LMP7, and MECL1 colocalized with proteasomal immunoreactivity and abnormal protein accumulation. Immunohistochemistry revealed focal MHC class I immunoreactivity in the cytoplasmic membrane of muscle fibers in IBM and in association with protein aggregates in IBM, and to a lesser degree, in MM. The present findings provide a link between abnormal protein accumulation and altered proteasomal expression in IBM and MM.

Intravenous Immunoglobulin in Autoimmune Neuromuscular Diseases
Dalakas, MC
JAMA. May 19, 2004; Volume 291, pages: 2367-2375.
From page 2373: Inclusion Body Myositis. Inclusion body myositis is the most common acquired inflammatory myopathy in patients older than 50 years. Patients present with distal and proximal muscle weakness, frequent falls, and dysphagia. Immunopathologically, inclusion body myositis is identical to polymyositis, but histologically, it is differentiated by the presence of vacuolated fibers and amyloid deposits. Inclusion body myositis is notoriously resistant to treatment with immunosuppressive medications. Therefore, the efficacy of IVIG was tested in 19 patients in a placebo-controlled crossover trial similar to that of the dermatomyositis trial. Although muscle strength scores improved more in IVIG randomized patients, the differences were not statistically significant, except for regional differences, most notable in the muscles used for swallowing. A second study showed similar results. A third study, carried out to investigate the potential synergistic effect of IVIG and prednisone, randomly assigned 36 patients to IVIG, 2 g/kg, or placebo once a month for 3 months. All patients received prednisone concurrently (tapered from 60 mg/d). After 3 months of treatment, there was no significant difference in muscle strength between the IVIG-plus prednisone group and the placebo-plus prednisone group. Despite these negative findings, some inclusion body myositis patients may derive modest, transient benefit from IVIG therapy, sufficient to justify a 2- to 3-month trial, especially in those with severe dysphagia, as recently noted.

Inclusion body myopathy associated with Paget disease of bone and frontotemporal dementia is caused by mutant valosin-containing protein
Authors: Giles D J Watts, Jill Wymer, Margaret J Kovach, Sarju G Mehta, Steven Mumm, Daniel Darvish, Alan Pestronk, Michael P Whyte & Virginia E Kimonis
Nature Genetics 36, 377 - 381 (April 2004)
Abstract: Inclusion body myopathy associated with Paget disease of bone and frontotemporal dementia (IBMPFD) is a dominant progressive disorder that maps to chromosome 9p21.1-p12. We investigated 13 families with IBMPFD linked to chromosome 9 using a candidate-gene approach. We found six missense mutations in the gene encoding valosin-containing protein (VCP, a member of the AAA-ATPase superfamily) exclusively in all 61 affected individuals. Haplotype analysis indicated that descent from two founders in two separate North American kindreds accounted for IBMPFD in about 50% of affected families. VCP is associated with a variety of cellular activities, including cell cycle control, membrane fusion and the ubiquitin-proteasome degradation pathway. Identification of VCP as causing IBMPFD has important implications for other inclusion-body diseases, including myopathies, dementias and Paget disease of bone (PDB), as it may define a new common pathological ubiquitin-based pathway.
From page 377: Hereditary inclusion body myopathy (IBM) associated with PDB and frontotemporal dementia (FTD), or IBMPFD, is a rare, complex and ultimately lethal autosomal dominant disorder (OMIM 605382; ref. 1 [see link below]). IBMPFD features adult-onset proximal and distal muscle weakness (clinically resembling limb girdle muscular dystrophy), early-onset PDB in most cases and 'premature' FTD2. Although the disorder was mapped to chromosome 9p21-p12, the genetic basis was not known.
From page 379: VCP is commonly present in aggregates from muscle with IBMPFD and s-IBM, although the predominant pattern of localization for VCP differs between IBMPFD and s-IBM. . . .
VCP is essential in the cell cycle and apoptosis pathways, neither of which seems to be disrupted in IBMPFD, as affected individuals are obviously viable. Clues about the nature of the mutations we identified in VCP can be drawn from pathways that have been implicated in other aggresome-associated degenerative disorders, which all involve protein quality control and the ubiquitin protein degradation pathways ref 21-24. A number of independent studies support the fact that disruption of a specific function of VCP leads to inclusion body formation. First, experiments identifying the involvement of VCP in endoplasmic reticulum-associated degradation showed that dysfunction of VCP caused vacuole and inclusion body formation, ultimately leading to cell death ref 18, 19, 25. Second, VCP interacts directly with polyubiquitinated proteins ref 18-20. Third, VCP colocalizes with ubiquitin-containing nuclear inclusions in the cerebral cortex in a number of neuronal degenerative disorders involving protein quality control and the ubiquitin protein degradation pathways, such as Huntington, Alzheimer, Creutzfeldt-Jakob and Parkinson disease (in particular the Lewy bodies), as well as in motor neuron disease with dementia ref 26. Fourth, mutations in the ubiquitin-binding domain of sequestosome 1 (SQSTMl, also called p62; refs. 27,28) cause autosomal dominant PDB (PDB3). We propose that mutations in VCP, as in SQSTMl, cause PDB by compromising ubiquitin-binding and target similar cellular pathways or proteins. Furthermore, p62 colocalizes with inclusion bodies in a number of degenerative disorders ref 22, 23. Thus, IBMPFD is probably a new member of the family of aggresome-associated disorders, and mutations in VCP may represent a new link in the pathway that leads to aggresome formation. Because IBMPFD is a dominant progressive syndrome, the mutations we identified are probably relatively subtle and aging, oxidative stress and endoplasmic reticulum stress probably define a threshold at which the IBMPFD phenotype becomes manifest. Rather than the mutations disrupting a normal function of VCP, they may add new, toxic functions that result in new VCP actions. Alternatively, the mutations could be dominant negative and disrupt normal hexamer formation of VCP.
Also see: http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=167320

Inflammatory Myopathies.
Grogan PM, Katz JS.
Department of Neurology, Wilford Hall Medical Center, 2200 Bergquist Drive, Suite 1,San Antonio, TX 78236, USA.
Curr Treat Options Neurol. 2004 Mar;6(2):155-161.
Therapies that suppress or modify the immune system remain the primary treatment for the idiopathic inflammatory myopathies. Dermatomyositis (DM) and polymyositis (PM) are the two conditions that respond best to immunotherapy. Although there are no randomized controlled trials, corticosteroids, specifically high-dose oral prednisone, remain the cornerstone of management. Recent controlled clinical trials show that intravenous immunoglobulin (IVIg) is an efficacious treatment in DM. Expert clinicians are generally using this as a second-line agent or as an adjunct to prednisone. IVIg has a relatively benign side effect profile compared with chronic steroid use, but the cost of treatment, the need for repetitive treatment cycles, and the potential for serious adverse effects have kept it from being a first-line agent in DM. There have been no trials performed using IVIg in PM. Chronic immunosuppressant medications, including azathioprine, cyclosporine, and methotrexate, are also available for long-term management in patients with recalcitrant disease or side effects from extended corticosteroid use. These agents lack the troubling side effects of prednisone and are less costly than IVIg, but require close medical monitoring for adverse reactions to blood, kidney, lung, or liver. Newer medications with potentially more benign side effect profiles, such as mycophenolate mofetil and etanercept, are currently being studied, but knowledge of how effective they are and how quickly they work are not yet available. Inclusion body myositis has proven resistant to immunosuppressive medications. The response has been so consistently poor and so easily contrasted with DM that the authors wonder why these conditions are so routinely lumped together in chapters and review articles. Clearly, this is based solely on the common pathologic feature of inflammation, rather than a clear understanding of how these conditions occur, or why they do or do not respond to treatment.

[ I think this is a very important article. I think the next articles in this chain of research will be critical. I have included a pdf file of the whole article here.]

Upregulated inducible co-stimulator (ICOS) and ICOS-ligand in inclusion body myositis muscle: significance for CD8+ T cell cytotoxicity.
Schmidt J, Rakocevic G, Raju R, Dalakas MC.
Neuromuscular Diseases Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, USA.
Brain. 2004 Mar 26 Advance Access published on March 26, 2004 DOI: 10.1093/brain/awh148
Interactions between inducible co-stimulatory molecule (ICOS) and ICOS-ligand (ICOS-L) are crucial for T-cell co-stimulation, effector cell differentiation and memory CD8(+) T-cell activation. Because in the muscle of patients with sporadic inclusion body myositis (sIBM) clonally expanded CD8(+) T cells invade major histocompatibility complex (MHC) class I-expressing muscle fibres, we investigated ICOS.ICOS-L interactions and correlated their expression with perforin, a marker for cytotoxic effector function by autoinvasive CD8(+) T cells. The mRNA from 20 muscle biopsies of sIBM, 20 non-inflammatory or dystrophic controls, two dermatomyositis (DM) and two polymyositis (PM) patients was reverse transcribed and reamplified by semi-quantitative and quantitative reverse transcription-polymerase chain reaction (RT-PCR), using primers for ICOS, ICOS-L and perforin. The lyceraldehyde 3-phosphate dehydrogenase (GAPDH)-normalized ratio of ICOS, ICOS-L and perforin expression was compared with the degree of endomysial inflammation. protein expression of ICOS, ICOS-L and perforin was confirmed by immunohistochemistry. We demonstrate that ICOS-L mRNA was upregulated in sIBM (arbitrary units, median +/- SEM: 48.6 +/- 14.9) compared with controls (6.2 +/- 17.8, P < 0.05) and significantly correlated with the expression of ICOS (53.9 +/- 16.6 versus 6.7 +/- 8.9 in controls, P < 0.001). By triple labelling immunohistochemistry, the CD8(+) T cells in sIBM and PM were found to invade ICOS-L- and MHC class I-co-expressing muscle fibres. Among the autoinvasive CD8(+) T cells, however, only a subset of approximately 5-10% were ICOS positive, and thereby perceptive for ICOS.ICOS-L signalling at the immunological synapse. In contrast, in Duchenne muscular dystrophy and DM, although ICOS and ICOS-L mRNA expression was also increased, the majority of ICOS-L- and ICOS-positive cells were in the perimysial regions and connective tissue. The mRNA for perforin was increased in sIBM (28.1 +/- 8.7) compared with controls (4.3 +/- 11.2, P = 0.18), and significantly correlated with mRNA of ICOS, ICOS-L and the degree of endomysial inflammation as assessed in coded haematoxylin/eosin tissue sections. By triple immunohistochemical staining and cell counting, perforin granules were found in 71% of the autoinvasive CD8(+) T cells that were also ICOS positive. Our data indicate that in sIBM there is upregulation of ICOS.ICOS-L co-stimulatory signalling in association with enhanced perforin expression by the autoinvasive CD8(+) T cells. The findings support previous suggestions that in IBM, the muscle fibres have the capacity for antigen presentation, thereby activating a specific subset among the autoinvasive CD8(+) T cells to exert a cytotoxic effect. The observations strengthen the immunopathogenesis of sIBM, and offer the basis for future therapeutic interventions targeting ICOS.ICOS-L co-stimulatory interactions.

"Coles Notes" condensation by Bill:
In the muscles of patients with sporadic inclusion body myositis (sIBM) some of the muscle fibres come to display an immune "flag" (antigen presentation), on their surface. This flag acts as a signal to the immune system. A chain of events is triggered that leads to the activation of a response from the immune system. Immune cells invade the muscle cell and kill it.
There are several steps in the chain of events between the immune system seeing the flag and the invasion. One step is that other molecules (co-stimulators) need to be present to help stimulate the immune cell invaders. Researchers are discovering these links in the chain and how they interact with each other. Part of the immune chain involved consists of interactions between the inducible co-stimulatory molecule (ICOS) and its "partner" (called a ligand), the ICOS-ligand (ICOS-L). These interactions are crucial for co-stimulation of the immune invader cells (T-cells) and for other critical steps in the immune response (effector cell differentiation and memory CD8+ T-cell activation).
This research strengthens the idea that sIBM is caused by some sort of problem in the immune system. It offers the chance that medications can be developed to target breaking a link in the chain of these immune interactions. It is possible that if a medication could break the interaction of ICOS and ICOS-L, the destruction of the muscle cells could be stopped.

Overexpression of semicarbazide-sensitive amine oxidase in human myopathies.
Olive M, Unzeta M, Moreno D, Ferrer I.
Institut de Neuropatologia, Hospital Universitari de Bellvitge, 08907 Hospitalet de Llobregat, Barcelona, Spain. 25169mop@comb.es
Muscle Nerve. 2004 Feb;29(2):261-6.
Oxidative stress has been implicated in the pathogenesis of several muscle diseases. Semicarbazide-sensitive amine oxidase (SSAO) metabolizes oxidative deamination of primary aromatic and aliphatic amines. In the oxidative reactions, amine substrates are converted into the aldehyde, followed by the production of ammonia and H(2)O(2). Although normal levels in muscle are very low, SSAO is expressed in almost all mammalian tissues. In this study, we examined the possible implication of SSAO as an additional source of oxidative stress in the pathogenesis of muscle disorders. The expression of SSAO was examined immunohistochemically in muscle biopsy specimens from patients with inclusion-body myositis (IBM; n = 5), desmin-related myopathy (DRM; n = 3), dermatomyositis (n = 3), granulomatous (sarcoid) myopathy (n = 2), muscle denervation-reinnervation (n = 3), and rhabdomyolysis (n = 2), as well as from control subjects (n = 3). Strong SSAO immunoreactivity was present in vacuolated and nonvacuolated fibers in IBM, in abnormal fibers in DRM, and in degenerating and regenerating fibers in dermatomyositis and rhabdomyolysis. In addition, SSAO overexpression was observed in muscle fibers adjacent to granulomas in sarcoid myopathy. These results suggest that SSAO is a source of oxidative stress in diseased human skeletal muscle and that it contributes to oxidative stress-induced damage in various inflammatory and other myopathies. Alternatively, the expression of SSAO in muscle fibers may be a consequence of muscle fiber injury.

BOOK REVIEW: Living with Myositis
J. Fenton (Ed.); Thoughtful Publications, 2003; ISBN
0-9545307-0-5 (£13.99)
Neuromuscular Disorders Volume14 (February, 2004) page. 179
"I've got IBM". "Really. I'm still using my old Apple-Mac" or "Oh!. What operating system are you running?" Almost without exception the people I meet have never heard of Inclusion Body Myositis (IBM). Not surprising really, why should they have? Until I was diagnosed as having IBM and until I had read this book I had never heard of it or of dermatomyositis (DM) or polymyositis (PM). The first 140 or so pages are in part ego trip and in part an attempt to put a positive spin on what are truly awful diseases. They may indeed help health-care professionals (horrible collective term) to appreciate the torments myositis sufferers can go through, but I would not recommend that the book be offered to early diagnosed patients. They have enough to worry them with the diagnosis without reading what they may have to go through during the next indeterminate number of years. However, taking the book as a whole there is no doubt in my mind that all doctors will benefit from reading it, especially those practitioners who have not yet had a patient who has presented the symptoms of myositis. One message that seems to come through, not always overtly, is that drugs are not the only treatment. Listening, understanding, accepting that the patient knows his or her own body better than anyone and tender loving care, go a long way in coping with the disease. Another is that proof positive is necessary for the correct treatment to be started; don't be embarrassed to make an early referral. My doctor did and my wife and I are grateful for although, as yet, there is no treatment for IBM, knowing the prognosis we are able to plan accordingly. Keep a copy of this book on your shelf ready to lend out, but remember, to some it will be a horror story, to others a good reference book, choose carefully.
After reading it I'm glad I only have IBM!
J. Hutson
3 York Road, Headington,
Oxford OX3 8NW, UK

Endoplasmic reticulum stress and unfolded protein response in inclusion body myositis muscle.
Vattemi G, Engel WK, McFerrin J, Askanas V.
Department of Neurology, University of Southern California Neuromuscular Center, University of Southern California Keck School of Medicine, Good Samaritan Hospital, Los Angeles, California 90017-1912, USA.
Am J Pathol. 2004 Jan;164(1):1-7.
Proteins in the endoplasmic reticulum (ER) require an efficient system of molecular chaperones whose role is to assure their proper folding and to prevent accumulation of unfolded proteins. The response of cells to accumulation of unfolded proteins in the ER is termed "unfolded protein response" (UPR). UPR is a functional mechanism by which cells attempt to protect themselves against ER stress, resulting from the accumulation of the unfolded/misfolded proteins. Because intracellular inclusions, containing either amyloid-beta (Abeta) or phosphorylated tau, are the characteristic feature of sporadic inclusion body myositis (s-IBM) muscle biopsies, we studied expression and immunolocalization of five ER chaperones, calnexin, calreticulin, GRP94, BiP/GRP78, and ERp72, in s-IBM and control muscle biopsies. Physical interaction of the ER chaperones with amyloid-beta precursor protein (AbetaPP) was studied by a combined immunoprecipitation/immunoblotting technique in s-IBM and control muscle biopsies, and in AbetaPP-overexpressing cultured human muscle fibers. In all s-IBM muscle biopsies, all five of the ER chaperones were immunodetected in the form of inclusions that co-localized with amyloid-beta. By immunoblotting, expression of ER chaperones was greatly increased as compared to the controls. By immunoprecipitation/immunoblotting experiments, ER chaperones co-immunoprecipitated with AbetaPP. Our studies provide evidence of the UPR in s-IBM muscle and demonstrate for the first time that the ER chaperones calnexin, calreticulin, GRP94, BiP/GRP78, and ERp72 physically associate with AbetaPP in s-IBM muscle, suggesting their playing a role in AbetaPP folding and processing.

Creutzfeldt-Jakob disease and inclusion body myositis: abundant disease-associated prion protein in muscle.
Kovacs GG, Lindeck-Pozza E, Chimelli L, Araujo AQ, Gabbai AA, Strobel T, Glatzel M, Aguzzi A, Budka H.
Institute of Neurology, University of Vienna, and Austrian Reference Centre for Human Prion Diseases, Vienna, Austria.
Ann Neurol. 2004 Jan;55(1):121-5.
Pathologicalprion protein (PrP(Sc)) is the hallmark of prion diseases affecting primarily the central nervous system. Using immunohistochemistry, paraffin-embedded tissue blot, and Western blot, we demonstrated abundant PrP(Sc) in the muscle of a patient with sporadic Creutzfeldt-Jakob disease and inclusion body myositis. Extraneural PrP(C)-PrP(Sc) conversion in Creutzfeldt-Jakob disease appears to become prominent when PrP(C) is abundantly available as substrate, as in inclusion body myositis muscle.
Comment: In ONE case study of a 68 year old man who had CJD AND IBM, the abnormal form of prion protein was found OUTSIDE his neural tissues and was found in the muscle. Pathological prion protein (PrPSc) is the hallmark of prion diseases affecting primarily the central nervous system. Researchers demonstrated abundant (abnormal prion) PrPSc in the muscle of a patient with sporadic Creutzfeldt-Jakob disease and inclusion body myositis. Extraneural PrPC-PrPSc conversion in Creutzfeldt-Jakob disease appears to become prominent when PrPC is abundantly available as substrate, as in inclusion body myositis muscle.
The exact relationships between CJD and prion abnormalities in the muscle and IBM are unknown at present. This line of research will be interesting to watch.

Diagnostic value of MHC class I staining in idiopathic inflammatory myopathies.
van der Pas J, Hengstman GJ, ter Laak HJ, Borm GF, van Engelen BG. Neuromuscular Centre, Institute of Neurology, University Medical Centre, Nijmegen, Netherlands.
J Neurol Neurosurg Psychiatry. 2004 Jan;75(1):136-9.
BACKGROUND: Identification of mononuclear cellular infiltrates in skeletal muscle tissue is the histological cornerstone of the diagnosis of idiopathic inflammatory myopathy (IIM). However, these infiltrates are not always present.
OBJECTIVE: To determine whether MHC class I antigen expression on the sarcolemma, which is absent in normal muscle tissue, is upregulated in IIM and could serve as an additional diagnostic test.
METHODS: Expression of MHC class I antigens was studied in 224 muscle samples of 61 adult patients with IIM (9 dermatomyositis, 23 polymyositis, 29 inclusion body myositis) and 163 controls (normal subjects and patients with various neuromuscular disorders) in a prospective blinded manner.
RESULTS: The sensitivity of the test for diagnosing IIM was 78% (95% confidence interval (CI), 66% to 88%), with a specificity of 95% (91% to 98%). The sensitivity before the start of immunosuppressive treatment was 89% (76% to 96%). The sensitivity was not changed by including all patients who had been on immunosuppressive treatment for less than four weeks before muscle biopsy (sensitivity 90% (79% to 97%)). False positive results were found in only seven controls (4%), six of whom had a muscular dystrophy.
CONCLUSIONS: Detection of sarcolemmal MHC class I is a valid test for IIM. It is not affected by the short term use of immunosuppressive agents (less than four weeks) and it should be incorporated in the histological evaluation when the diagnosis of IIM is under consideration or needs to be excluded.