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The recently discovered aging-dependent large accumulation of point mutations in the human fibroblast mtDNA control region raised the question of their occurrence in postmitotic tissues. In the present work, analysis of biopsied or autopsied human skeletal muscle revealed the absence or only minimal presence of those mutations. By contrast, surprisingly, most of 26 individuals 53 to 92 years old, without a known history of neuromuscular disease, exhibited at mtDNA replication control sites in muscle an accumulation of two new point mutations, i.e., A189G and T408A, which were absent or marginally present in 19 individuals younger than 34 years. These two mutations were not found in fibroblasts from 22 subjects 64 to 101 years of age (T408A), or were present only in three subjects in very low amounts (A189G). Furthermore, in several older individuals exhibiting an accumulation in muscle of one or both of these mutations, they were nearly absent in other tissues, whereas the most frequent fibroblast-specific mutation (T414G) was present in skin, but not in muscle. Among eight additional individuals exhibiting partial denervation of their biopsied muscle, four subjects >80 years old had accumulated the two muscle-specific point mutations, which were, conversely, present at only very low levels in four subjects <40 years old. The striking tissue specificity of the muscle mtDNA mutations detected here and their mapping at critical sites for mtDNA replication strongly point to the involvement of a specific mutagenic machinery and to the functional relevance of these mutations. R ecently, the discovery of an aging-dependent large accumulation of point mutations in the control region for mtDNA replication of human skin fibroblasts has been reported (1). Particularly striking was the demonstration, in a generally high proportion of molecules (up to 50%), of a T to G transversion at position 414 in the original Cambridge sequence (2), within the promoter for the synthesis of the RNA primer of mtDNA heavy (H)-strand synthesis (3) and for light (L)-strand transcription (4). This mutation was present in more than 50% of the individuals above 65 years of age and absent in younger individuals. In the present work, to investigate the occurrence of these aging-dependent mutations in other cell types, in particular, in postmitotic cells, a screening was carried out for the detection in human muscle of aging-related specific point mutations in the DLP4 and DLP6 segments of the main mtDNA control region, which were the segments previously found to carry the fibroblast mtDNA mutations (1). These two mtDNA segments correspond to one of the hypervariable portions of the main control region (5), and were chosen, for the purpose of analysis, as containing each a uniform melting domain (Y.M., unpublished data). They carry critical sequences for mtDNA replication (Fig. 1A). In particular, DLP4 contains the primary origin of H-strand mtDNA synthesis (O H1 ) (3), whereas DLP6 contains the promoter and start site for H-strand replication RNA...

Sporadic inclusion-body myositis (s-IBM), the most common muscle disease of older persons, is of unknown cause and there is no successful treatment. We summarize our most recent findings, which provide a better understanding of the steps in the pathogenetic cascade. We suggest that s-IBM is primarily a myodegenerative disease. Intriguing are the phenotypic similarities between s-IBM muscle fibers and the brains of Alzheimer disease, the most common neurodegenerative disease of older persons. In s-IBM, abnormal accumulation of the amyloid-beta (Abeta) precursor protein and its proteolytic fragment, Abeta, associated with the aging intracellular milieu of the muscle fiber, appear to be key upstream pathogenic events. We propose that the identified abnormal accumulation, misfolding, and aggregation of proteins, perhaps provoked by the aging milieu and aggravated by the oxidative stress, lead to the s-IBM-specific vacuolar degeneration and atrophy of muscle fibers.

FUS gene mutations are not an uncommon cause in patients with FALS from diverse genetic backgrounds, and have a prevalence of 5.6% in non-SOD1 and non-TARDBP FALS, and approximately 4.79% in all FALS. The pathogenicity of some of these novel mutations awaits further studies. Patients with FUS mutations manifest earlier symptom onset, a higher rate of bulbar onset, and shorter duration of symptoms.

The 26S proteasome system is involved in eliminating various proteins, including ubiquitinated misfolded/unfolded proteins, and its inhibition results in cellular accumulation of protein aggregates. Intramuscle-fiber ubiquitinated multiprotein-aggregates are characteristic of sporadic inclusion-body myositis (s-IBM) muscle fibers. Two major types of aggregates exist, containing either amyloid-beta (Abeta) or phosphorylated tau (p-tau). We have now asked whether abnormalities of the 26S proteasome contribute to s-IBM pathogenesis and whether the multiprotein aggregates have features of aggresomes. Using cultured human muscle fibers we also studied the effect of amyloid-beta precursor protein (AbetaPP) overexpression on proteasome function and the influence of proteasome inhibition on aggresome formation. We report that in s-IBM muscle biopsies 26S proteasome subunits were immunodetected in the gamma-tubulin-associated aggresomes, which also contained Abeta, p-tau, ubiquitin, and HSP70. In addition, a) expression of proteasome subunits was greatly increased, b) the 20Salpha proteasome subunit co-immunoprecipitated with AbetaPP/Abeta, and c) the three major proteasomal proteolytic activities were reduced. In cultured muscle fibers, AbetaPP-overexpressing fibers displayed diminished proteasomal proteolytic activities, and addition of proteasome inhibitor strikingly increased aggresome formation. Accordingly, proteasome dysfunction in s-IBM muscle fibers may play a role in accumulation of misfolded, potentially cytotoxic proteins and may be induced by increased intracellular AbetaPP/Abeta.

p62, also known as sequestosome1, is a shuttle protein transporting polyubiquitinated proteins for both the proteasomal and lysosomal degradation. p62 is an integral component of inclusions in brains of various neurodegenerative disorders, including Alzheimer disease (AD) neurofibrillary tangles (NFTs) and Lewy bodies in Parkinson disease. In AD brain, the p62 localized in NFTs is associated with phosphorylated tau (p-tau). Sporadic inclusion-body myositis (s-IBM) is the most common progressive muscle disease associated with aging, and its muscle tissue has several phenotypic similarities to AD brain. Abnormal accumulation of intracellular multiprotein inclusions, containing p-tau in the form of paired helical filaments, amyloid-beta, and several other "Alzheimer-characteristic proteins", is a characteristic feature of the s-IBM muscle fiber phenotype. Diminished proteasomal and lysosomal protein degradation appear to play an important role in the formation of intra-muscle-fiber inclusions. We now report that: (1) in s-IBM muscle fibers, p62 protein is increased on both the protein and the mRNA levels, and it is strongly accumulated within, and as a dense peripheral shell surrounding, p-tau containing inclusions, by both the light- and electron-microscopy. Accordingly, our studies provide a new, reliable, and simple molecular marker of p-tau inclusions in s-IBM muscle fibers. The prominent p62 immunohistochemical positivity and pattern diagnostically distinguish s-IBM from polymyositis and dermatomyositis. (2) In normal cultured human muscle fibers, experimental inhibition of either proteasomal or lysosomal protein degradation caused substantial increase of p62, suggesting that similar in vivo mechanisms might contribute to the p62 increase in s-IBM muscle fibers.

We review the newest advances related to seeking the pathogenic mechanism(s) of sporadic inclusion-body myositis (s-IBM) and present the pathologic diagnostic criteria of s-IBM. We discuss the possible pathogenic role of several themes, such as 1) increased amyloid-beta precursor protein (AbetaPP) and of its fragment Abeta; 2) phosphorylation of tau protein; 3) oxidative stress; 4) abnormal a) signal-transduction, b) transcription, and c) RNA accumulation; 5) "junctionalization" and myogenous" denervation; and 6) lymphocytic inflammation. Evidence is provided supporting our hypothesis that overexpression of AbetaPP within the aging muscle fibers is an early upstream event causing the subsequent pathogenic cascade. The remarkable pathologic similarities between s-IBM muscle and Alzheimer disease (AD) brain are discussed, and the possible cause and significance are addressed.

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