Unique fiber phenotype composition and metabolic properties of the stapedius and tensor tympani muscles in the human middle ear.

The middle ear muscles have vital roles, yet their precise function in hearing and protection remains unclear. To better understand the function of these muscles in humans, the morphology, fiber composition, and metabolic properties of nine tensor tympani and eight stapedius muscles were analyzed with immunohistochemical, enzyme-histochemical, biochemical, and morphometric techniques. Human orofacial, jaw, extraocular, and limb muscles were used as references. The immunohistochemical analysis showed that the stapedius and tensor tympani muscles were markedly dominated by fibers expressing fast contracting myosin heavy chain MyHC-2A and MyHC-2X (79 ± 6% vs. 86 ± 9%, respectively, p = 0.04). In fact, the middle ear muscles had one of the highest proportions of MyHC-2 fibers ever reported for human muscles. Interestingly, the biochemical analysis revealed a MyHC isoform of unknown identity in both the stapedius and tensor tympani muscles. Muscle fibers containing two or more MyHC isoforms were relatively frequently observed in both muscles. A proportion of these hybrid fibers expressed a developmental MyHC isoform that is normally absent in adult human limb muscles. The middle ear muscles differed from orofacial, jaw, and limb muscles by having significantly smaller fibers (220 vs. 360 µm2 , respectively) and significantly higher variability in fiber size, capillarization per fiber area, mitochondrial oxidative activity, and density of nerve fascicles. Muscle spindles were observed in the tensor tympani muscle but not in the stapedius muscle. We conclude that the middle ear muscles have a highly specialized muscle morphology, fiber composition, and metabolic properties that generally showed more similarities to orofacial than jaw and limb muscles. Although the muscle fiber characteristics in the tensor tympani and stapedius muscles suggest a capacity for fast, fine-tuned, and sustainable contractions, their difference in proprioceptive control reflects different functions in hearing and protection of the inner ear.

Skeletal muscle morphology, satellite cells, and oxidative profile in relation to physical function and lifelong endurance training in very old men.

In the current study, we compared muscle morphology in three advanced aging cohorts that differed in physical function, including a unique cohort of lifelong endurance athletes. Biopsies from the vastus lateralis muscle of seven lifelong endurance athletes (EAs) aged 82-92 yr, and 19 subjects from the Uppsala Longitudinal Study of Adult Men (ULSAM) aged 87-91 yr were analyzed. ULSAM subjects were divided into high- (n = 9, HF) and low- (n = 10, LF) function groups based on strength and physical function tests. The analysis included general morphology, fiber type and cross-sectional area, capillarization, deficient cytochrome c oxidase (COX) activity, number of myonuclei and satellite cells, and markers of regeneration and denervation. Fibers with central nuclei and/or nuclear clumps were observed in all groups. EA differed from LF and HF by having a higher proportion of type I fibers, 52% more capillaries in relation to fiber area, fewer COX-negative fibers, and less variation in fiber sizes (all P < 0.05). There were no differences between the groups in the number of myonuclei and satellite cells per fiber, and no significant differences between LF and HF (P > 0.05). In conclusion, signs of aging were evident in the muscle morphology of all groups, but neither endurance training status nor physical function influenced signs of regeneration and denervation processes. Lifelong endurance training, but not higher physical function, was associated with higher muscle oxidative capacity, even beyond the age of 80.NEW & NOTEWORTHY Here we show that lifelong endurance training, but not physical function, is associated with higher muscle oxidative capacity, even beyond the age of 80 yr. Neither endurance training status nor physical function was significantly associated with satellite cells or markers of regeneration and denervation in muscle biopsies from these very old men.

Longitudinal Muscle and Myocellular Changes in Community-Dwelling Men Over Two Decades of Successful Aging-The ULSAM Cohort Revisited.

Participants of the population-based Uppsala longitudinal study of adult men (ULSAM) cohort reaching more than 88 years of age (survivors, S) were investigated at age 70, 82, and 88-90 and compared at 70 years with non-survivors (NS) not reaching 82 years. Body composition, muscle mass and muscle histology were remarkably stable over 18 years of advanced aging in S. Analysis of genes involved in muscle remodeling showed that S had higher mRNA levels of myogenic differentiation factors (Myogenin, MyoD), embryonic myosin (eMyHC), enzymes involved in regulated breakdown of myofibrillar proteins (Smad2, Trim32, MuRF1,) and NCAM compared with healthy adult men (n = 8). S also had higher mRNA levels of eMyHC, Smad 2, MuRF1 compared with NS. At 88 years, S expressed decreased levels of Myogenin, MyoD, eMyHC, NCAM and Smad2 towards those seen in NS at 70 years. The gene expression pattern of S at 70 years was likely beneficial since they maintained muscle fiber histology and appendicular lean body mass until advanced age. The expression pattern at 88 years may indicate a diminished muscle remodeling coherent with a decline of reinnervation capacity and/or plasticity at advanced age.

Fibre typing of intrafusal fibres.

The first descriptions of muscle spindles with intrafusal fibres containing striated myofibrils and nervous elements were given approximately 150 years ago. It took, however, another 100 years to establish the presence of two types of intrafusal muscle fibres: nuclear bag and nuclear chain fibres. The present paper highlights primarily the contribution of Robert Banks in fibre typing of intrafusal fibres: the confirmation of the principle of two types of nuclear bag fibres in mammalian spindles and the variation in occurrence of a dense M-band along the fibres. Furthermore, this paper summarizes how studies from the Umeå University group (Laboratory of Muscle Biology in the Department of Integrative Medical Biology) on fibre typing and the structure and composition of M-bands have contributed to the current understanding of muscle spindle complexity in adult humans as well as to muscle spindle development and effects of ageing. The variable molecular composition of the intrafusal sarcomeres with respect to myosin heavy chains and M-band proteins gives new perspectives on the role of the intrafusal myofibrils as stretch-activated sensors influencing tension/stiffness and signalling to nuclei.

Re-evaluation of sarcolemma injury and muscle swelling in human skeletal muscles after eccentric exercise.

The results regarding the effects of unaccustomed eccentric exercise on muscle tissue are often conflicting and the aetiology of delayed onset muscle soreness (DOMS) induced by eccentric exercise is still unclear. This study aimed to re-evaluate the paradigm of muscular alterations with regard to muscle sarcolemma integrity and fibre swelling in human muscles after voluntary eccentric exercise leading to DOMS. Ten young males performed eccentric exercise by downstairs running. Biopsies from the soleus muscle were obtained from 6 non-exercising controls, 4 exercised subjects within 1 hour and 6 exercised subjects at 2-3 days and 7-8 days after the exercise. Muscle fibre sarcolemma integrity, infiltration of inflammatory cells and changes in fibre size and fibre phenotype composition as well as capillary supply were examined with specific antibodies using enzyme histochemistry and immunohistochemistry. Although all exercised subjects experienced DOMS which peaked between 1.5 to 2.5 days post exercise, no significant sarcolemma injury or inflammation was detected in any post exercise group. The results do not support the prevailing hypothesis that eccentric exercise causes an initial sarcolemma injury which leads to subsequent inflammation after eccentric exercise. The fibre size was 24% larger at 7-8 days than at 2-3 days post exercise (p<0.05). In contrast, the value of capillary number per fibre area tended to decrease from 2-3 days to 7-8 days post exercise (lower in 5 of the 6 subjects at 7-8 days than at 2-3 days; p<0.05). Thus, the increased fibre size at 7-8 days post exercise was interpreted to reflect fibre swelling. Because the fibre swelling did not appear at the time that DOMS peaked (between 1.5 to 2.5 days post exercise), we concluded that fibre swelling in the soleus muscle is not directly associated with the symptom of DOMS.

Intrafusal myosin heavy chain expression of human masseter and biceps muscles at young age shows fundamental similarities but also marked differences.

Muscle spindles are skeletal muscle mechanoreceptors that provide proprioceptive information to the central nervous system. The human adult masseter muscle has greater number, larger and more complex muscle spindles than the adult biceps. For a better knowledge of muscle diversity and physiological properties, this study examined the myosin heavy chain (MyHC) expression of muscle spindle intrafusal fibres in the human young masseter and young biceps muscles by using a panel of monoclonal antibodies (mAbs) against different MyHC isoforms. Eight MyHC isoforms were detected in both muscles-slow-tonic, I, IIa, IIx, foetal, embryonic, α-cardiac and an isoform not previously reported in intrafusal fibres, termed IIx'. Individual fibres co-expressed 2-6 isoforms. MyHC-slow tonic separated bag1, AS-bag1 and bag2 fibres from chain fibres. Typically, bag fibres also expressed MyHC-I and α-cardiac, whereas chain fibres expressed IIa and foetal. In the young masseter 98 % of bag1 showed MyHC-α cardiac versus 30 % in the young biceps, 35 % of bag2 showed MyHC-IIx' versus none in biceps, 17 % of the chain fibres showed MyHC-I versus 61 % in the biceps. In conclusion, the result showed fundamental similarities in intrafusal MyHC expression between young masseter and biceps, but also marked differences implying muscle-specific proprioceptive control, probably related to diverse evolutionary and developmental origins. Finding of similarities in MyHC expression between young and adult masseter and biceps muscle spindles, respectively, in accordance with previously reported similarities in mATPase fibre type composition suggest early maturation of muscle spindles, preceding extrafusal fibres in growth and maturation.

Remarkable heterogeneity in myosin heavy-chain composition of the human young masseter compared with young biceps brachii.

Adult human jaw muscles differ from limb and trunk muscles in enzyme-histochemical fibre type composition. Recently, we showed that the human masseter and biceps differ in fibre type pattern already at childhood. The present study explored the myosin heavy-chain (MyHC) expression in the young masseter and biceps muscles by means of gel electrophoresis (GE) and immuno-histochemical (IHC) techniques. Plasticity in MyHC expression during life was evaluated by comparing the results with the previously reported data for adult muscles. In young masseter, GE identified MyHC-I, MyHC-IIa MyHC-IIx and small proportions of MyHC-fetal and MyHC-α cardiac. Western blots confirmed the presence of MyHC-I, MyHC-IIa and MyHC-IIx. IHC revealed in the masseter six isomyosins, MyHC-I, MyHC-IIa, MyHC-IIx, MyHC-fetal, MyHC α-cardiac and a previously not reported isoform, termed MyHC-IIx'. The majority of the masseter fibres co-expressed two to four isoforms. In the young biceps, both GE and IHC identified MyHC-I, MyHC-IIa and MyHC-IIx. MyHC-I predominated in both muscles. Young masseter showed more slow and less-fast and fetal MyHC than the adult and elderly masseter. These results provide evidence that the young masseter muscle is unique in MyHC composition, expressing MyHC-α cardiac and MyHC-fetal isoforms as well as hitherto unrecognized potential spliced isoforms of MyHC-fetal and MyHC-IIx. Differences in masseter MyHC expression between young adult and elderly suggest a shift from childhood to adulthood towards more fast contractile properties. Differences between masseter and biceps are proposed to reflect diverse evolutionary and developmental origins and confirm that the masseter and biceps present separate allotypes of muscle.

Differences in fibre type composition between human masseter and biceps muscles in young and adults reveal unique masseter fibre type growth pattern.

The human jaw system is different from those of other primates, carnivores, ruminants, and rodents in temporomandibular joint and muscle anatomy. In adults, jaw muscles also differ markedly from limb and trunk muscles in composition and distribution of fibre types. It can be assumed that age-related changes between young age to adulthood in terms of craniofacial growth, teeth eruption, and improvement of jaw functions are paralleled by alterations also in composition and distribution of jaw muscle fibre types. To address this question, we have examined the fibre type composition of the human masseter, a jaw closing muscle, at young age. For comparison, the young biceps brachii was examined. The results were compared with previous data for adult masseter and biceps muscles. Young masseter and biceps were similar in that type I fibres outnumbered other fibre types and were of the same diameter. However, they differed in composition of other fibre types. Young masseter contained fibre types I, IM, IIC, IIAB, IIB, and scarce IIA, with regional differences, whereas young biceps showed types I, IIA, IIAB, and few IIB. Young masseter differed from young biceps also by smaller type II fibre diameter and by containing fetal MyHC. In addition, the masseter and biceps differed in age-related changes of composition and distribution of fibre types between young age and adulthood. We conclude that the human masseter is specialized in fibre types already at young age and shows a unique fibre type growth pattern, in concordance with being a separate allotype of muscle.

Muscle spindle composition and distribution in human young masseter and biceps brachii muscles reveal early growth and maturation.

Significant changes in extrafusal fiber type composition take place in the human masseter muscle from young age, 3-7 years, to adulthood, in parallel with jaw-face skeleton growth, changes of dentitions and improvement of jaw functions. As motor and sensory control systems of muscles are interlinked, also the intrafusal fiber population, that is, muscle spindles, should undergo age-related changes in fiber type appearance. To test this hypothesis, we examined muscle spindles in the young masseter muscle and compared the result with previous data on adult masseter spindles. Also muscle spindles in the young biceps brachii muscle were examined. The result showed that muscle spindle composition and distribution were alike in young and adult masseter. As for the adult masseter, young masseter contained exceptionally large muscle spindles, and with the highest spindle density and most complex spindles found in the deep masseter portion. Hence, contrary to our hypothesis, masseter spindles do not undergo major morphological changes between young age and adulthood. Also in the biceps, young spindles were alike adult spindles. Taken together, the results showed that human masseter and biceps muscle spindles are morphologically mature already at young age. We conclude that muscle spindles in the human young masseter and biceps precede the extrafusal fiber population in growth and maturation. This in turn suggests early reflex control and proprioceptive demands in learning and maturation of jaw motor skills. Similarly, well-developed muscle spindles in young biceps reflect early need of reflex control in learning and performing arm motor behavior.

Tissue-specific splicing of ISCU results in a skeletal muscle phenotype in myopathy with lactic acidosis, while complete loss of ISCU results in early embryonic death in mice.

Hereditary myopathy with lactic acidosis (HML) is caused by an intron mutation in the iron-sulphur cluster assembly gene (ISCU) leading to incorporation of intron sequence into the mRNA. This results in a deficiency of Fe-S cluster proteins, affecting the TCA cycle and the respiratory chain. The proteins involved in the Fe-S machinery are evolutionary conserved and shown to be fundamental in all organisms examined. ISCU is expressed at high levels in numerous tissues in mammals, including high metabolic tissues like the heart, suggesting that a drastic mutation in the ISCU gene would be damaging to all energy-demanding organs. In spite of this, the symptoms in patients with HML are restricted to skeletal muscle, and it has been proposed that splicing events may contribute to the muscle specificity. In this study we confirm that a striking difference in the splicing pattern of mutant ISCU exists between different tissues. The highest level of incorrectly spliced ISCU mRNA was found in skeletal muscle, while the normal splice form predominated in patient heart. The splicing differences were also reflected at a functional level, where loss of Fe-S cluster carrying enzymes and accumulation of iron were present in muscle, but absent in other tissues. We also show that complete loss of ISCU in mice results in early embryonic death. The mice data confirm a fundamental role for ISCU in mammals and further support tissue-specific splicing as the major mechanism limiting the phenotype to skeletal muscle in HML.

Sarcopenic obesity: satellite cells in the aging muscle.

PURPOSE OF REVIEW: Current knowledge on satellite cells in relation to suggested mechanisms of loss of muscle mass and strength, induction of fat infiltration, and countermeasures is highlighted. RECENT FINDINGS: Consensus on the definition of sarcopenia and sarcopenic obesity is proposed. Human satellite cell heterogeneity has now unequivocally been verified in situ as well as an adipogenic potential, though in mice other muscle stem cells are the hot topic to induce adipogenesis upon muscle damage. Inflammation, oxidative stress, proteolytic degradation, and nuclear apoptosis are discussed as pathogenetic mechanisms of sarcopenia, although little evidence exists that they are important in human muscle. In rodents, exercise-induced muscle injury is a hallmark for sequential events leading to muscle fiber necrosis and sarcopenia. Exercise in humans, on the contrary, is the key event to countermeasure sarcopenia. Cautions to extrapolate observation in rodents to explain human conditions have been presented. SUMMARY: Human satellite cells are indispensable for maintenance of human muscle mass, but their implications in the pathogenesis of sarcopenia and sarcopenic obesity are still under debate. Nevertheless, satellite cell activation upon exercise seems unequivocally together with adequate nutrition to be the most effective countermeasure for sarcopenia and sarcopenic obesity.

Satellite cell heterogeneity with respect to expression of MyoD, myogenin, Dlk1 and c-Met in human skeletal muscle: application to a cohort of power lifters and sedentary men.

Human satellite cells (SCs) are heterogeneous with respect to markers for their identification in the niche between the muscle fibre plasma membrane and its basal lamina. We have previously shown that, in biopsies from highly competitive power lifters, power lifters with long-term use of anabolic steroids and a population of healthy sedentary men, antibodies against the neuronal cell adhesion molecule (NCAM) and the paired box transcription factor Pax7 together label 94% of the SCs, NCAM alone labels 4% and Pax7 alone labels 1%. In the present study, we have further studied these biopsies with four markers related to SC activation and differentiation. Our study unequivocally shows that staining for MyoD and myogenin are present in nuclei of SCs and of myoblasts and myotubes in areas of muscle fibre regeneration. Staining for c-Met was observed in a proportion of Pax7+ SCs. However, widespread labelling of the sarcolemma precluded the quantification of c-Met+/Pax7+ SCs and the use of c-Met as a reliable SC marker. Pax7+ SCs labelled by anti-Delta like1 (Dlk1) were present in all samples but in variable proportions, whereas muscle progenitor cells related to repair were Dlk1⁻. Staining for Dlk1 was also observed in Pax7⁻ interstitial cells and in the cytoplasm of some small muscle fibres. Interestingly, the proportion of Dlk1+/Pax7+ SCs was significantly different between the groups of power lifters. Thus, our study confirms that human SCs show marked heterogeneity and this is discussed in terms of SC activation, myonuclei turnover, muscle fibre growth and muscle fibre damage and repair.

Skeletal muscle telomere length in healthy, experienced, endurance runners.

Measuring the DNA telomere length of skeletal muscle in experienced endurance runners may contribute to our understanding of the effects of chronic exposure to endurance exercise on skeletal muscle. This study compared the minimum terminal restriction fragment (TRF) length in the vastus lateralis muscle of 18 experienced endurance runners (mean age: 42 +/- 7 years) to those of 19 sedentary individuals (mean age: 39 +/- 10 years). The runners had covered almost 50,000 km in training and racing over 15 years. Minimum TRF lengths measured in the muscle of both groups were similar (P = 0.805) and within the normal range. Minimum TRF length in the runners, however, was inversely related to their years spent running (r = -0.63, P = 0.007) and hours spent training (r = -0.52, P = 0.035). Therefore, since exposure to endurance running may influence minimum TRF length, and by implication, the proliferative potential of the satellite cells, chronic endurance running may be seen as a stressor to skeletal muscle.

Intragenic deletion of TRIM32 in compound heterozygotes with sarcotubular myopathy/LGMD2H.

In 2005 the commonality of sarcotubular myopathy (STM) and limb girdle muscular dystrophy type 2H (LGMD2H) was demonstrated, as both are caused by the p D487N missense mutation in TRIM32 originally found in the Manitoba Hutterite population. Recently, three novel homozygous TRIM32 mutations have been described in LGMD patients. Here we describe a three generation Swedish family clinically presenting with limb girdle muscular weakness and histological features of a microvacuolar myopathy. The two index patients were compound heterozygotes for a frameshift mutation in TRIM32 (c.1560delC ) and a 30 kb intragenic deletion, encompassing parts of intron 1 and the entire exon 2 of TRIM32. In these patients, no full-length or truncated TRIM32 could be detected. Interestingly, heterozygous family members carrying only one mutation showed mild clinical symptoms and vacuolar changes in muscle. In our family, the phenotype encompasses additionally a mild demyelinating polyneuropathic syndrome. Thus STM and LGMD2H are the result of loss of function mutations that can be either deletions or missense mutations.

New multiple labelling method for improved satellite cell identification in human muscle: application to a cohort of power-lifters and sedentary men.

Presently applied methods to identify and quantify human satellite cells (SCs) give discrepant results. We introduce a new immunofluorescence method that simultaneously monitors two SC markers (NCAM and Pax7), the basal lamina and nuclei. Biopsies from power-lifters, power-lifters using anabolic substances and untrained subjects were re-examined. Significantly different results from those with staining for NCAM and nuclei were observed. There were three subtypes of SCs; NCAM(+)/Pax7(+) (94%), NCAM(+)/Pax7(-) (4%) and NCAM(-)/Pax7(+) (1%) but large individual variability existed. The proportion of SCs per nuclei within the basal lamina of myofibres (SC/N) was similar for all groups reflecting a balance between the number of SCs and myonuclei to maintain homeostasis. We emphasise that it is important to quantify both SC/N and the number of SCs per fibre. Our multiple marker method is more reliable for SC identification and quantification and can be used to evaluate other markers of muscle progenitor cells.

New aspects of obscurin in human striated muscles.

Obscurin is a giant protein (700-800 kDa) present in both skeletal muscles and myocardium. According to animal studies, obscurin interacts with myofibrillar Z-discs during early muscle development, but is translocalised to be predominantly associated with the M-bands in mature muscles. The proposed function for obscurin is in the assembly and organisation of myosin into regular A-bands during formation of new sarcomeres. In the present study, the precise localisation of obscurin in developing and mature normal human striated muscle is presented for the first time. We show that obscurin surrounded myofibrils at the M-band level in both developing and mature human skeletal and heart muscles, which is partly at variance with that observed in animals. At maturity, obscurin also formed links between the peripheral myofibrils and the sarcolemma, and was a distinct component of the neuromuscular junctions. Obscurin should therefore be regarded as an additional component of the extrasarcomeric cytoskeleton. To test this function of obscurin, biopsies from subjects with exercise-induced delayed onset muscle soreness (DOMS) were examined. In these subjects, myofibrillar alterations related to sarcomerogenesis are observed. Our immunohistochemical analysis revealed that obscurin was never lacking in myofibrillar alterations, but was either preserved at the M-band level or diffusely spread over the sarcomeres. As myosin was absent in such areas but later reincorporated in the newly formed sarcomeres, our results support that obscurin also might play an important role in the formation and maintenance of A-bands.

Myopathy with lactic acidosis is linked to chromosome 12q23.3-24.11 and caused by an intron mutation in the ISCU gene resulting in a splicing defect.

We describe the mapping and identification of the gene for hereditary myopathy with lactic acidosis (HML). HML is characterized by low physical performance, resulting in physical exertion that causes early exhaustion, dyspnoea and palpitations. Using an autosomal recessive mode of inheritance, we mapped the trait to chromosome 12q23.3-24.11, with a maximum lod score of 5.26. The 1.6-Mb disease-critical region contained one obvious candidate gene-ISCU-specifying a protein involved in iron-sulphur cluster assembly. IscU is produced in two isoforms; one cytosolic and one mitochondrial, coded for by different splice variants of the ISCU gene. Mutational analysis of all exon and intron sequences as well as 1000 bp of the promoter of the ISCU gene revealed one intron mutation that was specific for the disease haplotype. The mutation is located in a region with homology to the interferon-stimulated response element (ISRE), but we could not see any effect of the mutation on expression levels in vitro or in vivo. We did, however, observe a drastic difference in the splicing pattern between patients and controls. In controls the mRNA was, as expected, mainly in the mitochondrial form, while in the patients a larger mRNA transcript was predominant. Sequencing of the product revealed that the mutation activates cryptic splice sites in intron 5 resulting in aberrant mRNA containing 100 bp of the intron. To conclude, our data strongly suggest that an intron mutation in the ISCU gene, leading to incorrectly spliced mRNA, is the cause of myopathy with lactic acidosis in this family.

Myotilin: a prominent marker of myofibrillar remodelling.

Myofibrillar remodelling with insertion of sarcomeres is a typical feature of biopsies taken from persons suffering of exercise-induced delayed onset muscle soreness. Here we studied the presence of the sarcomeric protein myotilin in eccentric exercise related lesions. Myotilin is a component of sarcomeric Z-discs and it binds several other Z-disc proteins, i.e. alpha-actinin, filamin C, F-actin and FATZ. Myotilin has previously been shown to be present in nemaline rods and central cores and to be mutated in limb girdle muscular dystrophy 1A (LGMD1A) and in a subset of myofibrillar myopathies, indicating an important role in Z-disc maintenance. Our findings on non-diseased muscle affected by eccentric exercise give new information on how myotilin is associated to myofibrillar components upon remodelling. We show that myotilin was present in increased amount in lesions related to Z-disc streaming and events leading to insertion of new sarcomeres in pre-existing myofibrils and can therefore be used as a marker for myofibrillar remodelling. Interestingly, myotilin is preferentially associated with F-actin rather than with the core Z-disc protein alpha-actinin during these events. This suggests that myotilin has a key role in the dynamic molecular events mediating myofibrillar assembly in normal and diseased skeletal muscle.

Premature proliferative arrest of cricopharyngeal myoblasts in oculo-pharyngeal muscular dystrophy: Therapeutic perspectives of autologous myoblast transplantation.

Cultures of myoblasts isolated from cricopharyngeal muscles from patients with oculopharyngeal muscular dystrophy (OPMD) have been performed to study the effect of the expanded (GCG)8-13 repeat, located on the poly(A) binding protein nuclear-1 (PABPN1), on satellite cell phenotype. Cell cultures exhibited a reduced myogenicity, as well as a rapid decrease in proliferative lifespan, as compared to controls. The incorporation of BrdU decreased during the proliferative lifespan, due to a progressive accumulation of non-dividing cells. A lower fusion index was also observed, but myoblasts were able to form large myotubes when OPMD cultures were purified, although a rapid loss of myogenicity during successive passages was also observed. Myoblasts isolated from unaffected muscles did not show the defects observed in cricopharyngeal muscle cultures. The PABPN1 was predominantly located in nuclei of myoblasts and in both the nuclei and cytoplasm of myotubes in OPMD cultures. In vivo analysis of OPMD muscles showed that the number of satellite cells was slightly higher than that observed in age matched controls. Mutation of the PABPN1 in OPMD provokes premature senescence in dividing myoblasts, that may be due to intranuclear toxic aggregates. These results suggest that myoblast autografts, isolated from unaffected muscles, and injected into the dystrophic pharyngeal muscles, may be a useful therapeutic strategy to restore muscular function. Its tolerance and feasibility has been preclinically demonstrated in the dog.