Duchenne muscular dystrophy remains an untreatable genetic disease that severely limits motility and life expectancy in affected children. The only animal model specifically reproducing the alterations in the dystrophin gene and the full spectrum of human pathology is the golden retriever dog model. Affected animals present a single mutation in intron 6, resulting in complete absence of the dystrophin protein, and early and severe muscle degeneration with nearly complete loss of motility and walking ability. Death usually occurs at about 1 year of age as a result of failure of respiratory muscles. Here we report that intra-arterial delivery of wild-type canine mesoangioblasts (vessel-associated stem cells) results in an extensive recovery of dystrophin expression, normal muscle morphology and function (confirmed by measurement of contraction force on single fibres). The outcome is a remarkable clinical amelioration and preservation of active motility. These data qualify mesoangioblasts as candidates for future stem cell therapy for Duchenne patients.

Recent evidence points to a strong relationship between increased mitochondrial biogenesis and increased survival in eukaryotes. Branched-chain amino acids (BCAAs) have been shown to extend chronological life span in yeast. However, the role of these amino acids in mitochondrial biogenesis and longevity in mammals is unknown. Here, we show that a BCAA-enriched mixture (BCAAem) increased the average life span of mice. BCAAem supplementation increased mitochondrial biogenesis and sirtuin 1 expression in primary cardiac and skeletal myocytes and in cardiac and skeletal muscle, but not in adipose tissue and liver of middle-aged mice, and this was accompanied by enhanced physical endurance. Moreover, the reactive oxygen species (ROS) defense system genes were upregulated, and ROS production was reduced by BCAAem supplementation. All of the BCAAem-mediated effects were strongly attenuated in endothelial nitric oxide synthase null mutant mice. These data reveal an important antiaging role of BCAAs mediated by mitochondrial biogenesis in mammals.

Preclinical or clinical trials for muscular dystrophies have met with modest success, mainly because of inefficient delivery of viral vectors or donor cells to dystrophic muscles. We report here that intra-arterial delivery of wild-type mesoangioblasts, a class of vessel-associated stem cells, corrects morphologically and functionally the dystrophic phenotype of virtually all downstream muscles in adult immunocompetent alpha-sarcoglycan (alpha-SG) null mice, a model organism for limb-girdle muscular dystrophy. When mesoangioblasts isolated from juvenile dystrophic mice and transduced with a lentiviral vector expressing alpha-SG were injected into the femoral artery of dystrophic mice, they reconstituted skeletal muscle in a manner similar to that seen in wild-type cells. The success of this protocol was mainly due to widespread distribution of donor stem cells through the capillary network, a distinct advantage of this strategy over previous approaches.

1. A large population (n = 151) of human skinned skeletal muscle fibres has been studied.Force-velocity curves of sixty-seven fibres were obtained by load-clamp manoeuvres at 12 'C. In each fibre maximum shortening velocity (Vmax), maximum power output (Tmax), optimal velocity (velocity at which Wmax is developed, Vopt), optimal force (force at which Wmax is developed, P.pt), specific tension (P0/CSA, isometric tension/cross-sectional area)were assessed. Unloaded shortening velocity (1VO) was also determined at 12 'C in a different

Contractile and energetic properties of human skeletal muscle have been studied for many years in vivo in the body. It has been, however, difficult to identify the specific role of muscle fibres in modulating muscle performance. Recently it has become possible to dissect short segments of single human muscle fibres from biopsy samples and make them work in nearly physiologic conditions in vitro. At the same time, the development of molecular biology has provided a wealth of information on muscle proteins and their genes and new techniques have allowed analysis of the protein isoform composition of the same fibre segments used for functional studies. In this way the histological identification of three main human muscle fibre types (I, IIA and IIX, previously called IIB) has been followed by a precise description of molecular composition and functional and biochemical properties. It has become apparent that the expression of different protein isoforms and therefore the existence of distinct muscle fibre phenotypes is one of the main determinants of the muscle performance in vivo. The present review will first describe the mechanisms through which molecular diversity is generated and how fibre types can be identified on the basis of structural and functional characteristics. Then the molecular and functional diversity will be examined with regard to (1) the myofibrillar apparatus; (2) the sarcolemma and the sarcoplasmic reticulum; and (3) the metabolic systems devoted to producing ATP. The last section of the review will discuss the advantage that fibre diversity can offer in optimizing muscle contractile performance.

SUMMARY1. This study was performed to assess whether muscle contractile properties are related to the presence of specific myosin heavy chain (MHC) isoforms.2. Force-velocity relations and MHC isoform composition were determined in seventy-four single skinned muscle fibres from rat soleus, extensor digitorum longus and plantaris muscles.3. Four groups of fibres were identified according to their MHC isoform composition determined by monoclonal antibodies: type 1 (slow), and types 2A, 2B and 2X (fast).4. With respect to maximum velocity of shortening (V0), the fibres formed a continuum between 035 and 2 84 L/s (muscle lengths per second) at 12 'C. V0 in type 1 fibres (slow fibres) was between 0-35 and 095 L/s (0-639 + 0038 L/s; mean + S.E. of mean). V0 in type 2 fibres (fast fibres) was consistently higher than 091 L/s. Ranges of V0 in the three fast fibre types mostly overlapped. Type 2A and 2X fibres had similar mean V1 values (1P396+0±084 and 1-451 +0066 L/s respectively); type 2B fibres showed a higher mean V1 value (1 800+±0109 L/s) than type 2A and 2X fibres.5. Mean values of a/P*, an index of the curvature of force-velocity relations, allowed us to identify two groups of fibres: a high curvature group comprised of type 1 (mean a/PO, 0066+0007) and 2A (0066±+0024) fibres and a low curvature group comprised of type 2B (0 113+0-013) and 2X (0 132+O0008) fibres.6. Maximal power output was lower in slow fibres than in fast fibres, and among fast fibres it was lower in type 2A fibres than in type 2X and 2B.7. Force per unit cross-sectional area was less in slow fibres than in fast fibres. There was no relation between fibre type and cross-sectional area.8. The results suggest that MHC composition is just one of the determinants of shortening velocity and of other muscle contractile properties.

Biopsy samples were taken from vastus lateralis muscle of seven young (YO, age 30.2 +/- 2.2 years), and seven elderly (EL, age 72.7 +/- 2.3 years) subjects and two elderly subjects whose right leg had been immobilized for 3.5 months (EL-IMM, ages 70 and 75). The following main parameters were studied: (1) myosin heavy chain (MHC) isoform distribution of the samples, determined by SDS-PAGE; (2) cross-sectional area (CSA), specific force (Po/CSA) and maximum shortening velocity (Vo) of a large population (n = 593) of single skinned muscle fibres, classified on the basis of MHC isoform composition determined by SDS-PAGE; (3) actin sliding velocity (Vf) on pure myosin isoforms determined by in vitro motility assays; (4) myosin concentration in single fibres determined by quantitative SDS-PAGE. MHC isoform distribution was shifted towards fast isoforms in EL and to a larger extent in EL-IMM. In EL and, more consistently, in EL-IMM we observed a higher percentage of hybrid fibres than in YO, and noted the presence of MHC-neonatal and of unusual hybrid fibres containing more than two MHC isoforms. Po/CSA significantly decreased in type 1 and 2A fibres in the order YO EL EL-IMM. Vo of type 1 and 2A fibres was significantly lower in EL and higher in EL-IMM than in YO, i.e. immobilization more than counteracted the age-dependent decrease in Vo. The latter phenomenon was not observed for Vf. Vf on myosin 1 was lower in both EL and EL-IMM than in YO. Vf on myosin 2X was lower in EL than in YO, and a similar trend was observed for myosin 2A. Myosin concentration decreased in type 1 and 2A fibres in the order YO EL EL-IMM and was linearly related to the Po/CSA values of corresponding fibre types from the same subjects. The experiments suggest that (1) myosin concentration is a major determinant of the lower Po/CSA of single fibres in ageing and especially following immobilization and (2) ageing is associated with lower Vo of single fibres due to changes in the properties of myosin itself, whereas immobilization is associated with higher Vo in the absence of a change in myosin function.

1. This study aims to assess the role of myosin heavy chain (MHC) and alkali myosin light chain (MLC) isoforms in determining maximum velocity of shortening in fast skeletal muscle fibres. Sreter & Gergely, 1988; Bottinelli, Schiaffino & al. 1988;Rome, Sosnicki & Goble, 1990). A more recent study Reggiani, 1991). It is less well understood whether the large (Bottinelli et al. 1991) has for the first time also determined differences in maximum velocity of shortening among fast contractile properties of IIX MHC-containing fibres, and fibres are due to the presence of different fast MHCs, or on has shown that II B fibres have mean maximum shortening NS 2808, pp. 341-349 341J