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We present two cases of patients with juvenile amyotrophic lateral sclerosis (ALS), who had no history of familial ALS. The symptoms of both patients started as weakness of the unilateral upper limb and neck, and extended to bulbar and respiratory weakness in a relatively short period. Of note, the first patient was mentally retarded before the onset of weakness. Fused in sarcoma/translocated in liposarcoma (FUS/TLS) gene analyses revealed mutations of p. G492EfsX527 (c. 1475delG), which is a novel deletion/frameshift mutation, in the first patient and p. R514S mutation (c. 1542G > T) in the second patient. Molecular analysis revealed that the mutant FUS/TLS, especially the deletion/frameshift mutation, showed significant cytoplasmic localization in transfected motor neuron-like cells. Our findings suggest the association of mental retardation with the FUS/TLS mutation. Further investigation, including the effect of FUS/TLS on cognitive function, would aid better understanding of FUS/TLS proteinopathies.

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Skeletal muscle expressing Pro104Leu mutant caveolin 3 (CAV3 P104L ) in mouse becomes atrophied and serves as a model of autosomal dominant limb-girdle muscular dystrophy 1C. We previously found that caveolin 3-deficient muscles showed activated intramuscular transforming growth factor beta (TGF-b) signals. However, the cellular mechanism by which loss of caveolin 3 leads to muscle atrophy is unknown. Recently, several small-molecule inhibitors of TGF-b type I receptor (TbRI) kinase have been developed as molecular-targeting drugs for cancer therapy by suppressing intracellular TGF-b1, -b2, and -b3 signaling. Here, we show that a TbRI kinase inhibitor, Ki26894, restores impaired myoblast differentiation in vitro caused by activin, myostatin, and TGF-b1, as well as CAV3 P104L . Oral administration of Ki26894 increased muscle mass and strength in vivo in wild-type mice, and improved muscle atrophy and weakness in the CAV3 P104L mice. The inhibitor restored the number of satellite cells, the resident stem cells of adult skeletal muscle, with suppression of the increased phosphorylation of Smad2, an effector, and the upregulation of p21 (also known as Cdkn1a), a target gene of the TGF-b family members in muscle. These data indicate that both TGF-b-dependent reduction in satellite cells and impairment of myoblast differentiation contribute to the cellular mechanism underlying caveolin 3-deficient muscle atrophy. TbRI kinase inhibitors could antagonize the activation of intramuscular anti-myogenic TGF-b signals, thereby providing a novel therapeutic rationale for the alternative use of this type of anticancer drug in reversing muscle atrophy in various clinical settings.

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Duchenne muscular dystrophy (DMD) is an inherited severe muscle wasting disorder with, thus far, no effective therapy. DMD causes respiratory and cardiac failure as well as muscle wastage. Among the various symptoms, respiratory insufficiency is a major cause of death in DMD patients at about 20 years of age. So, naturally, the improvement of respiratory function will extend the patient's life. We report here, for the first time, a sensitive procedure using whole-body plethysmography to monitor respiratory parameters detected in the utrophin/dystrophin double knockout mouse (dko mouse), showing quite similar systemic symptoms to human DMD including restrictive ventilatory impairment. Furthermore, we show that a highly efficient dystrophin-transduction to the dko's diaphragm--achieved by simple intraperitoneal injection of a helper-dependent adenovirus vector (HDAdv) containing the full-length dystrophin expression cassette--provided beneficial results. In spite of dystrophin expression only in the diaphragm, this focal gene transfer could result in the rescue from ventilatory impairment (increased tidal volume (TV) and improvement of compensatory hyperpnea). Our result suggests that a DMD patient's mortal ventilatory impairment may be improved via technically easy means through the intraperitoneal injection of HDAdv.

Variations in gene promoter/enhancer activity in different muscle fiber types after gene transduction was noticed previously, but poorly analyzed. The murine stem cell virus (MSCV) promoter drives strong, stable gene expression in hematopoietic stem cells and several other cells, including cerebellar Purkinje cells, but it has not been studied in muscle. We injected a lentiviral vector carrying an MSCV-EGFP cassette (LvMSCV-EGFP) into tibialis anterior muscles and observed strong EGFP expression in muscle fibers, primary cultured myoblasts, and myotubes isolated from injected muscles. We also generated lentiviral-mediated transgenic mice carrying the MSCV-EGFP cassette and detected transgene expression in striated muscles. LvMSCV-EGFP transgenic mice showed fiber type-dependent variations in expression: highest in types I and IIA, intermediate in type IID/X, and lowest in type IIB fibers. The soleus and diaphragm muscles, consisting mainly of types I and IIA, are most severely affected in the mdx mouse model of muscular dystrophy. Further analysis of this promoter may have the potential to achieve certain gene expression in severely affected muscles of mdx mice. The Lv-mediated transgenic mouse may prove a useful tool for assessing the enhancer/promoter activities of a variety of different regulatory cassettes.

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