Sarcolemmal fragility secondary to the degradation of dystrophin in dilated cardiomyopathy, as estimated by electron microscopy

A common gene deletion or mutation of delta-sarcoglycan (δ-SG) in dystrophin-related proteins (DRPs) is identified in both TO-2 strain hamsters and human families with dilated cardiomyopathy. We have succeeded in the long-lasting in vivo supplementation of a normal δ-SG gene by recombinant adeno-associated virus vector, restoration of the morphological and functional degeneration, and improvement in the prognosis of the TO-2 hamster. To evaluate the integrity of the sarcolemma (SL) and the subsequent change of organelles in cardiomyocytes of the TO-2 strain hamster, we examined electron microscopy (EM) images focusing on the sarcolemmal stability at the end stage of heart failure. Two types of sarcolemmal degradation were detected: the widened and locally thickened SL, and blurred and discontinuous SL. Bizarrely formed mitochondria of varying sizes were also observed. Immuno-EM revealed clear expression of dystrophin in the SL and intense expression at the costamere as well as at the T-tubules in the control F1B strain hearts, but a patchy deposition of dystrophin was observed along the SL without the transgene of δ-SG. In contrast to the previous reports that dystrophin’s integrity was intact, the present results suggest that the gene deletion of δ-SG and the loss of δ-SG protein in the SL cardioselectively cause the morphological and functional deterioration of dystrophin and the resultant instability of the SL. The sarcolemmal fragility may be similar to Duchenne-type progressive muscular dystrophy in skeletal muscle. In addition to the mechanical role, another aspect of DRPs for the intracellular signal transmission is also discussed.     

Dystrophin and a dystrophin-related protein in intrafusal muscle fibers,and neuromuscular and myotendinous junctions

Summary To determine whether or not and how dystrophin exists in neuromuscular junctions (NMJs) and myotendinous junctions (MTJs), we studied the mid-belly and peripheral portions of control and mdx muscles, immunohistochemically and immunoelectrophoretically, using six kinds of polyclonal antibodies, and an antibody against a dystrophin-related protein (DRP). In controls these regions and the polar region of intrafusal muscle fibers showed a rather clearer immunohistochemical dystrophin reaction than those of extrafusal muscle fibers with all antibodies used. In the muscles of mdx mice NMJs only showed a positive dystrophin reaction with the c-terminal antibody, that is, no reaction with the other five antibodies, and MTJs in mdx showed a positive reaction with the c-terminal antibody and a faint to negative reaction with the other five antibodies. In biopsied human muscles NMJs and MTJs also showed a clear reaction with all ten antibodies, i.e., six polyclonal and four monoclonal ones. Although an immunohistochemical DRP reaction was clearly seen at NMJs, only a faint or no reaction was seen on MTJs and on intrafusal muscle fibers in both mouse and human materials. Western blot analysis of control mouse muscle for dystrophin showed a clearer band for the peripheral portion, which contains many MTJs, than for the mid-belly portion. These data suggest that dystrophin really exists on MTJs, and that dystrophin and DRP exist on NMJs in mouse and human muscles.     

Subsarcolemmal expression of utrophin in neuromuscular disorders: an immunohistochemical study of 80 cases

The immunohistochemical expression of utrophin in 80 muscle biopsies from patients with dystrophinopathies and other neuromuscular disorders is reported. All biopsy specimens were routinely studied by a battery of 12 histoenzymatic techniques, and immunohistochemistry was performed for spectrin, three domains of dystrophin and two domains of utrophin. Abnormal utrophin expression was observed in all dystrophinopathic muscles compared with normal controls or biopsy samples from several other muscular diseases. Inflammatory myopathies presented abnormal overexpression of utrophin and an abnormal dystrophin immunolabeling pattern. This overexpression of utrophin appears to be directly related to the decrease in dystrophin. We conclude that the study of utrophin is important for the histological interpretation and differential diagnosis of dystrophin-related muscular disorders. Received: 13 November 1997 / Revised: 6 February 1998, 20 May 1998 / Accepted: 22 May 1998     

Dystrophin and dystrophin-related protein in the central nervous system of normal controls and Duchenne muscular dystrophy

To clarify the localization and characterization of dystrophin and dystrophin-related protein (DRP) in the human central nervous system (CNS), we carried out immunoblotting and immunostaining studies using three region-specific anti-dystrophin and one anti-DRP antibodies. With immunostaining, punctate immuno-reactivity of dystrophin was seen along the cell bodies and dendrites of the cerebral cortical neurons and cerebellar Purkinje cells in the normal controls autopsied. By contrast, dystrophin was not detected at all in the CNS of Duchenne muscular dystrophy (DMD) patients with intellectual disturbance. Immunoreactivity of DRP was observed in the vascular walls of both normal and DMD brains, but not in the neuronal cells. Compensatory increase of DRP was not noted in DMD brains. This study suggests that in DMD the brain-type dystrophin originally present in neurons is absent and may be related to the intellectual disturbance.Supported by a grant (2-A) from the National Center of Neurology and Psychiatry of the Ministry of Health and Welfare, Japan     

Dystrophin is replaced by utrophin in frog heart; implications for muscular dystrophy

The possibility of using utrophin upregulation as a treatment for dystrophin-deficient muscular dystrophies has focused attention on the question of how many of dystrophin's various functions can be performed by the closely-related protein, utrophin. In Xenopus heart, little or no dystrophin was found on Western blots but the dystrophin-related protein, utrophin, was abundant. This utrophin was shown by immunofluorescence microscopy to be associated with cardiac muscle membranes and its distribution was similar to that of dystrophin in rabbit heart. The utrophin distribution pattern in the frog heart was shared by β-dystroglycan, a transmembrane protein responsible for localizing both dystrophin and utrophin at cell membranes. The results suggest that utrophin in Xenopus heart can perform similar functions to dystrophin in mammalian heart, lending further support to the possibility of utrophin upregulation therapy in muscular dystrophy. In skeletal muscle, however, Xenopus resembles mammals in expressing dystrophin at the sarcolemma and very little utrophin.     

Dystrophin-related protein in skeletal muscles in neuromuscular disorders: immunohistochemical study

Summary Thirty-four biopsied muscles of Duchenne, Becker and congenital muscular dystrophy, congenital myotonic dystrophy and amyotrophic lateral sclerosis were exmined by an immunocytochemical method with an anti-dystrophin-related protein (DRP) antibody. Strongly positive immunoreaction to DRP at the neuromuscular junctions in all biopsied specimens and faint reaction on the surface membrane of atrophic fibers in amyotrophic lateral sclerosis suggest that DRP is an anchor protein of the acetylcholine receptor. Additionally, the surface membrane of muscle fibers of Duchenne muscular dystrophy was positively stained. DRP is, therefore, thought to be expressed to compensate for dystrophin deficiency in these muscle fibers.Supported in part by a grant-in-aid from National Center of Neurology and Psychiatry (NCNP) of the Ministry of Health and Welfare, Japan