Erroneous diagnosis corrected after 28 years. Not spinal muscular atrophy with ophthalmoplegia but minicore myopathy.

OBJECTIVE: To correct, after 28 years, the previously reported diagnosis of ophthalmoplegia in a patient with presumed childhood spinal muscular atrophy. DESIGN: Clinical follow-up, laboratory, electrophysiologic, and muscle biopsy data are provided. RESULTS: The findings of clinical follow-up examination, electrophysiologic tests, and histologic examination of muscle specimens led to a revised diagnosis of minicore myopathy. CONCLUSIONS: Spinal muscular atrophy was diagnosed in 1967, before histochemical techniques for examining muscle tissue and quantitative electromyography became widely available. Modern laboratory techniques later made the diagnosis of minicore myopathy possible. Progressive external ophthalmoplegia has been described in 24% of patients with minicore myopathy, but there have been only 7 reports of ophthalmoplegia with spinal muscular atrophy since 1954, and some of these diagnoses have been questioned.

Scleromyxedema with myopathy and hyperthyroidism.

We describe a 62-year-old woman who developed extensive papular skin eruption with dysphagia and proximal muscle weakness. Laboratory studies showed a progressive increase of muscle enzymes, lambda monoclonal gammopathy, and elevated serum thyroid hormones. Several skin and muscle biopsies were necessary to reach the correct diagnosis of scleromyxedema in association with hyperthyroidism. Muscle biopsies contained rimmed vacuoles with some necrosis and regeneration, but no increased mucopolysaccharides. Hyperthyroidism was treated without appreciable improvement of the skin and muscle lesions. Myopathy is an increasingly recognized feature of scleromyxedema; its pathogenesis is still unexplained.

Chromosome 12-linked autosomal dominant scapuloperoneal muscular dystrophy.

Scapuloperoneal syndromes are characterized by their distribution of muscle weakness and wasting. The reported pattern of inheritance has been variable. Both neurogenic and myopathic forms of autosomally dominantly inherited scapuloperoneal syndrome have been described. It has been suggested that these are variants of other neuromuscular diseases. We examined 44 members from a family with 14 members affected with a scapuloperoneal syndrome. Physiological and histological analysis implied that this condition is predominantly myopathic. Linkage analysis was done to confirm the genetic etiology of the disease in this family and to evaluate the possibility that it is a allelic variant of other neuromuscular diseases. Genetic analysis demonstrated linkage of the disease to chromosome 12, which makes it genetically distinct from other loci known to cause neuromuscular disease. Muscle fibers with hyaline desmin-containing cytoplasmic inclusions in combination with focal myopathic changes may be a disease-specific morphological marker of the disease.

Muscle is electrically inexcitable in acute quadriplegic myopathy.

We directly stimulated muscle in three patients with acute quadriplegic myopathy to determine whether paralyzed muscle in this syndrome is electrically excitable. Two of the patients had been treated with neuromuscular blocking agents and corticosteroids, and one patient had been treated with corticosteroids alone. We found that paralyzed muscle is electrically inexcitable in affected patients. Muscle regained electrical excitability over weeks to months. The recovery of muscle excitability paralleled the clinical recovery of patients, suggesting that paralysis in this syndrome is secondary to electrical inexcitability of muscle membrane.

Expression of sodium channel subtypes during development in rat skeletal muscle.

This study contrasts the developmental patterns of expression of 2 subtypes of the voltage-dependent sodium channel in rat muscle that are differentiated by their immunoreactivity with monoclonal antibodies raised to the purified muscle sodium channel protein. One subtype is found in the transverse tubular (T) system of slow twitch fibers as well as the plasma membrane of fast and slow twitch fibers in the anterior tibial and soleus muscles. The second is present in the plasma membrane in all fibers of both muscles. The transverse tubular subtype exhibits 2 immunocytochemical staining patterns within muscle fibers, reticular and homogeneous, which may represent labeling of the developing T tubular system and of a cytoplasmic pool of alpha subunits of the sodium channel respectively. The reticular pattern eventually disappears in fast twitch fibers but persists into the adult stage in slow twitch fibers. The homogeneous pattern is also seen with antibodies to the plasma membrane subtype and disappears in early development as immunoreactivity to both subtypes gradually appears in the surface membrane. A reticular pattern is never seen with the plasma membrane subtype. The factors that modulate the expression of these subtypes is unknown.

Normal dystrophin in McLeod myopathy.

Dystrophin and its gene were studied in a patient with McLeod syndrome. This X-linked recessive myopathy has been localized to Xp21, as has the Duchenne muscular dystrophy gene locus, which codes for dystrophin. Histopathological study of the patient's muscle showed mild subclinical myopathy. Immunological studies of dystrophin in two separate biopsy specimens and analysis of dystrophin gene DNA from a blood sample did not detect an abnormality. This suggests that the Duchenne muscular dystrophy gene, albeit close to the McLeod locus, is not involved in McLeod myopathy.

Topological localization of a segment of the eel voltage-dependent sodium channel primary sequence (AA 927-938) that discriminates between models of tertiary structure.

Antibodies were raised against a synthetic peptide corresponding to residues 927-938 of the eel electroplax sodium channel primary sequence. This segment, lying between putative internal repeat domains II and III, is postulated to be exposed on the cytoplasmic surface of the membrane in several recent models of channel tertiary structure and on the external surface in another. The antiserum and affinity-purified IgG derived from it specifically recognize the peptide and the eel sodium channel in a solid-phase radioimmunoassay and bind to a single diffuse band of 260-280 kDa on Western blots of eel electroplax membrane proteins. All reactions are blocked by co-incubation of the antibodies with the synthetic peptide (1 microM). The antibody immunoprecipitates the solubilized channel in a form that retains its characteristic high-affinity binding of saxitoxin. In eel electroplax, the antibodies label only the innervated membrane known to contain sodium channels; at the ultrastructural level, this labeling is exclusively associated with the cytoplasmic surface of the membrane. Sodium channels containing the epitope are not seen in the postsynaptic membrane or in the membrane of the presynaptic nerve terminal. Segment 927-938 of the eel sodium channel is accessible on the surface of the protein in its solubilized form and is exposed in the cytoplasmic face of the innervated membrane of the electroplax in situ. This location is consistent with 3 models of channel structure but not with a fourth.

Freeze fracture analysis of muscle plasma membrane in bupivacaine HCl-induced degeneration and regeneration.

We did freeze fracture analysis of bupivacaine HCl-induced muscle fiber damage. The muscle plasma membrane in rat extensor digitorum longus and soleus muscle was analyzed for the density of orthogonal arrays and caveolae at periods of five minutes to 60 days after application of the myotoxin. In degenerating muscle a decrease in the density of caveolae was observed. In regenerating extensor digitorum longus muscle, a decrease in the density of orthogonal arrays and an increase in the density of caveolae was present. The results show that freeze fracture alterations in fast twitch regenerating muscle are similar to those seen in the muscles of patients with Duchenne muscular dystrophy.

Localization of sodium channel subtypes in adult rat skeletal muscle using channel-specific monoclonal antibodies.

Five monoclonal antibodies specific for the 260 kDa subunit of the rat skeletal muscle sodium channel were used to probe the distribution of this channel in adult muscle. All the antibodies reacted with the surface membrane of fast- and slow-twitch fibers in the rat anterior tibial and soleus muscles. Immunoreactivity was also present in the endplate region; this was significantly more intense than that in the surrounding extrajunctional membrane. At the electron microscopic level, this junctional immunoreactivity could be traced uniformly throughout the secondary folds of the post-synaptic membrane. Three of the monoclonal antibodies (A/B2, F/E4, and I/E3) exhibited an additional distinct immunoreactivity pattern, staining the interior of selected fibers in the anterior tibial muscle that were subsequently identified as slow-twitch fibers. An identical reactivity pattern was observed with most of the soleus muscle fibers. In longitudinal sections of slow fibers examined at the light microscopic level, transversely oriented, regularly spaced doublets of fluorescence were localized at the junction of the A and I bands in each sarcomere. In permeabilized slow fibers exposed to A/B2 and examined at the electron microscopic level, internal reactivity was associated exclusively with the membranes of the T-tubular system. A/B2 also strongly stained a transversely oriented pattern within cardiac muscle fibers exhibiting the characteristics of the T-tubular system in that tissue. We conclude that at least 3 subpopulations of sodium channels are present in adult skeletal muscle: those in the sarcolemma of fast and slow fibers, those in slow-twitch fiber T-tubular membranes, and those in the T-tubular system of fast fibers. The channels in the slow fiber T-system apparently share common epitopes with those in the T-system of cardiac fibers.

Topographical localization of the C-terminal region of the voltage-dependent sodium channel from Electrophorus electricus using antibodies raised against a synthetic peptide.

A peptide corresponding to amino acid residues 1783-1794 near the C terminus of the electric eel sodium channel primary sequence of the eel (Electrophorus electricus) sodium channel has been synthesized and used to raise an antiserum in rabbits. This antiserum specifically recognized the peptide in a solid-phase radioimmunoassay. Specificity of the antiserum for the native channel protein was shown by its specific binding to a 280-kDa protein in immunoblots of eel electroplax membrane proteins. The antiserum also specifically labeled the innervated membrane of the eel electroplax in immunofluorescent studies; noninnervated membrane was not labeled, consistent with the known distribution of sodium channels in this tissue. The membrane topology of the peptide recognized by this antiserum was probed in binding studies using oriented electroplax membrane vesicles. These vesicles were 98% "right-side-out" as determined by [3H]saxitoxin binding. Binding of the antipeptide antiserum to this fraction was measured before and after permeabilization with 0.01% saponin. Specific binding to intact vesicles was low, but this binding increased 10-fold after permeabilization, implying a cytoplasmic orientation for the peptide. Confirmation for this orientation was then sought by localizing the antibody bound to intact electroplax cells with immunogold electron microscopy. Gold particles identifying the antibody were found almost exclusively associated with the cytoplasmic surface of the innervated membrane. Our data imply that the region of the sodium channel primary sequence near the C terminus that is recognized by our antiserum is localized on the cytoplasmic side of the membrane; this localization provides some further constraints on models of sodium channel tertiary structure.

Antimuscle and antiacetylcholine receptor antibodies in myasthenia gravis.

The sera of 134 patients were examined for antimuscle antibodies by immunofluorescence (IF). These derived from 77 myasthenics, 30 myasthenics with thymoma, 6 patients with thymoma and no clinical evidence of myasthenia, and 21 patients with other autoimmune or neuromuscular diseases. Three separate patterns of antimuscle antibodies could be identified in the myasthenic sera by examination of the relaxed glycerinated myofibrils by both IF and phase-contrast optics: A-band (9 with thymoma, 1 without), I-band (11 with thymoma, 17 without), and a mixed A plus I pattern (5 with thymoma, 3 without). Seventy-seven myasthenic serum samples (24 with thymoma, 53 without) were available for evaluation of antibodies to acetylcholine receptor (anti-AChR) by radioimmunoassay. Ninety-one percent reacted with crude human receptor extract and 80% with receptor extracted from denervated rat muscle. There was no correlation between the titers of anti-AChR and the presence or staining patterns of antimuscle antibodies, but patients without anti-AChR did not have antimuscle antibodies. Myasthenics with thymoma had the highest prevalence of anti-AChR (23/24) and of antimuscle antibodies (25/30), and 15 of the 20 positives stained A-bands alone or with I-band, as compared to 4 of 21 positive reactions in those without tumor. Immunoabsorption, which removed or significantly reduced anti-AChR, did not alter antimuscle reactivity. The discrepancies between anti-AChR levels and the presence and types of antimuscle antibodies suggest that these are independent autoantibodies. Current theories of immunopathogenesis implicate altered thymic antigens or a major breakdown in immune regulation, either of which could explain their production.

Monoclonal antibodies against the voltage-sensitive Na+ channel from mammalian skeletal muscle.

A panel of 13 monoclonal antibodies against the voltage-sensitive Na+ channel of rat skeletal muscle has been characterized. Each of these antibodies reacted with the purified Na+ channel protein in a solid-phase radioimmunoassay. Nine antibodies specifically immunoprecipitated the Na+ channel in a form that retained its characteristic high affinity for saxitoxin, and 11 recognized the channel in a crude mixture of solubilized membrane proteins separated on a Sepharose CL-6B column. Six antibodies specifically labeled skeletal muscle in immunofluorescence techniques. In each case, antibody was localized only to the surface membrane of the muscle fibers. Eleven antibodies produced detectable reaction on immunoblot transfers of sarcolemmal membrane proteins; each of these bound to a diffuse 160- to 200-kDa band that comigrated with the large glycoprotein subunit of the purified Na+ channel. Further studies were carried out with one of these antibodies, L/D3. In immunoblots of a glycoprotein fraction prepared from muscle that had been homogenized rapidly in a solution containing detergent, EGTA, and protease inhibitors, L/D3 recognized only a single 260-kDa band. Incubation of solubilized muscle proteins at 4 degrees C for 24 hr without EGTA prior to isolation of the glycoprotein fraction resulted in partial conversion of this 260-kDa component to a smaller component between 160 and 200 kDa that comigrated with the principal immunoreactive component of sarcolemma. Based on its immunoreactivity with monoclonal antibodies, the large subunit of the rat skeletal muscle Na+ channel appears to be approximately equal to 260 kDa in its native state but may be sensitive to proteolysis during the isolation of sarcolemmal membranes.

Distribution of freeze-fracture particle sizes in Duchenne muscle plasma membrane.

We studied the distribution of freeze-fracture intramembranous particle (IMP) sizes in Duchenne muscle biopsies and controls. We found that the previously reported decrease in muscle plasma membrane particle density in Duchenne includes a variety of different IMPs, but does not preferentially involve IMPs of any one size.

Freeze-fracture study of muscle plasma membrane in obligate carriers of Duchenne muscular dystrophy.

Diminution of intramembranous particles and increase in the number of caveolae have been reported in freeze-fractured muscle plasma membrane of patients with Duchenne muscular dystrophy. We searched for similar alterations in a population of muscle plasma membranes from obligate carriers of Duchenne muscular dystrophy. There were no significant changes in muscle plasma membrane morphology in carriers and normal controls.

Muscle plasma membrane abnormalities in infants with Duchenne muscular dystrophy.

Muscle biopsies from one preclinical case, one early case, four symptomatic cases of Duchenne muscular dystrophy, and six controls were investigated for the presence of delta lesions by phase optics and conventional electronmicroscopy. By phase microscopy the frequency of delta lesions was 4.9% in the preclinical and early cases and 4.0% in the four symptomatic cases. The incidence of delta lesions in the controls was 0.4%. Electronmicroscopic studies of delta lesions revealed similar findings in the preclinical, early, and symptomatic cases. The common denominator of the delta lesions was disruption of the muscle plasma membrane. Plasma membrane lesions occur early in Duchenne muscular dystrophy.

Freeze-fracture analysis of plasma membrane cholesterol in Duchenne muscle.

We estimated cholesterol in Duchenne muscle plasma membrane using the sterol-specific ligand digitonin. In measuring the extent of digitonin-cholesterol complex formation with freeze-fracture electron microscopy, we found a significantly greater proportion of surface area taken up by complexes in Duchenne muscle (60% +/- 9%) than in controls (36% +/- 4%). This finding suggests that a population of Duchenne muscle fibers has a higher concentration of cholesterol in the plasma membrane. Membrane cholesterol is known to affect membrane deformability and the activity of membrane-bound enzymes and transport proteins; thus a difference in membrane cholesterol might account for structural and functional abnormalities in Duchenne plasma membrane.