Saiyid A. Shafiq

Delayed degeneration of dystrophic and normal muscle cell cultures treated with pepstatin, leupeptin, and antipain.

Abstract The effect of the proteinase inhibitors, pepstatin, leupeptin, and antipain, on dystrophic and normal embryonic chicken muscle cells growing in tissue culture was determined. The three inhibitors are effective against lysosomal cathepsins as well as other proteinases. The inhibitors appeared to delay atrophy and degeneration of dystrophic muscle fibers markedly; the effect on the normal muscle fibers was less striking. Catheptic activity and acidic autolysis are known to increase in the dystrophic chicken. These results support the suggestion that lysosomal proteases are involved, by an unknown mechanism, in the degradative process in dystrophic tissue. Delay in the process of degradation of muscle tissue suggests that these low molecular weight, nontoxic inhibitors offer some prospects as therapeutic agents for treatment of muscular dystrophy and other degenerative muscle diseases.

Characterization of a monoclonal antibody to myosin specific for mammalian and human type II muscle fibers.

We have characterized a monoclonal antibody (McAb), ALD-19, generated against slow myosin from chicken anterior latissimus dorsi (ALD) muscle for use in studies of human and animal muscle fiber types. This McAb bound selectively to the 200 kDa myosin heavy chain band in immunoblots against chicken, rat and human myosins and showed selective staining of A bands in the myofibrils. The reactivity of ALD-19 with various myosin types was quantitated by radioimmunoassays. Fiber type analysis revealed unexpected specificity of McAb ALD-19 for type II mammalian muscle fibers. This antibody should, therefore be useful for identification and quantification of normal type II fibers in human muscle biopsy specimens.

In vivo effect of protease inhibitors in denervation atrophy

The protease inhibitors leupeptin and pepstatin were used in vivo to examine their effect on denervation-induced atrophy in chicken pectoralis muscle. Comparison of muscle weights, fiber diameters, and light meromyosin paracrystals in treated versus untreated animals indicated that these inhibitors had a significant effect in delaying denervation atrophy. The studies suggest that protease inhibitors may have potential therapeutic capability in a variety of neuromuscular degenerative disorders.

Reduced density of intramembrane particles in erythrocytes of dystrophic chickens

The intramembrane particles in erythrocytes of chickens with hereditary muscular dystrophy and normal controls were compared by the freeze-fracture technique. Reduction of particles in both fracture faces and greater clustering of them was found in dystrophic samples. The concept of a generalized membrane abnormality in dystrophic chickens is supported.

An immunocytochemical study of type I muscle fibres in developing human skeletal muscles

An immunocytochemical study was done on the skeletal muscles of human fetuses (19-36 weeks gestation), infants and adults using a new monoclonal antibody (McAb) ALD-47. The antibody was generated against slow myosin of chicken and is specific for myosin heavy chain (MHC). In human infants and adults the type I muscle fibres are strongly reactive with this McAb and the type II fibres uniformly non-reactive. In the fetuses from 19-20 weeks gestation (in whom the fibre types are not distinguishable by the histochemical myosin ATPase test) a proportion of muscle fibres react specifically with ALD-47. Other muscle fibres at this stage react positively with a fast specific MHC McAb HM-1.2 or are negative to both ALD-47 and HM-1.2 antibodies. These McAbs, thus, identify three distinct fibre populations in the early fetal muscle which by histochemical staining appears homogeneous. The percentage of ALD-47 positive fibres increases in fetuses at later gestational periods; at all stages these fibres lack reactivity with the HM-1.2 antibody. Because of its selective fibre type reactivity in differentiating muscles, the McAb ALD-47 in conjunction with HM-1.2 should be useful in immunoaffinity fractionation and biochemical studies of myosin isoforms in developing human muscles.

A freeze-fracture study of fiber types in normal human muscle ☆

The sarcoplasmic reticulum (SR) and plasma membranes of Type 1 and Type 2 fibers of normal human muscle were examined by the freeze-fracture technique. Total particle counts in the SR appeared much lower than in other mammals and a packing density of about 1200 particles/micrometer 2 was found in both longitudinal and cisternal components of SR. There was no difference in particle density of Type 1 and Type 2 fibers. In freeze-fracture replicas of plasma membranes several fiber type differences were seen. The surface caveolae were uniformly distributed in Type 1 fibers whereas in Type 2 they were clustered preferentially at the I-band levels. Total density of intramembranous particles was greater in Type 1 fibers (347 +/- 68/micrometer 2 in P-face, 58 +/- 11/micrometer 2 in E face) than in Type 2 fibers (207 +/- 30/micrometer 2 in P-face, 80 +/- 9/micrometer 2 in E-face). There was a striking difference in respect to rectilinear arrays which were virtually absent in Type 1 fibers (0--2/micrometer 2) and numberous (up to 50--70/micrometer 2) in Type 2 fibers.

Avian adductor profundus muscle: characterization of a pure slow tonic region by histochemical, monoclonal antibody and peptide mapping studies

SummaryThe fibre type composition of the avian adductor profundus (AP) muscle which is composed of a thick white posterior part (Post. AP) and a thin red anterior part (Ant. AP) was investigated. Using the histochemical ATPase technique, monoclonal antibody analysis of myosin and C-protein isoforms, and electrophoretic and peptide mapping analyses of myosin, we have established that the Post. AP is composed of essentially pure slow tonic fibres similar to those of the anterior latissimus dorsi muscle (ALD). The Ant. AP, on the other hand, is shown to contain a mixture of slow and fast fibres, the latter giving immunocytochemical reactions atypical of the fast fibres. The larger size of the Post. AP in comparison with the ALD muscle should provide significantly more tissue for biochemical studies of tonic fibres than was previously available.

I-cell disease (mucolipidosis II): Differential expression in satellite cells and mature muscle fibers

A well documented case of I-cell disease is presented. Light- and electron-microscopic studies of muscle revealed marked accumulation of characteristic I-cell inclusions in satellite cells and only scattered autophagic vacuoles in muscle fibers. Correlation with previous tissue culture studies indicated an amelioration of structural abnormalities with differentiation from satellite cell to mature muscle fiber. Histochemically, the muscle demonstrated paucity of type I fibers without evidence of denervation thus suggesting a developmental disturbance in motor unit organization. Selective type I fiber dysfunction and reduced satellite cell regenerative capacity may be related factors in the neuromuscular disability of patients with I-cell disease.

Fine Structure of Aging Skeletal Muscle

Diminution of motor performance and decrease in muscle weight relative to body weight are prominent features in old age, but the mechanisms involved in these changes are not known. The histological alterations in skeletal muscles that might account for these deficits have been examined in both human and animal studies, and several reviews have been published recently (e.g., Gutmann, 1977; McComas, 1977; Kaldor and DiBattista, 1978). In general, it has been found that there is a reduction in both the number and the diameter of muscle fibers (Tucek and Gutmann, 1973; Adams and Victor, 1977) as well as an increased variation of fiber sizes, with some fibers becoming hypertrophic perhaps to compensate for the loss and atrophy of others (Rowe, 1969; Jennekens et al., 1971).

Freeze-fracture analysis of the effects of age and 2,4-dinitrophenol on the morphology of flight muscle mitochondria of Musca Domestica L. ☆

Morphometric comparison of freeze-fractured mitochondria in flight muscles of adult (37-day-old) and old (68-day-old) houseflies revealed a 28% decrease of cristae in the old flies. The major membrane change with age was an increase in the 90-120-A particles in the inner membrane external face concomitant with a loss of particle clusters associated with the openings of the cristae on to the inner membrane. In vitro treatment of flight muscle with 2,4-dinitrophenol, and uncoupler of mitochondrial respiration, did not produce this change but resulted in the formation of smooth particle-free vesicular swellings in the mitochondria. Such swelling were infrequent in the old muscle. The cause for the aging change is not clear, but a reduction in the ability of the intramembranous particles to aggregate, either through modification of altered synthesis, is indicated.