Synthesis of apolipoprotein A1 in skeletal muscles of normal and dystrophic chickens

We recently observed that, around the time of hatching, chick skeletal muscles synthesize and secrete apolipoprotein A1 (apo-A1) at high rates and that reinitiation of synthesis of this serum protein to high levels occurs in mature chicken breast muscle following surgical denervation (Shackelford, J. E., and Lebherz, H. G. (1983) J. Biol. Chem. 258, 7175-7180; 14829-14833). In the present work we investigate the effect of avian muscular dystrophy on the synthesis of apo-A1 in chicken muscles. The relative rate of synthesis of apo-A1 and levels of apo-A1 RNA in mature dystrophic breast (fast-twitch) muscle were about 6-fold higher than normal, while synthesis of apo-A1 in breast muscles derived from 2-day-old dystrophic chicks was close to normal. These observations suggest that the elevated apo-A1 synthetic rate in mature dystrophic breast muscle results from a failure of the diseased tissue to "shut down" apo-A1 synthesis to the normal level during postembryonic maturation. Apo-A1 synthesis in the "slow-twitch" lateral adductor muscle of dystrophic chickens was found to be normal. Our work is discussed in terms of the apparent similarities between the effects of surgical denervation and muscular dystrophy on the protein synthetic programs expressed by chicken skeletal muscles.     

Acetylcholinesterase and butyrylcholinesterase released from normal and dystrophic muscles by treatment with proteolytic enzymes

1. Skeletal muscle from C57BL dystrophic mice demonstrated decreased activities of acetylcholinesterase with increased activities of butyrylcholinesterase. These changes were less distinct when compared to those observed with 129 ReJ mice. 2. Collagenase or trypsin treatment solubilized less acetylcholinesterase activity but more butyrylcholinesterase activity from muscle of C57BL dystrophic mice than from muscle of control mice. 3. These treatments resulted in similar pattern of release of acetylcholinesterase activity from muscle of 129 ReJ mice, except that more acetylcholinesterase activity was released from dystrophic muscle (129 ReJ) than from control by pepsin treatment. 4. The acetylcholinesterase activities released by proteolytic enzymes were characterized by sucrose density gradient centrifugation.     

Myosin isoforms in normal and dystrophic chickens

The myosin isoform content in the affected fibers of chickens with inherited muscular dystrophy has been investigated with a new high-performance liquid chromatographic procedure for separation of the tryptic fragments of myosin subfragment 1 (S-1). The results indicate that dystrophic muscle contains substantial amounts of normal adult myosin, together with various myosin species present in normal 5-day posthatch chickens. Confirmation was obtained by comparative peptide mapping of the S-1 tryptic fragments and by N-terminal sequencing of 20-kDa species. Together with data on other contractile proteins and certain metabolic enzymes [Obinata, T., Takano-Ohmura, H., & Matsuda, R. (1980) FEBS Lett. 120, 195-198; Mikasa, T., Takeda, S., Shimizu, T., & Kitaura, T. (1981) J. Biochem. (Tokyo) 89, 1951-1962; Feit, H., & Domke, R. (1982) Cell Motil. 2, 309-315; Cosmos, E. (1966) Dev. Biol. 13, 163-181; Cosmos, E., & Butler, J. (1967) in Exploratory Concepts in Muscular Dystrophy and Related Disorders (Milhorat, A. R., Ed.) pp 197-204, Excerpta Medica, Amsterdam], the results are consistent with the hypothesis that there is a general defect in muscle maturation in avian dystrophy.     

Protein synthesis in muscles from normal and dystrophic hamsters.

Homogenates of hindleg muscle were obtained from control and dystrophic male hamsters, 30 and 190 days of age, and were used to prepare the postmicrosomal pH5-supernatant fraction. The activity of this fraction in the incorporation of [14C]phenylalanyl-tRNA into peptides was increased in the dystrophic-muscle preparations. No such increase was found in brain or liver preparations from dystrophic hamsters. The increased capacity for aminoacyl-tRNA binding that was observed in preparations from dystrophic animals is discussed.     

Content and synthesis of glycolytic enzymes and creatine kinase in skeletal muscles and normal and dystrophic chickens

A number of workers have reported that avian muscular dystrophy causes alterations in the levels of certain enzyme activities in "fast-twitch" muscle fibers but has little effect on enzyme activities in "slow-twitch" muscle fibers. In the present work, the effects of this disease on the content and relative rates of synthesis of a number of glycolytic enzymes and the skeletal muscle-specific MM isoenzyme of creatine kinase in chicken muscles was investigated. It was shown that (i) the approximate 50% reductions in steady-state concentrations of three glycolytic enzymes (aldolase, enolase, and glyceraldehyde-3-P dehydrogenase) in dystrophic breast (fast-twitch) muscle result predominantly from decreases in relative rates of synthesis, rather than accelerations in relative rates of degradation, of these proteins in the diseased tissue; (ii) in contrast to the situation with the glycolytic enzymes, muscular dystrophy has only minor effects (25% or less) on the content and relative rate of synthesis of MM creatine kinase in breast muscle fibers; (iii) the muscular dystrophy-associated alterations in content and synthesis of the glycolytic enzymes in breast muscle fibers become apparent only during postembryonic maturation of this tissue; and (iv) as expected, muscular dystrophy has no significant effect on the content or relative rates of synthesis of glycolytic enzymes in slow-twitch lateral adductor muscles of the chicken. These results are discussed in terms of the apparent similarities between the effects of muscular dystrophy and surgical denervation on the protein synthetic programs expressed by mature fast-twitch muscle fibers.     

Collagenase-releasable and-resistant cholinesterases in normal and dystrophic muscles

The release of acetylcholinesterase activity by collagenase from the particulate fraction of mouse muscle homogenate into the soluble fraction was dependent on the time of incubation of muscle homogenate with collagenase. The collagenase-stimulated release of acetylcholinesterase was inhibited by 1,10-phenanthroline, an inhibitor of collagenase. Differential effects of inhibitors of specific acetylcholinesterase and nonspecific cholinesterase were observed in both collagenase extract and collagenase-resistant fraction derived from homogenate of muscle of normal and dystrophic mice. The collagenase extract of dystrophic muscle contained distinctly lower activity of acetylcholinesterase than that of normal muscle, while both collagenase extract and collagenase-resistant fraction of dystrophic muscle showed much higher activity of butyrylcholinesterase activity than those from normal muscle.     

Lipogenesis in muscle, liver, and adipose tissue of normal and dystrophic chickens.

The in vitro rate of oxidation and incorporation of [U-14C]glucose into lipids of muscle, liver, and adipose tissue of dystrophic and control chickens at 1 week, 1, 6, and 12 months of age were determined. The muscle and liver from dystrophic birds showed a high rate of [U-14C]glucose oxidation and incorporation into lipids during 1 week, 6 and 12 months. Insulin did not have any consistent stimulatory effect in all the tissues on the rate of glucose oxidation and incorporation into lipids. The contributory role played by the liver in the accumulation of lipids in dystrophic muscle is discussed.     

Superoxide dismutase activity in early stages of development in normal and dystrophic chickens

Changes in superoxide dismutase activities in early stages of chronological development were investigated in normal and dystrophic chickens. Both cupro-zinc and manganese superoxide dismutase activities were significantly elevated in the dystrophic chickens studied as early as one week after hatching compared to those in the control. In control chickens, both cupro-zinc and manganese superoxide dismutase activities declined as they grew older. In dystrophic chickens, manganese superoxide dismutase activity declined gradually as they grew older as in the control. However, cupro-zinc superoxide dismutase activity increased until four weeks of age. The latter activity was still twice as high as that of the control at four months of age. Increased activities in superoxide dismutases in early stages of the development suggest presence of increased turnover of active oxygen species from the early stage of the disease in this avian muscular dystrophy. And the distinct time course of cupro-zinc superoxide dismutase activity suggests involvement of active oxygen species in pathogenesis of this disorder.