Biomedical subjects

M A McElligott

Publications and source records attributed to M A McElligott.

23 records · Page 2Linked to original sources

PubMed1984

Intracellular protein degradation in cultures of dystrophic muscle cells and fibroblasts.

We have analysed protein degradation in primary cultures of normal and dystrophic chick muscle, in fibroblasts derived from normal and dystrophic chicks, and in human skin fibroblasts from normal donors and from patients with Duchenne muscular dystrophy (DMD). Our results indicate that degradative rates of both short- and long-lived proteins are unaltered in dystrophic muscle cells and in dystrophic fibroblasts. Longer times in culture and co-culturing chick fibroblasts with the chick myotubes do not expose any dystrophy-related abnormalities in protein catabolism. Furthermore, normal and dystrophic muscle cells and fibroblasts are equally able to regulate proteolysis in response to serum and insulin. We conclude that cultures of chick myotubes, chick fibroblasts, and fibroblasts derived from humans afflicted with DMD are not appropriate models for studying the enhanced protein degradation observed in dystrophy.

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PubMed1983

Erythrocyte-mediated microinjection, a technique to study protein degradation in muscle cells.

The technique of erythrocyte-mediated microinjection has been successfully adapted for use with cultured muscle cells. Erythrocytes were fused with primary chick myotube cultures with poly(ethylene glycol), and fluorescent antibodies to haemoglobin demonstrated that this protein was injected into the sarcoplasm of myotubes. The microinjection treatment did not significantly alter protein metabolism in the muscle cells as monitored by rates of synthesis and degradation of muscle proteins. 125I-labelled ribonuclease A and bovine serum albumin were degraded with the expected exponential decay kinetics after microinjection into muscle cells, and the half-life of ribonuclease A (40 h) was approximately twice that of bovine serum albumin (17 h). The degradation of ribonuclease A in the muscle cells was enhanced 1.6-fold in the absence of horse serum and chick-embryo extract, whereas the degradation of bovine serum albumin was not altered during deprivation. These results are characteristic of the breakdown of microinjected ribonuclease A and bovine serum albumin in other cell types. Therefore, our experiments indicate the erythrocyte-mediated microinjection is a valid technique to study protein degradation in primary chick muscle cultures.

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PubMed1981

Muscle proteolytic enzyme activities in diabetic rats.

Proteolytic enzyme activities were measured in skeletal muscle of Sprague-Dawley rats with streptozotocin-induced diabetes [tail vein injection of streptozotocin (100 mg/kg), under ether anesthesia]. Assay of rat muscle homogenates from diabetic rats revealed a significant increase in alkaline serine protease activity as compared to untreated control rats and diabetic rats given insulin. There were no significant changes in lysosomal cathepsin activities in diabetic muscle as compared to controls. Gel studies of myofibrils isolated from the three groups of rats, subjected to autolysis, revealed that the serine protease had copurified with the myofibrils. Treatment of rats with compound 48/80, which degranulates mast cells, abolished the alkaline protease activity. There was no serine protease activity associated with the myofibrils isolated from compound 48/80-treated rats. Results from this study indicate that serine proteases are not involved in muscle protein breakdown in diabetes and are of mast cell origin.

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PubMed1981

Lysosomes and proteolytic enzyme activities in cultured striated muscle cells.

Primary cell cultures prepared from chick embryonic skeletal muscle and the rat myogenic line L6 were examined morphologically and biochemically during several stages of development. The L6 cells were cultured to provide three morphologically distinct populations: prefusion, postfusion, and a subclone of cells that did not fuse even at high density. Ultrastructural studies revealed the characteristic morphology of healthy myoblasts. Acridine orange staining and cytochemical localization of acid phosphatase suggest the presence of presumptive lysosomal material. Enzymatic studies of lysosomal cathepsins B, D, H, and L revealed unusually high enzyme specific activities in these homogeneous myoblast populations. No activity was detected for the two nonlysosomal enzymes Ca2+-proteinase and serine proteinase. It is suggested that the lysosomal apparatus and its complement of enzymes play a significant role in the differentiation of muscle myotubes.

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PubMed1981

Properties of the lysosomal apparatus during differentiation of cultured striated muscle cells.

A technique for the preparation of relatively pure myoblasts from chick primary culture is described. Cultured rat myogenic cells (L6) were plated and grown to provide three morphologically distinct cell populations: perfusion, postfusion, and nonfusion. Homogenates of L6 cells demonstrated two major peaks of proteolytic activity at pH 3.0 and 5.5. The activity could be partially inhibited by leupeptin or pepstatin. Differential centrifugation indicated significant acid hydrolase activity in the "H" fraction of prefused cells, which shifted to the "L" fraction after fusion of the cells. Two populations of lysosomes were resolved in the myoblasts and myotubes after isopycnic centrifugation in Percoll. The equilibrium densities were 1.044 and 1.060-1.068. Cells were incubated with several protease inhibitors. Only chloroquine caused a large inhibition of protein degradation.

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