Reciprocal amplification of ROS and Ca(2+) signals in stressed mdx dystrophic skeletal muscle fibers.
Muscular dystrophies are among the most severe inherited muscle diseases. The genetic defect is a mutation in the gene for dystrophin, a cytoskeletal protein which protects muscle cells from mechanical damage. Mechanical stress, applied as osmotic shock, elicits an abnormal surge of Ca(2+) spark-like events in skeletal muscle fibers from dystrophin deficient (mdx) mice. Previous studies suggested a link between changes in the intracellular redox environment and appearance of Ca(2+) sparks in normal mammalian skeletal muscle. Here, we tested whether the exaggerated Ca(2+) responses in mdx fibers are related to oxidative stress. Localized intracellular and mitochondrial Ca(2+) transients, as well as ROS production, were assessed with confocal microscopy. The rate of basal cellular but not mitochondrial ROS generation was significantly higher in mdx cells. This difference was abolished by pre-incubation of mdx fibers with an inhibitor of NAD(P)H oxidase. In addition, immunoblotting showed a significantly stronger expression of NAD(P)H oxidase in mdx muscle, suggesting a major contribution of this enzyme to oxidative stress in mdx fibers. Osmotic shock produced an abnormal and persistent Ca(2+) spark activity, which was suppressed by ROS-reducing agents and by inhibitors of NAD(P)H oxidase. These Ca(2+) signals resulted in mitochondrial Ca(2+) accumulation in mdx fibers and an additional boost in cellular and mitochondrial ROS production. Taken together, our results indicate that the excessive ROS production and the simultaneous activation of abnormal Ca(2+) signals amplify each other, finally culminating in a vicious cycle of damaging events, which may contribute to the abnormal stress sensitivity in dystrophic skeletal muscle.
Shkryl
Vyacheslav M
VM
Department of Pharmacology and Physiology, University of Medicine and Dentistry of New Jersey, New Jersey Medical School, 185 South Orange Avenue, Newark, NJ 07103, USA.
Martins
Adriano S
AS
Ullrich
Nina D
ND
Nowycky
Martha C
MC
Niggli
Ernst
E
Shirokova
Natalia
N
D000818:Animals; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016166:Free Radical Scavengers / Q000494:pharmacology; D005609:Free Radicals / Q000378:metabolism; D008297:Male; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D053078:Membrane Potential, Mitochondrial / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D018101:Mice, Inbred mdx; D008928:Mitochondria / Q000378:metabolism; D018485:Muscle Fibers, Skeletal / Q000378:metabolism*; D020388:Muscular Dystrophy, Duchenne / Q000378:metabolism* / Q000503:physiopathology; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009997:Osmotic Pressure / Q000502:physiology; D018384:Oxidative Stress / Q000502:physiology*; D026361:Reactive Nitrogen Species / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*
D004791:Enzyme Inhibitors; D016166:Free Radical Scavengers; D005609:Free Radicals; D008562:Membrane Glycoproteins; D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; C488842:Cybb protein, mouse; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases