scholarly journals Aging increases the oxidation of dichlorohydrofluorescein in single isolated skeletal muscle fibers at rest, but not during contractions

2013 ◽  
Vol 305 (4) ◽  
pp. R351-R358 ◽  
Author(s):  
Jesus Palomero ◽  
Aphrodite Vasilaki ◽  
Deborah Pye ◽  
Anne McArdle ◽  
Malcolm J. Jackson
Keyword(s):  
Skeletal Muscle ◽  
Reactive Nitrogen Species ◽  
Contractile Activity ◽  
Muscle Fibers ◽  
Reactive Nitrogen ◽  
Nitrogen Species ◽  
Regulatory Processes ◽  
Skeletal Muscle Fibers ◽  
Old Mice ◽  
Young Mice

An increase in the activity of reactive oxygen species (ROS) has been implicated in the mechanisms of loss of skeletal muscle that occurs during aging, but few studies have attempted to directly assess activities in intact muscle fibers. The current project used the nonspecific fluorescent probe for ROS and reactive nitrogen species, 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein (CM-DCFH), in single, isolated, mature skeletal muscle fibers from adult and old mice in addition to biochemical measurements of key regulatory proteins for ROS in muscles of these animals. Data confirmed the changes in key regulatory processes for ROS (increased glutathione peroxidase 1 and catalase activities and reduced total glutathione content) previously reported in muscle from old mice and showed increased CM-DCFH oxidation in muscle fibers from old mice at rest and indicate that these changes are likely due to an increase in generation of oxidants rather than a lack of scavenging capacity. The increased CM-DCFH oxidation persisted even when cellular defenses against oxidants were increased by loading fibers from young and old mice with glutathione. During contractile activity, and in contrast to the increase observed in fibers from young mice, there was no further increase in CM-DCFH oxidation in muscle fibers from old mice. These data also suggest that the defect in short-term adaptations to contractions that occurs in old mice may be related to a diminished, or absent, increase in the muscle generation of ROS and/or reactive nitrogen species that normally accompanies contractile activity in young mice.

scholarly journals Exercise-induced alterations and loss of sarcomeric M-line organization in the diaphragm muscle of obscurin knockout mice

2017 ◽  
Vol 312 (1) ◽  
pp. C16-C28 ◽  
Author(s):  
D. Randazzo ◽  
B. Blaauw ◽  
C. Paolini ◽  
E. Pierantozzi ◽  
S. Spinozzi ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Muscle Fibers ◽  
Detailed Examination ◽  
Diaphragm Muscle ◽  
Hindlimb Muscles ◽  
Skeletal Muscle Fibers ◽  
Exercise Induced ◽  
Old Mice ◽  
Ankyrin B ◽  
Deficient Mice

We recently reported that skeletal muscle fibers of obscurin knockout (KO) mice present altered distribution of ankyrin B (ankB), disorganization of the subsarcolemmal microtubules, and reduced localization of dystrophin at costameres. In addition, these mice have impaired running endurance and increased exercise-induced sarcolemmal damage compared with wild-type animals. Here, we report results from a combined approach of physiological, morphological, and structural studies in which we further characterize the skeletal muscles of obscurin KO mice. A detailed examination of exercise performance, using different running protocols, revealed that the reduced endurance of obscurin KO animals on the treadmill depends on exercise intensity and age. Indeed, a mild running protocol did not evidence significant differences between control and obscurin KO mice, whereas comparison of running abilities of 2-, 6-, and 11-mo-old mice exercised at exhaustion revealed a progressive age-dependent reduction of the exercise tolerance in KO mice. Histological analysis indicated that heavy exercise induced leukocyte infiltration, fibrotic connective tissue deposition, and hypercontractures in the diaphragm of KO mice. On the same line, electron microscopy revealed that, in the diaphragm of exercised obscurin KO mice, but not in the hindlimb muscles, both M-line and H-zone of sarcomeres appeared wavy and less defined. Altogether, these results suggest that obscurin is required for the maintenance of morphological and ultrastructural integrity of skeletal muscle fibers against damage induced by intense mechanical stress and point to the diaphragm as the skeletal muscle most severely affected in obscurin-deficient mice.

scholarly journals Rapid dispersal of clustered postsynaptic nuclei following dissociation of skeletal muscle fibers.

1996 ◽  
Vol 199 (11) ◽  
pp. 2359-2367
Author(s):  
C Brösamle ◽  
D P Kuffler
Keyword(s):  
Skeletal Muscle ◽  
Synapse Formation ◽  
Contractile Activity ◽  
Muscle Fibers ◽  
Skeletal Muscle Fibers ◽  
Synaptic Region ◽  
Nuclear Clusters ◽  
Specialized Structure

The vertebrate neuromuscular junction is a highly specialized structure containing many unique proteins and an underlying cluster of nuclei. Part of this specialization results from the expression of the genes for these proteins in nuclei clustered in the postsynaptic region. Contractile activity, as well as molecules located in the synaptic extracellular matrix (ECM), have been implicated in the induction of gene expression in these clustered nuclei. The present experiments were aimed at examining whether the presence of the synaptic ECM and presynaptic cells play a role in maintaining the clustering of the nuclei. We describe the normal distribution of nuclei clustered in the synaptic region of intact adult frog, Rana pipiens, skeletal muscle fibers and show that innervation is not required to maintain the nuclear clusters. Even after long-term (4 week) denervation, the clusters remain unchanged. Dissociation of the muscle fibers with proteases that remove ECM, Schwann cells and other satellite cells from the synaptic sites is followed by a rapid (within approximately 1.5 h) and almost complete dispersal of the clustered nuclei. Attempts to recluster the postsynaptic nuclei by the application of ECM components to muscle fibers in vitro were not successful. We propose that a factor or factors, localized in the synaptic ECM as a result of synapse formation and acting via the transmembrane or cytoplasmic domains of their respective receptors, induces the formation of a specialized cytoskeleton in the postsynaptic region that is capable of pulling in or 'trapping' nuclei. The removal of these factors from the ECM by proteases brings about the disorganization of the cytoskeleton and the freeing of the 'trapped' nuclei.

Induction of GLUT-1 protein in adult human skeletal muscle fibers

2000 ◽  
Vol 279 (5) ◽  
pp. E1191-E1195 ◽  
Author(s):  
M. Gaster ◽  
J. Franch ◽  
P. Staehr ◽  
H. Beck-Nielsen ◽  
T. Smith ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Human Skeletal Muscle ◽  
Contractile Activity ◽  
Muscle Fibers ◽  
Insulin Dependent Diabetes Mellitus ◽  
Human Muscle ◽  
Dependent Diabetes Mellitus ◽  
Glut 1 ◽  
Adult Human ◽  
Skeletal Muscle Fibers

Prompted by our recent observations that GLUT-1 is expressed in fetal muscles, but not in adult muscle fibers, we decided to investigate whether GLUT-1 expression could be reactivated. We studied different stimuli concerning their ability to induce GLUT-1 expression in mature human skeletal muscle fibers. Metabolic stress (obesity, non-insulin-dependent diabetes mellitus), contractile activity (training), and conditions of de- and reinnervation (amyotrophic lateral sclerosis) could not induce GLUT-1 expression in human muscle fibers. However, regenerating muscle fibers in polymyositis expressed GLUT-1. In contrast to GLUT-1, GLUT-4 was expressed in all investigated muscle fibers. Although the significance of GLUT-1 in adult human muscle fibers appears limited, GLUT-1 may be of importance for the glucose supplies in immature and regenerating muscle.

Hypoxia-induced skeletal muscle fiber dysfunction: role for reactive nitrogen species

2006 ◽  
Vol 290 (1) ◽  
pp. L127-L135 ◽  
Author(s):  
Coen A. C. Ottenheijm ◽  
Leo M. A. Heunks ◽  
Maartje C. P. Geraedts ◽  
P. N. Richard Dekhuijzen
Keyword(s):  
Skeletal Muscle ◽  
Reactive Nitrogen Species ◽  
Single Fiber ◽  
Contractile Protein ◽  
Reactive Nitrogen ◽  
Nitrogen Species ◽  
Muscle Bundle ◽  
Skinned Muscle ◽  
Skinned Muscle Fibers ◽  
Protein Dysfunction

Hypoxia impairs skeletal muscle function, but the precise mechanisms are incompletely understood. In hypoxic rat diaphragm muscle, generation of peroxynitrite is elevated. Peroxynitrite and other reactive nitrogen species have been shown to impair contractility of skinned muscle fibers, reflecting contractile protein dysfunction. We hypothesized that hypoxia induces contractile protein dysfunction and that reactive nitrogen species are involved. In addition, we hypothesized that muscle reoxygenation reverses contractile protein dysfunction. In vitro contractility of rat soleus muscle bundles was studied after 30 min of hyperoxia (Po2 ∼90 kPa), hypoxia (Po2 ∼5 kPa), hypoxia + 30 μM NG-monomethyl-l-arginine (l-NMMA, a nitric oxide synthase inhibitor), hyperoxia + 30 μM l-NMMA, and hypoxia (30 min) + reoxygenation (15 min). One part of the muscle bundle was used for single fiber contractile measurements and the other part for nitrotyrosine detection. In skinned single fibers, maximal Ca2+-activated specific force (Fmax), fraction of strongly attached cross bridges (αfs), rate constant of force redevelopment ( ktr), and myofibrillar Ca2+ sensitivity were determined. Thirty minutes of hypoxia reduced muscle bundle contractility. In the hypoxic group, single fiber Fmax, αfs, and ktr were significantly reduced compared with hyperoxic, l-NMMA, and reoxygenation groups. Myofibrillar Ca2+ sensitivity was not different between groups. Nitrotyrosine levels were increased in hypoxia compared with all other groups. We concluded that acute hypoxia induces dysfunction of skinned muscle fibers, reflecting contractile protein dysfunction. In addition, our data indicate that reactive nitrogen species play a role in hypoxia-induced contractile protein dysfunction. Reoxygenation of the muscle bundle partially restores bundle contractility but completely reverses contractile protein dysfunction.

Markers of protein oxidation by hydroxyl radical and reactive nitrogen species in tissues of aging rats

1998 ◽  
Vol 274 (2) ◽  
pp. R453-R461 ◽  
Author(s):  
Christiaan Leeuwenburgh ◽  
Polly Hansen ◽  
Aviv Shaish ◽  
John O. Holloszy ◽  
Jay W. Heinecke
Keyword(s):  
Skeletal Muscle ◽  
Young Adult ◽  
Hydroxyl Radical ◽  
Reactive Nitrogen Species ◽  
Butylated Hydroxytoluene ◽  
Gas Chromatography Mass Spectrometry ◽  
Reactive Nitrogen ◽  
Nitrogen Species ◽  
Old Rats ◽  
Antioxidant Supplements

Many lines of evidence implicate oxidative damage in aging. Possible pathways include reactions that modify aromatic amino acid residues on proteins. o-Tyrosine is a stable marker for oxidation of protein-bound phenylalanine by hydroxyl radical, whereas 3-nitrotyrosine is a marker for oxidation of protein-bound tyrosine by reactive nitrogen species. To test the hypothesis that proteins damaged by hydroxyl radical and reactive nitrogen accumulate with aging, we used isotope dilution gas chromatography-mass spectrometry to measure levels of o-tyrosine and 3-nitrotyrosine in heart, skeletal muscle, and liver from young adult (9 mo) and old (24 mo) female Long-Evans/Wistar hybrid rats. We also measured these markers in young adult and old rats that received antioxidant supplements (α-tocopherol, β-carotene, butylated hydroxytoluene, and ascorbic acid) from the age of 5 mo. We found that aging did not significantly increase levels of protein-bound o-tyrosine or 3-nitrotyrosine in any of the tissues. Antioxidant supplementation had no effect on the levels of protein-bound o-tyrosine and 3-nitrotyrosine in either young or old animals. These observations indicate that the o-tyrosine and 3-nitrotyrosine do not increase significantly in heart, skeletal muscle, and liver in old rats, suggesting that proteins damaged by hydroxyl radical and reactive nitrogen species do not accumulate in these tissues with advancing age.

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