Elsevier

Free Radical Biology and Medicine

Volume 43, Issue 12, 15 December 2007, Pages 1584-1593
Free Radical Biology and Medicine

Original Contribution
Oxidation and structural perturbation of redox-sensitive enzymes in injured skeletal muscle

https://doi.org/10.1016/j.freeradbiomed.2007.08.019Get rights and content

Abstract

Molecular events that control skeletal muscle injury and regeneration are poorly understood. However, inflammation associated with oxidative stress is considered a key player in modulating this process. To understand the consequences of oxidative stress associated with muscle injury, inflammation, and regeneration, hind-limb muscles of C57Bl/6J mice were studied after injection of cardiotoxin (CT). Within 1 day post-CT injection, polymorphonuclear neutrophilic leukocyte accumulation was extensive. Compared to baseline, tissue myeloperoxidase (MPO) activity was elevated eight- and fivefold at 1 and 7 days post-CT, respectively. Ubiquitinylated protein was elevated 1 day postinjury and returned to baseline by 21 days. Cysteine residues of creatine kinase (CK) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were irreversibly oxidized within 1 day post-CT injection and were associated with protein conformational changes that fully recovered after 21 days. Importantly, protein structural alterations occurred in conjunction with significant decreases in CK activity at 1, 3, and 7 days post-CT injury. Interestingly, elevations in tissue MPO activity paralleled the time course of conformational changes in CK and GAPDH. In combination, these results demonstrate that muscle proteins in vivo are structurally and functionally altered via the generation of reactive oxygen species produced during inflammatory events after muscle injury and preceding muscle regeneration.

Section snippets

Mouse CT model

C57Bl/6J male mice were obtained from The Jackson Laboratory (Bar Harbor, ME, USA) and used when 13-21 weeks of age. All procedures complied with the National Institutes of Health animal use and care guidelines and were approved by the Institutional Animal Care and Use Committees at The University of Texas Health Science Center at San Antonio and at the South Texas Veterans Health Care System. Hair was removed with a depilatory cream from bilateral hind limbs 1-2 days before induction of

Time course of histological alterations in CT-injured skeletal muscle

The architecture of normal, mature skeletal muscle consists of multinucleated muscle fibers with peripherally located nuclei (Fig. 1A); inflammatory cells are rarely present. After NS injection into the left AC (negative control), small, focal areas of muscle injury and regeneration were occasionally observed; otherwise, most of these specimens and all baseline samples were entirely normal (data not shown). In contrast, within 1 day after CT injection, toxic injury of skeletal muscle was

Discussion

Skeletal muscle injury and regeneration are a complex process that involves the coordination of diverse cell types, including inflammatory cells [25]. However, the relationships of inflammatory-induced oxidative stress and muscle regeneration are poorly understood. The current study examined the effects of inflammation and oxidative stress on CK and GAPDH cysteine-oxidation state, conformational alterations, and activity in baseline and post-CT-injury muscles. Interestingly, elevations in MPO

Acknowledgments

The authors acknowledge the expert technical assistance of Richard Castillo. This work was supported by grants from the National Institutes of Health (HL070158, HL074236, AG013319) to P.K.S. and the Veterans Administration to both P.K.S. and A.C.

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