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J Appl Physiol 105: 352-357, 2008. First published May 22, 2008; doi:10.1152/japplphysiol.90320.2008
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HIGHLIGHTED TOPIC
Biology of Physical Activity in Youth

Role of the calcium-calpain pathway in cytoskeletal damage after eccentric contractions

Bao-Ting Zhang,1 Simon S. Yeung,1 David G. Allen,2 Ling Qin,3 and Ella W. Yeung1

1Muscle Physiology Laboratory, Department of Rehabilitation Sciences, Hong Kong Polytechnic University, Hung Hom, Kowloon, Hong Kong; 2Muscle Cell Function Laboratory, School of Medical Sciences and Bosch Institute, University of Sydney, New South Wales, Australia; and 3Musculoskeletal Research Laboratory, Department of Orthopaedics and Traumatology, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong

Submitted 26 February 2008 ; accepted in final form 20 May 2008


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
The mechanism(s) underlying eccentric damage to skeletal muscle cytoskeleton remain unclear. We examined the role of Ca2+ influx and subsequent calpain activation in eccentric damage to cytoskeletal proteins. Eccentric muscle damage was induced by stretching isolated mouse muscles by 20% of the optimal length in a series of 10 tetani. Muscle force and immunostaining of the cytoskeletal proteins desmin, dystrophin, and titin were measured at 5, 15, 30, and 60 min after eccentric contractions and compared with the control group that was subjected to 10 isometric contractions. A Ca2+-free solution and leupeptin (100 µM), a calpain inhibitor, were applied to explore the role of Ca2+ and calpain, respectively, in eccentric muscle damage. After eccentric contractions, decreases in desmin and dystrophin immunostaining were apparent after 5 min that accelerated over the next 60 min. Increased titin immunostaining, thought to indicate damage to titin, was evident 10 min after stretch, and fibronectin entry, indicating membrane disruption, was evident 20 min after stretch. These markers of damage also increased in a time-dependent manner. Muscle force was reduced immediately after stretch and continued to fall, reaching 56 ± 2% after 60 min. Reducing extracellular calcium to zero or applying leupeptin minimized the changes in immunostaining of cytoskeletal proteins, reduced membrane disruption, and improved the tetanic force. These results suggest that the cytoskeletal damage and membrane disruption were mediated primarily by increased Ca2+ influx into muscle cells and subsequent activation of calpain.

cytoskeleton; muscle damage; calcium-activated protease; calpain

MUSCLE DAMAGE following eccentric contractions (EC) is characterized by an immediate, prolonged loss of muscle strength and plays an important role in exercise training (9) and sports injuries (29). Many factors contribute to the loss of muscle force, including abnormalities in excitation-contraction coupling (2, 18, 36), damage to force-generating structures such as actin and myosin (31), and damage to the force-transmitting system that is composed of cytoskeletal proteins (3, 7, 21, 22).

Muscle cytoskeletal proteins maintain the structural integrity of the myofiber, transmit force generated by actomyosin interactions, and are ubiquitous throughout the muscle cell. These proteins are susceptible to damage during eccentric exercise. For example, in extensor digitorum longus (EDL) muscle, immunolabeling of desmin is reduced within 5 min of eccentric exercise, and this loss is correlated with a decline in contractile force (21). Increased titin immunostaining, thought to indicate damage (see DISCUSSION), has also been observed ~30 min after EC, and at a similar time fibronectin, which is normally extracellular, appears inside the muscle cells, indicating defects in membrane integrity (14, 21). In rat muscle, the cytoskeletal protein dystrophin is disrupted even after a single EC experimental protocol (22). In dystrophin-deficient (mdx) mice, stretch-induced muscle fiber disruption and force decline are much more severe (10, 11, 37). Although cytoskeletal proteins have been implicated in stretch-induced muscle injury (3, 7, 14, 21), the underlying mechanisms remain unclear.

It has been proposed that calcium plays a role in EC-induced muscle damage (19, 21, 34, 37). Resting levels of intracellular calcium ([Ca2+]i) increase significantly after EC protocols in single muscle fibers (2) and whole muscle (23). Furthermore, [Ca2+]i is elevated in resting muscle fibers from mdx mice, and even more so after mechanical stretch (16, 37). Removing extracellular calcium prevents the rise in resting [Ca2+]i and ameliorates the decline in muscle force, suggesting that Ca2+ flux contributes to eccentric damage (37). Calcium increases protein turnover in muscle through activation of proteases, phospholipases, and lysosomal enzymes (8). However, it is more likely that calpain, a calcium-activated neutral protease, mediates the muscle damage that follows EC (6). Activities of µ- and m-calpain are increased in rat skeletal muscle following prolonged treadmill running (5). Autolytic activation of calpain-3 is increased in human muscle biopsies subjected to eccentric exercise (25). Application of a calpain inhibitor, leupeptin, largely prevents the excitation-contraction uncoupling and the titin disruption that follows elevated [Ca2+]i in toad muscle fibers (35). In mdx muscles, application of leupeptin decreases central nuclei and normalizes the cross-sectional area of muscle fiber (1). Thus it seems likely that increased Ca2+ influx and subsequent calpain activation are involved in eccentric-related muscle damage. How calpain relates to cytoskeletal damage, however, remains unknown.

In the present study, we confirm the time-dependent decline in force and disruption of the cytoskeleton in mouse EDL muscle following EC. These abnormalities were alleviated by removing extracellular calcium or by sustained application of leupeptin to isolated muscle fibers. This study supports the hypothesis that damage to muscle cytoskeleton following EC is primarily attributed to calcium-activated calpain proteolysis.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Animals.   Male BALB/c mice (age 10–14 wk, body wt 30–40 g) were used. Animal care procedures and the experimental protocol were approved by the Animal Ethics Committee of the Hong Kong Polytechnic University.

Muscle dissection and mounting.   Mice were killed by cervical dislocation. The skin overlying the lateral aspect of the hindlimb was then dissected from the thigh to the ankle. The proximal and distal tendons of the EDL muscle were carefully isolated and gripped with T-shaped aluminum foil microclips. The muscle was then transferred to an experimental chamber (capacity 1.5 ml). One end of the tendon was attached to a hook connected to the lever arm of a position feedback motor (300B-LC, Aurora Scientific), and the other end was attached to a force transducer (BG-10g, Kulite Semiconductor Products). The force transducer was clamped to a mechanical micromanipulator that allowed the muscle length to be adjusted.

Solutions.   During the experiments, EDL muscle was superfused with Krebs solution composed of (in mM) 121 NaCl, 5 KCl, 1.8 CaCl2, 0.5 MgCl2, 0.4 NaH2PO4, 24 NaHCO3, 5.5 glucose, and 0.1 EDTA. This solution was bubbled with 95% O2-5% CO2, maintaining a pH of 7.4. All experiments were performed at room temperature (~22°C). Krebs solution without added CaCl2 constituted the zero-extracellular Ca2+ solution.

Experimental protocol.   The muscle was stimulated with 0.5-ms pulses at 1.2x supramaximal intensity and adjusted to the length that produced maximal tetanic force (optimum length, Lo). Tetanic stimulation was delivered at 100 Hz for 400 ms with a rest period of 3.6 s between tetani (cycle, 4 s). Muscles in the control group were subjected to 10 isometric contractions. Muscles in the eccentric group were subjected to 10 EC, stretch was applied 250 ms after the start of stimulation, and the muscle was stretched from Lo to Lo + 20% over 150 ms (stretching velocity, 2 x Lo/s). The muscle was returned to Lo between tetani. In some experiments, within 5 s after the last EC, the extracellular solution was changed to zero Ca. The first test tetanus was measured at 60 s, and the muscles were then snap-frozen ~5 min after the last EC. Leupeptin (100 µM) (Sigma Aldrich) was applied 60 min before EC to block calpain activity.

The maximal isometric tension at 100 Hz was measured at 1, 5, 15, 30, and 60 min after isometric or eccentric stimulation (all at the original Lo). All force data are presented as a percentage of the pretetanus level, and n = 5 for every time point in every group.

Immunohistochemistry of muscle cytoskeletal proteins.   After the test tetanus, muscle cytoskeletal proteins were evaluated by immunohistochemistry at 5, 10, 20, 30, and 60 min post-tetanic stimulation. Each EDL muscle was snap-frozen in isopentane prechilled with liquid nitrogen, embedded in OCT embedding medium (Tissue-Tek), and stored at –80°C for further analysis.

Cryosections (10 µm thick) were cut from the midportion of the muscle, collected on gelatin-coated slides, and stored at –80°C. After fixation in cold acetone (4°C) for 10 min, endogenous peroxidase activity was blocked with 3% H2O2 in PBS. Sections were incubated for 20 min in 2% normal rabbit serum to block nonspecific binding. The sections were then incubated overnight at 4°C with primary antibodies against dystrophin, desmin, fibronectin, or titin (dilution 1:50 in 2% BSA/PBS). Following three washes with PBS, sections were incubated with biotinylated secondary antibody and then reacted with avidin-biotin complex reagent (Vector Laboratories) for 30 min. After washing again with PBS, sections were incubated with 0.96 mg/ml 3,3'-diaminobenzidine (Vector Laboratories) to visualize antibody staining. Negative control staining was performed by omitting primary antibody. All primary antibodies used in this study were affinity-purified goat polyclonal antibodies (Santa Cruz Biotechnology). Other immunochemistry-related chemicals were purchased from Sigma-Aldrich.

Bright-field images were captured using a Zeiss Axioplan 2 confocal microscope (Zeiss) equipped with a Spots RT digital camera (Zeiss). All sections were imaged with fixed optical parameters, light intensity, and magnification to ensure identical background levels. All cells were included in the analysis; (~1,100 fibers in the EDL muscle cross section), and the acquired images were evaluated using digital image morphometry program (ImageJ 1.32j, National Institutes of Health). Fibers were considered abnormal if the staining intensity was less than (desmin) or exceeded (titin and fibronectin) the predetermined threshold, which was determined by the intensity of negative control staining. In the case of dystrophin, abnormal fibers were identified by discontinuous or patchy dystrophin labeling. Changes were evaluated semiquantitatively by scoring the percentage of abnormal cells to the total number of fibers per muscle cross section. To determine observer reliability, at least 20 images from each experiment were counted by a skilled observer blinded to the treatment groups. The intraclass correlation coefficient (ICC) between the two raters was 0.95, suggesting excellent reliability.

Statistics.   The effect of EC on tension deficit at different time points in Ca2+-containing, zero Ca2+, or leupeptin-containing solution was analyzed by repeated-measures ANOVA. The percentage of cells showing abnormal protein immunostaining in Ca2+-containing, zero Ca2+, or leupeptin-containing solution was analyzed by one-way ANOVA with Bonferroni correction for multiple comparisons. P < 0.05 is considered significant. Values are expressed as means ± SE.


   RESULTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 GRANTS
 REFERENCES
 
Effect of stretched contractions on tetanic force in isolated EDL mouse muscle.   The changes in force following isometric and eccentric contractions were determined (Fig. 1). Tetanic force was transiently depressed immediately after 10 isometric contractions, due to muscle fatigue, but recovered to the control level within 5 min. Following 10 eccentric contractions, the force at 1 min declined to 76 ± 4% (n = 5, P < 0.01), and continued to decline, reaching 56 ± 2% at 60 min (n = 5, P < 0.001).


Figure 1
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  		Fig. 1. Effect of removing extracellular Ca2+ or applying leupeptin, a calpain blocker, on mouse extensor digitorum longus (EDL) muscle tension following eccentric contractions (EC). Muscle force was normalized to the prestretch level (baseline, {blacksquare}). After stretched contractions (bullet), muscle force was significantly lower than after isometric contractions (Isom, {blacktriangleup}). Application of leupeptin ({blacktriangledown}) or solution lacking Ca2+ (zero Ca2+, {circ}) partially recovered relative muscle force after stretched contractions. *P < 0.05, significant difference between untreated and treated (Ca2+-lacking solution, or leupeptin treated) groups after stretch. Values are expressed as means ± SE.

 
Effects of a calpain inhibitor and zero Ca2+ on tetanic force.   To test the hypothesis that cellular damage induced by EC is mediated by Ca2+ influx and subsequent calpain activation, EC were performed in the presence of 100 µM leupeptin, a calpain inhibitor, or zero-Ca2+ solution applied after the last EC. Application of leupeptin 1 h before and after stretch significantly attenuated the decline in muscle force. Force at 30 min and at 60 min (69 ± 4%) poststretch were larger than in the absence of leupeptin (n = 5, P < 0.05). Similarly, tetanic force was also greater in the absence of extracellular Ca2+ at 15, 30, and 60 min (76 ± 3%) (n = 5, P < 0.05) (Fig. 1).

Effects of a calpain inhibitor and zero Ca2+ on the levels of cytoskeletal proteins.   Representative examples of the effects of leupeptin and zero Ca2+ on EC-induced changes in the staining of cytoskeletal proteins dystrophin, desmin, and titin, and the extracellular matrix protein fibronectin are shown in Fig. 2A. Figure 2B shows the time course of average changes after stretched contractions: either the loss of dystrophin and desmin staining or the increase of fibronectin and titin staining. Desmin staining is completely absent in some cells and patchily absent in others and quite normal in others. Dystrophin staining is located in the surface membrane and becomes disrupted in an irregular fashion. Titin and fibronectin staining is not detectable within the muscle cell under control conditions and becomes detectable throughout the muscle cell or in a patchy way after stretch. Note that increased loss of desmin staining and dystrophin disruption were evident at 5 min, whereas the increase in staining for titin and fibronectin was not observed until 10 min and 20 min, respectively. For example, Fig. 3 shows the dystrophin, desmin, and fibronectin immunostaining at 5 min post-eccentric contractions and illustrates abnormal dystrophin staining, absence of desmin staining, but no increase in fibronectin staining. Reducing extracellular Ca2+ to zero prevented the loss of desmin staining (2.1 ± 0.7%, P < 0.05) as did application of leupeptin (1.7 ± 0.6%, P < 0.01). EC-induced changes in dystrophin, titin, and fibronectin staining were not significantly affected by zero extracellular Ca2+ or leupeptin until 10–20 min.


Figure 2
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  		Fig. 2. Effects of leupeptin and lack of extracellular calcium on eccentric contraction-induced changes in the staining pattern of mouse EDL muscle cytoskeletal proteins. A: cross sections of EDL muscle immunostained for different cytoskeletal proteins 30 min post-isometric contractions or eccentric contractions (EC) in standard Krebs solution, solution lacking Ca2+ (zero Ca2+) or leupeptin-containing solution. *Same fiber in serial sections. Arrows indicate dystrophin rupture. Scale bar, 50 µm. B: stretch-induced changes in muscle cytoskeletal proteins expressed as the percentage of muscle fibers affected (affected fibers/total no. of fibers per section). *P < 0.05, significant difference between untreated and treated (solution lacking Ca2+, or leupeptin treated) groups after stretch. Values are expressed as means ± SE.

 

Figure 3
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  		Fig. 3. Changes in the staining pattern of mouse EDL muscle cytoskeletal proteins dystrophin and desmin and extracellular matrix fibronectin 5 min post- EC. Absence of desmin and disruption of dystrophin staining was evident at 5 min post-EC, whereas fibronectin staining was not detectable intracellularly within the same fiber. *Same fiber in serial sections. Arrows indicate dystrophin rupture. Scale bar, 50 µm.

 
At 30 and 60 min after stretch, all the changes in protein immunostaining were highly significant, and application of zero extracellular Ca2+ solution or leupeptin prevented all the immunostaining abnormalities (desmin, P < 0.001; dystrophin, P < 0.001; titin, P < 0.01; and fibronectin, P < 0.01). These data confirm previous studies that EC can lead to significant loss or damage to cytoskeletal proteins. Desmin and dystrophin were damaged earlier and more severely than titin. Not all desmin- or dystrophin-abnormal fibers displayed fibronectin staining, perhaps representing differences in the cytoskeletal damage pathway compared with the membrane damage pathway. The calpain inhibitor leupeptin and zero-extracellular Ca2+ solution prevented stretch-induced abnormalities of cytoskeletal proteins, including desmin, dystrophin, and titin, as well as extracellular matrix fibronectin.


   DISCUSSION
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
GRANTS
 REFERENCES
 
The aim of this study was to determine whether the calcium-activated protease calpain contributes to damage of cytoskeletal components in mouse skeletal muscle following EC. Following repeated EC, loss of immunolabeling for desmin and dystrophin and increase in titin and fibronectin staining were largely prevented and the decline in force was partly prevented by applying the calpain inhibitor leupeptin or removing extracellular calcium.

Muscle cytoskeleton in stretched muscles.   Muscle cytoskeleton contains a transverse (consisting primarily of desmin) and longitudinal (consisting of titin and nebulin) filament system (28, 32), by which the generated force is transmitted from the sarcomere to the fiber surface. In addition there is the dystrophin-glycoprotein complex, which completes these connections through the sarcolemma to the extracellular matrix. Changing this complex by lengthening active muscles may also affect the function of other membrane-associated proteins, including ion channels, which may in turn lead to elevated [Ca2+]i (33, 37). Consistent with previous studies (7, 21, 22), immunolabeling for desmin and dystrophin declined rapidly, whereas the increase in immunological staining of titin and in intracellular fibronectin staining was somewhat slower. Temporal differences in the damage to these cytoskeletal proteins probably relate to their sensitivity to calcium-protease activity, although differences in spatial distribution or functional role during eccentric contractions may also contribute. The increase in titin staining after stretch was first described by Lieber et al. (21) and was thought to result from greater exposure of antigenic sites after partial hydrolysis. Titin is very readily proteolyzed by calpains (17) in response to increased Ca2+ concentration or stretch (35). The protease calpain-3 cleaves titin at the PEVK region near the COOH terminus (20), which might involve the binding site of titin antibody used in this study. Of course, a possibility that progressive disruption of extramyofibrilar network would make antibody accessible to intramyofibrillar components more easily is not excluded. Absence of fibronectin staining in fibers that had reduced desmin staining suggests that many fibers with damaged cytoskeletal proteins retained their membrane integrity (Fig. 3). We also confirmed earlier studies showing that eccentric damage leads to a reduction in muscle force (2, 15, 21, 37).

Intracellular calcium and cytoskeletal damage in stretched muscles.   Although [Ca2+]i was not measured in the present study, other reports show that [Ca2+]i was significantly elevated after 10 min and remains elevated 1.2- to 2.5-fold for hours (2, 18) and even days (13, 23) following EC protocols. In mdx fibers, EC increases the resting [Ca2+]i, decreases tetanic [Ca2+]i, and decreases muscle force, and these changes could be alleviated by excluding Ca2+ from the medium (37). Excitation-contraction can become uncoupled, and titin degradation occurs following elevation of [Ca2+] in skinned rat and toad muscles (35). These findings suggest that increasing resting [Ca2+]i may play a role in structural and functional muscle defects, particularly after EC. In our study, the changes in immunostaining of desmin, dystrophin, and titin after EC were largely prevented in Ca2+-free solution. Treatment with zero Ca2+ and leupeptin also reduced fibronectin entry into muscle cells, indicating that membrane damage also has a Ca2+/calpain-dependent component. Furthermore, removing extracellular Ca2+ after the EC protocol improved the muscle force. Thus these data all support the hypothesis that, after eccentric exercise, intracellular calcium rises, activates calpain, and damages the muscle cytoskeleton.

Which calcium-activated protease causes cytoskeletal damage in stretched muscles?   Skeletal muscle fiber contains both the ubiquitous calpains, µ-calpain and m-calpain, as well as a muscle-specific calpain, calpain-3 (4, 17). The protease inhibitor leupeptin inhibits m-and µ-calpain (17) and also inhibits calpain-3 once the Ca2+-dependent autolysis process is activated (12, 27). In this study, sustained application of leupeptin largely prevented the changes in immunolabeling for desmin, dystrophin, and titin, suggesting a direct link between cytoskeleton damage and increased calpain activity. It is proposed that calpain-3 might be involved in this process because of its very high sensitivity to a small rise in [Ca2+]i, even in the nanomolar range (35). Although [Ca2+]i needs to be relatively high (≥2 µM) for ≥1 min to activate detectable levels of µ-calpain proteolysis (26), sensitivity increases, as with calpain-3, after its initial activation. Thus, a role for µ-calpain in stretch-induced cytoskeletal damage cannot be excluded. Distinguishing between the roles of the various calpains present in muscle will require more selective calpain inhibitors or calpain-knockout mice.

Time course of calpain activation and cytoskeletal protein damage.   The time course of activation of calpains following stretch-induced muscle injury also requires further investigation. In our previous studies (2, 37), resting [Ca2+]i was only significantly increased at 10 min after stretch. Activation of calpain is potentially fast, as in skinned fibers, in which [Ca2+]i can be raised instantly, calpain activity was detectable after 1–2 min after an increase in [Ca2+]i (35). However, in situ measurements of calpain-3 activity in human muscle biopsies showed that calpain-3 activation after eccentric exercise was increased but not until 24 h after the ECs (25). Thus it appears surprising that in the present experiments we were able to detect the consequences of Ca2+ activation of calpain after only 5 min. There are several possible explanations for this discrepancy. One possibility is that the Ca2+ rise occurs very rapidly in restricted regions around the Ca2+ influx channels and might not have been detected in the above experiments. Similarly, the detection of calpain-3 activation in muscle biopsies may require homogeneous activation, whereas localized activation may occur much earlier but be difficult to detect. Another possibility is that the loss or disruption of desmin and dystrophin has a mechanical origin although this must be modulated in some way by Ca2+/calpain to be consistent with our results. Finally the difference of time course of Ca2+ elevation, activation of calpain, and loss of desmin could result from variations in the experimental approaches, such as species (Balb/c, C57, mdx mouse, rat, toad, and human), degree of muscle activation, or other details of the experimental approach.

Role of cytoskeletal proteins in the reduced force following eccentric contractions.   The mechanism of the reduced force after EC has long been of interest, and it is often suggested that cytoskeletal damage is one component. Two relevant observations in the present study are 1) that force fell to 76%, 1 min after EC, at a time when no changes in cytoskeletal proteins were detectable; and 2) when cytoskeletal damage was largely eliminated by either zero Ca2+ or leupeptin, force, instead of falling to 56% after 1 h, declined to only 69–76%. These findings suggest that the role of the cytoskeletal proteins in force transmission and in maintaining structural integrity required for force production are relatively modest at least in the short term. This agrees with the desmin knockout data where desmin knockout muscles generate lower stress and are less vulnerable to injury compared with wild-type muscles (30). Instead, other mechanisms of force reduction, such as reduced sarcoplasmic reticulum Ca2+ release or reduced Ca2+-sensitivity of the contractile proteins or overstretched sarcomeres, are probably more important (2, 24).

In conclusion, this study demonstrates clearly that calpain, a Ca2+-activated protease, is involved in early damage to muscle cytoskeleton after EC. Removal of calcium or application of leupeptin largely prevented the eccentric damage to muscle cytoskeleton but had only moderate effects on the muscle force. These results suggest two potential therapeutic targets for reducing cytoskeletal damage after EC: 1) by lowering Ca2+ entry (e.g., stretch-activated channel blockers), or 2) by preventing the activation of proteases.


   GRANTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
REFERENCES
 
The work described in this paper was supported by Hong Kong Polytechnic University Central Research Grant G-YF11, and by a grant from the Research Grants Council of the Hong Kong Special Administrative Region, China (PolyU 5468/05M).


   FOOTNOTES
 

Address for reprint requests and other correspondence: E. W. Yeung, Muscle Physiology Laboratory, Dept. of Rehabilitation Sciences, Hong Kong Polytechnic Univ., Hung Hom, Kowloon, Hong Kong (e-mail: rsella{at}polyu.edu.hk)

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


   REFERENCES
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 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 

  1. Badalamente MA, Stracher A. Delay of muscle degeneration and necrosis in mdx mice by calpain inhibition. Muscle Nerve 23: 106–111, 2000.[CrossRef][Web of Science][Medline]
  2. Balnave CD, Allen DG. Intracellular calcium and force in single mouse muscle fibers following repeated contractions with stretch. J Physiol 488: 25–36, 1995.[Abstract/Free Full Text]
  3. Barash IA, Peters D, Friden J, Lutz GJ, Lieber RL. Desmin cytoskeletal modifications after a bout of eccentric exercise in the rat. Am J Physiol Regul Integr Comp Physiol 283: R958–R963, 2002.[Abstract/Free Full Text]
  4. Bartoli M, Richard I. Calpains in muscle wasting. Int J Biochem Cell Biol 37: 2115–2133, 2005.[CrossRef][Web of Science][Medline]
  5. Belcastro AN. Skeletal muscle calcium-activated neutral protease (calpain) with exercise. J Appl Physiol 74: 1381–1386, 1993.[Abstract/Free Full Text]
  6. Belcastro AN, Shewchuk LD, Raj DA. Exercise induced muscle injury: a calpain hypothesis. Mol Cell Biochem 179: 135–145, 1998.[CrossRef][Web of Science][Medline]
  7. Biral D, Jakubiec-Puka A, Ciechomska I, Sandri M, Rossini K, Carraro U, Betto R. Loss of dystrophin and some dystrophin-associated proteins with concomitant signs of apoptosis in rat leg muscle overworked in extension. Acta Neuropathol (Berl) 100: 618–626, 2000.[CrossRef][Medline]
  8. Byrd SK. Alterations in the sarcoplasmic reticulum: a possible link to exercise-induced muscle damage. Med Sci Sports Exerc 24: 531–536, 1992.[Web of Science][Medline]
  9. Colliander EB, Tesch PA. Effects of eccentric and concentric muscle actions in resistance training. Acta Physiol Scand 140: 31–39, 1990.[Web of Science][Medline]
  10. Deconinck N, Ragot T, Marechal G, Perricaudet M, Gillis JM. Functional protection of dystrophic mouse (mdx) muscles after adenovirus-mediated transfer of a dystrophin minigene. Proc Natl Acad Sci USA 93: 3570–3574, 1996.[Abstract/Free Full Text]
  11. DelloRusso C, Scott JM, Hartigan-O'Connor D, Salvatori G, Barjot C, Robinson AS, Crawford RW, Brooks SV, Chamberlain JS. Functional correction of adult mdx mouse muscle using gutted adenoviral vectors expressing full-length dystrophin. Proc Natl Acad Sci USA 99: 12979–12984, 2002.[Abstract/Free Full Text]
  12. Diaz BG, Moldoveanu T, Kuiper MJ, Campbell RL, Davies PL. Insertion sequence 1 of muscle-specific calpain, p94, acts as an internal propeptide. J Biol Chem 279: 27656–27666, 2004.[Abstract/Free Full Text]
  13. Duan C, Delp MD, Hayes DA, Armstrong RB. Rat skeletal muscle mitochondrial [Ca2+] and injury from downhill walking. J Appl Physiol 68: 1241–1251, 1990.[Abstract/Free Full Text]
  14. Fridén J, Lieber RL. Segmental muscle fiber lesions after repetitive eccentric contractions. Cell Tissue Res 293: 165–171, 1998.[CrossRef][Web of Science][Medline]
  15. Fridén J, Lieber RL. Eccentric exercise-induced injuries to contractile and cytoskeletal muscle fibre components. Acta Physiol Scand 171: 321–326, 2001.[CrossRef][Web of Science][Medline]
  16. Gailly P, De Backer F, Van Schoor M, Gillis JM. In situ measurements of calpain activity in isolated muscle fibres from normal and dystrophin-lacking mdx mice. J Physiol 582: 1261–1275, 2007.[Abstract/Free Full Text]
  17. Goll DE, Thompson VF, Li H, Wei W, Cong J. The calpain system. Physiol Rev 83: 731–801, 2003.[Abstract/Free Full Text]
  18. Ingalls CP, Warren GL, Williams JH, Ward CW, Armstrong RB. E-C coupling failure in mouse EDL muscle after in vivo eccentric contractions. J Appl Physiol 85: 58–67, 1998.[Abstract/Free Full Text]
  19. Jones DA, Jackson MJ, McPhail G, Edwards RH. Experimental mouse muscle damage: the importance of external calcium. Clin Sci 66: 317–322, 1984.[Web of Science][Medline]
  20. Kramerova I, Kudryashova E, Tidball JG, Spencer MJ. Null mutation of calpain 3 (p94) in mice causes abnormal sarcomere formation in vivo and in vitro. Hum Mol Genet 13: 1373–1388, 2004.[Abstract/Free Full Text]
  21. Lieber RL, Thornell LE, Fridén J. Muscle cytoskeletal disruption occurs within the first 15 minutes of cyclic eccentric contraction. J Appl Physiol 80: 278–284, 1996.[Abstract/Free Full Text]
  22. Lovering RM, De Deyne PG. Contractile function, sarcolemma integrity, and the loss of dystrophin after skeletal muscle eccentric contraction-induced injury. Am J Physiol Cell Physiol 286: C230–C238, 2004.[Abstract/Free Full Text]
  23. Lynch GS, Fary CJ, Williams DA. Quantitative measurement of resting skeletal muscle [Ca2+]i following acute and long-term downhill running exercise in mice. Cell Calcium 22: 373–383, 1997.[CrossRef][Web of Science][Medline]
  24. Morgan DL, Allen DG. Early events in stretch-induced muscle damage. J Appl Physiol 87: 2007–2015, 1999.[Abstract/Free Full Text]
  25. Murphy RM, Goodman CA, McKenna MJ, Bennie J, Leikis M, Lamb GD. Calpain-3 is autolyzed and hence activated in human skeletal muscle 24 h following a single bout of eccentric exercise. J Appl Physiol 103: 926–931, 2007.[Abstract/Free Full Text]
  26. Murphy RM, Verburg E, Lamb GD. Ca2+ activation of diffusible and bound pools of µ-calpain in rat skeletal muscle. J Physiol 576: 595–612, 2006.[Abstract/Free Full Text]
  27. Ono Y, Kakinuma K, Torii F, Irie A, Nakagawa K, Labeit S, Abe K, Suzuki K, Sorimachi H. Possible regulation of the conventional calpain system by skeletal muscle-specific calpain, p94/calpain 3. J Biol Chem 279: 2761–2771, 2004.[Abstract/Free Full Text]
  28. Price MG. Striated muscle endosarcomeric, and exosarcomeric lattices. In: Advances in Structural Biology, edited by Malhotra SF. New York: JAI Press, 1991.
  29. Proske U, Morgan DL, Brockett CL, Percival P. Identifying athletes at risk of hamstring strains and how to protect them. Clin Exp Pharmacol Physiol 31: 546–550, 2004.[CrossRef][Web of Science][Medline]
  30. Sam M, Shah S, Fridén J, Milner DJ, Capetanaki Y, Lieber RL. Desmin knockout muscles generate lower stress and are less vulnerable to injury compared with wild-type muscles. Am J Physiol Cell Physiol 279: C1116–C1122, 2000.[Abstract/Free Full Text]
  31. Thompson JL, Balog EM, Fitts RH, Riley DA. Five myofibrillar lesion types in eccentrically challenged, unloaded rat adductor longus muscle-a test model. Anat Rec 254: 39–52, 1999.[CrossRef][Medline]
  32. Thornell LE, Price MG. The cytoskeleton in muscle cells in relation to function. Biochem Soc Trans 19: 1116–1120, 1991.[Web of Science][Medline]
  33. Turner PR, Fong PY, Denetclaw WF, Steinhardt RA. Increased calcium influx in dystrophic muscle. J Cell Biol 115: 1701–1712, 1991.[Abstract/Free Full Text]
  34. Turner PR, Westwood T, Regen CM, Steinhardt RA. Increased protein degradation results from elevated free calcium levels found in muscle from mdx mice. Nature 335: 735–738, 1988.[CrossRef][Web of Science][Medline]
  35. Verburg E, Murphy RM, Stephenson DG, Lamb GD. Disruption of excitation-contraction coupling and titin by endogenous Ca2+-activated proteases in toad muscle fibres. J Physiol 564: 775–790, 2005.[Abstract/Free Full Text]
  36. Warren GL, Ingalls CP, Lowe DA, Armstrong RB. Excitation-contraction uncoupling: major role in contraction-induced muscle injury. Exerc Sport Sci Rev 29: 82–87, 2001.[CrossRef][Medline]
  37. Yeung EW, Whitehead NP, Suchyna TM, Gottlieb PA, Sachs F, Allen DG. Effects of stretch-activated channel blockers on [Ca2+]i and muscle damage in the mdx mouse. J Physiol 562: 367–380, 2005.[Abstract/Free Full Text]




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