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Published ahead of print on April 2, 2003, doi:10.1164/rccm.200209-1028OC
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American Journal of Respiratory and Critical Care Medicine Vol 167. pp. 1664-1669, (2003)
© 2003 American Thoracic Society

Original Article

Exercise-induced Quadriceps Oxidative Stress and Peripheral Muscle Dysfunction in Patients with Chronic Obstructive Pulmonary Disease

Annabelle Couillard, François Maltais, Didier Saey, Richard Debigaré, Annie Michaud, Christelle Koechlin, Pierre LeBlanc and Christian Préfaut

UPRES-EA 701, Laboratory of Physiologie des Interactions, Service Central de Physiologie Clinique, Hôpital Arnaud de Villeneuve, Montpellier, France; and Centre de Recherche, Hôpital Laval, Institut Universitaire de Cardiologie et de Pneumologie de l'Université Laval, Sainte-Foy, Quebec, Canada

Correspondence and requests for reprints should be addressed to Correspondence and requests for reprints should be addressed to Annabelle Couillard, Laboratory of Physiologie des Interactions, Service Central de Physiologie Clinique, Hôpital Arnaud de Villeneuve, 34295 Montpellier cedex 5, France. E-mail: annabelle_couillard{at}yahoo.fr


   ABSTRACT
TOP
 ABSTRACT
METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Exercise-induced muscle oxidative stress may be involved in the myopathy associated with chronic obstructive pulmonary disease (COPD). This study was designed to look at whether local exercise induces muscle oxidative stress and whether this oxidative stress may be associated with the reduced muscle endurance in patients with COPD. Quadriceps endurance was measured in 12 patients with COPD (FEV1 = 0.96 ± 0.14 SEM) and 10 healthy sedentary subjects by repeated knee extensions of the dominant leg. Biopsies of the vastus lateralis muscle were obtained before and 48 hours after exercise. Muscle oxidative stress was measured by lipid peroxidation and oxidized proteins. Muscle antioxidant was evaluated by peroxidase glutathion activity. Quadriceps endurance was significantly reduced in patients with COPD when compared with the healthy control subjects (p < 0.01). Forty-eight hours postexercise, only patients with COPD had a significant increase in muscle lipid peroxidation (p < 0.05) and oxidized proteins (p < 0.05), whereas increased peroxidase glutathion activity was only observed in control subjects (p < 0.05). Both increases in muscle lipid peroxidation and oxidized proteins were significantly and inversely correlated with quadriceps endurance capacity in COPD (p < 0.05). In summary, local exercise induced muscle oxidative stress in patients with COPD, whereas it failed to raise antioxidant activity. In these individuals, muscle oxidative stress was associated with a reduced quadriceps endurance.

Key Words: malondialdehyde • skeletal muscle • glutathion peroxidase • myopathy • chronic obstructive pulmonary disease

The peripheral muscle dysfunction observed in patients with chronic obstructive pulmonary disease (COPD) is, in part, characterized by a marked reduction in quadriceps endurance (1, 2). Because this peripheral myopathy is associated with reduced exercise capacity (3) and quality of life (4), the documentation of its underlying mechanisms is likely to be clinically relevant.

Deconditioning related to progressive reduction of daily activities is often quoted as being one of the main reasons why patients with COPD have peripheral myopathy (5). Recent studies, however, have suggested that other factors such as exposure to systemic corticosteroids (6), malnutrition (7), hypoxia (8), and apoptosis (9) may also contribute to the alteration in peripheral muscle function. Oxidative stress, resulting from an inability of the antioxidant systems to cope with elevated oxidant production, is also believed to play an important role in altering peripheral muscle function in patients with COPD (10, 11). Indeed, our group has recently documented evidences of lipid peroxidation, a marker of oxidative stress, in the plasma of patients with COPD but not in healthy subjects after local quadriceps exercise performed to exhaustion (2). Although we hypothesized that the contracting quadriceps was the source of this oxidative stress, it was not confirmed because no muscle biopsies were done. This is an important question because muscle oxidative stress causes noticeable myocyte damage (12, 13) and may potentially be detrimental to muscle function, thus contributing to muscle fatigue (14) and reduced endurance (13). On the basis of this observation, we hypothesized that muscle oxidative stress could be associated with reduced peripheral muscle endurance in patients with COPD.

The objectives of the present study were therefore to determine (1) whether local muscle exercise performed to exhaustion induces oxidative stress within the quadriceps itself and (2) whether this oxidative stress is associated with reduced quadriceps endurance capacity in patients with COPD.

Some of the results of these studies have been reported previously in the form of an abstract (15).


   METHODS
TOP
 ABSTRACT
 METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Study Population
This study involved 22 male subjects, with 12 subjects having COPD as determined by moderate to severe irreversible airflow obstruction (16) (FEV1 < 60% predicted and FEV1/FVC < 70% and < 10% improvement in FEV1 after ß2-agonist inhalation). These 12 patients were all ex-smokers, had no resting hypoxemia, and were clinically stable at the time of the evaluation. They had neither experienced respiratory tract infection nor exacerbation of their disease for at least 4 weeks before being studied. All were receiving inhaled anticholinergic and/or ß2 agonists, and some also received inhaled steroids. None was treated with oral corticosteroids. To avoid potentially confounding factors, we excluded from the study subjects with known muscle disorders, cardiac failure, diabetes mellitus, or alcoholism, as well as those with any other comorbidity that could have impaired their capacity to exercise. Ten age-matched nonsmoker healthy males with only sedentary activities were recruited as control subjects through newspaper advertisements. All participants had a body mass index smaller than 35 kg/m2, and all were questioned on their dietary habits so that those taking antioxidants or vitamin supplements could be excluded. None of the subjects involved in a previous study on systemic oxidative stress and local muscle exercise in COPD (2) participated in the present investigation. The study was approved by the institutional ethics committee, and written consent was obtained after subjects had received a complete explanation of the objectives of the study protocol.

Study Protocol
Pulmonary function test.
All subjects underwent spirometry including measurements of FVC and FEV1. The FEV1/FVC ratio was also calculated. The results of pulmonary function testing were related to the normal values of Knudson and coworkers (17)

Physical activity.
Levels of physical activity were assessed through a physical activity questionnaire adapted for older retired adults (18). This questionnaire provides a reliable and valid method for classifying the activity level of older subjects as high, medium, or low with a score of 9 or more indicating a low physical activity level, thus classifying the subjects as being sedentary. Additional details on this measurement are provided in the online supplement.

Midthigh muscle cross-sectional area measurement.
A computed tomography of the dominant (the stronger one) thigh halfway between the pubic symphisis and the inferior condyle of the femur was performed using a fourth-generation Toshiba Scanner 900S (Toshiba Inc., Tokyo, Japan) (19). Additional details on this measurement are provided in the online supplement.

Quadriceps strength measurement.
The quadriceps maximal voluntary strength was measured while subjects performed dynamic knee extensions against a hydraulic resistance (HF STAR, Hydrafitness Total Power; Henley Health Care, Belton, TX) that could be adjusted to six different levels of resistance. Starting with the lowest level of resistance, the subjects were asked to perform three sets of movements at 5-minute intervals until they reached a level of resistance at which they could no longer complete the full range of movement. Under these conditions, the generated strength reached a plateau, defined as maximal voluntary contraction. Additional details on this measurement are provided in the online supplement.

Quadriceps endurance measurement.
To assess the quadriceps endurance of the dominant leg, we used an exercise bench and followed the method described by Serres and coworkers (1). Subjects were asked to perform repeated knee extensions of the leg against weights corresponding to 30% of their maximal voluntary capacity at a pace of six movements per minute imposed by audio signals (metronome) until exhaustion. The dynamic knee extension was performed for 3 seconds, immediately followed by active leg return (eccentric flexion) against resistance for 3 seconds, and by rest before the next extension. The test was concluded when subjects could no longer perform maximal extension or if they could not sustain, despite verbal encouragement, the required frequency two consecutive times. The duration of the test was then recorded as the quadriceps endurance time. The dyspnea score was measured at rest and immediately after exercise on an analogic visual scale ranging from 0 to 10.

Twitch quadriceps force assessment.
To determine whether local exercise induced contractile muscle fatigue, we quantified the force of the dominant quadriceps during maximal magnetic stimulation of the femoral nerve before and 10 minutes after exercise. This was done using a commercially available magnetic stimulator (Magstim 200; Magstim Co Ltd, Whitland, Wales, UK) equipped with a figure-of-eight 42-mm coil according to the method of Mador and coworkers (20). Additional details on this method are available in the online supplement.

Muscle biopsies.
Percutaneous biopsy specimens of the vastus lateralis muscle of the nondominant leg at baseline and of the dominant leg 48 hours after exercise were taken at midthigh (15 cm above the patella) as described by Bergström (21). The biopsy specimens were first dissected free of visible connective tissue and fat and the remaining muscle tissue was immediately frozen in isopentane cooled to freezing point with liquid nitrogen and stored at -80°C until analysis.

Skeletal Muscle Data Analysis
Fiber typing determination.
All muscle specimens were coded and analyzed without knowledge of the clinical data. Muscle samples were cut into 10-µm thick transverse sections in a cryostat at -20°C. One of these sections was stained for myofibrillar adenosine triphosphatase activity according to the single-step ethanol-modified technique (22). Depending on the staining intensity, fibers were labeled as being type I (nonstained), type IIa (lightly stained), or type IIx (darkly stained). A small proportion of fibers stained intermediate between IIa and IIx and they were classified as IIab, but because of their relative scarcity they were not included in the analysis. Muscle sections were magnified and transmitted to an image analysis software (Photoshop L5) for fiber counting and classification. For each subject, the fiber-type composition was calculated as the total number of fibers of a given type divided by the total number of fibers.

Determination of markers of oxidative stress.
Muscle thiobarbituric acid reactive substances (TBARs) were used as markers of muscle lipid peroxidation and were determined fluorimetrically using the method described by Ohkawa and coworkers (23). The final results were expressed in nmol/g wet weight. The reproducibility of TBARs, calculated as coefficients of variation, was less than 10%. In subjects in whom sufficient amount of muscle tissue was still available (COPD, n = 8; control subjects, n = 7), we also measured protein oxidation by evaluating the levels of protein carbonyls using immunoblotting (Oxyblot kit; Serologicals Corporation, Norcross, GA). Muscle protein carbonyl contents were calculated by adding the integrated density of individual protein bands (Alpha Innotech Corporation, San Leandro, CA) obtained by Western blot analysis (24). Additional details on this method are available in the online supplement.

Determination of muscle antioxidant activity.
Muscle activity of glutathion peroxidase (GPX) was quantified spectrophotometrically according to the method of Nakamura and coworkers (25). The reproducibility of GPX activity, calculated as coefficients of variation, was less than 10%.

Venous Blood Analysis
Determination of muscle damage.
Blood samples were drawn in heparinized tubes, and plasma was obtained by centrifugation (2,500 rpm for 10 minutes at 4°C) and stored at -80°C until analysis. The plasma activity of creatine kinase (CK) was determined at rest as well as 6 and 48 hours after exercise using biochemical assay kits.

Study Design
Subjects were instructed to abstain from strenuous physical activity 4 days before and 2 days after being studied. On Day 1, the subjects underwent spirometry and a computed tomography of the thigh, and they had to answer the physical activity questionnaire. After the first venous blood sample was obtained at rest, strength was evaluated in each leg and a baseline muscle biopsy was performed on the nondominant leg. On Day 2, the subjects were familiarized with the endurance test procedures by performing five consecutive dynamic knee extensions of the dominant leg. They then performed the local muscle endurance exercise test. A second venous blood sample was obtained 6 hours after the end of exercise. On Day 4 (48 hours after local exercise), a third venous blood sample and a biopsy of the dominant quadriceps were obtained. This second biopsy was taken 48 hours postexercise because previous investigations have shown that peak increases in TBARs content and enzymatic antioxidant activity in muscle samples occur in this time frame (26, 27).

Statistical Analysis
Results are expressed as mean ± SEM. According to the homogeneity and normality of the data, comparisons between groups before and after local exercise were performed using the Student's test or the Mann–Whitney test. Comparisons within groups were performed with the Student's paired t test or the Wilcoxon test. Possible correlations between variables were evaluated using Spearman correlation coefficients. The results were considered statistically significant with p values less than or equal to 0.05.


   RESULTS
TOP
 ABSTRACT
 METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Subjects' Characteristics
The anthropometric data did not show any significant differences between patients with COPD and control subjects (Table 1) . Spirometric values showed that patients with COPD had on average severe airflow obstruction (Table 1). All subjects had a low level of physical activity (Table 1).


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  		TABLE 1. Characteristics of the study population

 
Morphologic and Functional Muscle Characteristics
In patients with COPD, there were fewer type I fibers, but a larger proportion of type II fibers than in control subjects (Table 2) . Midthigh muscle cross-sectional area was considerably lesser in patients with COPD (Table 2). Peripheral muscle performance as determined by quadriceps strength also showed significant intergroup difference (Table 2). A significant positive correlation was found between midthigh muscle cross-sectional area and quadriceps strength (r2 = 0.88; p < 0.05), but the strength/midthigh muscle cross-sectional area ratio did not significantly differ between patients with COPD and control subjects. Quadriceps endurance time was twofold lower in the patients with COPD compared with control subjects (Table 2). Resting and post-exercise dyspnea scores varied from 0 to 2.6 ± 0.1 and from 0 to 2.7 ± 0.2 in patients with COPD and healthy subjects, respectively. Resting twitch quadriceps force/midthigh muscle cross-sectional area ratio was not significantly different between both groups. When compared with the baseline values, there was a 25 ± 7% and 27 ± 8% decrease in twitch quadriceps force after local muscle exercise in patients with COPD and healthy subjects, respectively.


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  		TABLE 2. Histologic and functional characteristics of the quadriceps

 
Oxidative Stress and Muscle Damages at Rest and after Local Muscle Exercise
No difference between patients and control subjects was found in the resting levels of muscle lipid peroxidation (TBARs) (Figure 1) , protein carbonyl contents (Figure 2) , or GPX activity (Figure 3) . Representative immunoblots of muscle protein carbonyl groups obtained in a control subject and in a patient with COPD before and after exercise are shown in Figure 2. Four to five bands whose molecular weight varied from 27 to 68 kD were detected in patients with COPD and in control subjects. The migration pattern of these bands was similar between the two groups. After quadriceps exercise, these bands became more intense in patients with COPD, but no new bands were detected. Quadriceps exercise induced a significant increase in muscle TBARs (Figure 1) and in muscle protein carbonyl contents (Figure 2) in patients with COPD (p < 0.05) but not in control subjects. The difference between postexercise and baseline muscle TBARs (Figure 4) and between postexercise and baseline muscle protein carbonyls significantly and inversely correlated with the quadriceps endurance time in patients with COPD (r =-0.66, p < 0.05 and r = -0.70, p = 0.05, respectively). The quadriceps endurance time did not significantly correlate with FEV1 or the physical activity score (r = -0.27 and r = -0.32, respectively, p > 0.05). There was no significant correlation between the proportion of type II fibers and the increase in muscle TBARs or muscle protein carbonyl contents.



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  		Figure 1. Local quadriceps endurance exercise induced a significant increase in the muscle levels of thiobarbituric acid reactive substances (TBARs) in patients with chronic obstructive pulmonary disease (COPD) (black bars; p < 0.05) but not in the healthy subjects (white bars). *p value less than 0.05 compared with baseline values.

 


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  		Figure 2. Representative immunoblots of muscle protein carbonyl groups in a control subject and a patient with COPD before (pre) and after (post) quadriceps exercise (A). Four to five bands whose molecular weight varied from 27 to 68 kD were detected in both subjects. Compared with the preexercise levels, these bands became more intense after quadriceps exercise in the patient with COPD, but no new bands were detected. Negative controls (underivatized protein) are also shown. As indicated by the group mean data (B), local quadriceps endurance exercise induced a significant increase in the muscle protein carbonyl contents in patients with COPD (black bars; n = 8) but not in the healthy subjects (white bars; n = 7). *p value less than 0.05 compared with baseline values. ID = integrated density.

 


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  		Figure 3. Local quadriceps endurance exercise induced a significant increase in glutathion peroxidase (GPX) activity in the healthy subjects but not in patients with COPD. *p value less than 0.05 compared with baseline values, {dagger}p value less than 0.05 compared with values of patients with COPD 48 hours after exercise.

 


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  		Figure 4. There was an inverse and significant correlation between quadriceps endurance time and exercise-induced increased muscle TBARs in patients with COPD (r = -0.66; p < 0.05).

 
Exercise induced a significant increase in muscle GPX activity only in the healthy subjects (p < 0.05) in such a way that muscle GPX activity was significantly different between both groups 48 hours after exercise (Figure 3).

Plasma CK levels were significantly increased 6 hours after local exercise in both groups (from 133 ± 36 IU/L to 169 ± 34 in COPD, p < 0.01 and from 126 ± 20 to 181 ± 28 in healthy subjects, p < 0.01). However, the increase in CK from baseline to 6 hours postexercise ({delta} CK) only correlated with the quadriceps endurance time in healthy subjects (r = 0.83; p < 0.01).


   DISCUSSION
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
On the basis of previous studies that have shown increased systemic oxidized glutathion and lipid peroxidation after exercise in patients with COPD, several investigators have suggested that this oxidative stress may originate, at least in part, within the contracting muscles (2, 11, 28). With muscle biopsy, the current study extends these results by demonstrating that local quadriceps exercise of sufficient intensity to cause fatigue can induce oxidative stress within the contracting quadriceps in patients with COPD but not in healthy subjects. Furthermore, local exercise failed to induce the expected physiologic increase in antioxidant defenses, as assessed by GPX activity, in patients with COPD, a possible explanation for the greater susceptibility to muscle oxidative stress in these individuals. Another important finding of the present study was the relationship between exercise-induced oxidative stress and quadriceps endurance, supporting the concept that muscle oxidative stress may have important functional consequences in patients with COPD.

Oxidative Stress and Antioxidant Defenses at Rest and after Local Quadriceps Exercise
Like Rabinovich and coworkers (29), we were unable to document elevated muscle lipid peroxidation in resting patients with COPD. Those findings, however, contrast with those of a recent study that showed greater accumulation of lipofuscin, a marker of lipid peroxidation, in the vastus lateralis muscle of patients with COPD when compared with healthy subjects (30). This discrepancy may be explained by the different methods that were used to assess lipid peroxidation; whereas TBARs are markers of acute oxidative stress, lipofuscin, which accumulates in the cells, is a reflector of cumulative stress. One may thus speculate that TBARs muscle content returns to normal levels between repeated short courses of oxidative stress, provided the recuperation period is adequate. On the other hand, repeated bursts of oxidative stress are likely to generate long-term accumulation of lipofuscin.

At rest, GPX activity was similar in both groups despite muscle histologic changes in favor of type II fiber in patients with COPD. This result is somewhat surprising because type II muscle fibers have lower levels of GPX activity when compared with type I fiber (31). Thus, normal resting GPX activity in COPD may reflect an adaptive mechanism in COPD to repeated bursts of oxidant production (27). However, this possible adaptive mechanism seems to be insufficient as indicated by the lack of increase in GPX activity after exercise.

The present study confirms the hypothesis (2) that local muscle exercise performed until exhaustion can produce oxidative stress within the contracting muscles. Conversely, Rabinovich and colleagues (29) were unable to document such evidence after whole body exercise in patients with COPD. This may be explained by differences in patient characteristics and exercise intensity, which in the Rabinovich study was probably insufficient to induce muscle oxidative stress. Another significant difference between the two studies is that our local exercise protocol was purposely designed to minimize other possible sources of oxidative stress (lungs, liver, etc.). When analyzed together, the results of these two studies indicate that intensity and type of exercise are important factors in the genesis of muscle oxidative stress in patients with COPD.

Potential Mechanisms of Exercise-induced Oxidative Stress
Because the results of this study suggest that there is a role for oxidative stress as a mediator of peripheral muscle dysfunction in patients with COPD, it would be clinically relevant to try to determine its underlying mechanisms. Exercise can lead to oxidative stress either by increasing prooxidant activity or because of an inadequate antioxidant activity. Xanthine oxidase, within the muscle or its capillary endothelium, can also be an important source of oxidant production during exercise in patients with COPD in the presence of O2 and of adenosine triphosphate breakdown products, including inosine monophosphate. In support of this hypothesis, Heunks and coworkers (11) were recently able to demonstrate that exercise-induced increase in lipid peroxides observed during whole body exercise in patients with COPD can be prevented by pretreating the patients with allopurinol, a potent xanthine oxidase inhibitor. The potential role of this enzyme in generating oxidative stress has been further substantiated by Pouw and coworkers (32) who reported, in patients with COPD, elevated muscle inosine monophosphate levels, as a precursor of hypoxanthine that serves as a substrate for xanthine oxidase activity.

An inefficient mitochondrial handling of oxygen could also contribute to an increase in oxidant production by the respiratory chain. On the basis of studies that have shown, in the quadriceps of patients with COPD, a low citric acid cycle and fatty acid ß-oxidation enzyme activities (33) and also on the basis of other studies that have demonstrated increased cytochrome oxidase activity (8), an enzyme of the mitochondrial electron transport chain, it can be speculated that mitochondrial uncoupling may also enhance oxidant production during exercise in COPD.

The possibility that increased muscle lipid peroxidation and protein oxidation observed 48 hours after exercise in patients with COPD could be related to exercise-induced inflammatory response cannot be excluded. Muscle trauma during exercise can cause injury, which may lead to a prolonged inflammation response, and accumulation of neutrophils and macrophages within the capillary bed of damaged muscle tissues (34). It is therefore possible that those inflammatory cells, which are capable of producing free oxygen radicals, may contribute to the increased muscle lipid peroxidation observed after exercise in patients with COPD (35). We, however, believe that postexercise muscle inflammation is unlikely to be the main mechanism of oxidative stress based on the modest rise in CK increase and on the fact that no patient reported muscle pain 48 hours postexercise, indicating only mild muscle damage. In previous experiments done in our laboratory, we were also unable to show any evidence muscle inflammatory cell infiltration after local quadriceps exercise in patients with COPD (Maltais and Debigaré, unpublished observation).

There is increasing evidence that oxidant production may be used by cells to signal an adaptive enzymatic antioxidant response. Indeed, a recent report (27) suggested that exercise-induced oxidant production serves to raise enzymatic antioxidant responses. In the current study, the finding of a significant postexercise increase in GPX activity in healthy subjects supports these observations. In contrast, muscle GPX activity did not increase appropriately in response to the increase in oxidant production in patients with COPD. A depletion in muscle glutathion, a substrate required for GPX catalyzed reactions, has been reported previously in patients with COPD (36) and is a possible explanation for this observation. Other important antioxidant systems such as vitamin E blood levels may also be deficient in patients with COPD (2). In summary, the available information suggests that the increased susceptibility to muscle oxidative stress in COPD is due to an imbalance between oxidant production and antioxidant defenses in favor of the former.

Peripheral Muscle Dysfunction in Patients with COPD
Because subjects in both groups had similar levels of physical activity, our results suggest that in COPD, factors other than inactivity may be involved in the development of peripheral muscle dysfunction. In this context, there is increasing evidence that suggests a role for oxidative stress as a mediator of peripheral muscle dysfunction in patients with COPD. In the present study, inverse and significant relationships were found between markers of exercise-induced muscle oxidative stress and quadriceps endurance. These findings support the role of cytotoxic oxidant and oxidative stress in the development of muscle fatigue and altered endurance capacity in patients with COPD. Indeed, muscle oxidative stress causes noticeable damage to myocytes organelles such as DNA, proteins, and lipids, resulting in excessive rise in intracellular free calcium, mitochondrial dysfunction, and bioenergetic enzymes downregulation (12, 13). Additional investigations evaluating, for example, the effects of antioxidants on peripheral muscle function are required to further demonstrate the specific role of muscle oxidative stress in mediating peripheral myopathy in patients with COPD.

The release of CK in the extracellular environment is influenced by various factors including magnitude of mechanical cell injury, increase in body temperature, content of high-energy substrate within the cells, and duration of exercise (37). Hence, the positive and significant relationship observed between exercise-induced CK release and quadriceps endurance time in the control subjects was not unexpected. However, we were surprised to find similar exercise-induced CK increases in both groups, despite a much shorter duration of exercise in patients with COPD. This probably indicates that other mechanisms may have initiated the rise in CK in patients with COPD. Given that products of peroxidation decrease membrane fluidity, increase membrane fragility and, therefore, susceptibility to rupture (38), the increase in peroxidative damage observed after local exercise could have contributed to the CK release.

Methodologic Considerations
Physical activity.
A specific physical activity questionnaire adapted for older retired adults able to carry on usual daily activities was selected for this study. Although this questionnaire does not provide a direct quantification of the number and intensity of daily activities, it has been validated against other activity measures such as 24-hour activity self-reporting and pedometer scores (18). We are therefore confident that both groups had similar low levels of physical activity.

Quadriceps endurance.
To study local muscle endurance, we used a protocol similar to the one described by Andersen and coworkers (39). This protocol minimizes the ventilatory response and provides highly reproducible values for muscle endurance (40), whereas electromyographic recordings confirm that the quadriceps is the main active muscle during the local exercise. The increase in dyspnea and presumably in ventilation after local exercise was of small amplitude in this study, and we, therefore, assume that quadriceps exhaustion was the main factor limiting exercise. The 25% decrease in quadriceps twitch force 10 minutes postexercise also supports this contention. Interestingly, the reductions in quadriceps twitch force observed after local exercise were of greater magnitude than those reported by Mador and coworkers (20) after whole body cycling exercise, suggesting that a greater degree of muscle fatigue may be reached after local exercise than after whole body exercise.

Muscle Markers of Oxidative Stress
Tissue lipid peroxidation measurement (TBARs), which is an indicator of reactive oxygen species molecular reactions, was used to assess global oxidative stress. One potential limitation of TBARs is that under oxidative stress conditions, malondialdehyde, hydroperoxides, and some carbohydrates and amino acids may yield products that are able to react with thiobarbituric acid (41). To circumvent this limitation, the oxidation of muscle protein was also evaluated. The fact that a similar conclusion about oxidative stress was reached with both methods is reassuring in terms of the validity of our findings.

In summary, this study shows that exhaustive local exercise may induce oxidative stress in the quadriceps of patients with COPD and that the expected increase in antioxidant defenses after exercise is compromised in these individuals. From a clinical perspective, this oxidant/antioxidant imbalance may contribute to the reduction of muscle endurance. Further studies are needed to determine more precisely the mechanisms involved in the exercise-induced muscle oxidative stress in COPD and to confirm its implication in the myopathy of these patients.


   Acknowledgments
 
The authors thank Marthe Belanger, Marie-Josée Breton, Brigitte Jean, and Catherine Scott-Carmeni for valuable technical assistance and Dr. Jean Deslauriers for editing the manuscript.


   FOOTNOTES
 
Supported in part by Canadian Institutes of Health Research grant MOP-53135. A.C. was supported by a traveling grant from la Coopération Franco-Québécoise. F.M. is a research scholar of the Fonds de la Recherche en Santé du Québec. R.D. and D.S. are recipients of a Ph.D. training award of Fonds de la Recherche en Santé du Québec.

This article has an online supplement, which is accessible from this issue's table of contents online at www.atsjournals.org

Received in original form September 11, 2002; accepted in final form March 25, 2003


   REFERENCES
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
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