BLOOMER, RICHARD J.1; SCHILLING, BRIAN K.1; KARLAGE, ROBYN E.1; LEDOUX, MARK S.2; PFEIFFER, RONALD F.2; CALLEGARI, JONATHAN1
Although the pathology of Parkinson disease (PD) is indeed complex, one biochemical factor implicated in both neurodegeneration (9,10,18) and muscle fatigue/weakness (17) is excessive production of reactive oxygen and nitrogen species (RONS). The presence of increased RONS coupled with decreased antioxidant protection leads to a state of oxidative stress (11), a condition that is well documented in subjects with PD (18). The increase in oxidative stress biomarkers in persons with PD (7,28) seems related to both increased RONS production (13) and lowered antioxidant capacity (1,7,21,27). This ultimately has the potential to damage cellular structures, including phospholipid membranes, mitochondria, and DNA (29).
Several reports indicate that RONS directly impair muscle contractile function and force via defects in excitation-contraction coupling (8), while leading to greater fatigue rates in skeletal muscle (17,25) by promoting the oxidation of cellular lipids and proteins (3). Lipid peroxidation impairs cell membrane integrity and results in a loss of membrane fluidity (2), whereas protein oxidation affects structural and contractile proteins (20), as well as enzymatic proteins (12), all of which are important for skeletal muscle contraction.
One potential intervention aimed at minimizing oxidative stress in persons with PD is exercise. Structured physical activity involving either aerobic (24) or anaerobic (5) exercise has proven effective to both increase endogenous enzymatic [e.g., superoxide dismutase (SOD), glutathione peroxidase (GPx)] and nonenzymatic (e.g., glutathione) antioxidants and decrease RONS production. Together, such adaptations have been noted in numerous studies to decrease oxidative stress [for reviews, please see References (15,16,22)]. In neurologically normal adults, significant adaptations have been reported to occur in as short as 8 wk. However, it is unknown if similar findings can be observed in those with PD, especially considering that exercise needs to be of sufficient intensity (19,23) and duration (4) to promote such adaptations, and subjects with PD may be physically incapable of exercising at the required level.
Possibly, an improvement in a subject's oxidative status may be related to enhanced muscle strength and physical performance, as well as attenuation in PD progression. Surprisingly, no study to date has investigated the role of structured exercise to improve oxidative status in persons with PD. It was our purpose in this pilot study to determine the impact of resistance exercise in a sample of PD subjects on common biomarkers of oxidative stress and antioxidant status. It was also our objective to determine whether resistance exercise is well tolerated in persons with PD. We hypothesized that resistance training would be well tolerated and result in enhanced antioxidant enzyme activity and decreased oxidative stress in persons with PD compared to no exercise. Such information is important when designing long-term, more intensive exercise regimens aimed at improving oxidative status in this subject population.
Eight men and eight women with mild to moderate PD (Hoehn and Yahr stages I and II) were recruited for our study. All subjects completed a medical screening by a board-certified neurologist (M.S.L. or R.F.P.) to determine eligibility. None of the subjects used tobacco products and did not use antioxidant supplements for at least 2 wk preceding the start of the intervention until the end of the intervention. In addition, subjects did not use cardiovascular drugs that are known to have antioxidant properties (e.g., statins, angiotensin-converting enzyme inhibitors). We did not exclude subjects on the basis of PD-related drug use. Two of the women reported using estrogen replacement therapy and were matched to training and control groups. None of the men reported using testosterone replacement therapy, and no subject reported using growth hormone; however, serum levels of these hormones were not examined. All subjects were capable of walking a 20-ft path, turning, and returning to the start without the use of an assistive device. Subjects with orthostatic hypotension, dementia, or other significant comorbidities (e.g., stroke, severe degenerative osteoarthritis) were excluded. All experimental procedures were performed in accordance with the policy statement of the American College of Sports Medicine on research with human subjects. The University of Memphis Human Subjects Committee approved all experimental procedures (H06107). All subjects provided both verbal and written consent before participating in this investigation.
Before randomization and at the end of the 8-wk intervention period, all subjects performed an assessment of maximum strength for the lower body using a leg press machine. After the warm-up, subjects were provided several (three to five) attempts to achieve their one-repetition maximum (1RM), with 2-3 min of rest between each. The maximum weight lifted one time was recorded as the 1RM.
After this assessment, subjects were randomly assigned to either a resistance exercise training group (n = 8, 4 men and 4 women: 61 ± 2 yr; 172 ± 3 cm; 82 ± 7 kg) or a control group (n = 8, 4 men and 4 women: 57 ± 3 yr; 167 ± 4 cm; 77 ± 7 kg) for an 8-wk period. Randomization was based on disease stage (Hoehn and Yahr stages I and II) and sex, utilizing an random number table. A group of healthy control subjects without PD (n = 9, 5 men and 4 women: 60 ± 2 yr; 170 ± 4 cm; 76 ± 3 kg) were included for baseline comparison. Subjects with PD assigned to the no-exercise control group were instructed to continue their usual activities for the 8-wk study period. Subjects with PD assigned to the resistance exercise training group performed three sets of five to eight repetitions for the leg press, seated leg curl, and calf press (Hammer Strength™) twice weekly for 8 wk under direct supervision of a Certified Strength and Conditioning Specialist. Subjects performed each set to a point of momentary muscular failure. Progression was planned so that when eight repetitions could be completed for all three sets, the weight was increased 5% to 10%. For each training session (as well as for the blood collection described below), subjects reported to the laboratory in the "on" state (typically 60 min after their first morning dose of antiparkinsonian medications). No medication-specific restrictions were given to subjects or to the monitoring physician.
Blood collection and biochemistry.
Before beginning the 8-wk intervention, and within 3 d of completing the intervention (with the majority of postintervention samples collected on day 2 after the final training session), fasting venous blood samples (approximately 20 mL) were taken from subjects in the morning (0800-1000 h) after a 10-min period of quiet rest. Blood was processed immediately and stored in multiple aliquots at −80°C until analyzed. All assays were performed in duplicate and on first thaw. Hydrogen peroxide (H2O2) and catalase activity were measured in plasma using the Amplex Red reagent method as described by the reagent provider (Molecular Probes, Invitrogen Detection Technologies, Eugene, OR). The coefficient of variation for these assays was 3.9% and 4.8%, respectively. Malondialdehyde (MDA), a marker of lipid peroxidation, was measured in plasma using the method described by Jentzsch et al. (14). The coefficient of variation for this assay was 4.7%. Antioxidant capacity was measured in serum using the trolox-equivalent antioxidant capacity (TEAC) assay using procedures outlined by the reagent provider (Sigma Chemical, St. Louis, MO) and as previously described (26). The coefficient of variation for this assay was 5.2%. Serum SOD activity was measured using enzymatic procedures as described by the reagent provider (Cayman Chemical, Ann Arbor, MI), where 1 unit of SOD is the amount of enzyme needed to exhibit 50% dismutation of the superoxide radical. The coefficient of variation for this assay was 5.1%. Plasma GPx activity was measured using enzymatic procedures as described by the reagent provider (Cayman Chemical). Values for GPx were calculated using the nicotinamide adenine dinucleotide phosphate (NADPH) extinction coefficient and are presented in nanomoles per minute per milliliter where 1 unit is defined as the amount of enzyme needed to oxidize 1.0 nmol of NADPH to NADP+. The coefficient of variation for this assay was 7.1%.
For the main analysis, all dependent variables were analyzed using a 2 (group) × 2 (pre- and postintervention) ANOVA. Single degree of freedom contrasts were used to compare the two PD groups before and after intervention. Cohen's d and effect size calculations were performed. Data for healthy control subjects were compared to those of PD subjects using a one-way ANOVA. All analyses were performed using JMP statistical software (Version 4.0.3; SAS Institute, Cary, NC). Statistical significance was set at α < 0.05. Data are presented as means ± SEM.
Of the 16 PD subjects enrolled in the intervention, 13 (n = 6, resistance training group; n = 7, control group) successfully completed all aspects of the study. Of the three subjects who were not included in the final analysis, two failed to complete all training sessions (due to reasons unrelated to the actual exercise protocol) and one failed to provide blood samples both before and after intervention. No differences were noted for age, height, and weight between PD subjects and healthy controls. Leg press 1RM values were not different between resistance training (165 ± 30 kg) and control (160 ± 32 kg) groups before intervention (P > 0.05), but resistance training resulted in an 18% increase in leg press 1RM in the training group, with no change noted in the control group. Healthy control subjects had similar 1RM values (164 ± 21 kg) compared with PD subjects (P > 0.05).
At baseline, no differences in oxidative stress or antioxidant biomarkers were noted between exercising and nonexercising PD participants or between PD and healthy controls (Figs. 1 and 2; Table 1). No main effects were noted for MDA (P > 0.05), but the interaction effect approached statistical significance (P = 0.06). A group main effect was noted for H2O2 (P = 0.01), with lower values for the resistance training group compared with the control group. The interaction effect for H2O2 approached statistical significance (P = 0.13). Resistance training resulted in a decrease in both MDA (15%; Cohen's d = 0.76; effect size = 0.36) and H2O2 (16%; Cohen's d = 0.63; effect size = 0.30), with a 14% increase observed in both biomarkers in the PD control group (Fig. 1 and Fig. 2). With these changes, a postintervention difference was apparent between the resistance training and control groups for H2O2 (P = 0.007), with a trend for difference noted for MDA (P = 0.06). Post hoc power analyses noted that a total sample size of 16 and 22 subjects (n = 8 and n = 11 subjects per group) would be needed to observe a statistically detected interaction effect for MDA and H2O2, respectively. Resistance training also resulted in a modest increase in both SOD (9%; Cohen's d = 0.69; effect size = 0.33) and GPx (15%; Cohen's d = 0.94; effect size = 0.42) activity, although these changes did not reach statistical significance (P > 0.05). Post hoc power analyses noted that a total sample size of 70 and 44 subjects (n = 35 and n = 22 subjects per group) would be needed to observe a statistically detected interaction effect for SOD and GPx activity, respectively. No changes were noted for TEAC or catalase as a result of resistance training (P > 0.05). Data for antioxidant enzymes and TEAC are shown in Table 1.
To our knowledge, this is the first investigation to study the effect of supervised resistance exercise on oxidative stress and antioxidant biomarkers in persons with PD. We noted modest decreases in both MDA and H2O2 in the PD resistance training group. In contrast, a slight increase was noted in both biomarkers in the PD control group. We also noted a modest increase in both SOD and GPx activity in the PD resistance training group. However, these changes were not statistically significant, likely due to our small sample size. Interestingly, preintervention values for all biomarkers were not different between PD subjects and healthy control subjects, a finding that opposes some work (1,27) but is in agreement with the findings of Sudha et al. (28). This may be because all of the PD subjects in this investigation had early disease stages, and healthy control subjects were age-matched.
Despite our inability to detect statistically significant findings in all measured variables, we believe that these data provide evidence that resistance exercise has the potential to improve oxidative status in persons with PD. It is noteworthy that all subjects tolerated the moderate volume exercise protocol very well (nine sets performed twice weekly), with no reported problems related to muscle pain or recovery. Possibly, a longer-duration, more rigorous, and higher-volume exercise protocol may yield more robust changes in the biomarkers investigated. It is important to understand that adaptations in endogenous antioxidant protection are dependent on the magnitude of RONS production during prior bouts of acute exercise (24). That is, the production of RONS serves as the "trigger" to promote adaptations within the antioxidant defense system. Therefore, more strenuous acute exercise sessions that increase RONS to a greater extent may lead to a further improvement in endogenous antioxidant protection, potentially decreasing the oxidation of important macromolecules. Future work is needed within a sample of PD subjects to test this hypothesis and to determine whether more demanding exercise protocols, performed for longer than 8 wk, can be well tolerated.
In addition, future studies should take note of our post hoc power analyses in which we identify a sample size as small as 22 total subjects (n = 11 subjects per group) to achieve a statistically detected interaction effect for oxidative stress biomarkers (MDA and H2O2), with larger samples (e.g., n = 35 subjects per group) being needed to detect changes in antioxidant enzyme activity. This assumes similar variance and effect as we observed in our sample of PD subjects. Possibly, longer-duration, more strenuous protocols used to induce positive adaptations in our chosen biomarkers may yield a greater magnitude of effect, and hence decrease the sample size requirements.
Prior studies involving healthy human subjects have reported similar findings as those noted here for increased antioxidant enzyme activity after exercise training [for review, see References (5,22)]. The increased antioxidant enzyme activity in the PD resistance training group may have been partly responsible for the decrease in both MDA and H2O2. That is, SOD catalyzes the dismutation of superoxide anion that has the potential to ultimately form the hydroxyl radical, directly promoting lipid peroxidation. Moreover, GPx directly catalyzes the reduction of H2O2, using glutathione as the electron donor.
Because of the short duration of our investigation, it is difficult to infer the potential clinical relevance of our findings. It is apparent that oxidative stress is involved in the pathogenesis of PD (9,10) and is further increased with PD progression (6,13). Therefore, any increase in antioxidant capacity and decrease in macromolecule oxidation may be viewed as positive. However, longer-term exercise intervention trials are needed to determine the relationship between subject oxidant status and functional/clinical parameters. Because PD is a progressive disease, possibly, regular exercise may simply serve the function of slightly maintaining or improving a subject's oxidant status (as observed in the PD resistance training group), as opposed to worsening oxidant status with the progression of the disease. Such an effect during the course of several years may have important clinical relevance for persons with PD. In this way, exercise training may serve to complement other medical and alternative therapies (e.g., diet, antioxidant nutrients such as vitamin E and coenzyme Q10) in persons with PD.
In conclusion, we report that short-term, low-volume resistance exercise training may be associated with reductions in markers of oxidative stress (MDA and H2O2) and increases in antioxidant capacity (SOD and GPx activity). Future studies with larger samples are needed to extend these findings. In such studies, considerations should be made for the inclusion of a longer-term and higher-volume resistance exercise training protocol, a variety of oxidative stress biomarkers (e.g., lipid, protein, and DNA oxidation), enrollment of PD subjects of different disease status, and correlation analyses between oxidant status, clinical, and functional end points. Through such work, it may be discovered that exercise training may be a successful adjunctive therapy for persons with PD.
This project was supported in part by Life Fitness, Inc. The authors thank Michael Falvo, Kelsey Fisher-Wellman, and Webb Smith for their assistance with different aspects of this work. The results of the present study do not constitute endorsement by ACSM.
1. Abraham S, Soundararajan CC, Vivekanandhan S, Behari M. Erythrocyte antioxidant enzymes in Parkinson's disease. Indian J Med Res
2. Alessio HM. Lipid peroxidation in healthy and diseased models: influence of different types of exercise. In: Sen CK, Packer L, Hanninen O, editors. Handbook of Oxidants and Antioxidants in Exercise
. Amsterdam (Netherlands): Elsevier Science; 2000. p. 115-27.
3. Andrade FH. Reactive oxygen species and skeletal muscle function. In: Radak Z, editor. Free Radicals in Exercise and Aging
. Champaign (IL): Human Kinetics; 2000. p. 1-33.
4. Bloomer RJ, Davis PG, Consitt LA, Wideman L. Plasma protein carbonyl response to increasing exercise duration in aerobically trained men and women. Int J Sports Med
5. Bloomer RJ, Goldfarb AH. Anaerobic exercise and oxidative stress: a review. Can J Appl Physiol
6. Bostantjopoulou S, Kyriazis G, Katsarou Z, Kiosseoglou G, Kazis A, Mentenopoulos G. Superoxide dismutase activity in early and advanced Parkinson's disease. Funct Neurol
7. Buhmann C, Arlt S, Kontush A, et al. Plasma and CSF markers of oxidative stress are increased in Parkinson's disease and influenced by antiparkinsonian medication. Neurobiol Dis
8. Goldhaber JI, Qayyum MS. Oxygen free radicals and excitation-contraction coupling. Antioxid Redox Signal
9. Hald A, Lotharius J. Oxidative stress and inflammation in Parkinson's disease: is there a causal link? Exp Neurol
10. Halliwell B. Role of free radicals in the neurodegenerative diseases: therapeutic implications for antioxidant treatment. Drugs Aging
11. Halliwell B, Gutteridge JMC. Free Radicals in Biology and Medicine
. 2nd ed. New York (NY): Clarendon Press, Oxford University Press; 1989. p. 1-85.
12. Haycock JW, Jones P, Harris JB, Mantle D. Differential susceptibility of human skeletal muscle proteins to free radical induced oxidative damage: a histochemical, immunocytochemical and electron microscopical study in vitro
. Acta Neuropathol
13. Ihara Y, Chuda M, Kuroda S, Hayabara T. Hydroxyl radical and superoxide dismutase in blood of patients with Parkinson's disease: relationship to clinical data. J Neurol Sci
14. Jentzsch AM, Bachmann H, Furst P, Biesalski HK. Improved analysis of malondialdehyde in human body fluids. Free Rad Biol Med
15. Ji LL. Exercise-induced modulation of antioxidant defense. Ann NY Acad Sci
16. Ji LL, Gomez-Cabrera MC, Vina J. Exercise and hormesis: activation of cellular antioxidant signaling pathway. Ann NY Acad Sci
17. Juel C. Muscle fatigue and reactive oxygen species. J Physiol
. 2006;576(Pt 1):1.
18. Kidd P. Parkinson's disease as multifactorial oxidative neurodegeneration: implications for integrative management. Altern Med Rev
19. Leaf DA, Kleinman MT, Hamilton M, Barstow TJ. The effect of exercise intensity on lipid peroxidation. Med Sci Sports Exerc
20. Liu DF, Wang D, Stracher A. The accessibility of the thiol groups on G- and F-actin of rabbit muscle. Biochem J
21. Paraskevas GP, Kapaki E, Petropoulou O, Anagnostouli M, Vagenas V, Papageorgiou C. Plasma levels of antioxidant vitamins C and E are decreased in vascular parkinsonism. J Neurol Sci
22. Powers SK, Ji LL, Leeuwenburgh C. Exercise training-induced alterations in skeletal muscle antioxidant capacity: a brief review. Med Sci Sports Exerc
23. Quindry JC, Stone WL, King J, Broeder CE. The effects of acute exercise on neutrophils and plasma oxidative stress. Med Sci Sports Exerc
24. Radak Z, Taylor AW, Ohno H, Goto S. Adaptation to exercise-induced oxidative stress: from muscle to brain. Exerc Immunol Rev
25. Reid MB. Nitric oxide, reactive oxygen species, and skeletal muscle contraction. Med Sci Sports Exerc
26. Rice-Evans CA. Measurement of total antioxidant activity as a marker of antioxidant status in vivo
: procedures and limitations. Free Radic Res
. 2000;33 Suppl:S59-66.
27. Serra J, Dominguez R, de Lustig E, et al. Parkinson's disease is associated with oxidative stress: comparison of peripheral antioxidant profiles in living Parkinson's, Alzheimer's, and vascular dementia patients. J Neural Transm
28. Sudha K, Rao A, Rao S, Rao A. Free radical toxicity and antioxidants in Parkinson's disease. Neurol India
29. Valko M, Leibfritz D, Moncol J, Cronin MT, Mazur M, Telser J. Free radicals and antioxidants in normal physiological functions and human disease. Int J Biochem Cell Biol