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Oxidative Stress Responses in Older Men
during Endurance Training and Detraining
IOANNIS G. FATOUROS1, ATHANASIOS Z. JAMURTAS2, VASILIKI VILLIOTOU3, SOFIA POULIOPOULOU3,PANAGIOTIS FOTINAKIS1, KIRIAKOS TAXILDARIS1, and GEORGE DELICONSTANTINOS3 1Democritus University of Thrace, Department of Physical Education and Sport Science, Komotini, GREECE; 2Universityof Thessaly, Department of Physical Education and Sports Sciences, Trikala, GREECE; and 3University of Athens,Medical School, Department of Experimental Physiology, Athens, GREECE ABSTRACT
FATOUROS, I. G., A. Z. JAMURTAS, V. VILLIOTOU, S. POULIOPOULOU, P. FOTINAKIS, K. TAXILDARIS, and G.
DELICONSTANTINOS. Oxidative Stress Responses in Older Men during Endurance Training and Detraining. Med. Sci. Sports Exerc., Vol. 36, No. 12, pp. 2065–2072, 2004. Purpose: Aging is associated with increased oxidative stress, whereas systematic exercise
training has been shown to improve quality of life and functional performance of the aged. This study aimed to evaluate responses of selected markers of oxidative stress and antioxidant status in inactive older men during endurance training and detraining. Methods:
Nineteen older men (65–78 yr) were randomly assigned into either a control (C, N ϭ 8) or an endurance-training (ET, N ϭ 11, threetraining sessions per week, 16 wk, walking/jogging at 50 – 80% of HR ) group. Before, immediately posttraining, and after 4 months of detraining, subjects performed a progressive diagnostic treadmill test to exhaustion (GXT). Plasma samples, collected before and immediately post-GXT, were analyzed for malondialdehyde (MDA) and 3-nitrotyrosine (3-NT) levels, total antioxidant capacity (TAC), and glutathione peroxidase activity (GPX). Results: ET caused a 40% increase in running time and a 20% increase in maximal
) (P Ͻ 0.05). ET lowered MDA (9% at rest, P Ͻ 0.01; and 16% postexercise, P Ͻ 0.05) and 3-NT levels (20% postexercise, P Ͻ 0.05), whereas it increased TAC (6% at rest, P Ͻ 0.01; and 14% postexercise, P Ͻ 0.05) and GPX (12%
postexercise, P Ͻ 0.05). However, detraining abolished these adaptations. Conclusions: ET may attenuate basal and exercise-induced
lipid peroxidation and increase protection against oxidative stress by increasing TAC and GPX activity. However, training cessation
may reverse these training-induced adaptations. Key Words: OXIDATIVE DAMAGE, ANTIOXIDANT CAPACITY, MDA, 3-NI-
TROTYROSINE, CARDIOVASCULAR EXERCISE, AGING Reactive oxygen and nitrogen species (ROS/NOS) Systematic exercise is believed to extend average life such as superoxide (O Ϫ), hydrogen peroxide span by reducing the incidence of cardiovascular and other degenerative diseases while increasing functional perfor- O) generated by either increased oxygen consumption, mance in the aged (7,15). However, the mechanisms by cytokines, inflammation processes, and alterations in blood which exercise produces its positive effects are unclear.
flow like ischemia reperfusion or other stressful conditions Whole-body and active muscle oxygen consumption is in- (trauma, inflammation, chronic diseases, etc.) have been creased 10 –20 times above resting value during acute in- implicated in aging development (21). The free radical tense physical exercise (4). Acute endurance exercise causes theory of aging suggests that an accumulation of oxidative significant ROS/NOS generation in several tissues (e.g., damage to DNA, proteins, and lipids takes place in aged muscle, heart, and liver (4)), increases oxidative stress bi-omarkers (i.e., protein carbonyls and MDA) (9), and alters tissues such as skeletal muscle (21). Aging has been asso- antioxidants’ and antioxidant enzymes’ levels in various ciated with increased generation of reactive oxygen- and tissues (6,9,14). It has been postulated that because endur- nitrogen-centered species, limited antioxidant capacity, de- ance exercise causes an augmented generation of oxidants in creased function of repair systems, and increased macro- muscle, systematic exercise training might upregulate mus- cle’s antioxidant defense system. Chronic exercise traininghas been suggested to induce positive adaptations to anti-oxidant defense systems (27). Recent evidence suggests thatthe amount of physical activity is associated with high Address for correspondence: Ioannis G. Fatouros, Dept. of Physical Edu-cation & Sport Science, 7th km of National Rd. Komotini-Xanthi, Komo- antioxidant enzyme activity level, and that this association is tini, 69100, Greece; E-mail: [email protected].
intensity-dependent (3). Liu et al. (19) demonstrated that Submitted for publication March 2004.
chronic exercise might induce a tissue-specific positive ad- aptation to oxidative stress development and antioxidantenzyme levels in rats. Radak et al. (24) also suggested that 0195-9131/04/3612-2065MEDICINE & SCIENCE IN SPORTS & EXERCISE® endurance exercise training exerts a beneficial effect in Copyright 2004 by the American College of Sports Medicine oxidative damage development independent of age. In fact, endurance exercise training has been shown to decrease TABLE 1. Physical characteristics, physical activity, treadmill time to exhaustion, and aerobic capacity levels of the subjects.
Detraining
Detraining
Variable
(N ؍ 8)
(N ؍ 8)
(N ؍ 5)
(N ؍ 11)
(N ؍ 11)
(N ؍ 6)
Values are means Ϯ SD; C, control group; ET, endurance training group.
a Subjects’ physical activity level according to Baecke Questionnaire for Older Adults, V˙O DNA damage, increase DNA repair in aged rat skeletal diately post-GXT. Thereafter, subjects were randomly as- muscle (25), and upregulate erythrocyte antioxidant enzyme signed to one of two groups: control group (C, N ϭ 8) and activities (15). In addition, 6-month resistance exercise endurance-training group (ET, N ϭ 11). Subjects in ET training was shown to attenuate aerobic exercise-induced trained for 16 wk. After training, subjects underwent a lipid peroxidation in the elderly (34). In contrast, other 16-wk detraining period in which no training was per- studies have shown no change (15) or a decrease in antiox- formed. A GXT and blood sampling was repeated at the end idant enzyme activity in muscle with endurance training (18). Most of the studies conducted in this field used mostly Exclusionary criteria. Subjects’ participation was
younger individuals and various training protocols. There- based upon the following criteria: 1) were available to fore, it is unclear whether systematic exercise training can participate in measurements for 32 wk; 2) were completely alter oxidative stress status induced by ROS/RNOS and antioxidant function in the aged. Although a number of and a score below 9 in the Modified Baecke Questionnaire studies examined training-induced adaptations of oxidative for Older Adults were used as indices of physical inactivity) stress and antioxidant status markers in young and older (Table 1) (1,36); 3) were free of musculoskeletal problems adults, there is a lack of information on adaptations caused and potentially orthopaedic/neuromuscular limitations; 4) by prolonged cessation of training stimulus (detraining).
had a resting blood pressure below 160/100 mm Hg (sub- Therefore, the objective of the present study was to in- jects on antihypertensive medications (six men) maintained vestigate: a) whether a long-term endurance exercise train- their medication throughout the study); 5) had no signs of ing protocol was able to alter oxidative stress biomarkers as cardiovascular/respiratory complications (at rest and during well as antioxidant status responses in aged individuals, and GXT); 6) reported no tobacco use within the 6 months b) how cessation of training stimulus affects possible train- before the study or during the training and detraining period, ing-induced adaptation of oxidative stress development in had normal dietary habits (diet recalls were administered throughout the study), did not consume aspirin (cyclo-oxy-genase can affect oxidant/antioxidant status) or alcoholicbeverages at least 1 wk before exercise testing, and were not consuming antioxidant compounds including vitamins, min- Subjects. Nineteen apparently healthy older men be-
erals, and medications (i.e., probucol, nebivolol, and anti- tween 65 and 78 yr volunteered to participate in a 16-wk training study. A written informed consent was signed by all Measurement of anthropometric variables. Sub-
participants regarding their participation after being in- jects’ body weight was measured while they were wearing formed of all risks, discomforts and benefits involved in the underclothes on a balance scale (Seca) calibrated to the study. Procedures were in accordance with the Helsinki nearest 0.1 kg after an 8-h fast. Barefoot standing height was Declaration of 1975, and institutional review board approval measured to the nearest 0.1 cm by using a wall-mounted was received for this study. The physical characteristics of stadiometer. Subcutaneous skinfold thickness was measured sequentially, in triplicate, at the chest, biceps, triceps, sub- A two-group randomized repeated-measures design was scapula, abdomen, ilium, calf, and thigh by the same inves- followed. Subjects visited the laboratory three times at base- tigator using a skinfold caliper (Harpenden, HSK-BI, British line. During their first visit, subjects were examined by a Indicators, UK) and standard technique (1). The average of trained physician for limiting health complications, given a three measures for each skinfold was used, and the sum of physical activity questionnaire to complete (36), and asked the eight skinfolds was used as an index of body fatness.
to sign an informed consent. In their second visit, subjects Measurement of maximal oxygen consumption
had their body height/weight and skinfolds measured, and underwent a progressive diagnostic treadmill test to exhaus- or jogging) using a modified version of the Bruce protocol tion (GXT) to evaluate their maximal oxygen consumption (1) before training, immediately posttraining, and after 4 ). Blood samples were collected before and imme- months of detraining. A 12-lead ECG, heart rate, and bra- Official Journal of the American College of Sports Medicine chial artery cuff pressure were monitored continuously dur- guard column (Merck Ltd., Poole, Dorset, UK, with a C-8 ing the test and the recovery period. Ratings of perceived cartridge) (12). The eluant was 500 mM KH PO -H PO exertion (1) were determined at the end of each minute of (pH 3.01) with 10% methanol (v/v) at a flow rate of 1 exercise and recovery. A SensorMedics Vmax 29 pulmo- mL·minϪ1 through a Polymer Laboratories Isocratic pump nary gas exchange system (Yorba Linda, CA) was used to and an Ultraviolet detector set at 274 nm. Using this tech- nique, the detection limit was 0.2 ␮M. 3-NT and other reagents was measured continuously via breath-by-breath analysis were purchased from Sigma Chemical Co. (Poole). All sam- with the use of a computerized system. To ascertain that ples were determined in duplicate, while inter- and intra-coef- had been attained, the following three criteria had to ficients of variation were 4.9% and 5.8%, respectively.
be met: 1) no further increase in O uptake with an increase Measurement of total antioxidant capacity (TAC).
in work rate (leveling-off criterion), 2) attainment of the TAC was measured in duplicate in plasma by chemolumi- age-predicted maximal heart rate, and 3) respiratory ex- nescence using a luminometer (Burthold, Autoluminat, LB953, U.S.). This assay is based on the ability of antioxi- Diet records. To examine whether dietary changes in-
dants present in the plasma to inhibit the oxidation of fluenced lipid peroxidation and antioxidant status outcomes 2,2'-azinobis (3-ethylbenzithiazoline) sulfonic acid (ABTS, (especially on TAC), 5-d diet recalls were completed before Sigma) to the radical cation ABTSϩ by a peroxidase (5).
and during training and detraining (once every 2 wk). A TAC was evaluated by a trolox (6-hydroxy-2,5,7,8-tetram- trained dietitian taught the subjects how to complete diet ethylchroman-2-carboxylic acid, Aldrich) standard curve, recall questionnaires and determine food serving and sizes.
and is expressed as trolox equivalent antioxidant capacity Diet records were analyzed using the computerized nutri- concentration (mmol·LϪ1). All samples were determined in tional analysis system Science Fit Diet 200A (Sciencefit, duplicate, while inter- and intra-coefficients of variation were 5.1% and 5.6%, respectively.
Blood sampling. Each subject reported to the labora-
Determination of glutathione peroxidase (GPX).
tory at 7:00 a.m. after an overnight fast on three separate Whole-blood GPX activity (U·LϪ1) was determined as pre- occasions (pretraining, posttraining, and 4 months after viously described (3). GPX activity was measured in a training cessation) for blood sampling. Subjects abstained Hitachi 2001 UV/VIS spectrophotometer (Hitachi Instru- from alcohol and caffeine consumption for at least 24 h, and ments Inc., U.S.) at 37°C. GPX activity was determined did not perform physical exercise for the last 48 h before using cumene hydroperoxide as the oxidant of glutathione testing. Peripheral blood samples were drawn with subjects (Ransel RS 505, Randox, Crumlin, UK). All samples were in a seated position. Blood samples were obtained from the determined in duplicate, while inter- and intra-coefficients antecubital region of the arm by Vacutainers containing of variation were 4.8% and 3.8%, respectively.
either SST Gel and Clot Activator, dipotassium ethylene Exercise training. Subjects in the control group did not
diamine tetraacetic acid (EDTA), or heparin as anticoagu- train, and participated only in the measurement procedures.
lants to obtain serum and plasma samples and immediately Subjects in ET exercised 3ϫ wkϪ1 for 16 wk. Subjects were placed on ice. Blood samples were centrifuged (Beck- always completed a 3–5 min warm-up, which consisted of man centrifuge, Fullerton, CA) at 4°C and 1500 ϫ g for 15 light walking at approximately 40% of their maximal heart min, and serum or plasma was obtained. These samples rate (attained during the exercise test), before starting their were stored frozen in multiple aliquots (ϳ0.25 mL in Ep- training routine. The maximum duration of each training microtubes) at Ϫ80°C until assayed. Samples session never exceeded 60 min, and training sessions were were thawed only once before analysis.
always supervised. The cardiovascular training protocol MDA measurement. Plasma MDA was determined
consisted of walking/jogging on a treadmill (Startrac TR with the method developed by Londero et al. (20). MDA 3500, Tustin, CA) as previously described (7). Briefly, was obtained by acid hydrolysis of 1,1,3,3,-tetrame- subjects walked/jogged at 50 – 80% of HR thoxypropane (TMP) as previously described (20). HPLC 12– 42 min each time (duration increased 2 min every week).
separation of MDA-TBA adducts was performed with a Blood arterial pressure and heart rate were monitored.
spectrofluorometric detector set at 532 nm excitation and Detraining. After the completion of the training period,
553 nm emission. The mobile phase consisted of 40:60 (v/v) a number of subjects (N ϭ 7) of the exercise group returned 0.05 M methanol-potassium phosphate buffer, pH 6.8; the to their pretraining physical activity levels becoming com- flow rate was 1.0 mL·minϪ1, with an average retention time pletely inactive (Table 1), whereas four of the subjects in of 1.7 min. A 10-␮m (250 ϫ 4.6 mm) particil SCX was used this group continued with their training regimens, and were for the separation (with a detection limit of 0.8 pmol per 30 excluded from detraining measurements.
␮L injected). TBA and TMP were purchased from Sigma Statistical analyses. Data were analyzed using the
Chemical Co. (Poole, Dorset, UK). All samples were deter- SPSS PC program for Windows. Values reported are means mined in duplicate, while inter- and intra-coefficients of Ϯ SD. One-way (grouping was the only factor) ANOVA variation were 7.9% and 8.8%, respectively.
was conducted initially to examine whether there were dif- Measurement of 3-nitro-L-tyrosine (3-NT). HPLC
ferences between groups in the premeasurement values of separation of 3-NT was achieved using an HPLC technol- each dependent variable. MANOVA repeated measures (3 ogy Nucleosil 5 ␮ C-18 column (250 ϫ 4.6 mm) with a ϫ 2, time by treatment) were performed on each dependent TRAINING AND OXIDATIVE STRESS IN THE ELDERLY Medicine & Science in Sports & Exerciseா variable to detect differences in each group for each time levels were significantly (P Ͻ 0.05) increased (40 – 45%) in point. When F-ratios were significant, post hoc comparisons both groups after exercise, independent of treatment or time of means were analyzed with Scheffe´’s multiple comparison of measurement. After training, C demonstrated similar tests. To compare treatments’ effectiveness in changing the MDA responses both at rest and postexercise. In contrast, E status of each dependent variable at each time point of demonstrated a significant (P Ͻ 0.1) decrease of MDA at measurement, a one-way ANOVA (with treatment as the rest (10%) and attenuation of MDA response to exercise independent factor) was applied on the delta differences (16%) posttraining. MDA levels returned to pretraining between different time point measurements. When F-ratios levels after 4 months of detraining.
were significant, post hoc comparisons of means were an- 3-Nintrotyrosine
responses. Values
alyzed with Scheffe´ tests. Statistical significance was ac- 3-NT are shown in Table 3 and were similar in both groups, both at rest and after GXT. Exercise induced a significant(P Ͻ 0.05) augmentation (Ͼ45%) of 3-NT, independent oftreatment and time of measurement. However, training elic- ited a significant (P Ͻ 0.05) reduction (20%) of 3-NT Subjects. Thirty-five men volunteered to participate.
concentration after GXT, but not at rest. 3-NT levels re- Eight men were excluded because they did not meet the turned to pretraining levels after 4 months of detraining.
selection criteria (two were too frail, four had medical Glutathione peroxidase (GPX) and total antioxi-
limitations, and two were too fit), whereas three declined dant capacity (TAC). Tables 4 and 5 present GPX and
participation. During training, three more men (one from C TAC adaptations, respectively. TAC did not differ between and two from ET) were asked to stop because they had groups, either at rest or after exercise at pretraining. Exer- missed more than three training sessions up to that point, cise caused a substantial (P Ͻ 0.05) increase (Ͼ9%) of whereas two more (one from C and one from ET) stopped TAC, independent of treatment and time of measurement.
because of injury or illness (all stopped within the first 4 Endurance training elicited a significant increase of TAC at wk). There were no differences between individuals who rest (6%, P Ͻ 0.1) and postexercise (13%, P Ͻ 0.05).
dropped out of the study and those who completed the study However, detraining returned TAC values at baseline. GPX activity was significantly (P Ͻ 0.05) increased (6%) by groups were required to complete 48 training sessions exercise, independent of treatment and time of measure- (100% compliance was achieved). In the detraining mea- ment. GPX activity demonstrated a significant (P Ͻ 0.05) surements, only seven subjects from the exercise group and increase (12%) after GXT posttraining. There were no dif- six subjects from the control group participated. All partic- ferences between groups for GPX activity at pretraining.
ipants demonstrated a low level of maximal oxygen con- Both TAC and GPX activity levels returned to pretraining Ͻ 20 mL·kgϪ1·minϪ1), with no differ- levels after 4 months of detraining.
ences detected between groups (Table 1).
Subjects’ physical characteristics are shown in Table 1.
DISCUSSION
No significant differences were noted at baseline betweengroups for age, height, and fitness level at baseline of the The present investigation presents evidence that endur- study and after 4 months of detraining. There were also no ance exercise training may attenuate exercise-induced oxi- differences between groups at any time point relative to dative stress in older individuals while simultaneously en- dietary intake (antioxidant, nutrient, and fat consumption).
hancing antioxidant defense, both at rest and after exercise.
Body weight and sum of skinfolds were similar across A 16-wk progressive training program increased partici- groups at baseline (Table 1). The endurance-training pro- pants’ fitness level, reduced their body fat content, and gram resulted in a significant (P Ͻ 0.05) reduction in body enhanced their ability to defend against reactive oxygen and weight (6.6%) and sum of skinfolds (6%) (Table 1) at the nitrogen species. Results of the present investigation suggest end of training, compared with baseline measurements. Af- that systematic exercise may offer a protective mechanism ter 4 months of detraining, body weight and skinfold sum against oxidative damage as well as enhancement of func- tional performance of aged men. However, it is likely that Aerobic endurance. Subjects in both groups exhibited
a very low initial level of aerobic fitness (V mL·kgϪ1·minϪ1), with no significant differences between TABLE 2. MDA (␮M) levels at rest and after a GXT after a period of endurance groups at baseline (Table 1). ET demonstrated a significant training and detraining in older men.
increase (45%) in time to exhaustion, whereas V Exercise
increased by 26.3% by the end of training (Table 1, P Ͻ 0.05). C group showed no change in these two variables.
tion all returned to pretraining levels.
Values are means Ϯ SD; MDA, malondialdehyde; C, control group; ET, endurance- MDA responses. Lipid peroxidation data are shown in
training group; P Ͻ .05 vs pre values.
* Significant difference from corresponding resting value.
Table 2. Pretraining, MDA levels were similar in both † Significant difference between control and exercise group at a given time point.
groups at rest and after the GXT for both groups. MDA # Significant difference as compared with pretraining value.
Official Journal of the American College of Sports Medicine TABLE 3. 3-NT (␮M) levels at rest and after a GXT after a period of endurance TABLE 5. Total antioxidant capacity (␮M Trolox) at rest and after a GXT after a training and detraining in older men.
period of endurance training and detraining in older men.
Exercise
Exercise
Values are means Ϯ SD; 3-NT, 3-nitrotyrosine; C, control group; ET, endurance training Values are means Ϯ SD. C, control group; ET, endurance training group; P Ͻ 0.05 vs group; P Ͻ 0.05 vs pre values.
* Significant difference from corresponding resting value.
* Significant difference from corresponding resting value.
† Significant difference between control and exercise group at a given time point.
† Significant difference between control and exercise group at a given time point.
# Significant difference as compared with pretraining value.
# Significant difference as compared to pretraining value.
cessation of training reverses training-induced adaptations ing markers associated with oxidative stress (22). In the of aerobic fitness as well as oxidative stress and antioxidant present investigation, exhausting exercise increased lipid peroxidation regardless of training. This is in accordance Enhanced fitness levels with endurance training.
with previous findings indicating that acute aerobic- or Subjects in the present investigation demonstrated a signif- resistance-type exercise is capable of inducing an increase of lipid peroxidation in both young and old subjects (34).
exhaustion (ϳ45%), indicating a substantial increase of Subjects in the present study demonstrated higher basal their aerobic capacity. Previous reports have reported en- MDA values compared with previously reported values for durance-training–induced increases of V older adults (34), but this could be attributed to participants’ 10 to 25%, even in individuals 70 – 80 yr old who had never greater age and their lower fitness level.
exercised before (10). Furthermore, subjects’ body compo- Reduced lipid peroxidation levels were observed in the sition was significantly altered because their skinfold sum endurance-trained group. Previous research has produced was reduced (ϳ6%), and this was paralleled by a similar controversial results regarding exercise-training effects on response of their body weight as previously described (16).
basal lipid peroxidation levels. It was shown that systematic Reduction of lipid peroxidation at rest and pos-
exercise training decreases resting lipid peroxidation levels texercise. Recent studies employing molecular genetics
(18). Others argued that even a long-term, high-intensity and pharmacological interventions strongly suggest that aerobic training program (ranging from 60 to 90 min·dϪ1, ROS may modulate the aging process, and that endogenous with a frequency of 5 d·wkϪ1) is not associated with low- production of ROS by normal physiological processes is ered basal lipid peroxidation levels (34). Resistance training probably a significant obstacle to lifespan (21). A number of with weights was unable to decrease basal lipid peroxidation studies have shown an age-related increase of lipid peroxi- in older adults (33). It is not certain whether exercise train- dation in several tissues (18) associated with mitochondrial ing mainly lowers baseline levels of lipid peroxidation or ROS production, such as hydrogen peroxide (30). The mi- decreases exercise-induced lipid peroxidation. However, in tochondrial hypothesis of aging suggests that free radical this study unfit older individuals were used instead of ani- generation in the mitochondria produce free radical reac- mals, whereas endurance training is known to cause far tions that damage specific vital macromolecules in this more significant metabolic adaptations in mitochondria and organelle (11). Therefore, mitochondria are the main source aerobic metabolism than resistance training, which was used and target of cellular free radicals. An increase in free in previous studies. Most previous studies trained younger radical production or a decrease in antioxidant enzyme participants, with intensity, duration, and training volume content leads to oxidative stress and cellular dysfunction.
varying among them. It has been suggested that exercise Furthermore, the activity of certain enzymes (NADH-dehy- training lowers resting lipid peroxidation by upregulating drogenase and cytochrome oxidase) located in the inner antioxidant enzyme levels in tissues engaged in systematic membrane of the mitochondria, as well as other mitochon- exercise (23). However, it must be kept in mind that this drial proteins (i.e., adenine nucleotide translocase, acyl car- study recorded only a tendency toward lower lipid peroxi- nitine transferase, and citrate synthase), are considered ag- dation, and that additional data are needed in order to beconclusive about this.
TABLE 4. GPX activity (U⅐LϪ1) changes at rest and after a GXT after a period of Endurance training for 16 wk increased aerobic capacity endurance training and detraining in older men.
and decreased exercise-induced lipid peroxidation. Exer- Exercise
cise-trained men may be able to produce fewer ROS com- pared with sedentary controls, because endurance training has been shown to upregulate mitochondrial respiratory chain proteins, leading to a reduced electron flux through each electron transport chain, electron leakage, and the Ϯ SD; GPX, glutathione peroxidase; C, control group; ET, endurance training group; P Ͻ 0.05 vs pre values.
resulting radical formation (4). During aerobic exercise, * Significant difference from corresponding resting value.
mitochondria are in state 3. The reduction of the respiratory † Significant difference between control and exercise group at a given time point.
# Significant difference as compared with pretraining value.
chain is substantially limited during the state 4 to 3 transi- TRAINING AND OXIDATIVE STRESS IN THE ELDERLY Medicine & Science in Sports & Exerciseா tion, while at the same time electron flow through the ONOOϪ effect on modified biological tissue, such as mus- respiratory chain is increased (13). Lower levels of reactive intermediates produced in state 3 may offer a protective Aged skeletal muscle’s increased relaxation time has mechanism against oxidative stress by enhancing electron been correlated with a potential age-related dysfunction of coupling (17). Improved coupling of electron transport and the sarcoplasmic reticulum (SR) Ca-ATPase (35). Previous oxygen reduction to water in state 3 may constitute a sig- research has shown an in vivo presence of ONOOϪ in nificant mechanism for controlling free radical production, skeletal muscle and a significantly greater 3-NT content in because skeletal muscle raises its oxygen consumption up to the SERCA2a slow-twitch isoform of the Ca-ATPase in 20-fold during its transition from rest to exercise. However, aged muscle that could serve as an indication of aged- more work is needed to support the claim that upregulation associated increase in susceptibility to oxidation by species of electron transport chain complexes reduces oxidant for- such as ONOOϪ (35). 3-NT increase in aged skeletal muscle mation. Furthermore, mitochondria may produce other ox- has been observed both during relaxation and contraction of idants such as nitric oxide (17). Mitochondrial nitric oxide muscle through simultaneous generation of O and superoxide react to form peroxynitrate (a strong oxi- (35). Posttranslational modification of proteins by ROS/ dant), making mitochondria a significant source of harmful NOS has been recognized as an important feature of bio- oxidants during aerobic exercise (32). In this work, 3-NT (a logical oxidative stress (35), as significant levels of such O–mediated tissue damage) was significantly modified proteins may accumulate in aged biological tissue reduced by endurance exercise training. These adaptations (35), accompanied by modified enzyme activities caused by are very important to older individuals, as aging seems to covalent protein modification. Aged skeletal muscle would increase mitochondria-generated free radicals (23). Findings be exposed to ONOOϪ, as indicated by the presence of a in this study agree with previous observations that suggest significant amount of 3-NT, caused by simultaneous gener- that chronic exercise training may decrease MDA levels in brain mitochondria, indicating a beneficial adaptation of In the present study, acute intense exercise increased exercise training on oxidative stress development (19). An 3-NT, independent of treatment. In a limited number of improvement of antioxidant status by training, as seen in the previous studies, 3-NT has been found to increase in urine, present investigation, may attenuate ROS production in serum, and the liver by acute exhausting exercise (26). This finding may reflect the presence of oxidative stress and 3-NT response to training. Nitric oxide and its de-
augmentation of serum proteins’ nitration and carbonylation rivatives are detectable in both intracellular and extracellu- during exercise. In contrast with MDA, 3-NT was not re- duced at rest after training. Previous studies that used animal synthase (NOS) is upregulated (50 –200%) during muscular subjects revealed either an increase or no change in 3-NT O, its derivatives, and ROS demonstrate sim- after training (31). However, endurance training was able to ilar production and distribution patterns in muscle fiber decrease exercise-induced 3-NT response after exhausting cytosol and extracellular space (28). N ˙ exercise, indicating a lowering effect of NOS production muscle is likely to be affected by ROS because N ˙ and protein oxidation. It seems that systematic endurance undergoes electron exchange reactions with ROS as they training offers a protective mechanism against 3-NT accu- both compete for the same redox-sensitive molecular tar- mulation during exercise in inactive older individuals. The biochemical mechanisms underlying this type of adaptation sulting in peroxynitrite (ONOOϪ) synthesis (28). Due to the Effects of endurance training on antioxidant sta-
exercise, we expect that skeletal muscle will be systemati- tus. It has been postulated that aging causes increased
cally exposed to ONOOϪ. 3-NT has been identified as a protein breakdown through oxidative damage and selective stable end product formed upon reaction of free or protein- protein degradation, along with reduced protein synthesis, bound tyrosine with NO , such as ONOOϪ (28). Because of gradually leading to decreased antioxidant enzyme levels in its stability, and because several studies have shown a dose- aging tissue, and especially in skeletal muscles (21). Endur- dependent increase of 3-NT in serum proteins in experimen- ance training can restore the age-associated reduction of tal mammals treated with tetranitromethane (a specific pro- muscle protein content, as well as mitochondrial oxidative tein nitrating agent), this molecule has been proposed as a capacity (8). Results of the present study indicate an in- creased TAC at rest and enhanced TAC and GPX activity the assessment of the exposure of tissue to oxidative stress levels postexercise after training. These findings agree with by NO in general (5). A recent report further underlines the previous reports (14), suggesting a significant increase of practical implications of the 3-NT measurement by indicat- GPX activity in rats, although others failed to observe such ing that nitrotyrosine levels are related to coronary artery an adaptation (18). The discrepancy seen in these studies disease (CAD) and may be regulated by statin treatment, may be largely due to differences in the training threshold suggesting a role for nitric oxide oxidants as inflammatory employed. Earlier work with younger subjects support the mediators in CAD, with implications in atherosclerosis risk notion that systematic training induces positive adaptations assessment (29). 3-NT could be a potential marker of in antioxidant enzyme activities (6,23) associated with in- Official Journal of the American College of Sports Medicine creased protein content and mRNA levels in muscle, and exercise-induced adaptations (and to what extent), and to this adaptation may be fiber-type specific (23). GPX has determine which training protocol is more effective in main- been consistently found to increase by training (18,23) in taining these adaptations. This preliminary evidence on de- several muscles conducted mostly on younger participants.
training suggests that exercise effects in older individuals Because GPX facilitates H O and lipid peroxides removal may be temporary, and that uninterrupted exercise is crucial produced in the mitochondrial inner membrane, its increase to maintain any positive adaptations. This study examined with training coincides with the reduction in oxidative stress only a 4-month detraining period on oxidative stress re- attenuation seen in this study, both at rest and after exercise.
sponse. Well-designed studies are needed to investigate Most of these adaptations were seen in studies using shorter and longer periods of detraining.
younger or middle-aged individuals, and only a few studies In summary, 4 months of systematic endurance exercise (34) have been conducted with aging populations. Future training lowered oxidative stress levels, whereas it increased research should examine the adaptations of the antioxidant antioxidant status in inactive older individuals. Prolonged system after prolonged training in older humans.
training cessation abolished these effects. The decrease in Detraining responses. After 4 months of cessation of
oxidative stress capacity, the enhanced total antioxidant the training stimulus, all training-induced adaptations were capacity, and the improvements in body composition and completely reversed, with oxidative stress and antioxidant cardiorespiratory endurance could have important health status markers returning to pretraining values. There are no previous reports on detraining effects in the elderly, and thusit is difficult to be conclusive regarding this aspect. More The authors would like to thank Mrs. Thalia Mageiria for her studies are needed to examine whether detraining attenuates REFERENCES
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