Muscle protein changes following eccentric exercise in humans

This study characterized changes in the protein composition of human muscle tissue after eccentric exercise. Four subjects performed 70 maximum eccentric, isokinetic actions of the forearm flexors with one arm. The other arm served as control. A biopsy of the biceps muscle of each arm was taken 2 days after exercise when muscles were very sore (mean = 8.0; 1 = normal; 10 = very, very sore), and muscle damage was documented by a mean decrease of 0.2 radians in the relaxed elbow angle. Proteins from the biopsy tissue were solubilized in a high ionic strength buffer containing several proteolytic inhibitors. Protein concentrations of the extracts were determined and identical amounts loaded onto sodium dodecyl sulfate (SDS) polyacrylamide gels (7.5, 12.5, and 17.5%). Densitometric analysis of the Coomassie brilliant blue stained gels revealed alterations in the amounts of three protein bands in the exercised tissue relative to the control. These changes were in the linear portion of the graph of absorbance versus protein amount. Wilcoxon's signed rank test showed the first two of the following bands to increase significantly in amount (P less than 0.062). The average percentage changes [mean (SEM)] for these bands were 63 (21), 39 (5), and 82 (35). The corresponding molecular weights determined from known standards were 76300 (860), 33200 (310), and 12000 (80) daltons, respectively. These changes imply that the increased synthesis, decreased degradation, or some combination thereof, of these three proteins may be necessary for the repair or regeneration response to exercise-induced muscle damage.     

Expression of tissue remodelling,inflammation- and angiogenesis-related factors after eccentric exercise in humans

Molecular Biology Reports - Eccentric exercise has been extensively used as a model to study the contraction-induced muscle damage and its consequent processes. This study aimed at examining...     

Gender differences in muscle inflammation after eccentric exercise.

Unaccustomed exercise is followed by delayed-onset muscle soreness and morphological changes in skeletal muscle. Animal studies have demonstrated that women have an attenuated response to muscle damage. We studied the effect of eccentric exercise in untrained male (n = 8) and female (n = 8) subjects using a unilateral exercise design [exercise (Ex) and control (Con) legs]. Plasma granulocyte counts [before (Pre) and 48 h after exercise (+48h)] and creatine kinase activity [Pre, 24 h after exercise (+24h), +48h, and 6 days after exercise (+6d)] were determined before (Pre) and after (+24h, +48h, +6d) exercise, with biopsies taken from the vastus lateralis of each leg at +48h for determination of muscle damage and/or inflammation. Plasma granulocyte counts increased for men and decreased for women at +48h (P < 0.05), and creatine kinase activity increased for both genders at +48h and +6d (P < 0.01). There were significantly greater areas of both focal (P < 0.001) and extensive (P < 0.01) damage in the Ex vs. Con leg for both genders, which was assessed by using toluidine blue staining. The number of leukocyte common antigen-positive cells/mm(2) tissue increased with exercise (P < 0.05), and men tended to show more in their Ex vs. Con leg compared with women (P = 0.052). Men had a greater total (Ex and Con legs) number of bcl-2-positive cells/mm(2) tissue vs. women (P < 0.05). Atrophic fibers with homogeneous bcl-2-positive staining were seen only in men (n = 3). We conclude that muscle damage is similar between genders, yet the inflammatory response is attenuated in women vs. men. Finally, exercise may stimulate the expression of proteins involved in apoptosis in skeletal muscle.     

Cytokine response to eccentric exercise in young and elderly humans

To examine the plasma interleukin(IL)-6 response in elderly (E) and young (Y) humans, 10 E and 10 Ysubjects completed 60 min of eccentric lower limb exercise at the samerelative oxygen uptake. Plasma IL-6 was measured before, immediatelyafter, and 5 days into recovery from exercise, as were the biochemicalmarkers of muscle damage, creatine kinase (CK), and myoglobin. In both groups, IL-6 increased (P < 0.05) immediately afterexercise and peaked 4 h after exercise at 4.35 ± 1.7 vs.5.05 ± 3.17 pg/ml for E and Y subjects, respectively. However,the increase in IL-6 in both groups was modest relative to theincreases in CK peaking at 539 ± 413 vs. 10,301 ± 5,863 U/lfor E and Y subjects, respectively. In addition, the increase in IL-6was less pronounced (P < 0.05) in E subjects comparedwith Y subjects. These results suggest that IL-6 increasesprogressively after eccentric exercise, suggesting that this increaseis related to muscle damage. However, the modest increase in IL-6,despite large increases in CK, suggests that the IL-6 response tomuscle damage does not make an important contribution to the largeincrease in IL-6 observed during concentric exercise of long duration.Our data also suggest that aging may be associated with impaired repairmechanisms for exercise-induced muscle damage.

     

Time course of glycogen accumulation after eccentric exercise.

This study examined the time course of glycogen accumulation in skeletal muscle depleted by concentric work and subsequently subjected to eccentric exercise. Eight men exercised to exhaustion on a cycle ergometer [70% of maximal O2 consumption (VO2max)] and were placed on a carbohydrate-restricted diet. Approximately 12 h later they exercised one leg to subjective failure by repeated eccentric action of the knee extensors against a resistance equal to 120% of their one-repetition maximum concentric knee extension force (ECC leg). The contralateral leg was not exercised and served as a control (CON leg). During the 72-h recovery period, subjects consumed 7 g carbohydrate.kg body wt-1.day-1. Moderate soreness was experienced in the ECC leg 24-72 h after eccentric exercise. Muscle biopsies from the vastus lateralis of the ECC and CON legs revealed similar glycogen levels immediately after eccentric exercise (40.2 +/- 5.2 and 47.6 +/- 6.4 mmol/kg wet wt, respectively; P greater than 0.05). There was no difference in the glycogen content of ECC and CON legs after 6 h of recovery (77.7 +/- 7.9 and 85.1 +/- 4.9 mmol/kg wet wt, respectively; P greater than 0.05), but 18 h later, the ECC leg contained 15% less glycogen than the CON leg (90.2 +/- 8.2 vs. 105.8 +/- 8.9 mmol/kg wet wt; P less than 0.05). After 72 h of recovery, this difference had increased to 24% (115.8 +/- 8.0 vs. 153.0 +/- 12.2 mmol/kg wet wt; P less than 0.05). These data confirm that glycogen accumulation is impaired in eccentrically exercised muscle.(ABSTRACT TRUNCATED AT 250 WORDS)     

Skeletal muscle collagen content in humans after high-force eccentric contractions.

The purpose of this study was to investigate the effects of high-force eccentric muscle contractions on collagen remodeling and on circulating levels of matrix metalloproteinases (MMP) and tissue inhibitors of metalloproteinases (TIMP) in humans. Nine volunteers [5 men and 4 women, mean age 23 (SD 4) yr] each performed a bout of 100 maximum voluntary eccentric contractions of the knee extensors. Muscle biopsies were taken before exercise and on days 4 and 22 afterward. Image analysis of stained tissue sections was used to quantify endomysial collagen staining intensity. Maximum voluntary contractile isometric force was recorded preexercise and on days 1, 2, 3, 4, 8, 11, and 14 postexercise. Venipuncture blood samples were also drawn on these days for measurement of serum creatine kinase activity and concentrations of MMP-9, TIMP-1, TIMP-2, and the MMP-2/TIMP-2 complex. Maximum voluntary contractile force declined by 39 +/- 23% (mean +/- SD) on day 2 postexercise and recovered thereafter. Serum creatine kinase activity peaked on day 4 postexercise (P < 0.01). Collagen type IV staining intensity increased significantly on day 22 postexercise to 126 +/- 29% (mean +/- SD) of preexercise values (P < 0.05). Serum MMP-9 levels increased on day 8 postexercise (P < 0.01), and serum TIMP-1 was also significantly elevated on days 1, 2, 3, 4, and 14 postexercise (P < 0.05). These results suggest that a single bout of eccentric muscle contractions results in remodeling of endomysial type IV collagen, possibly via the MMP pathway.     

Impaired substrate oxidation in healthy elderly men after eccentric exercise.

The metabolic response to eccentric exercise in healthy older adults is unknown. Therefore, substrate metabolism was examined in the basal state and after sustained hyperglycemia (180 min, 10 mM) in eight healthy, sedentary older [66 +/- 2 yr; body mass index (BMI) of 25.5 +/- 1.2 kg/m] and nine younger (23 +/- 1 yr; BMI of 23.6 +/- 1.7 kg/m) men, under control conditions and 48 h after eccentric exercise. Indirect calorimetry was performed to evaluate carbohydrate and lipid oxidation (C(ox) and L(ox), respectively). Eccentric exercise caused muscle soreness and increased plasma creatine kinase in both groups of men (P < 0.02). Although a similar level of hyperglycemia was maintained in the two groups, glucose infusion rates were lower (P < 0.001) in the older men. Compared with basal levels, hyperglycemia stimulated an increase in C(ox) and a decrease in L(ox) during the control and exercise trials in the younger group (P < 0.03), but only during the control trial in the older subjects (P < 0.007). C(ox) was unchanged after eccentric exercise in the younger men [4.00 +/- 0.30 vs. 3.54 +/- 0.44 mg x kg fat-free mass (FFM)(-1) x min(-1); exercise vs. control] but was suppressed by 20% in the older group (3.37 +/- 0.37 vs. 4.21 +/- 0.23 mg x kg FFM(-1) x min(-1); P < 0.04). Moreover, L(ox) was reduced by 38% in the younger subjects (0.47 +/- 0.09 vs. 0.76 +/- 0.10 mg x kg FFM(-1) x min(-1); P< 0.03) but was augmented by 89% in the older group (0.68 +/- 0.11 vs. 0.36 +/- 0.08 mg x kg FFM(-1) x min(-1); P < 0.04). In addition, hyperglycemia-stimulated C(ox), L(ox), and respiratory exchange ratio responses to eccentric exercise were related to abdominal adiposity (r = -0.57, P < 0.04, r = 0.68, P < 0.02 and r = -0.60, P < 0.02, respectively). Despite normal glucose tolerance and the absence of obesity per se, older men experience a reduction in carbohydrate oxidation in response to hyperglycemia after eccentric exercise.     

Impaired muscle glycogen resynthesis after eccentric exercise

Eight men performed 10 sets of 10 eccentric contractions of the knee extensor muscles with one leg [eccentrically exercised leg (EL)]. The weight used for this exercise was 120% of the maximal extension strength. After 30 min of rest the subjects performed two-legged cycling [concentrically exercised leg (CL)] at 74% of maximal O2 uptake for 1 h. In the 3 days after this exercise four subjects consumed diets containing 4.25 g CHO/kg body wt, and the remainder were fed 8.5 g CHO/kg. All subjects experienced severe muscle soreness and edema in the quadriceps muscles of the eccentrically exercised leg. Mean (+/- SE) resting serum creatine kinase increased from a preexercise level of 57 +/- 3 to 6,988 +/- 1,913 U/l on the 3rd day of recovery. The glycogen content (mmol/kg dry wt) in the vastus lateralis of CL muscles averaged 90, 395, and 592 mmol/kg dry wt at 0, 24, and 72 h of recovery. The EL muscle, on the other hand, averaged 168, 329, and 435 mmol/kg dry wt at these same intervals. Subjects receiving 8.5 g CHO/kg stored significantly more glycogen than those who were fed 4.3 g CHO/kg. In both groups, however, significantly less glycogen was stored in the EL than in the CL.     

Increase in blood bradykinin concentration after eccentric weight-training exercise in men.

The purpose of this study was to verify the possible appearance in the blood of bradykinin (BK) and des-Arg(9)-bradykinin (des-Arg(9)-BK) after eccentric exercise in 13 male subjects. Eccentric exercise (5 x 10 leg presses at 120% maximal voluntary concentric contraction) resulted in muscle damage and inflammation, as suggested by the significant increase in serum creatine kinase activity (from 204 +/- 41 to 322 +/- 63 U/l 12 h postexercise) and by severe lasting pain, which also peaked at 12 h postexercise. Blood BK and des-Arg(9)-BK concentrations were measured by competitive enzyme immunoassays using highly specific polyclonal rabbit IgG. Des-Arg(9)-BK concentration was not modified (preexercise: 44 +/- 14 pmol/l; pooled postexercise: 47 +/- 4 pmol/l). In contrast, BK concentration significantly increased immediately after the exercise session (68 +/- 9 vs. 42 +/- 3 pmol/l preexercise) and returned to basal values at 12, 24, and 48 h (pooled value: 40 +/- 4 pmol/l). This observation suggests that the inflammatory process due to eccentric exercise-induced muscle damage could be mediated in part by BK.     

Supplementation with vitamin C and N-acetyl-cysteine increases oxidative stress in humans after an acute muscle injury induced by eccentric exercise

There has been no investigation to determine if the widely used over-the-counter, water-soluble antioxidants vitamin C and N-acetyl-cysteine (NAC) could act as pro-oxidants in humans during inflammatory conditions. We induced an acute-phase inflammatory response by an eccentric arm muscle injury. The inflammation was characterized by edema, swelling, pain, and increases in plasma inflammatory indicators, myeloperoxidase and interleukin-6. Immediately following the injury, subjects consumed a placebo or vitamin C (12.5 mg/kg body weight) and NAC (10 mg/kg body weight) for 7 d. The resulting muscle injury caused increased levels of serum bleomycin-detectable iron and the amount of iron was higher in the vitamin C and NAC group. The concentrations of lactate dehydrogenase (LDH), creatine kinase (CK), and myoglobin were significantly elevated 2, 3, and 4 d postinjury and returned to baseline levels by day 7. In addition, LDH and CK activities were elevated to a greater extent in the vitamin C and NAC group. Levels of markers for oxidative stress (lipid hydroperoxides and 8-iso prostaglandin F2alpha; 8-Iso-PGF2alpha) and antioxidant enzyme activities were also elevated post-injury. The subjects receiving vitamin C and NAC had higher levels of lipid hydroperoxides and 8-Iso-PGF2alpha 2 d after the exercise. This acute human inflammatory model strongly suggests that vitamin C and NAC supplementation immediately post-injury, transiently increases tissue damage and oxidative stress.     

MR measurements of muscle damage and adaptation after eccentric exercise.

The purposes of this study were, first, to clarify the long-term pattern of T2 relaxation times and muscle volume changes in human skeletal muscle after intense eccentric exercise and, second, to determine whether the T2 response exhibits an adaptation to repeated bouts. Six young adult men performed two bouts of eccentric biceps curls (5 sets of 10 at 110% of the 1-repetition concentric maximum) separated by 8 wk. Blood samples, soreness ratings, and T2-weighted axial fast spin-echo magnetic resonance images of the upper arm were obtained immediately before and after each bout; at 1, 2, 4, 7, 14, 21, and 56 days after bout 1; and at 2, 4, 7 and 14 days after bout 2. Resting muscle T2 [27.6 +/- 0.2 (SE) ms] increased immediately postexercise by 8 +/- 1 ms after both bouts. T2 peaked 7 days after bout 1 at 47 +/- 4 ms and remained elevated by 2.5 ms at 56 days. T2 peaked lower (37 +/- 4 ms) and earlier (2-4 days) after bout 2, suggesting an adaptation of the T2 response. Peak serum creatine kinase values, pain ratings, and flexor muscle swelling were also significantly lower after the second bout (P < 0.05). Total volume of the imaged arm region increased transiently after bout 1 but returned to preexercise values within 2 wk. The exercised flexor compartment swelled by over 40%, but after 2 wk it reverted to a volume 10% smaller than that before exercise and maintained this volume loss through 8 wk, consistent with partial or total destruction of a small subpopulation of muscle fibers.     

Potential biomarkers of muscle injury after eccentric exercise

Proteomics was utilized to identify novel potential plasma biomarkers of exercise-induced muscle injury. Muscle injury was induced in nine human volunteers by eccentric upper extremity exercise. Liquid chromatography–mass spectrometry identified 30 peptides derived from nine proteins which showed significant change in abundance post-exercise. Four of these proteins, haemoglobin α chain, haemoglobin β chain, α1-antichymotrypsin (ACT) and plasma C-1 protease inhibitor (C1 Inh), met the criterion for inclusion based on changes in at least two distinct peptides. ACT and C1 Inh peptides peaked earlier post-exercise than creatine kinase, and thus appear to provide new information on muscle response to injury.     

Protein metabolism in rat tibialis anterior muscle after stimulated chronic eccentric exercise.

In another study (J. Appl. Physiol. 69: 1709-1717, 1990) we reported that gastrocnemius (GAST) muscle enlargement failed to occur after 10 wk of 192 contractions performed every 3rd or 4th day. This result was surprising because increased protein synthesis rates were determined after an initial acute exercise bout with the same paradigms. In the same set of animals, tibialis anterior (TA) muscles were enlarged 16-30% compared with sedentary control muscles after the same chronic training regimen. This indicated that the regulation of protein expression may be different between the GAST and TA muscles. The present experiment attempted to define and explain these differences by comparing changes in various indexes of protein metabolism in TA with the same parameters determined in the accompanying study for the GAST. As in the GAST, results showed that TA protein synthesis rates are increased by acute exercise and principally regulated by translational and possibly posttranslational mechanisms. The differential response in muscle mass between the GAST and TA muscles after training may be due, in part, to greater relative resistances imposed on the TA than on the GAST that result in a more-prolonged effect on protein synthesis rates, with lower numbers of stimulated contractions required to stimulate increases in protein synthesis. Data also revealed that although as little as 1 min of total contractile duration (24 repetitions) increased TA protein synthesis rate by 30%, 8 min of total contractile duration (192 repetitions) further increased TA protein synthesis rates to only 45% above control.     

Low-frequency depression of tension in the cat gastrocnemius muscle after eccentric exercise.

Subjecting a muscle to a series of eccentric contractions in which the contracting muscle is lengthened results in a number of changes in its mechanical properties. These include a fall in isometric tension that is particularly pronounced during low-frequency stimulation, a phenomenon known as low-frequency depression (LFD). Reports of LFD have not taken into account the shift in optimum length for active tension generation to longer muscle lengths that takes place after eccentric contractions. Given the length dependence of the stimulation frequency-tension curve, we tested the hypothesis that the change in this relationship after eccentric exercise is due to the shift in optimum length. We measured LFD by recording tension in response to a linearly increasing rate of stimulation of the nerve to medial gastrocnemius of anesthetized cats, over the range 0-100 pulses per second. Tension responses were measured before and after 50 eccentric contractions consisting of 6-mm stretches starting at 3 mm below optimum length and finishing at 3 mm above it. An index of LFD was derived from the tension responses to ramp stimulation. It was found that LFD after the eccentric contractions was partly, but not entirely, due to changes in the muscle's optimum length. An additional factor was the effect of fatigue. These observations led to the conclusion that the muscle length dependence of LFD was reduced by eccentric contractions. All of this means that after eccentric exercise the tension deficit at low rates of muscle activation is likely to be less severe than first thought.     

Sex differences in oxidative stress after eccentric and concentric exercise

Objectives: Comparison of redox balance changes in the blood of women and men as a result of submaximal eccentric (ECC) and concentric (CONC) efforts.

Methods: 10 women and 10 men performed three 45-minute submaximal treadmill runs at constant velocities (downhill run – ECC, uphill run – CONC and level run). Prior to the 45-minute exercises, after their completion and following 24 hours of recovery, the concentration of lactate, oxidized low-density lipoprotein (ox-LDL), 3-nitrotyrosine, uric acid (UA) and the white blood cell count (WBC), neutrophil (NEUT), lymphocyte (LYMPH) and monocyte content in the blood were determined.

Results: In women, the ox-LDL increased significantly 10 minutes and 24 hours following ECC (P?P?P?P?Discussion: ECC cause impaired redox balance only in women. Due to the increase in antioxidant capacity of the blood without accompanying oxidative damage to macromolecules, for both sexes, it is recommended to perform concentric running efforts at the highest possible subliminal intensity.     

Normal forces and myofibrillar disruption after repeated eccentric exercise

Hortobágyi, Tibor, Joseph Houmard, David Fraser,Ronald Dudek, Jean Lambert, and James Tracy. Normalforces and myofibrillar disruption after repeated eccentric exercise.J. Appl. Physiol. 84(2): 492-498, 1998.To investigate the "rapid-adaptation" phenomenon, weexamined force, neural, and morphological adaptations in 12 subjectswho performed 100 eccentric contractions with the quadriceps muscle(bout 1) and repeated the sameexercise after a 2-wk hiatus (bout2). Two days after bout1, quadriceps muscle strength and surfaceelectromyographic (EMG) activity declined ~37 and 28%, respectively,in the control group (n = 6). Atday 2 after bout 1, significant increases occurred in patellar tendonreflex amplitude (~25%), muscle soreness (fivefold), and serumcreatine kinase (220%), and 65 ± 12% of the total number of pixelsin the EMG indicated myofibrillar disruption. At day7 after bout 1, all variables returned to normal. At day 2 after bout 2, no significant changesoccurred in force, EMG, creatine kinase, or soreness, but reflexamplitude increased, and 23 ± 4% of the total number of pixels inthe EMG still indicated myofibrillar disruption. The results suggestthat the rapid force recovery following eccentric exercise is mediatedat least in part by neural factors and that this recovery may occurindependently of cell disruption.

     

Appetite and blood glucose profiles in humans after glycogen-depleting exercise.

Regulatory functions of glycogen stores and blood glucose on human appetite, particularly relating to exercise, are not fully understood. Ten men (age 20-31 yr) performed glycogen-depleting exercise in an evening, ate a low-carbohydrate dinner, and stayed overnight in the laboratory. The next day, blood glucose was monitored continuously for 517 +/- 23 (SE) min. Subjects had access to high-fat and high-carbohydrate foods after baseline glucose and respiratory quotient were determined. In the afternoon, 1 h of moderate exercise was performed. Baseline respiratory quotient was 0. 748 +/- 0.008, plasma free fatty acids were 677 +/- 123 micromol/l, insulin was 4.8 +/- 0.5 microU/ml, and leptin was 1.9 +/- 0.3 ng/ml. Postabsorptively, 8 of 10 meals were initiated during stability in blood glucose. Postprandially, the association between meal initiation and blood glucose declines became significant (chi(2) = 7. 82). During moderate exercise, blood glucose initially decreased but recovered before completion. When the glycogen buffer is depleted, meal initiation can occur during blood glucose stability; the relationship between blood glucose declines and meal initiation reestablishes with refeeding.     

Work rate-dependent lactate kinetics after exercise in humans

Arterial blood lactate concentrations were measured on 19 subjects before, during, and after a 3-min bicycle exercise at several work rates, and the concentrations during the recovery phases were fitted to a biexponential time function consisting of a rapidly increasing and a slowly decreasing component. Highly significant correlations with the work rate of the exercise preceding the recovery were found for all the parameters of the fitted equation. The two velocity constants show inverse linear relationships, whereas the other parameters vary according to a definite power function. A functional meaning has been given to the two velocity constants, namely the ability of the tissues to exchange and to remove lactate. For the group of subjects studied, after exercises at work rates below about 3.5 W/kg, the tissue's ability to utilize, and possibly to exchange lactate, increases over values generally reported for resting conditions, whereas after exercises at higher work rates the inverse occurs. Lactate kinetics during recovery appear to be the result of two underlying processes, one enhancing the ability of the tissues to exchange and remove lactate and the other restraining it.     

Enhanced procollagen processing in skeletal muscle after a single bout of eccentric loading in humans.

Increases in procollagen processing within skeletal muscle have previously been reported in small animal models only. While indirect measurements in humans have suggested an increase procollagen processing, no intra-skeletal muscle measurements have confirmed these findings. In this study, eight young healthy male subjects performed a single bout of unaccustomed high intensity eccentric exercise on one leg, with the contralateral leg being the control. A significant increase in the muscle interstitial concentration of the N-terminal propeptide of procollagen type I (PINP) was observed (day 0: 1.96 +/- 0.44 ng ml(-1), day 2: 1.94 +/- 0.32 ng ml(-1), day 4: 3.90 +/- 1.03 ng ml(-1), day 8: 7.23 +/- 2.34 ng ml(-1)*, *P < 0.05 vs. basal and control) with no change being noted in the control leg. By day 2 post-exercise, an increase in the histological immunoreactivity of PINP and the N-terminal propeptide of procollagen type III (PIIINP) was also shown in the exercising leg only. Further, from day 2 post-exercise, immunoreactivity for tenascin C and reactive macrophages (CD68+ cells) was seen within the perimysial and endomysial connective tissue of the exercising leg only, indicating a high mechanical load and inflammation. This study shows that following a single bout of high intensity eccentric exercise there is an increase in procollagen processing within skeletal muscle in humans.