Changes in spontaneous and LPS-induced ex vivo cytokine production and mRNA expression in male and female athletes following prolonged exhaustive exercise.

PURPOSE: The capacity of whole blood cultures to produce cytokines in response to endotoxin (LPS) was studied in athletes before, 30 min after, 3 h after and 24 h after a half-marathon run. METHODS: Eight well trained men and 8 well trained women (6 of them in the late luteal phase of their cycle) participated. EDTA blood was incubated with or without LPS for 1 h, and cytokine concentration and gene expression were determined. To quantify LPS-dependent release on a per monocyte basis (LDR), the mean values of the difference (delta) between cytokine concentration in stimulated and unstimulated cultures, normalized to monocyte numbers, were calculated. RESULTS: LDR of TNF-alpha was significantly reduced by exercise with identical kinetic in men and women. TNF-alpha mRNA expression was slightly down-regulated following exercise (P < 0.05), but significantly so only in women. LDR of IL-6 was also reduced, but with a faster kinetic in women than in men. Similarly, 30 min post-exercise; LDR and spontaneous release of IL-1ra were significantly less in women than men. Concomitantly, IL-Ira mRNA was significantly elevated in unstimulated and in stimulated cultures in men only. IL-10 and IL-10 mRNA were significantly induced 30 min following exercise in absence of any detectable LDR. Women showed significantly lower levels than men. LDR and spontaneous release of IL-8 was enhanced in men and TGF-beta1 in women. A significant up-regulation was seen in unstimulated IL-8 mRNA for women and LPS-stimulated IL-8 mRNA expression for men following exercise. CONCLUSION: Altogether, LPS-dependent ex vivo cytokine release was strongly influenced by exercise and these changes could only in part be attributed to changes in messenger RNA. Results for IL-1ra, IL-6 and IL-10 pointed to a less pronounced anti-inflammatory response in women as compared with men. Our results also indicate an early production of IL-10 by peripheral blood cells in response to exercise.

Medium chain acylcarnitines dominate the metabolite pattern in humans under moderate intensity exercise and support lipid oxidation.

BACKGROUND: Exercise is an extreme physiological challenge for skeletal muscle energy metabolism and has notable health benefits. We aimed to identify and characterize metabolites, which are components of the regulatory network mediating the beneficial metabolic adaptation to exercise. METHODOLOGY AND PRINCIPAL FINDINGS: First, we investigated plasma from healthy human subjects who completed two independent running studies under moderate, predominantly aerobic conditions. Samples obtained prior to and immediately after running and then 3 and 24 h into the recovery phase were analyzed by a non-targeted (NT-) metabolomics approach applying liquid chromatography-qTOF-mass spectrometry. Under these conditions medium and long chain acylcarnitines were found to be the most discriminant plasma biomarkers of moderately intense exercise. Immediately after a 60 min (at 93% V(IAT)) or a 120 min run (at 70% V(IAT)) a pronounced, transient increase dominated by octanoyl-, decanoyl-, and dodecanoyl-carnitine was observed. The release of acylcarnitines as intermediates of partial beta-oxidation was verified in skeletal muscle cell culture experiments by probing (13)C-palmitate metabolism. Further investigations in primary human myotubes and mouse muscle tissue revealed that octanoyl-, decanoyl-, and dodecanoyl-carnitine were able to support the oxidation of palmitate, proving more effective than L-carnitine. CONCLUSIONS: Medium chain acylcarnitines were identified and characterized by a functional metabolomics approach as the dominating biomarkers during a moderately intense exercise bout possessing the power to support fat oxidation. This physiological production and efflux of acylcarnitines might exert beneficial biological functions in muscle tissue.

Gender- and menstrual phase dependent regulation of inflammatory gene expression in response to aerobic exercise.

The immunological reaction to exercise has been investigated with increasing intensity in the last 10-20 years, with most human studies performed in male subjects. Recently, gender-specific aspects have received growing attention, but studies carefully monitoring the influence of gender including the menstrual cycle, are rare. Here, we report gene expression patterns in response to a run at 93% of the individual anaerobic threshold of 9 women with regular menstrual cycles and no use of oral contraceptives who ran both at day 10 (follicular phase, F) and at day 25 (luteal phase, L) of their cycle. 12 male subjects (M) served as controls. The mRNA was pooled group wise and processed on a gene expression microarray encompassing 789 genes, including major genes of the inflammatory and anti-inflammatory reaction. The differences of gene expression between time points to (before run) and t1 (after run) were analyzed. Females in L showed a higher extent of regulation than females in F or men. Among those genes which were up-regulated above 1.5 fold change (log2) pro-inflammatory genes were significantly enriched (p = 0.033, after Bonferroni correction) in L, while this was not the case in F or M. Conversely, women in L showed a strong trend towards downregulation of anti-inflammatory genes. Some prominent genes like IL6 (coding for interleukin-6), and IL1RN (also termed IL1RA, coding for interleukin-1 receptor antagonist) were clearly regulated in opposite directions in L as opposed to F and M. In conclusion, women in L showed a distinctly different pattern of gene regulation in response to exercise, compared with women in F or M. The overall direction of gene expression changes of women in L is clearly pro-inflammatory. This finding accentuates a need for careful consideration of the female cyclic phase when investigating women in exercise immunology studies. Our results may also have implications relevant to other forms of stress in females.

Trauma-induced systemic inflammatory response versus exercise-induced immunomodulatory effects.

Accidental trauma and heavy endurance exercise, both induce a kind of systemic inflammatory response, also called systemic inflammatory response syndrome (SIRS). Exercise-related SIRS is conditioned by hyperthermia and concomitant heat shock responses, whereas trauma-induced SIRS manifests concomitantly with tissue necrosis and immune activation, secondarily followed by fever. Inflammatory cytokines are common denominators in both trauma and exercise, although there are marked quantitative differences. Different anti-inflammatory cytokines may be involved in the control of inflammation in trauma- and exercise-induced stress. Exercise leads to a balanced equilibrium between inflammatory and anti-inflammatory responses. Intermittent states of rest, as well as anti-oxidant capacity, are lacking or minor in trauma but are high in exercising individuals. Regular training may enhance immune competence, whereas trauma-induced SIRS often paves the way for infectious complications, such as sepsis.

Regulation of immediate early gene expression by exercise: short cuts for the adaptation of immune function.

Onset of physical activity induces a wide variety of rapid biophysical and biochemical responses that act on cells and lead to a wide range of phenotypic adaptations. Here we elucidate the role of immediate early gene (IEG) expression as a first line of transcriptional response that mediates protein neosynthesis and leads to adaptation of immune function. New posttranscriptional mechanisms have been described that speed up transcriptional responses. These include RNA-RNA interactions such as those exploited by miRNAs and stimulus-dependent cytoplasmic polyadenylation. We describe these shortcuts that modulate expression and discuss the challenges of accurately measuring them using various transcriptomic screening and quantification approaches. Although there is high complexity of the upstream as well as the downstream pathways that lead to IEG expression, IEG expression itself may only show a limited number of response patterns. Focusing transcriptomic approaches in exercise immunology at the IEG-level may facilitate the discovery of exercise-specific transcriptional signatures.

cDNA microarray analysis reveals novel candidate genes expressed in human peripheral blood following exhaustive exercise.

It is generally accepted that exhausting endurance exercise exhibits strong effects on the immune system. Such effects have been attributed to changes in the cellular composition of peripheral blood as well as to changes in the expression of plausible candidate genes. The list of candidate genes is far from being complete, since this issue has not yet been investigated in a systematic way. In this study, we used a custom-made cDNA microarray focused on inflammation as a screening approach to study gene expression in eight one-half marathon runners before, immediately after, and 24 h after exercise. Significant differential gene expression was verified by quantitative real-time PCR. Linear regression analysis showed that microarray expression analysis of cell type-specific surface molecules reflects the observed individual cellular shifts in peripheral blood cells with high statistical significance. In line with the results of former studies, we observed an upregulation of mitogen-activated protein kinase-activated protein kinase-2 (MAPKAP-K2), L-selectin, and IL-1 receptor antagonist (IL-1ra) after exhaustive exercise. The main results of this study report, for the first time, the downregulation of CD81; the upregulation of thioredoxin, which may play an important part in anti-oxidative defense; and, surprisingly, the downregulation of the anti-carcinogenic gene glutathione-S-transferase-3 (GSTM3) in peripheral blood. The study shows cDNA microarray expression analysis as a reliable systematic instrument to complete the list of candidate genes that may play a role in exhaustive exercise-induced modulation of the immune response.

CDNA-microarray analysis as a research tool for expression profiling in human peripheral blood following exercise.

Exhausting endurance exercise has strong effects on the immune system. Changes have been shown in the cellular composition of peripheral blood and in gene expression within those cells. In this study, custom-made cDNA microarrays focused on inflammation were used to analyze gene expression blood cells obtained from eight half-marathon runners before (t0), immediately after (t1) and 24 hours after exercise (t2). The microarrays that were used contained 384 different cDNAs spotted in triplicate. Differentially-regulated gene expression was analyzed using a simple rule-based clustering. Comparing t1 vs. t0, and t2 vs. t0, 36 and 21 sequences respectively, showed a consistent pattern of changes in all eight athletes. Taken together, the pattern of these modified genes can be viewed as a "gene expression fingerprint" for each time point in response to a half marathon. The known and novel genes identified here represent targets for further molecular characterization of the complex reaction of the body to an exhaustive challenge. These data suggest that gene expression fingerprints can serve as a powerful research tool to design novel strategies for diagnosis and treatment of exercise related injury and stress.

Inverse response of leukocyte heat shock proteins and DNA damage to exercise and heat.

Elevated ambient temperature may exert an additional impact on the exercise-induced expression of heat shock proteins (HSP) and DNA damage in leukocytes. The protective functions of HSP include antioxidative and antiapoptotic effects and may prevent damage to DNA. Twelve athletes completed a continuous run (75% VO2max) on the treadmill, six at 28 degrees C and six at 18 degrees C room temperature. Leukocyte expression of HSP27 and inducible HSP70 was analyzed on mRNA- (RT-PCR) and protein-level (flow cytometry), while DNA damage was quantified by the comet assay. High ambient temperature induced an additional accumulation of HSP-mRNA and -protein in leukocytes compared with the exercise-induced expression at 18 degrees C. HSP27 showed a special heat sensitivity. Surprisingly, the increase of DNA damage was less pronounced after exercise at 28 degrees C compared to 18 degrees C although heat shock in vitro clearly induced DNA damage. The inverse relation between HSP and DNA damage may indicate functions of HSP which protect against exercise-induced DNA-damage in terms of thermotolerance or apoptosis.

Vitamin E supplementation does not increase the vitamin C radical concentration at rest and after exhaustive exercise in healthy male subjects.

BACKGROUND: Extensive exercise may promote the formation of reactive oxygen species and subsequently contribute to tissue damage. A compound which can protect cells and tissues is vitamin E. The vitamin E radical, formed during the radical scavenging process, can be reduced by vitamin C resulting in a higher level of the vitamin C radical (semidehydroascorbate free radical). An increase of the vitamin C radical, however, is assumed to exert prooxidative effects. AIM OF THE STUDY: To elucidate whether supplementation of vitamin E and exhaustive exercise lead to an increase of the vitamin C radical in human plasma. METHODS: A placebo controlled, cross over study with 13 male volunteers was carried out. After an 8 day supplementation period with 500 I.U. D-alpha-tocopherol, the subjects performed two exhaustive treadmill runs. Blood samples were collected at rest, 0, 0.25, 1, 3, 24 and 48 h after exercise. Serum was separated and concentrations of D-alpha-tocopherol and ascorbic acid were determined by HPLC. Vitamin C radical levels in plasma were assessed by electron paramagnetic resonance (EPR). RESULTS: Vitamin E and C both showed a tendency to decrease between 3 h and 24 h after exercise. Vitamin C radical level remained stable during the whole period. Neither vitamin E supplementation nor exercise had any influence on the plasma concentration of the vitamin C radical. CONCLUSIONS: Vitamin E supplementation under conditions of mild oxidative stress does not result in an increased vitamin C radical concentration.

Evaluation of stress responses to interval training at low and moderate altitudes.

PURPOSE: The purpose of the present field study was to explore whether extensive interval training (IT) performed with a similar behavior of blood lactate (LA) at an altitude of 1800 m (ALT) and near sea level (SL) goes along with a comparable hormonal, metabolic, and acute phase response in highly trained endurance athletes. METHODS: Twelve distance runners (VO2 64.6 +/- 6.9 mL.kg(-1) ) performed IT (10 x 1000 m, 2-min rest) at SL with a running velocity (V) corresponding to 112% of the individual anaerobic threshold (IAT). After an acclimatization period of 7 d, IT was repeated with a lower V (107% IAT) at ALT. Blood samples were drawn at rest, 0, 0.3, 3, and 24 h after IT. LA during IT was similar at SL and ALT (5.4 +/- 1.3/5.3 +/- 1.2 mmol.L(-1)), whereas HR tended to be higher at SL. RESULTS: Postexercise rises in plasma noradrenaline (NA), NA sulfate, adrenaline, glucose, interleukin-6 (IL-6), and neutrophils were significantly more pronounced at ALT. The increase of cortisol and human growth hormone showed an insignificant trend toward higher values at ALT. A slight but significant increase of plasma erythropoietin was only apparent after IT at ALT. No differences between either condition were observed for exercise-related changes in free fatty acids, IL-8, lympho-, or monocyte counts. CONCLUSIONS: In spite of a matched accumulation pattern of LA between ALT and N, stress responses, such as sympathetic activation and hepatic glucose release, still appear to be greater at ALT. This additional impact of moderate ALT on the stress response to IT should be taken into account if repeated training sessions are performed within a short period of time.