Peter Schjerling

Pro- and anti-inflammatory cytokine balance in strenuous exercise in humans

1 The present study investigates to what extent and by which time course prolonged strenuous exercise influences the plasma concentration of pro‐inflammatory and inflammation responsive cytokines as well as cytokine inhibitors and anti‐inflammatory cytokines. 2 Ten male subjects (median age 27.5 years, range 24–37) completed the Copenhagen Marathon 1997 (median running time 3:26 (h:min), range 2:40–4:20). Blood samples were obtained before, immediately after and then every 30 min in a 4 h post‐exercise recovery period. 3 The plasma concentrations of tumour necrosis factor (TNF)α, interleukin (IL)‐1β, IL‐6, IL‐1ra, sTNF‐r1, sTNF‐r2 and IL‐10 were measured by enzyme‐linked immunosorbent assay (ELISA). The highest concentration of IL‐6 was found immediately after the race, whereas IL‐1ra peaked 1 h post exercise (128‐fold and 39‐fold increase, respectively, as compared with the pre‐exercise values). The plasma level of IL‐1β, TNFα, sTNF‐r1 and sTNF‐r2 peaked in the first hour after the exercise (2.1‐, 2.3‐, 2.7‐ and 1.6‐fold, respectively). The plasma level of IL‐10 showed a 27‐fold increase immediately post exercise. 4 In conclusion, strenuous exercise induces an increase in the pro‐inflammatory cytokines TNFα and IL‐1β and a dramatic increase in the inflammation responsive cytokine IL‐6. This is balanced by the release of cytokine inhibitors (IL‐1ra, sTNF‐r1 and sTNF‐r2) and the anti‐inflammatory cytokine IL‐10. The study suggests that cytokine inhibitors and anti‐inflammatory cytokines restrict the magnitude and duration of the inflammatory response to exercise.

Muscle-derived interleukin-6: possible biological effects

Interleukin‐6 (IL‐6) is produced locally in working skeletal muscle and can account for the increase in plasma IL‐6 during exercise. The production of IL‐6 during exercise is related to the intensity and duration of the exercise, and low muscle glycogen content stimulates the production. Muscle‐derived IL‐6 is released into the circulation during exercise in high amounts and is likely to work in a hormone‐like fashion, exerting an effect on the liver and adipose tissue, thereby contributing to the maintenance of glucose homeostasis during exercise and mediating exercise‐induced lipolysis. Muscle‐derived IL‐6 may also work to inhibit the effects of pro‐inflammatory cytokines such as tumour necrosis factor α. The latter cytokine is produced by adipose tissue and inflammatory cells and appears to play a pathogenetic role in insulin resistance and atherogenesis.

Vitamin D controls T cell antigen receptor signaling and activation of human T cells

Phospholipase C (PLC) isozymes are key signaling proteins downstream of many extracellular stimuli. Here we show that naive human T cells had very low expression of PLC-γ1 and that this correlated with low T cell antigen receptor (TCR) responsiveness in naive T cells. However, TCR triggering led to an upregulation of ∼75-fold in PLC-γ1 expression, which correlated with greater TCR responsiveness. Induction of PLC-γ1 was dependent on vitamin D and expression of the vitamin D receptor (VDR). Naive T cells did not express VDR, but VDR expression was induced by TCR signaling via the alternative mitogen-activated protein kinase p38 pathway. Thus, initial TCR signaling via p38 leads to successive induction of VDR and PLC-γ1, which are required for subsequent classical TCR signaling and T cell activation.

Interleukin‐6 production in contracting human skeletal muscle is influenced by pre‐exercise muscle glycogen content

1 Prolonged exercise results in a progressive decline in glycogen content and a concomitant increase in the release of the cytokine interleukin‐6 (IL‐6) from contracting muscle. This study tests the hypothesis that the exercise‐induced IL‐6 release from contracting muscle is linked to the intramuscular glycogen availability. 2 Seven men performed 5 h of a two‐legged knee‐extensor exercise, with one leg with normal, and one leg with reduced, muscle glycogen content. Muscle biopsies were obtained before (pre‐ex), immediately after (end‐ex) and 3 h into recovery (3 h rec) from exercise in both legs. In addition, catheters were placed in one femoral artery and both femoral veins and blood was sampled from these catheters prior to exercise and at 1 h intervals during exercise and into recovery. 3 Pre‐exercise glycogen content was lower in the glycogen‐depleted leg compared with the control leg. Intramuscular IL‐6 mRNA levels increased with exercise in both legs, but this increase was augmented in the leg having the lowest glycogen content at end‐ex. The arterial plasma concentration of IL‐6 increased from 0.6 ± 0.1 ng l−1 pre‐ex to 21.7 ± 5.6 ng l−1 end‐ex. The depleted leg had already released IL‐6 after 1 h (4.38 ± 2.80 ng min−1 (P < 0.05)), whereas no significant release was observed in the control leg (0.36 ± 0.14 ng min−1). A significant net IL‐6 release was not observed until 2 h in the control leg. 4 This study demonstrates that glycogen availability is associated with alterations in the rate of IL‐6 production and release in contracting skeletal muscle.

A trauma‐like elevation of plasma cytokines in humans in response to treadmill running

1 Elevated levels of cytokines, especially interleukin (IL)‐6 and IL‐1ra, can be measured in the plasma of athletes after exhaustive long term exercise. 2 The present study investigates the kinetics of several cytokines and chemokines in ten male athletes before, during and after 2.5 h of treadmill running at 75% of maximal oxygen consumption (VO2,max). Blood was sampled before, every half‐hour during running and every hour in the following 6 h recovery period. 3 The plasma concentration of IL‐6 increased after 30 min of running, and peaked at the end of running with a 25‐fold increase compared with the pre‐exercise value. IL‐1ra increased only after running, and peaked after 2 h of rest with an 18‐fold increase compared with the pre‐exercise value. No changes were found in the concentrations of IL‐1β, tumour necrosis factor (TNF)α, IL‐15 and macrophage inflammatory protein (MIP)‐1β, and the concentrations of IL‐8 and MIP‐1α were below detection limits. 4 The results suggest that very early events in exercise trigger the release of IL‐6, and that the cytokine response to exercise has similarities to that observed after trauma.

Physical activity and plasma interleukin-6 in humans – effect of intensity of exercise

Abstract The present study included data from three marathon races to investigate the hypothesis that a relationship exists between running intensity and elevated concentrations of interleukin (IL)-6 in plasma. The study included a total of 53 subjects whose mean age was 30.6 [95% confidence interval (CI) 1.4] years, mean body mass 77.7 (95%CI 2.0) kg, mean maximal oxygen uptake (V˙O2max) 59.3 (95%CI 1.4) ml · min−1 · kg−1, and who had participated in the Copenhagen Marathons of 1996, 1997 or 1998, achieving a mean running time of 206 (95%CI 7) min. Running intensity was calculated as running speed divided by V˙O2max. The concentration of IL-6 in plasma peaked immediately after the run. There was a negative correlation between peak IL-6 concentration and running time (r=−0.30, P < 0.05) and a positive correlation between peak IL-6 concentration and running intensity (r=0.32, P < 0.05). The IL-1 receptor antagonist (IL-1ra) plasma concentration peaked 1.5 h after the run and there was a positive correlation between the peak plasma concentrations of IL-6 and IL-1ra (r=0.39, P < 0.01). Creatine kinase (CK) plasma concentration peaked on the 1st day after the run, but no association was found between peak concentrations of IL-6 and CK. In conclusion, the results confirmed the hypothesized association between plasma IL-6 concentration and running intensity, but did not confirm the previous finding of a connection between IL-6 plasma concentration and muscle damage.

The effects of heavy resistance training and detraining on satellite cells in human skeletal muscles

The aim of this study was to investigate the modulation of satellite cell content and myonuclear number following 30 and 90 days of resistance training and 3, 10, 30, 60 and 90 days of detraining. Muscle biopsies were obtained from the vastus lateralis of 15 young men (mean age: 24 years; range: 20–32 years). Satellite cells and myonuclei were studied on muscle cross‐sections stained with a monoclonal antibody against CD56 and counterstained with Mayers haematoxylin. Cell cycle markers CyclinD1 and p21 mRNA levels were determined by Northern blotting. Satellite cell content increased by 19% (P= 0.02) at 30 days and by 31% (P= 0.0003) at 90 days of training. Compared to pre‐training values, the number of satellite cells remained significantly elevated at 3, 10 and 60 days but not at 90 days of detraining. The two cell cycle markers CyclinD1 and p21 mRNA significantly increased at 30 days of training. At 90 days of training, p21 was still elevated whereas CyclinD1 returned to pre‐training values. In the detraining period, p21 and CyclinD1 levels were similar to the pre‐training values. There were no significant alterations in the number of myonuclei following the training and the detraining periods. The fibre area controlled by each myonucleus gradually increased throughout the training period and returned to pre‐training values during detraining. In conclusion, these results demonstrate the high plasticity of satellite cells in response to training and detraining stimuli and clearly show that moderate changes in the size of skeletal muscle fibres can be achieved without the addition of new myonuclei.

Effects of α-AMPK knockout on exercise-induced gene activation in mouse skeletal muscle

We tested the hypothesis that 5′AMP‐activated protein kinase (AMPK) plays an important role in regulating the acute, exercise‐induced activation of metabolic genes in skeletal muscle, which were dissected from whole‐body α2‐ and α1‐AMPK knockout (KO) and wild‐type (WT) mice at rest, after treadmill running (90 min), and in recovery. Running increased α1‐AMPK kinase activity, phosphorylation (P) of AMPK, and acetyl‐CoA carboxylase (ACC)β in α2‐WT and α2‐KO muscles and increased α2‐AMPK kinase activity in α2‐WT. In α2‐KO muscles, AMPK‐P and ACCβ‐P were markedly lower compared with α2‐WT. However, in α1‐WT and α1‐KO muscles, AMPK‐P and ACCβ‐P levels were identical at rest and increased similarly during exercise in the two genotypes. The α2‐KO decreased peroxisome‐proliferator‐activated receptor γ coactivator (PGC)‐1α, uncoupling protein‐3 (UCP3), and hexokinase II (HKII) transcription at rest but did not affect exercise‐induced transcription. Exercise increased the mRNA content of PGC‐1α, Forkhead box class O (FOXO)1, HKII, and pyruvate dehydrogenase kinase 4 (PDK4) similarly in α2‐WT and α2‐KO mice, whereas glucose transporter GLUT 4, carnitine palmitoyltransferase 1 (CPTI), lipoprotein lipase, and UCP3 mRNA were unchanged by exercise in both genotypes. CPTI mRNA was lower in α2‐KO muscles than in α2‐WT muscles at all time‐points. In α1‐WT and α1‐KO muscles, running increased the mRNA content of PGC‐1α and FOXOl similarly. The α2‐KO was associated with lower muscle adenosine 5′‐triphosphate content, and the inosine monophosphate content increased substantially at the end of exercise only in α2‐KO muscles. In addition, subcutaneous injection of 5‐aminoimidazole‐4‐carboxamide‐1‐β‐4‐ribofuranoside (AICAR) increased the mRNA content of PGC‐1α, HKII, FOXO1, PDK4, and UCP3, and α2‐KO abolished the AICAR‐induced increases in PGC‐1α and HKII mRNA. In conclusion, KO of the α2‐ but not the α1‐AMPK isoform markedly diminished AMPK activation during running. Nevertheless, exercise‐induced activation of the investigated genes in mouse skeletal muscle was not impaired in α1‐ or α2‐AMPK KO muscles. Although it cannot be ruled out that activation of the remaining α‐isoform is sufficient to increase gene activation during exercise, the present data do not support an essential role of AMPK in regulating exercise‐induced gene activation in skeletal muscle.

Muscle contractions induce interleukin-6 mRNA production in rat skeletal muscles

1 The present study explored the hypothesis that interleukin‐6 (IL‐6) might be locally produced in response to skeletal muscle contractions and whether the production might reflect the type of muscle contraction performed. Rats were anaesthetized and the calf muscles of one limb were stimulated electrically for concentric or eccentric contractions (4 × 10 contractions with 1 min of rest between the 4 series, 100 Hz). The contralateral muscles served as unstimulated controls. The mRNA levels for IL‐6, the glucose transport protein GLUT‐4 and β‐actin in the rat muscles (white and red gastrocnemius and soleus) were quantified by quantitative competitive RT‐PCR. 2 The IL‐6 mRNA level, measured 30 min after the stimulation, increased after both eccentric and concentric contractions and there were no significant differences in IL‐6 mRNA levels between the different muscle fibre types. No significant increase in IL‐6 mRNA level was seen in the unstimulated contralateral muscle fibres. 3 No increase in GLUT‐4 mRNA level was detected, indicating that the increase in IL‐6 mRNA level was not due to general changes in transcription. 4 We conclude that IL‐6 is locally produced after muscle contraction, with no significant differences between different muscle fibre types. This local production of IL‐6 is not due to general changes in transcription, since no changes in the level of GLUT‐4 mRNA were found. The fact that increased IL‐6 mRNA levels were seen after both concentric and eccentric contractions indicates that the production of IL‐6 is not solely due to muscle damage, seen primarily after eccentric exercise.

Expression of collagen and related growth factors in rat tendon and skeletal muscle in response to specific contraction types

Acute exercise induces collagen synthesis in both tendon and muscle, indicating an adaptive response in the connective tissue of the muscle–tendon unit. However, the mechanisms of this adaptation, potentially involving collagen‐inducing growth factors (such as transforming growth factor‐β‐1 (TGF‐β‐1)), as well as enzymes related to collagen processing, are not clear. Furthermore, possible differential effects of specific contraction types on collagen regulation have not been investigated. Female Sprague–Dawley rats were subjected to 4 days of concentric, eccentric or isometric training (n= 7–9 per group) of the medial gastrocnemius, by stimulation of the sciatic nerve. RNA was extracted from medial gastrocnemius and Achilles tendon tissue 24 h after the last training bout, and mRNA levels for collagens I and III, TGF‐β‐1, connective tissue growth factor (CTGF), lysyl oxidase (LOX), metalloproteinases (MMP‐2 and ‐9) and their inhibitors (TIMP‐1 and 2) were measured by Northern blotting and/or real‐time PCR. In tendon, expression of TGF‐β‐1 and collagens I and III (but not CTGF) increased in response to all types of training. Similarly, enzymes/factors involved in collagen processing were induced in tendon, especially LOX (up to 37‐fold), which could indicate a loading‐induced increase in cross‐linking of tendon collagen. In skeletal muscle, a similar regulation of gene expression was observed, but in contrast to the tendon response, the effect of eccentric training was significantly greater than the effect of concentric training on the expression of several transcripts. In conclusion, the study supports an involvement of TGF‐β‐1 in loading‐induced collagen synthesis in the muscle–tendon unit and importantly, it indicates that muscle tissue is more sensitive than tendon to the specific mechanical stimulus.