Takuya Osada

Production of interleukin-6 in contracting human skeletal muscles can account for the exercise-induced increase in plasma interleukin-6.

1 Plasma interleukin (IL)‐6 concentration is increased with exercise and it has been demonstrated that contracting muscles can produce IL‐ The question addressed in the present study was whether the IL‐6 production by contracting skeletal muscle is of such a magnitude that it can account for the IL‐6 accumulating in the blood. 2 This was studied in six healthy males, who performed one‐legged dynamic knee extensor exercise for 5 h at 25 W, which represented 40% of peak power output (Wmax). Arterial‐femoral venous (a‐fv) differences over the exercising and the resting leg were obtained before and every hour during the exercise. Leg blood flow was measured in parallel by the ultrasound Doppler technique. IL‐6 was measured by enzyme‐linked immunosorbent assay (ELISA). 3 Arterial plasma concentrations for IL‐6 increased 19‐fold compared to rest. The a‐fv difference for IL‐6 over the exercising leg followed the same pattern as did the net IL‐6 release. Over the resting leg, there was no significant a‐fv difference or net IL‐6 release. The work was produced by 2.5 kg of active muscle, which means that during the last 2 h of exercise, the median IL‐6 production was 6.8 ng min−1 (kg active muscle)−1 (range, 3.96‐9.69 ng min−1 kg−1). 4 The net IL‐6 release from the muscle over the last 2 h of exercise was 17‐fold higher than the elevation in arterial IL‐6 concentration and at 5 h of exercise the net release during 1 min was half of the IL‐6 content in the plasma. This indicates a very high turnover of IL‐6 during muscular exercise. We suggest that IL‐6 produced by skeletal contracting muscle contributes to the maintenance of glucose homeostasis during prolonged exercise.

Transcriptional activation of the IL-6 gene in human contracting skeletal muscle: influence of muscle glycogen content

In humans, the plasma interleukin 6 (IL‐6) concentration increases dramatically during low‐intensity exercise. Measurements across the working limb indicate that skeletal muscle is the source of IL‐6 production. To determine whether energy availability influences the regulation of IL‐6 expression during prolonged exercise, six male subjects completed two trials consisting of 180 min of two‐legged dynamic knee extensor with either normal or low (~60% of control) pre‐exercise muscle glycogen levels. Increases in plasma IL‐6 during exercise were significantly higher (P < 0.05) in the low‐glycogen (16‐fold) trial verses the control (10‐fold) trial. Transcriptional activation of the IL‐6 gene in skeletal muscle was also higher in the low‐glycogen trial; it increased by about 40‐fold after 90 min of exercise and about 60‐fold after 180 min of exercise. Muscle IL‐6 mRNA followed a similar but delayed pattern, increasing by more than 100‐fold in the low‐glycogen trial and by about 30‐fold in the control trial. These data demonstrate that exercise activates transcription of the IL‐6 gene in working skeletal muscle, a response that is dramatically enhanced when glycogen levels are low. These findings also support the hypothesis that IL‐6 may be produced by contracting myofibers when glycogen levels become critically low as a means of signaling the liver to increase glucose production.

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.

Brain and central haemodynamics and oxygenation during maximal exercise in humans

During maximal exercise in humans, fatigue is preceded by reductions in systemic and skeletal muscle blood flow, O2 delivery and uptake. Here, we examined whether the uptake of O2 and substrates by the human brain is compromised and whether the fall in stroke volume of the heart underlying the decline in systemic O2 delivery is related to declining venous return. We measured brain and central haemodynamics and oxygenation in healthy males (n= 13 in 2 studies) performing intense cycling exercise (360 ± 10 W; mean ±s.e.m.) to exhaustion starting with either high (H) or normal (control, C) body temperature. Time to exhaustion was shorter in H than in C (5.8 ± 0.2 versus 7.5 ± 0.4 min, P < 0.05), despite heart rate reaching similar maximal values. During the first 90 s of both trials, frontal cortex tissue oxygenation and the arterial–internal jugular venous differences (a‐v diff) for O2 and glucose did not change, whereas middle cerebral artery mean flow velocity (MCA Vmean) and cardiac output increased by ∼22 and ∼115%, respectively. Thereafter, brain extraction of O2, glucose and lactate increased by ∼45, ∼55 and ∼95%, respectively, while frontal cortex tissue oxygenation, MCA Vmean and cardiac output declined ∼40, ∼15 and ∼10%, respectively. At exhaustion in both trials, systemic declined in parallel with a similar fall in stroke volume and central venous pressure; yet the brain uptake of O2, glucose and lactate increased. In conclusion, the reduction in stroke volume, which underlies the fall in systemic O2 delivery and uptake before exhaustion, is partly related to reductions in venous return to the heart. Furthermore, fatigue during maximal exercise, with or without heat stress, in healthy humans is associated with an enhanced rather than impaired brain uptake of O2 and substrates.

Acute interleukin‐6 administration does not impair muscle glucose uptake or whole‐body glucose disposal in healthy humans

The cytokine interleukin (IL)‐6 has recently been linked with type 2 diabetes mellitus and has been suggested to affect glucose metabolism. To determine whether acute IL‐6 administration affects whole‐body glucose kinetics or muscle glucose uptake, 18 healthy young men were assigned to one of three groups receiving a high dose of recombinant human IL‐6 (HiIL‐6; n= 6), a low dose of IL‐6 (LoIL‐6; n= 6) or saline (Con; n= 6) infused into one femoral artery for 3 h. The stable isotope [6,6‐2H2] glucose was infused into a forearm vein throughout the 3 h infusion period and for a further 3 h after the cessation of infusion (recovery) to determine endogenous glucose production and whole‐body glucose disposal. Infusion with HiIL‐6 and LoIL‐6 resulted in a marked (P < 0.05) increase in systemic IL‐6 concentration throughout the 3 h of infusion (mean arterial plasma [IL‐6]s of 319 and 143 pg ml−1 for HiIL‐6 and LoIL‐6, respectively), followed by a rapid decline (P < 0.05) during the recovery period. Subjects experienced clinical symptoms such as shivering and discomfort during HiIL‐6 administration, but were asymptomatic during LoIL‐6 administration. In addition, only HiIL‐6 elevated (P < 0.05) plasma adrenaline (epinephrine). IL‐6 infusion, irrespective of dose, did not result in any changes to endogenous glucose production, whole‐body glucose disposal or leg‐ glucose uptake. These data demonstrate that acute IL‐6 administration does not impair whole‐body glucose disposal, net leg‐glucose uptake, or increase endogenous glucose production at rest in healthy young humans.

Cytochrome P450 2C9 plays an important role in the regulation of exercise‐induced skeletal muscle blood flow and oxygen uptake in humans

Previous studies show that exercise‐induced hyperaemia is unaffected by systemic inhibition of nitric oxide synthase (NOS) and it has been proposed that this may be due to compensation by other vasodilators. We studied the involvement of cytochrome P450 2C9 (CYP 2C9) in the regulation of skeletal muscle blood flow in humans and the interaction between CYP 2C9 and NOS. Seven males performed knee extensor exercise. Blood flow was measured by thermodilution and blood samples were drawn frequently from the femoral artery and vein at rest, during exercise and in recovery. The protocol was repeated three times on the same day. The first and the third protocols were controls, and in the second protocol either the CYP 2C9 inhibitor sulfaphenazole alone, or sulfaphenazole in combination with the NOS inhibitor Nω‐monomethyl‐l‐arginine (l‐NMMA) were infused. Compared with control there was no difference in blood flow at any time with sulfaphenazole infusion (P > 0.05) whereas with infusion of sulfaphenazole and l‐NMMA, blood flow during exercise was 16 ± 4 % lower than in control (9 min: 3.67 ± 0.31 vs. 4.29 ± 0.20 l min−1; P < 0.05). Oxygen uptake during exercise was 12 ± 3 % lower (9 min: 525 ± 46 vs. 594 ± 24 ml min−1; P < 0.05) with co‐infusion of sulfaphenazole and l‐NMMA, whereas oxygen uptake during sulfaphenazole infusion alone was not different from that of control (P > 0.05). The results demonstrate that CYP 2C9 plays an important role in the regulation of hyperaemia and oxygen uptake during exercise. Since inhibition of neither NOS nor CYP 2C9 alone affect skeletal muscle blood flow, an interaction between CYP 2C9 and NOS appears to exist so that a CYP‐dependent vasodilator mechanism takes over when NO production is compromised.

Muscle glycogen content and glucose uptake during exercise in humans: influence of prior exercise and dietary manipulation

There are many factors that can influence glucose uptake by contracting skeletal muscle during exercise and although one may be intramuscular glycogen content, this relationship is at present not fully elucidated. To test the hypothesis that muscle glycogen concentration influences glucose uptake during exercise, 13 healthy men were studied during two series of experiments. Seven men completed 4 h of two‐legged knee extensor exercise 16 h after reducing of muscle glycogen by completing 60 min of single‐legged cycling (Series 1). A further six men completed 3 h of two‐legged knee extensor exercise on two occasions: one after 60 min of two‐legged cycling (16 h prior to the experimental trial) followed by a high carbohydrate diet (HCHO) and the other after the same exercise followed by a low carbohydrate diet (LCHO) (Series 2). Muscle glycogen was decreased by 40 % when comparing the pre‐exercised leg (EL) with the control leg (CL) prior to exercise in Series 1. In addition, muscle glycogen was decreased by the same magnitude when comparing LCHO with HCHO in Series 2. In Series 1, glucose uptake was 3‐fold higher in the first 60 min of exercise, in the presence of unchanged pre‐exercise GLUT4 protein in EL compared with CL, suggesting that the lower glycogen, and not the exercise the day before, might have provided the stimulus for increased glucose uptake. Despite the same magnitude of difference in pre‐exercise glycogen concentration when comparing Series 1 with Series 2, neither direct‐nor isotopic tracer‐determined glucose uptake was higher in LCHO compared with HCHO in Series 2. However, arterial concentrations of insulin and glucose were lower, while free fatty acids and adrenaline were higher in LCHO compared with HCHO. These data suggest that pre‐exercise glycogen content may influence glucose uptake during subsequent exercise. However, this is only the case when delivery of substrates and hormones remains constant. When delivery of substrates and hormones is altered, the potential effect of glycogen on glucose uptake is negated.

Intramuscular fatty acid metabolism in contracting and non-contracting human skeletal muscle

The present study was undertaken to investigate the fate of blood‐borne non‐esterified fatty acids (NEFA) entering contracting and non‐contracting knee extensor muscles of healthy young individuals. [U‐13C]‐palmitate was infused into a forearm vein during 5 h of one‐legged knee extensor exercise at 40 % of maximal work capacity and the NEFA kinetics, oxidation and rate of incorporation into intramuscular triacylglycerol (mTAG) were determined for the exercising and the non‐exercising legs. During 4 h of one‐legged knee extensor exercise, mtag content decreased by 30 % (P < 0.05) in the contracting muscle, whereas it was unchanged in the non‐contracting muscle. The uptake of plasma NEFA, as well as the proportion directed towards oxidation, was higher in the exercising compared to the non‐exercising leg, whereas the rate of palmitate incorporation into mtag was fourfold lower (0.70 ± 0.14 vs. 0.17 ± 0.04 μmol (g dry wt)−1 h−1; P lt; 0.05), resulting in fractional synthesis rates of 1.0 ± 0.2 and 3.8 ± 0.9 % h−1 (P lt; 0.01) for the contracting and non‐contracting muscle, respectively. These findings demonstrate that mTAG in human skeletal muscle is continuously synthesised and degraded and that the metabolic fate of plasma NEFA entering the muscle is influenced by muscle contraction, so that a higher proportion is directed towards oxidation at the expense of storage in mTAG.

Quantification of ischemic muscle deoxygenation by near infrared time-resolved spectroscopy

The purpose of this study was to quantify muscle deoxygenation in human skeletal muscles using near infrared time-resolved spectroscopy (NIRTRS) and compare NIRTRS indicators and blood saturation. The forearm muscles of five healthy males (aged 27-32 yrs.) were monitored for changes in hemoglobin saturation (SO2) during 12 min of arterial occlusion and recovery. SO2 was determined by measuring the temporal profile of photon diffusion at 780 and 830 nm using NIRTRS, and was defined as SO2-TRS. Venous blood samples were also obtained for measurements of SvO2, and PvO2. Interstitial PO2(PintO2) was monitored by placing an O2 electrode directly into the muscle tissue. Upon the initiation of occlusion, all parameters fell progressively until reaching a plateau in the latter half of occlusion. It was observed at the end of occlusion that SO2-TRS (24.1 +/- 5.6%) agreed with SvO2 (26.2 +/- 6.4) and that PintO2 (14.7 +/- 1.0 Torr) agreed with PvO2 (17.3 +/- 2.2 Torr). The resting O2 store (oxygenated hemoglobin) and O2 consumption rate were 290 microM and 0.82 microM s-1, respectively, values which reasonably agree with the reported results. These results indicate that there was no O2 gradient between vessels and interstisium at the end of occlusion.

Noninvasive monitoring of deterioration in skeletal muscle function with forearm cast immobilization and the prevention of deterioration

BackgroundIn this research inactivity was simulated by immobilizing the forearm region in a plaster cast. Changes in skeletal muscle oxidative function were measured using near-infrared spectroscopy (NIRS), and the preventative effect of the training protocol on deterioration of skeletal muscle and the clinical utility of NIRS were examined.MethodsFourteen healthy adult men underwent immobilization of the forearm of the non-dominant arm by plaster cast for 21 days. Eight healthy adult subjects were designated as the immobilization group (IMM) and six were designated as the immobilization + training group (IMM+TRN). Grip strength, forearm circumference and dynamic handgrip exercise endurance were measured before and after the 21-day immobilization period. Using NIRS, changes in oxidative function of skeletal muscles were also evaluated. Muscle oxygen consumption recovery was recorded after the completion of 60 seconds of 40% maximum voluntary contraction (MVC) dynamic handgrip exercise 1 repetition per 4 seconds and the recovery time constant (TcVO2mus) was calculated.ResultsTcVO2mus for the IMM was 59.7 ± 5.5 seconds (average ± standard error) before immobilization and lengthened significantly to 70.4 ± 5.4 seconds after immobilization (p < 0.05). For the IMM+TRN, TcVO2mus was 78.3 ± 6.2 seconds before immobilization and training and shortened significantly to 63.1 ± 5.6 seconds after immobilization and training (p < 0.05).ConclusionsThe training program used in this experiment was effective in preventing declines in muscle oxidative function and endurance due to immobilization. The experimental results suggest that non-invasive monitoring of skeletal muscle function by NIRS would be possible in a clinical setting.