Iain T. Campbell

Muscle wasting and energy balance in critical illness.

BACKGROUND In nine patients with multiple organ failure ultrasound was able to identify muscle wasting despite the presence of oedema (Campbell et al., J Clin Nutr 62 (1995) 533). AIMS The purpose of the present study was twofold: one was to determine whether this technique was applicable to a much larger ICU population, many of whom were not as ill as the original subjects. The second reason was to determine whether a relationship could be identified between rates of wasting and energy balance. METHODS Serial measurements of both mid-upper arm circumference (MAC) and muscle thickness, using ultrasound, were made at 1-3 day intervals between 5 and 39 (median 7) days in 50 critically ill patients. RESULTS Muscle thickness decreased in 48 of the 50 patients at a median rate of 1.6%/day with a range of 0.2-5.7%/day. In 33 patients, in whom MAC did not change significantly with time, muscle thickness decreased by between 0.3 and 4.2 (median 1.6)%/day. In three patients MAC increased significantly with time but muscle thickness decreased by between 1.3 and 5.7 (median 2.6)%/day. Twelve patients showed a significant decrease in MAC with time and muscle thickness in this group decreased by between 0.2 and 4.0 (median 1.3)%/day. The percentage decrease in muscle thickness between the groups, in whom MAC decreased or did not change, was not significantly different (P = 0.475). CONCLUSION We have demonstrated that an ultrasound technique devised to identify muscle wasting in the presence of severe fluid retention works in the majority (48/50) of patients when applied to a wider ICU population. Energy balance made no difference to the rate of wasting.

Inhibition of Nitric Oxide Synthase by L‐NAME Speeds Phase II Pulmonary V̇O2 Kinetics in the Transition to Moderate‐Intensity Exercise in Man

There is evidence that the rate at which oxygen uptake (V̇O2) rises at the transition to higher metabolic rates within the moderate exercise intensity domain is modulated by oxidative enzyme inertia, and also that nitric oxide regulates mitochondrial function through competitive inhibition of cytochrome c oxidase in the electron transport chain. We therefore hypothesised that inhibition of nitric oxide synthase (NOS) by nitro‐L‐arginine methyl ester (L‐NAME) would alleviate the inhibition of mitochondrial V̇O2 by nitric oxide and result in a speeding of V̇O2 kinetics at the onset of moderate‐intensity exercise. Seven males performed square‐wave transitions from unloaded cycling to a work rate requiring 90 % of predetermined gas exchange threshold with and without prior intravenous infusion of L‐NAME (4 mg kg−1 in 50 ml saline over 60 min). Pulmonary gas exchange was measured breath‐by‐breath and V̇O2 kinetics were determined from the averaged response to four exercise bouts performed in each condition using a mono‐exponential function following elimination of the phase I response. There were no significant differences between the control and L‐NAME conditions for baseline V̇O2 (means ±s.e.m. 797 ± 32 vs. 794 ± 29), the duration of phase I (15.4 ± 0.8 vs. 17.2 ± 0.6), or the steady‐state increment in V̇O2 above baseline (1000 ± 83 vs. 990 ± 85 ml min−1), respectively. However, the phase II time constant of the V̇O2 response was significantly smaller following L‐NAME infusion (22.1 ± 2.4 vs. 17.9 ± 2.3; P < 0.05). These data indicate that inhibition of NOS by L‐NAME results in a significant (19 %) speeding of pulmonary V̇O2 kinetics in the transition to moderate‐intensity cycle exercise in man. At least part of the intrinsic inertia to oxidative metabolism at the onset of moderate‐intensity exercise may result from competitive inhibition of mitochondrial V̇O2 by nitric oxide at cytochrome c oxidase, although other mechanisms for the effect of L‐NAME on V̇O2 kinetics remain to be explored.

Genetic variation in proinflammatory and anti-inflammatory cytokine production in multiple organ dysfunction syndrome.

ObjectivesThe objectives of this study were to examine the prevalence of genetic variation for in vitro cytokine production (tumor necrosis factor [TNF]-&agr;, interleukin-10, transforming growth factor-&bgr;1) in patients with multiple organ dysfunction syndrome, to measure circulating cytokine levels and relate these to genotype, and to identify the relationship between genetic variation and outcome. DesignProspective analysis. SettingIntensive care unit of a university teaching hospital. PatientsEighty-eight critically ill patients with multiple organ dysfunction syndrome. Measurements and Main ResultsThe frequency of the different interleukin-10 genotypes (corresponding to high, intermediate, and low interleukin-10 production in vitro) were significantly different between controls and multiple organ dysfunction syndrome patients. High interleukin-10 producers were under-represented in the multiple organ dysfunction syndrome group: This genotype occurred in 30% of controls but in only 6% of patients (p < .001). There was no relationship between interleukin-10 genotype and mortality. The frequency of TNF-&agr; genotypes was also significantly different between patients and controls. Intermediate TNF-&agr; producers were under-represented (5.7% vs. 23%) and high TNF-&agr; producers over-represented (35.2% vs. 16%) in the patient group (p < .001). TNF-&agr; genotype was not related to mortality. The distribution of TNF-&bgr; genotypes (homozygous B1, homozygous B2, and heterozygotes) was also different between controls and patients (p = .008). The B2/B2 genotype (associated with high TNF-&agr; production) tended to occur less frequently in the intensive care unit population (31% vs. 50%) and was associated with a higher mortality rate than either the B1/B1 or B1/B2 genotypes (48% vs. 11% and 33% respectively, p = .115). The combination of proinflammatory (TNF-&agr;/TNF-&bgr;) and anti-inflammatory (interleukin-10/transforming growth factor-&bgr;1) cytokine genotypes was associated with prolonged patient survival time. Patients predisposed to produce a balanced cytokine response (e.g., intermediate interleukin-10/TNF-&agr; producers) demonstrated the longest survival times, although overall mortality was no different. ConclusionA genetic predisposition to high interleukin-10 production or intermediate TNF-&agr; production in vitro may be protective of admission to the intensive care unit, although once admitted, any protection provided by these genotypes seems to be lost. TNF-&bgr; genotype conferred no advantage to patients with multiple organ dysfunction syndrome, the TNFB2 allele being associated with increased mortality. The combination of proinflammatory and anti-inflammatory cytokine genotypes supports the idea that a balanced cytokine response is favorable and was associated with prolonged patient survival time.

Elevated core and muscle temperature to levels comparable to exercise do not increase heat shock protein content of skeletal muscle of physically active men.

Aim:  Exercise‐associated hyperthermia is routinely cited as the signal responsible for inducing an increased production of heat shock proteins (HSPs) following exercise. This hypothesis, however, has not been tested in human skeletal muscle. The aim of the present study was to therefore investigate the role of increased muscle and core temperature in contributing to the exercise‐induced production of the major HSP families in human skeletal muscle.

Metabolic responses to isoenergetic meals containing different proportions of carbohydrate and fat

The purpose of the present study was to investigate the interrelationship between carbohydrate and fat metabolism at rest after isoenergetic meals of varying proportions of carbohydrate and fat. Eight physically-active subjects (BMI 18.1-23.4 kg/m2) were studied at rest on three occasions after an overnight fast. In a balanced design they were given meals containing carbohydrate, protein and fat in the following amounts respectively (g/70 kg body weight): meal 1 121, 16, 48; meal 2 70, 16, 70; meal 3 50, 14, 80. All meals were isoenergetic, containing 4.0 MJ/70 kg body weight, and were of similar appearance. In addition, on a fourth occasion five of the eight subjects consumed meal 4 (g/70 kg body weight): carbohydrate 0, protein 0, fat 108. Blood samples were taken before eating the meal and at intervals following the meal to determine metabolic and hormonal responses. Energy expenditure and substrate oxidation were measured by indirect calorimetry and balance was calculated over the 5 h postprandial period. The incremental areas under the time curves for fat oxidation were greatest after meals 3 and 4 (P < 0.05), whereas incremental areas under the carbohydrate oxidation v. time curves were relatively reduced after these two meals (P < 0.05). This was accompanied by lesser suppression of plasma non-esterified fatty acid concentrations (P < 0.001) and reduced plasma insulin concentrations (P < 0.001) following these meals. Energy balance was almost identical after the three isoenergetic meals. In contrast, there was an inverse relationship between carbohydrate and fat balance following these meals, with carbohydrate balance decreasing as carbohydrate intake decreased and fat balance increasing as fat intake increased. We conclude that there is a close interrelationship between carbohydrate and fat metabolism following isoenergetic meals in resting subjects.

Influence of nitric oxide synthase inhibition on pulmonary O2 uptake kinetics during supra‐maximal exercise in humans

We have recently reported that inhibition of nitric oxide synthase (NOS) with NG‐nitro‐l‐arginine methyl ester (l‐NAME) accelerates the ‘phase II’ pulmonary O2 uptake kinetics following the onset of moderate and heavy intensity submaximal exercise in humans. These data suggest that the influence of nitric oxide (NO) on mitochondrial function is an important factor in the inertia to aerobic respiration that is evident in the transition from a lower to a higher metabolic rate. The purpose of the present study was to investigate the influence of l‐NAME on pulmonary kinetics following the onset of supra‐maximal exercise, where it has been suggested that O2 availability represents an additional limitation to kinetics. Seven healthy young men volunteered to participate in this study. Following an incremental cycle ergometer test for the determination of , the subjects returned on two occasions to perform a ‘step’ exercise test from a baseline of unloaded cycling to a work rate calculated to require 105%, preceded either by systemic infusion of l‐NAME (4 mg kg−1 in 50 ml saline) or 50 ml saline as a control (Con). Pulmonary gas exchange was measured on a breath‐by‐breath basis throughout the exercise tests. The duration of ‘phase I’ was greater with l‐NAME (Con: 14.0 ± 2.1 versusl‐NAME: 16.0 ± 1.6 s; P= 0.03), suggestive of a slower cardiovascular adaptation following the onset of exercise. However, the phase II time constant was reduced by 44% with l‐NAME (Con: 36.3 ± 17.3 versusl‐NAME: 20.4 ± 8.3 s; P= 0.01). The accumulation of blood lactate during exercise was also reduced with l‐NAME (Con: 4.0 ± 1.1 versusl‐NAME: 2.7 ± 2.1 mm; P= 0.04). These data indicate that skeletal muscle NO production represents an important limitation to the acceleration of oxidative metabolism following the onset of supra‐maximal exercise in humans.

Trained Men Display Increased Basal Heat Shock Protein Content of Skeletal Muscle

PURPOSE 1) To compare the baseline levels of heat shock and antioxidant protein content in the skeletal muscle of trained and untrained humans and 2) to characterize the exercise-induced stress response of aerobically trained human skeletal muscle to an acute exercise challenge. METHODS Resting muscle biopsies were obtained from the vastus lateralis muscle of six untrained and six aerobically trained young males. To characterize the stress response of a trained population, the trained subjects also performed a 45-min nondamaging running exercise protocol at an intensity corresponding to 75% of V O2max. Muscle biopsies were obtained from the vastus lateralis muscle at 48 h and 7 d after exercise. RESULTS Trained subjects displayed significantly higher (P<0.05) resting levels of heat shock protein 60 (HSP60, 25%), alphaB-crystallin (43%), and manganese superoxide (MnSOD, 45%) protein content compared with untrained subjects. Trained subjects also exhibited no significant change (P > 0.05) in resting levels of HSP70 (16%), HSC70 (13%), and total superoxide dismutase (SOD) activity (46%) compared with untrained subjects. Resting HSP27 levels were unaffected by exercise training (P > 0.05). In the trained subjects, exercise failed to induce significant increases (P>0.05)in muscle content of HSP70, HSC70, HSP60, HSP27, alphaB-crystallin, and MnSOD protein content or in the activity of SOD at any time point after exercise. CONCLUSION This study demonstrates for the first time that trained men display a selective up-regulation of basal heat shock and antioxidant protein content and do not exhibit a stress response to customary running exercise. It is suggested that an increase in these protective systems functions to maintain homeostasis during the stress of exercise by protecting against disruptions to the cytoskeleton/contractile machinery, by maintaining redox balance, and by facilitating mitochondrial biogenesis.

Retrospective study of temperature rhythms of intensive care patients.

The hourly rectal temperature record of 15 patients who spent greater than or equal to 8 days in the ICU were analyzed for circadian rhythmicity. Using cosinor analysis, a statistically significant rhythm in a period of 24 h was present on 80% of the patient days studied. The position of the acrophase, or peak, of the rhythm varied markedly both between patients and within patients, with changes of several hours from day to day. There was a tendency for the amplitude of the rhythm to be greater when the patients were unconscious than when they were conscious and in those patients who died compared with those who survived.

Nitric oxide synthase inhibition with l-NAME reduces maximal oxygen uptake but not gas exchange threshold during incremental cycle exercise in man

We hypothesized that the effective inhibition of nitric oxide synthase (NOS), achieved via systemic infusion of NG‐nitro‐l‐arginine methyl ester (l‐NAME), would reduce the gas exchange threshold (GET) and the maximal oxygen uptake during incremental cycle exercise in man if NO is important in the regulation of muscle vasodilatation. Seven healthy males, aged 18–34 years, volunteered to participate in this ethically approved study. On two occasions, the subjects completed an incremental exercise test to exhaustion on an electrically braked cycle ergometer following the infusion of either l‐NAME (4 mg kg−1 in 50 ml saline) or placebo (50 ml saline, CON). At rest, the infusion of l‐NAME resulted in a significant increase in mean arterial pressure (MAP; CON vs. l‐NAME, 89 ± 8 vs. 103 ± 11 mmHg (mean ±s.d.; P < 0.05)) and a significant reduction in heart rate (HR; CON vs. l‐NAME, 60 ± 12 vs. 51 ± 8 beats min−1; P < 0.01). At submaximal work rates, there was no significant difference in between the conditions and no difference in the GET (CON vs. l‐NAME, 1.94 ± 0.47 vs. 2.01 ± 0.41 l min−1). However, at higher work rates, differences in between the conditions became more pronounced such that was significantly lower with l‐NAME (CON vs. l‐NAME, 4.02 ± 0.41 vs. 3.80 ± 0.34 l min−1; P < 0.05). The reduction in was associated with a reduction in HRmax (CON vs. l‐NAME, 186 ± 10 vs. 178 ± 7 beats min−1; P < 0.01). These results demonstrate that NOS inhibition with l‐NAME has no effect on GET but reduces during large muscle group exercise in man, presumably by direct or indirect effects on cardiac output and muscle blood flow.

Application of the [γ-32P] ATP kinase assay to study anabolic signaling in human skeletal muscle

AMPK (AMP-dependant protein kinase)-mTORC1 (mechanistic target of rapamycin in complex 1)-p70S6K1 (ribosomal protein S6 kinase 1 of 70 kDa) signaling plays a crucial role in muscle protein synthesis (MPS). Understanding this pathway has been advanced by the application of the Western blot (WB) technique. However, because many components of the mTORC1 pathway undergo numerous, multisite posttranslational modifications, solely studying the phosphorylation changes of mTORC1 and its substrates may not adequately represent the true metabolic signaling processes. The aim of this study was to develop and apply a quantitative in vitro [γ-(32)P] ATP kinase assay (KA) for p70S6K1 to assess kinase activity in human skeletal muscle to resistance exercise (RE) and protein feeding. In an initial series of experiments the assay was validated in tissue culture and in p70S6K1-knockout tissues. Following these experiments, the methodology was applied to assess p70S6K1 signaling responses to a physiologically relevant stimulus. Six men performed unilateral RE followed by the consumption of 20 g of protein. Muscle biopsies were obtained at pre-RE, and 1 and 3 h post-RE. In response to RE and protein consumption, p70S6K1 activity as assessed by the KA was significantly increased from pre-RE at 1 and 3 h post-RE. However, phosphorylated p70S6K1(thr389) was not significantly elevated. AMPK activity was suppressed from pre-RE at 3 h post-RE, whereas phosphorylated ACC(ser79) was unchanged. Total protein kinase B activity also was unchanged after RE from pre-RE levels. Of the other markers we assessed by WB, 4EBP1(thr37/46) phosphorylation was the only significant responder, being elevated at 3 h post-RE from pre-RE. These data highlight the utility of the KA to study skeletal muscle plasticity.