Karen Maes

Interleukin-6 Causes Myocardial Failure and Skeletal Muscle Atrophy in Rats

Background—The impact of interleukin (IL)-6 on skeletal muscle function remains the subject of controversy. Methods and Results—The effects of 7-day subcutaneous administration of recombinant human IL-6 were examined at 3 doses, 50, 100, or 250 &mgr;g · kg−1 · d−1, in rats. Skeletal muscle mass decreased dose-dependently (with increasing dose: in the diaphragm, −10%, P=NS; −15%, P=0.0561; and −15% P<0.05; and in the gastrocnemius, −9%, P=NS; −9%, P=NS; and −18%, P<0.005) because of decreases in cross-sectional area of all fiber types without alterations in diaphragm contractile properties. Cardiovascular variables showed a dose−dependent heart dilatation (for end-diastolic volume: control, 78 &mgr;L; moderate dose, 123 &mgr;L; and high dose, 137 &mgr;L, P<0.001), reduced end–systolic pressure (control, 113 mm Hg; moderate dose, 87 mm Hg; and high dose, 90 mm Hg; P=0.037), and decreased myocardial contractility (for preload recruitable stroke work: control, 79 mm Hg; moderate dose, 67 mm Hg; and high dose, 48 mm Hg; P<0.001). Lung edema was confirmed by an increased wet-to-dry ratio (control, 4.2; moderate dose, 4.6; and high dose, 4.5; P<0.001) and microscopy findings. These cardiovascular alterations led to decreases in organ blood flow, particularly in the diaphragm (control, 0.56 mL · min−1 · g−1; moderate dose, 0.21 mL · min−1 · g−1; and high dose, 0.23 mL · min−1 · g−1; P=0.037). In vitro recombinant human IL–6 administration did not cause any alterations in diaphragm force or endurance capacity. Conclusions—IL-6 clearly caused ventilatory and peripheral skeletal muscle atrophy, even after short-term administration. Blood flow redistribution, resulting from the myocardial failure induced by IL-6, was likely responsible for this muscle atrophy, because IL-6 did not exert any direct effect on the diaphragm.

Intermittent spontaneous breathing protects the rat diaphragm from mechanical ventilation effects

Objective:Short-term mechanical ventilation has been proven to reduce diaphragm force and fiber dimensions. We hypothesized that intermittent spontaneous breathing during the course of mechanical ventilation would minimize the effects of mechanical ventilation on diaphragm force and expression levels of transcription factors (MyoD and myogenin). Design:Randomized, controlled experiment. Setting:Animal basic science laboratory. Subjects:Male Wistar rats, weighing 350–500 g. Interventions:Anesthetized and tracheotomized rats were submitted to either 24 hrs of spontaneous breathing (SB, n = 5), 24 hrs of continuous controlled mechanical ventilation (CMV, n = 7), or controlled mechanical ventilation with intermittent spontaneous breathing: 60 mins every 5 hrs of mechanical ventilation repeated four times (ISB60, n = 8) or 5 mins every 5 hrs 55 mins of mechanical ventilation repeated four times (SB5, n = 9). They were compared with control animals free from intervention (C, n = 5). Measurements and Main Results:The profile of the diaphragm force-frequency curve of the controls and SB group was significantly different from that of the ISB and CMV groups; especially, the mean asymptotic force was less in the ISB and CMV compared with controls and SB. CMV resulted in a significant decrease in the diaphragm type I (−26%, p < .05 vs. C) and type IIx/b (−39%, p < .005 vs. C and SB) cross-sectional area, whereas this was not observed in the ISB groups. Diaphragm MyoD protein expression was significantly decreased after ISB60 (−35%, p < .0001 vs. C and SB) and even more after CMV (−73%, p < .0001 vs. others). The same pattern was observed with myogenin protein levels. Positive relationships between diaphragm MyoD and myogenin protein levels and diaphragm force were observed. Conclusions:The data demonstrated that intermittent spontaneous breathing during the course of mechanical ventilation may minimize the deleterious effect of controlled mechanical ventilation on diaphragm force, fiber dimensions, and expression of transcription factors.

Rocuronium exacerbates mechanical ventilation-induced diaphragm dysfunction in rats

Objective:Nondepolarizing neuromuscular blocking agents are commonly used in the intensive care setting, but they have occasionally been associated with development of myopathy. In addition, diaphragmatic atrophy and a reduction in diaphragmatic force were reported after short-term controlled mechanical ventilation in animal models. We hypothesized that infusion of rocuronium, an aminosteroidal neuromuscular blocking agent, during 24 hrs of controlled mechanical ventilation would further alter diaphragm function and would enhance activation of the ubiquitin– proteasome pathway. Design:Randomized, controlled experiment. Setting:Basic animal science laboratory. Subjects:Male Wistar rats, 14 wks old. Interventions:Rats were divided into four groups: a control group, a group of anesthetized rats breathing spontaneously for 24 hrs, and two groups submitted to mechanical ventilation for 24 hrs, receiving a continuous infusion of either 0.9% NaCl or rocuronium. Measurements and Main Results:In vitro diaphragm force was decreased more significantly after 24 hrs of mechanical ventilation combined with rocuronium infusion than after mechanical ventilation alone (e.g., tetanic force, −27%; p < .001 vs. mechanical ventilation). Similarly, the decrease in diaphragm type IIx/b fiber dimensions was more pronounced after mechanical ventilation with rocuronium treatment than with saline treatment (−38% and −29%, respectively; p < .001 vs. control). Diaphragm hydroperoxide levels increased similarly in both mechanically ventilated groups. Diaphragm muscle RING-finger protein-1 (MURF-1) messenger RNA expression, an E3 ligase of the ubiquitin–proteasome pathway, increased after mechanical ventilation (+212%, p < .001 vs. control) and increased further with combination of rocuronium (+320%, p < .001 vs. control). Significant correlations were found between expression of MURF-1 messenger RNA, diaphragm force, and type IIx/b fiber dimensions. Conclusions:Infusion of rocuronium during controlled mechanical ventilation leads to further deterioration of diaphragm function, additional atrophy of type IIx/b fibers, and an increase in MURF-1 messenger RNA in the diaphragm, which suggests an activation of the ubiquitin–proteasome pathway. These findings could be important with regard to weaning failure in patients receiving this drug for prolonged periods in the intensive care unit setting.

Long-term nose-only cigarette smoke exposure induces emphysema and mild skeletal muscle dysfunction in mice.

SUMMARY Mouse models of chronic obstructive pulmonary disease (COPD) focus on airway inflammation and lung histology, but their use has been hampered by the lack of pulmonary function data in their assessment. Systemic effects such as muscle dysfunction are also poorly modeled in emphysematous mice. We aimed to develop a cigarette-smoke-induced emphysema mouse model in which serial lung function and muscular dysfunction could be assessed, allowing the disease to be monitored more appropriately. C57Bl6 mice were nose-only exposed to cigarette smoke or filtered air for 3–6 months. Lung function tests were repeated in the same mice after 3 and 6 months of cigarette smoke or air exposure and compared with lung histological changes. Contractile properties of skeletal muscles and muscle histology were also determined at similar time points in separate groups of mice. Serial lung function measurements documented hyperinflation after 3 and 6 months of cigarette smoke exposure, with a significant 31–37% increase in total lung capacity (TLC) and a significant 26–35% increase in compliance (Cchord) when compared with animals exposed to filtered air only (P<0.001 after 3 and after 6 months). These functional changes preceded the changes in mean linear intercept, which became only significant after 6 months of cigarette smoke exposure and which correlated very well with TLC (r=0.74, P=0.004) and Cchord (r=0.79, P=0.001). After 6 months of cigarette smoke exposure, a significant fiber-type shift from IIa to IIx/b was also observed in the soleus muscle (P<0.05), whereas a 20% reduction of force was present at high stimulation frequencies (80 Hz; P=0.09). The extensor digitorum longus (EDL) muscle was not affected by cigarette smoke exposure. These serial pulmonary function variables are sensitive outcomes to detect emphysema progression in a nose-only cigarette-smoke-exposed animal model of COPD. In this model, muscular changes became apparent only after 6 months, particularly in muscles with a mixed fiber-type composition.

Atrophy and hypertrophy signalling in the diaphragm of patients with COPD

We investigated whether atrophy and hypertrophy signalling were altered in the diaphragm of chronic obstructive pulmonary disease (COPD) patients. We studied diaphragm fibre dimensions and proportion, expression of markers of the ubiquitin-proteasome pathway, nuclear factor (NF)-κB pathways, muscle regulatory factors and myostatin in diaphragm biopsies from 19 patients with severe COPD and 13 patients without COPD. Type I proportion was significantly increased in the diaphragm of COPD patients while type II proportion was decreased. The cross-sectional area of all fibre types was reduced in the COPD patients. In addition, MAFbx mRNA was higher in the diaphragm of COPD patients while Nedd4 mRNA decreased. Cytoplasmatic levels of inhibitor protein IκBα and IκBβ were decreased in the COPD patients as was NF-κB p50 DNA-binding activity. MyoD mRNA and its nuclear protein content were decreased in the diaphragm of COPD patients and myogenin mRNA and protein levels remained unchanged. Myostatin mRNA was decreased but its protein levels in the nuclear and cytoplasmic fraction were significantly increased in the COPD patients. These data show that the ubiquitin-proteasome pathway, the NF-κB pathway and myostatin protein were up-regulated in the diaphragm of COPD patients while MyoD expression was reduced. These alterations may contribute to diaphragm remodeling in COPD.

Effects of Acute Administration of Corticosteroids during Mechanical Ventilation on Rat Diaphragm

RATIONALE Mechanical ventilation is known to induce ventilator-induced diaphragm dysfunction. Patients submitted to mechanical ventilation often receive massive doses of corticosteroids that may cause further deterioration of diaphragm function. OBJECTIVES To examine whether the combination of 24 hours of controlled mechanical ventilation with corticosteroid administration would exacerbate ventilator-induced diaphragm dysfunction. METHODS Rats were randomly assigned to a group submitted to 24 hours of controlled mechanical ventilation receiving an intramuscular injection of saline or 80 mg/kg methylprednisolone, a group submitted to 24 hours of spontaneous breathing receiving saline, or methylprednisolone and a control group. MEASUREMENTS AND MAIN RESULTS The diaphragm force-frequency curve was shifted downward in the mechanical ventilation group, but this deleterious effect was prevented when corticosteroids were administered. Diaphragm cross-sectional area of type I fibers was similarly decreased in both mechanical ventilation groups while atrophy of type IIx/b fibers was attenuated after corticosteroid administration. The mechanical ventilation-induced reduction in diaphragm MyoD and myogenin protein expression was attenuated after corticosteroids. Plasma cytokine levels were unchanged while diaphragm lipid hydroperoxides were similarly increased in both mechanical ventilation groups. Diaphragmatic calpain activity was significantly increased in the mechanical ventilation group, but calpain activation was abated with corticosteroid administration. Inverse correlations were found between calpain activity and diaphragm force. CONCLUSIONS A single high dose of methylprednisolone combined with controlled mechanical ventilation protected diaphragm function from the deleterious effects of controlled mechanical ventilation. Inhibition of the calpain system is most likely the mechanism by which corticosteroids induce this protective effect.

N-Acetylcysteine protects the rat diaphragm from the decreased contractility associated with controlled mechanical ventilation.

Objective:Controlled mechanical ventilation results in diaphragmatic dysfunction, and oxidative stress has been shown to be an important contributor to ventilator-induced diaphragm dysfunction. We hypothesized that the administration of an antioxidant, N-acetylcysteine, would restore the redox balance in the diaphragm and prevent against the deleterious effects of controlled mechanical ventilation. Design:Randomized, controlled experiment. Settings:Basic science animal laboratory. Subjects:Male Wistar rats, 14 wks old. Interventions:Anesthetized rats were submitted for 24 hrs to either spontaneous breathing receiving 150 mg/kg N-acetylcysteine (SBNAC) or saline (SBSAL) or to controlled mechanical ventilation receiving 150 mg/kg N-acetylcysteine (MVNAC) or saline (MVSAL). Measurements and Main Results:After 24 hrs of controlled mechanical ventilation, diaphragmatic force production was significantly lower in MVSAL compared with all groups. Importantly, administration of N-acetylcysteine completely abolished this controlled mechanical ventilation-induced diaphragmatic contractile dysfunction. Diaphragmatic protein oxidation was significantly increased after 24 hrs of controlled mechanical ventilation (+53%, p < .01) in MVSAL animals, whereas administration of N-acetylcysteine prevented this controlled mechanical ventilation-induced oxidative stress. Diaphragmatic 20S proteasome activity was increased in MVSAL (+62%, p < .05). Further, compared with SBSAL, diaphragm caspase-3 activity was significantly increased in MVSAL (+279%, p < .001), and N-acetylcysteine treatment provided partial protection against caspase-3 activation. Diaphragmatic calpain activity was significantly increased after controlled mechanical ventilation (+137%, p < .001) in MVSAL animals, but N-acetylcysteine treatment protected against this event. Finally, significant negative correlations existed between calpain activity and diaphragm force production (r from −0.56 to −0.49, p < .05). Conclusions:These data show that the administration of N-acetylcysteine protects the diaphragm from the deleterious effects of controlled mechanical ventilation. Specifically, N-acetylcysteine prevents against controlled mechanical ventilation-induced diaphragmatic oxidative stress and proteolysis and abolishes controlled mechanical ventilation-induced diaphragmatic contractile dysfunction.

Vitamin D supplementation during rehabilitation in COPD: a secondary analysis of a randomized trial

RationalePulmonary rehabilitation is an important treatment for patients with Chronic Obstructive Pulmonary Disease, who are often vitamin D deficient. As vitamin D status is linked to skeletal muscle function, we aimed to explore if high dose vitamin D supplementation can improve the outcomes of rehabilitation in Chronic Obstructive Pulmonary Disease.Material and methodsThis study is a post-hoc subgroup analysis of a larger randomized trial comparing a monthly dose of 100.000 IU of vitamin D with placebo to reduce exacerbations. 50 Subjects who followed a rehabilitation program during the trial are included in this analysis. We report changes from baseline in muscle strength and exercise performance between both study arms after 3 months of rehabilitation.ResultsVitamin D intervention resulted in significantly higher median vitamin D levels compared to placebo (51 [44-62] ng/ml vs 15 [13-30] ng/ml; p < 0.001). Patients receiving vitamin D had significantly larger improvements in inspiratory muscle strength (-11±12 cmH2O vs 0±14 cmH2O; p = 0.004) and maximal oxygen uptake (110±211 ml/min vs -20±187 ml/min; p = 0.029). Improvements in quadriceps strength (15±16 Nm) or six minutes walking distance (40±55 meter) were not significantly different from the effects in the placebo group (7±19 Nm and 11±74 meter; p>0.050).ConclusionHigh dose vitamin D supplementation during rehabilitation may have mild additional benefits to training.

Musculoskeletal Disorders in Chronic Obstructive Pulmonary Disease

Chronic obstructive pulmonary disease (COPD) is a lung disease characterized by airway obstruction and inflammation but also accompanied by several extrapulmonary consequences, such as skeletal muscle weakness and osteoporosis. Skeletal muscle weakness is of major concern, since it leads to poor functional capacity, impaired health status, increased healthcare utilization, and even mortality, independently of lung function. Osteoporosis leads to fractures and is associated with increased mortality, functional decline, loss of quality of life, and need for institutionalization. Therefore, the presence of the combination of these comorbidities will have a negative impact on daily life in patients with COPD. In this review, we will focus on these two comorbidities, their prevalence in COPD, combined risk factors, and pathogenesis. We will try to prove the clustering of these comorbidities and discuss possible preventive or therapeutic strategies.

Bortezomib partially protects the rat diaphragm from ventilator-induced diaphragm dysfunction.

Objective: Controlled mechanical ventilation leads to diaphragmatic contractile dysfunction and atrophy. Since proteolysis is enhanced in the diaphragm during controlled mechanical ventilation, we examined whether the administration of a proteasome inhibitor, bortezomib, would have a protective effect against ventilator-induced diaphragm dysfunction. Design: Randomized, controlled experiment. Settings: Basic science animal laboratory. Interventions: Anesthetized rats were submitted for 24 hrs to controlled mechanical ventilation while receiving 0.05 mg/kg bortezomib or saline. Control rats were acutely anesthetized. Measurements and Main Results: After 24 hrs, diaphragm force production was significantly lower in mechanically ventilated animals receiving an injection of saline compared to control animals (−36%, p < .001). Importantly, administration of bortezomib improved the diaphragmatic force compared to mechanically ventilated animals receiving an injection of saline (+15%, p < .01), but force did not return to control levels. Compared to control animals, diaphragm cross-sectional area of the type IIx/b fibers was significantly decreased by 28% in mechanically ventilated animals receiving an injection of saline (p < .01) and by 16% in mechanically ventilated animals receiving an injection of bortezomib (p < .05). Diaphragmatic calpain activity was significantly increased in mechanically ventilated animals receiving an injection of saline (+52%, p < .05) and in mechanically ventilated animals receiving an injection of bortezomib (+36%, p < .05). Caspase-3 activity was increased after controlled mechanical ventilation with saline by 55% (p < .05), while it remained similar to control animals in mechanically ventilated animals receiving an injection of bortezomib. Diaphragm 20S proteasome activity was slightly increased in both ventilated groups, and the amount of ubiquitinated proteins was significantly and similarly enhanced in mechanically ventilated animals receiving an injection of saline and mechanically ventilated animals receiving an injection of bortezomib. Conclusions: These data show that the administration of bortezomib partially protects the diaphragm from controlled mechanical ventilation–induced diaphragm contractile dysfunction without preventing atrophy. The fact that calpain activity was still increased after bortezomib treatment may explain the persistence of atrophy. Part of bortezomib effects might have been due to its ability to inhibit caspase-3 in this model.