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Introduction: Histological data on muscle fiber size and proportion in (very) young typically developing (TD) children is not well documented and data on capillarization and satellite cell content are also lacking. Aims: This study investigated the microscopic properties of the medial gastrocnemius muscle in growing TD children, grouped according to age and gender to provide normal reference values in healthy children. Methods: Microbiopsies of the medial gastrocnemius (MG) muscle were collected in 46 TD boys and girls aged 2-10 years subdivided into 4 age groups (2-4, 4-6, 6-8 and 8-10 years). Sections were immunostained to assess fiber type cross-sectional area (fCSA) and proportion, the number of satellite cells (SC), capillary to fiber ratio (C/F), capillary density for type I and II fiber (CFD), capillary domain, capillary-to-fiber perimeter exchange index (CFPE) and heterogeneity index. fCSA was normalized to fibula length2 and the coefficient of variation (CV) was calculated to reflect fCSA intrasubject variability. Results: Absolute fCSA of all fibers increased with age (r = 0.72, p < 0.001) but more in boys (+112%, p < 0.05) than in girls (+48%, p > 0.05) Normalized fCSA, CV and fiber proportion did not differ between age groups and gender. C/F was strongly correlated with age in boys (r = 0.83, p < 0.001), and to a lesser extent in girls (r = 0.37, p = 0.115), while other capillary parameters as well as the number of SC remained stable with increasing age in boys and girls. Discussion: This study provides reference values of histological measures in MG according to age in normally growing boys and girls. These data may be used as a reference to determine disease impact and efficacy of therapeutic approach on the muscle. [ABSTRACT FROM AUTHOR]

Inconsistent alterations in skeletal muscle histology have been reported in adolescents with cerebral palsy (CP) and whether alterations are present in young children and differ from older children is not yet known. This study aimed to define histological alterations in the medial gastrocnemius (MG) of ambulant CP (gross-motor classification system, GMFCS I-III) stratified in two age groups (preschool children, PS: 2–5 and school age children, SA: 6–9-yr old) compared with age-matched typically developing (TD) children. We hypothesized that alterations in muscle microscopic properties are already present in PS-CP and are GMFCS level specific. Ultrasound guided percutaneous microbiopsies were collected in 46 CP (24-PS) and 45 TD (13-PS) children. Sections were stained to determine fiber cross-sectional area (fCSA) and proportion, capillary, and satellite cell amount. Average absolute and normalized fCSA were similar in CP and TD, but a greater percentage of smaller fibers was found in CP. Coefficient of variation (CV) was significantly larger in PS-CP-GMFCS I-II and for type I fiber. In SA-CP, all fiber types contributed to the higher CV. Type IIx proportion was higher and type I was lower in PS-CP-GMFCS-III and for all SA-CP. Reduced capillaryto-fiber ratio was present in PS-CP-GMFCS II-III and in all SA-CP. Capillary fiber density was lower in SA-CP. Capillary domain was enhanced in all CP, but capillary spatial distribution was maintained as was satellite cell content. We concluded that MG histological alterations are already present in very young CP but are only partly specific for GMFCS level and age. NEW & NOTEWORTHY Inconsistent histological alterations have been reported in children with cerebral palsy (CP) but whether they are present in very young and ambulant CP children and differ from those reported in old CP children is not known. This study highlighted for the first time that enhanced muscle fiber size variability and loss of capillaries are already present in very young CP children, even in the most ambulant ones, and these alterations seem to extend with age. [ABSTRACT FROM AUTHOR]

Botulinum toxin-A (BoNT-A) injection is known to exert beneficial effects on muscle tone, joint mobility and gait in children with cerebral palsy (CP). However, recent animal and human studies have raised the concern that BoNT-A might be harmful to muscle integrity. In CP-children, the impact of BoNT-A on muscle structure has been poorly studied, and inconsistent results have been reported. This study was aimed at determining the time course effect of a single BoNT-A administration on medial gastrocnemius (MG) morphology in CP-children. MG microbiopsies from 12 ambulant and BoNT-A-naïve CP-children (age, 3.4 (2.3) years, ranging from 2.5 to 7.8 years; seven boys and five girls; GMFCS I = 5, II = 4 and III = 3) were collected before and 3 and 6 months after BoNT-A treatment to analyze the fiber cross-sectional area (fCSA) and proportion; capillarization; and satellite cell (SC) content. Compared with the baseline, the fCSA decreased at 3 months (−14%, NS) and increased at 6 months (+13%, NS). Fiber size variability was significantly higher at 3 months (type I: +56%, p = 0.032; type IIa: +37%, p = 0.032) and 6 months (type I: +69%, p = 0.04; type IIa: +121%, p = 0.032) compared with the baseline. The higher type I proportion seen at 3 months was still present and more pronounced at 6 months (type I: +17%, p = 0.04; type IIx: −65%, p = 0.032). The capillary fiber density was reduced at 3 months (type I: −43%, NS; type II: −44%, p = 0.0320) but normalized at 6 months. There was a non-significant increase in SC/100 fibers at 3 months (+75%, NS) and 6 months (+40%, NS) compared with the baseline. These preliminary data suggest that BoNT-A induced alterations in the MG of children with CP, which were still present 6 months after BoNT-A injection but with signs of muscle recovery. [ABSTRACT FROM AUTHOR]

Cerebral palsy (CP) is a heterogeneous group of motor disorders attributed to a non-progressive lesion in the developing brain. Knowledge on skeletal muscle properties is important to understand the impact of CP and treatment but data at the microscopic levels are limited and inconsistent. Currently, muscle biopsies are collected during surgery and are restricted to CP eligible for such treatment or they may refer to another muscle or older children in typically developing (TD) biopsies. A minimally invasive technique to collect (repeated) muscle biopsies in young CP and TD children is needed to provide insights into the early muscle microscopic alterations and their evolution in CP. This paper describes the protocol used to 1) collect microbiopsies of the medial gastrocnemius (MG) and semitendinosus (ST) in CP children and age-matched TD children, 2) handle the biopsies for histology, 3) stain the biopsies to address muscle structure (Hematoxylin & Eosin), fiber size and proportion (myosin heavy chain), counting of the satellite cells (Pax7) and capillaries (CD31). Technique feasibility and safety as well as staining feasibility and measure accuracy were evaluated. Two microbiopsies per muscle were collected in 56 CP (5.8±1.1 yr) and 32 TD (6±1.1 yr) children using ultrasound-guided percutaneous microbiopsy technique. The biopsy procedure was safe (absence of complications) and well tolerated (Score pain using Wong-Baker faces). Cross-sectionally orientated fibers were found in 86% (CP) and 92% (TD) of the biopsies with 60% (CP) and 85% (TD) containing more than 150 fibers. Fiber staining was successful in all MG biopsies but failed in 30% (CP) and 16% (TD) of the ST biopsies. Satellite cell staining was successful in 89% (CP) and 85% (TD) for MG and in 70% (CP) and 90% (TD) for ST biopsies, while capillary staining was successful in 88% (CP) and 100% (TD) of the MG and in 86% (CP) and 90% (TD) for the ST biopsies. Intraclass coefficient correlation showed reliable and reproducible measures of all outcomes. This study shows that the percutaneous microbiopsy technique is a safe and feasible tool to collect (repeated) muscle biopsies in young CP and TD children for histological analysis and it provides sufficient muscle tissue of good quality for reliable quantification. [ABSTRACT FROM AUTHOR]

Background: Evidence supports a critical role of vitamin D status on exacerbation in chronic obstructive pulmonary disease, indicating the need to avoid vitamin D deficiency in these patients. However, oral vitamin D supplementation is limited by the potential risk for hypercalcemia. In this study, we investigated if local delivery of vitamin D to the lungs improves vitamin D-mediated anti-inflammatory action in response to acute inflammation without inducing hypercalcemia.Methods: We studied vitamin D sufficient (VDS) or deficient (VDD) mice in whom 1α,25(OH)2D3 (0.2 μg/kg) or a vehicle followed by lipopolysaccharide (LPS 25 µg) were delivered to the lung as a micro-spray.Results: Local 1α,25(OH)2D3 reduced LPS-induced inflammatory cells in bronchoalveolar lavage (BAL) in VDS (absolute number of cells: - 57% and neutrophils - 51% p < 0.01) and tended to diminish LPS-increased CXCL5 BAL levels in VDS (- 40%, p = 0.05) while it had no effect on CXCL1 and CXCL2 in BAL and mRNA in lung of VDS and VDD. It also significantly attenuated the increased IL-13 in BAL and lung, especially in VDD mice (- 41 and - 75%, respectively). mRNA expression of Claudin-18 in lung was significantly lower in VDS mice with local 1α,25(OH)2D3 while Claudin-3, -5 and -8 mRNA levels remained unchanged. Finally, in VDD mice only, LPS reduced lung mRNA expression of adhesion junction Zona-occludens-1, in addition to increasing uric acid and total protein in BAL, which both were prevented by local 1α,25(OH)2D3.Conclusion: Under normal levels of vitamin D, local 1α,25(OH)2D3 nebulization into the lung efficiently reduced LPS induction of inflammatory cells in BAL and slightly attenuated LPS-increase in CXCL5. In case of severe vitamin D deficiency, although local 1α,25(OH)2D3 nebulization failed to significantly minimize cellular inflammation in BAL at this dose, it prevented epithelial barrier leakage and damage in lung. Additional research is needed to determine the potential long-term beneficial effects of local 1α,25(OH)2D3 nebulization on lung inflammation. [ABSTRACT FROM AUTHOR]

Vitamin D deficiency, which is highly prevalent in the general population, exerts similar deleterious effects on skeletal muscles to those induced by cigarette smoking. We examined whether cigarette smoke (CS) exposure and/or vitamin D deficiency impairs the skeletal muscle hypertrophic response to overload. Male C57Bl/6JolaH mice on a normal or vitamin D-deficient diet were exposed to CS or room air for 18 wk. Six weeks after initiation of smoke or air exposure, sham surgery or denervation of the agonists of the left plantaris muscle was performed. The right leg served as internal control. Twelve weeks later, the hypertrophic response was assessed. CS exposure instigated loss of body and muscle mass, and increased lung inflammatory cell infiltration (P < 0.05), independently of diet. Maximal exercise capacity, whole body strength, in situ plantaris muscle force, and key markers of hypertrophic signaling (Akt, 4EBP1, and FoxO1) were not significantly affected by smoking or diet. The increase in plantaris muscle fiber cross-sectional area in response to overload was attenuated in vitamin D-deficient CS-exposed mice (smoking × diet interaction for hypertrophy, P = 0.03). In situ fatigue resistance was elevated in hypertrophied plantaris, irrespective of vitamin D deficiency and/or CS exposure. In conclusion, our data show that CS exposure or vitamin D deficiency alone did not attenuate the hypertrophic response of overloaded plantaris muscles, but this hypertrophic response was weakened when both conditions were combined. These data suggest that current smokers who also present with vitamin D deficiency may be less likely to respond to a training program. NEW & NOTEWORTHY Plantaris hypertrophy caused by compensatory overload after denervation of the soleus and gastrocnemius muscles showed increased mass and fiber dimensions, but to a lesser extent when vitamin D deficiency was combined with cigarette smoking. Fatigue resistance was elevated in hypertrophied plantaris, irrespective of diet or smoking, whereas physical fitness, hypertrophic markers, and in situ plantaris force were similar. These data showed that the hypertrophic response to overload is attenuated when both conditions are combined. [ABSTRACT FROM AUTHOR]

Abstract Rationale Pulmonary 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 methods This 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. Results Vitamin 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). Conclusion High dose vitamin D supplementation during rehabilitation may have mild additional benefits to training.

Abstract Background High dose of corticosteroids has been previously shown to protect against controlled mechanical ventilation (CMV)-induced diaphragmatic dysfunction while inhibiting calpain activation. Because literature suggests that the calpain inhibiting effect of corticosteroid depends on the dose administered, we determined whether lower doses of corticosteroids would also provide protection of the diaphragm during CMV. This may be important for patients undergoing mechanical ventilation and receiving corticosteroids. Methods Rats were assigned to controls or to 24 hours of CMV while being treated at the start of mechanical ventilation with a single intramuscular administration of either saline, or 5 mg/kg (low MP) or 30 mg/kg (high MP) of methylprednisolone. Results Diaphragmatic force was decreased after CMV and this was exacerbated in the low MP group while high MP rescued this diaphragmatic dysfunction. Atrophy was more severe in the low MP group than after CMV while no atrophy was observed in the high MP group. A significant and similar increase in calpain activity was observed in both the low MP and CMV groups whereas the high dose prevented calpain activation. Expression of calpastatin, the endogenous inhibitor of calpain, was decreased in the CMV and low MP groups but its level was preserved to controls in the high MP group. Caspase-3 activity increased in all CMV groups but to a lesser extent in the low and high MP groups. The 20S proteasome activity was increased in CMV only. Conclusions Administration of 30 mg/kg methylprednisolone during CMV protected against CMV-induced diaphragm dysfunction while 5 mg/kg was more deleterious. The protective effect is due mainly to an inhibition of the calpain system through preservation of calpastatin levels and to a lesser extent to a caspase-3 inhibition.

Introduction: Apart from its adverse effects on the respiratory system, cigarette smoking also induces skeletal muscle atrophy and dysfunction. Whether short-term smoking cessation can restore muscle mass and function is unknown. We, therefore, studied the impact of 1- and 2-week smoking cessation on skeletal muscles in a mouse model.Methods: Male mice were divided into four groups: Air-exposed (14 weeks); cigarette smoke (CS)-exposed (14 weeks); CS-exposed (13 weeks) followed by 1-week cessation; CS-exposed (12 weeks) followed by 2 weeks cessation to examine exercise capacity, physical activity levels, body composition, muscle function, capillarization, mitochondrial function and protein expression in the soleus, plantaris, and diaphragm muscles.Results: CS-induced loss of body and muscle mass was significantly improved within 1 week of cessation due to increased lean and fat mass. Mitochondrial respiration and protein levels of the respiratory complexes in the soleus were lower in CS-exposed mice, but similar to control values after 2 weeks of cessation. Exposing isolated soleus muscles to CS extracts reduced mitochondrial respiration that was reversed after removing the extract. While physical activity was reduced in all groups, exercise capacity, limb muscle force, fatigue resistance, fiber size and capillarization, and diaphragm cytoplasmic HIF-1α were unaltered by CS-exposure. However, CS-induced diaphragm atrophy and increased capillary density were not seen after 2 weeks of smoking cessation.Conclusion: In male mice, 2 weeks of smoking cessation reversed smoking-induced mitochondrial dysfunction, limb muscle mass loss, and diaphragm muscle atrophy, highlighting immediate benefits of cessation on skeletal muscles.Implications: Our study demonstrates that CS-induced skeletal muscle mitochondrial dysfunction and atrophy are significantly improved by 2 weeks of cessation in male mice. We show for the first time that smoking cessation as short as 1 to 2 weeks is associated with immediate beneficial effects on skeletal muscle structure and function with the diaphragm being particularly sensitive to CS-exposure and cessation. This could help motivate smokers to quit smoking as early as possible. The knowledge that smoking cessation has potential positive extrapulmonary effects is particularly relevant for patients referred to rehabilitation programs and those admitted to hospitals suffering from acute or chronic muscle deterioration yet struggling with smoking cessation. [ABSTRACT FROM AUTHOR]

Cerebral palsy (CP), the single largest cause of childhood physical disability, is characterized firstly by a lesion in the immature brain, and secondly by musculoskeletal problems that progress with age. Previous research reported altered muscle properties, such as reduced volume and satellite cell (SC) numbers and hypertrophic extracellular matrix compared to typically developing (TD) children (>10 years). Unfortunately, data on younger CP patients are scarce and studies on SCs and other muscle stem cells in CP are insufficient or lacking. Therefore, it remains difficult to understand the early onset and trajectory of altered muscle properties in growing CP children. Because muscle stem cells are responsible for postnatal growth, repair and remodeling, multiple adult stem cell populations from young CP children could play a role in altered muscle development. To this end, new methods for studying muscle samples of young children, valid to delineate the features and to elucidate the regenerative potential of muscle tissue, are necessary. Using minimal invasive muscle microbiopsy, which was applied in young subjects under general anaesthesia for the first time, we aimed to isolate and characterize muscle stem cell-derived progenitors of TD children and patients with CP. Data of 15 CP patients, 3–9 years old, and 5 aged-matched TD children were reported. The muscle microbiopsy technique was tolerated well in all participants. Through the explant technique, we provided muscle stem cell-derived progenitors from the Medial Gastrocnemius. Via fluorescent activated cell sorting, using surface markers CD56, ALP, and PDGFRa, we obtained SC-derived progenitors, mesoangioblasts and fibro-adipogenic progenitors, respectively. Adipogenic, skeletal, and smooth muscle differentiation assays confirmed the cell identity and ability to give rise to different cell types after appropriate stimuli. Myogenic differentiation in CP SC-derived progenitors showed enhanced fusion index and altered myotube formation based on MYOSIN HEAVY CHAIN expression, as well as disorganization of nuclear spreading, which were not observed in TD myotubes. In conclusion, the microbiopsy technique allows more focused muscle research in young CP patients. Current results show altered differentiation abilities of muscle stem cell-derived progenitors and support the hypothesis of their involvement in CP-altered muscle growth. [ABSTRACT FROM AUTHOR]

Background: Patient counselling and addressing drug-related problems are the pharmacist’s key activities to ensure the safe and effective use of medicines. This study aimed to describe the dispensing practice of prescribed medicines in daily community pharmacy practice and to identify factors influencing counselling provision; Methods: An observational study was conducted in community pharmacies in Basel, Switzerland. One master student in pharmacy performed non-participatory observations for one day at each of the participating community pharmacies. Patient characteristics, counselling content, additional activities, and pharmaceutical interventions were documented on a structured checklist; Results: 556 prescription encounters (PE) in 18 participating community pharmacies were observed (269 first prescriptions; 287 refill prescriptions). Patients were regular customers (n = 523, 94.1%) and 53.8 ± 23.4 years old. Counselling was provided to 367 (66.0%) customers on 2.9 ± 3.1 themes per PE. Factors influencing counselling were dispensing by the pharmacist, new customer, customer who did not refuse counselling, customer with a first prescription, with a prescription resulting in a pharmaceutical intervention, and a prescription filled by carers. During 144 PEs, 203 interventions were documented. Pharmacists proposed few additional activities and performed no cognitive pharmaceutical service; Conclusions: Our study quantified counselling and additional services at the dispensing of prescribed medicines and identified influencing factors on counselling provision at the patient, prescription, and pharmacy level. [ABSTRACT FROM AUTHOR]

Object recognition improves with training. This training effect only partially generalizes to untrained images of the trained objects (new exemplars, orientation,: : :). The aim of this study is to investigate whether and to what extent the learning transfer improves when participants are trained with more exemplars of an object. Participants were trained to recognize two sets of stimuli using a backward masking paradigm. During training with the first set, only one exemplar of each object was presented. The second set was trained using four exemplars of each object. After 3 days of training, participants were tested on all the trained exemplars and a completely new exemplar of the same objects. In addition, recognition performance was compared to a set of completely new objects. For the objects of which four exemplars were used during training, participants showed more generalization toward new exemplars compared to when they were only trained with one exemplar. Part of the generalization effect extended to completely new objects. In conclusion, more variation during training leads to more generalization toward new visual stimuli. [ABSTRACT FROM AUTHOR]

Background: Mechanical ventilation (MV) is associated with diaphragm weakness, a phenomenon termed ventilator-induced diaphragmatic dysfunction. Weaning should balance diaphragmatic loading as well as prevention of overload after MV. The weaning methods pressure support ventilation (PSV) and spontaneous breathing trials (SBT) lead to gradual or intermittent reloading of a weak diaphragm, respectively. This study investigated which weaning method allows more efficient restoration of diaphragm homeostasis.Methods: Rats (n = 8 per group) received 12 h of MV followed by either 12 h of pressure support ventilation (PSV) or intermittent spontaneous breathing trials (SBT) and were compared to rats euthanized after 12 h MV (CMV) and to acutely euthanized rats (CON). Force generation, activity of calpain-1 and caspase-3, oxidative stress, and markers of protein synthesis (phosphorylated AKT to total AKT) were measured in the diaphragm.Results: Reduction of diaphragmatic force caused by CMV compared to CON was worsened with PSV and SBT (both p < 0.05 vs. CON and CMV). Both PSV and SBT reversed oxidative stress and calpain-1 activation caused by CMV. Reduced pAKT/AKT was observed after CMV and both weaning procedures.Conclusions: MV resulted in a loss of diaphragmatic contractility, which was aggravated in SBT and PSV despite reversal of oxidative stress and proteolysis. [ABSTRACT FROM AUTHOR]

Chronic obstructive pulmonary disease (COPD) is associated with skeletal muscle dysfunction. Vitamin D plays an important role in muscle strength and performance in healthy individuals. Vitamin D deficiency is highly prevalent in COPD, but its role in skeletal muscle dysfunction remains unclear. We examined the time-course effect of vitamin D deficiency on limb muscle function in mice with normal or deficient vitamin D serum levels exposed to air or cigarette smoke for 6, 12 or 18 weeks. The synergy of smoking and vitamin D deficiency increased lung inflammation and lung compliance from 6 weeks on with highest emphysema scores observed at 18 weeks. Smoking reduced body and muscle mass of the soleus and extensor digitorum longus (EDL), but did not affect contractility, despite type II atrophy. Vitamin D deficiency did not alter muscle mass but reduced muscle force over time, downregulated vitamin D receptor expression, and increased muscle lipid peroxidation but did not alter actin and myosin expression, fiber dimensions or twitch relaxation time. The combined effect of smoking and vitamin D deficiency did not further deteriorate muscle function but worsened soleus mass loss and EDL fiber atrophy at 18 weeks. We conclude that the synergy of smoking and vitamin D deficiency in contrast to its effect on lung disease, had different, independent but important noxious effects on skeletal muscles in a mouse model of mild COPD. [ABSTRACT FROM AUTHOR]

Background: Chronic obstructive pulmonary disease (COPD) is characterized by excessive inflammation and disturbed bacterial clearance in the airways. Although cigarette smoke (CS) exposure poses a major risk, vitamin D deficiency could potentially contribute to COPD progression. Many in vitro studies demonstrate important anti-inflammatory and antibacterial effects of vitamin D, but a direct contribution of vitamin D deficiency to COPD onset and disease progression has not been explored.Methods: In the current study, we used a murine experimental model to investigate the combined effect of vitamin D deficiency and CS exposure on the development of COPD-like characteristics. Therefore, vitamin D deficient or control mice were exposed to CS or ambient air for a period of 6 (subacute) or 12 weeks (chronic). Besides lung function and structure measurements, we performed an in depth analysis of the size and composition of the cellular infiltrate in the airways and lung parenchyma and tested the ex vivo phagocytic and oxidative burst capacity of alveolar macrophages.Results: Vitamin D deficient mice exhibited an accelerated lung function decline following CS exposure compared to control mice. Furthermore, early signs of emphysema were only observed in CS-exposed vitamin D deficient mice, which was accompanied by elevated levels of MMP-12 in the lung. Vitamin D deficient mice showed exacerbated infiltration of inflammatory cells in the airways and lung parenchyma after both subacute and chronic CS exposure compared to control mice. Furthermore, elevated levels of typical proinflammatory cytokines and chemokines could be detected in the bronchoalveolar lavage fluid (KC and TNF-α) and lung tissue (IP-10, MCP-1, IL-12) of CS-exposed vitamin D deficient mice compared to control mice. Finally, although CS greatly impaired the ex vivo phagocytic and oxidative burst function of alveolar macrophages, vitamin D deficient mice did not feature an additional defect.Conclusions: Our data demonstrate that vitamin D deficiency both accelerates and aggravates the development of characteristic disease features of COPD. As vitamin D deficiency is highly prevalent, large randomized trials exploring effects of vitamin D supplementation on lung function decline and COPD onset are needed. [ABSTRACT FROM AUTHOR]

Objectives: Diaphragm dysfunction develops during severe sepsis as a consequence of hemodynamic, metabolic, and intrinsic abnormalities. Similarly, 12 hours of controlled mechanical ventilation also promotes diaphragm dysfunction. Importantly, patients with sepsis are often treated with mechanical ventilation for several days. It is unknown if controlled mechanical ventilation exacerbates sepsis-induced diaphragm dysfunction, and this forms the basis for these experiments. We investigate the effects of 12-hour controlled mechanical ventilation on contractile function, fiber dimension, cytokine production, proteolysis, autophagy, and oxidative stress in the diaphragm of septic rats. Design: Randomized controlled experiment. Setting: Animal research laboratory. Subjects: Adult male Wistar rats. Interventions: Treatment with a single intraperitoneal injection of either saline or Escherichia coli lipopolysaccharide (5 mg/kg). After 12 hours, the saline-treated animals (controlled mechanical ventilation) and half of the septic animals (lipopolysaccharide + controlled mechanical ventilation) were submitted to 12 hours of controlled mechanical ventilation while the remaining septic animals (lipopolysaccharide) were breathing spontaneously for 12 hours. They were compared to a control group. All animals were studied 24 hours after saline or lipopolysaccharide administration. Measurements and Main Results: Twenty-four hours after saline or lipopolysaccharide administration, diaphragm contractility was measured in vitro. We also measured diaphragm muscle fiber dimensions from stained cross sections, and inflammatory cytokines were determined by proteome array. Activities of calpain, caspase-3, and proteasome, expression of 20S-proteasome α subunits, E2 conjugases, E3 ligases, and autophagy were measured with immunoblotting and quantitative polymerase chain reaction. Lipopolysaccharide and/or controlled mechanical ventilation independently decreased diaphragm contractility and fiber dimensions and increased diaphragm interleukin-6 production, protein ubiquitination, expression of Atrogin-1 and Murf-1, calpain and caspase-3 activities, autophagy, and protein oxidation. Compared with lipopolysaccharide alone, lipopolysaccharide + controlled mechanical ventilation worsened diaphragm contractile dysfunction, augmented diaphragm interleukin- 6 levels, autophagy, and protein oxidation, but exerted no exacerbating effects on diaphragm fiber dimensions, calpain, caspase-3, or proteasome activation. Conclusions: Twelve hours of controlled mechanical ventilation potentiates sepsis-induced diaphragm dysfunction, possibly due to increased proinflammatory cytokine production and autophagy and worsening of oxidative stress. [ABSTRACT FROM AUTHOR]

BACKGROUND: Mechanical ventilation is crucial for patients with respiratory failure. The mechanical takeover of diaphragm function leads to diaphragm dysfunction and atrophy (ventilator-induced diaphragmatic dysfunction), with an increase in oxidative stress as a major contributor. In most patients, a sedative regimen has to be initiated to allow tube tolerance and ventilator synchrony. Clinical data imply a correlation between cumulative propofol dosage and diaphragm dysfunction, whereas laboratory investigations have revealed that propofol has some antioxidant properties. The authors hypothesized that propofol reduces markers of oxidative stress, atrophy, and contractile dysfunction in the diaphragm. METHODS: Male Wistar rats (n = 8 per group) were subjected to either 24 h of mechanical ventilation or were undergone breathing spontaneously for 24 h under propofol sedation to test for drug effects. Another acutely sacrificed group served as controls. After sacrifice, diaphragm tissue was removed, and contractile properties, cross-sectional areas, oxidative stress, and proteolysis were examined. The gastrocnemius served as internal control. RESULTS: Propofol did not protect against diaphragm atrophy, oxidative stress, and protease activation. The decrease in tetanic force compared with controls was similar in the spontaneous breathing group (31%) and in the ventilated group (34%), and both groups showed the same amount of muscle atrophy. The gastrocnemius muscle fibers did not show atrophy. CONCLUSIONS: Propofol does not protect against ventilator-induced diaphragmatic dysfunction or oxidative injury. Notably, spontaneous breathing under propofol sedation resulted in the same amount of diaphragm atrophy and dysfunction although diaphragm activation per se protects against ventilator-induced diaphragmatic dysfunction. This makes a drug effect of propofol likely. [ABSTRACT FROM AUTHOR]

Chronic obstructive pulmonary disease (COPD) is a lung disease characterized by airway obstruction and inflammation but also accompanied by several extra-pulmonary 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. [ABSTRACT FROM AUTHOR]

Botulinum Neurotoxin type-A (BoNT-A) injections are widely used as first-line spasticity treatment in spastic cerebral palsy (SCP). Despite improved clinical outcomes, concerns regarding harmful effects on muscle morphology have been raised. Yet, the risk of initiating BoNT-A to reduce muscle growth remains unclear. This study investigated medial gastrocnemius (MG) morphological muscle growth in children with SCP (n = 26, median age of 5.2 years (3.5)), assessed by 3D-freehand ultrasound prior to and six months post-BoNT-A injections. Post-BoNT-A MG muscle growth of BoNT-A naive children (n = 11) was compared to (a) muscle growth of children who remained BoNT-A naive after six months (n = 11) and (b) post-BoNT-A follow-up data of children with a history of BoNT-A treatment (n = 15). Six months after initiating BoNT-A injection, 17% decrease in mid-belly cross-sectional area normalized to skeletal growth and 5% increase in echo-intensity were illustrated. These muscle outcomes were only significantly altered when compared with children who remained BoNT-A naive (+4% and −3%, respectively, p < 0.01). Muscle length growth persevered over time. This study showed reduced cross-sectional growth post-BoNT-A treatment suggesting that re-injections should be postponed at least beyond six months. Future research should extend follow-up periods investigating muscle recovery in the long-term and should include microscopic analysis. [ABSTRACT FROM AUTHOR]

Controlled mechanical ventilation (CMV) is known to result in rapid and severe diaphragmatic dysfunction, but the recovery response of the diaphragm to normal function after CMV is unknown. Therefore, we examined the time course of diaphragm function recovery in an animal model of CMV. Healthy rats were submitted to CMV for 24-27 h (n = 16), or to 24-h CMV followed by either 1 h (CMV + 1 h SB, n = 9), 2 h (CMV + 2 h SB, n = 9), 3 h (CMV + 3 h SB, n = 9), or 4-7 h (CMV + 4-7 h SB, n = 9) of spontaneous breathing (SB). At the end of the experiment, the diaphragm muscle was excised for functional and biochemical analysis. The in vitro diaphragm force was significantly improved in the CMV + 3 h SB and CMV + 4-7 h SB groups compared with CMV (maximal tetanic force: +27%, P < 0.05, and +59%, P < 0.001, respectively). This was associated with an increase in the type IIx/b fiber dimensions (P < 0.05). Neutrophil influx was increased in the CMV + 4-7 h SB group (P < 0.05), while macrophage numbers remained unchanged. Markers of protein synthesis (phosphorylated Akt and eukaryotic initiation factor 4E binding protein 1) were significantly increased (±40%, P < 0.001, and ±52%, P < 0.01, respectively) in the CMV + 3 h SB and CMV + 4-7 h SB groups and were positively correlated with diaphragm force (P < 0.05). Finally, also the maximal specific force generation of skinned single diaphragm fibers was increased in the CMV + 4-7 h SB group compared with CMV (+45%, P < 0.05). In rats, reloading the diaphragm for 3 h after CMV is sufficient to improve diaphragm function, while complete recovery occurs after longer periods of reloading. Enhanced muscle fiber dimensions, increased protein synthesis, and improved intrinsic contractile properties of diaphragm muscle fibers may have contributed to diaphragm function recovery. [ABSTRACT FROM AUTHOR]

Objective: Mechanical ventilation (MV) is a life saving intervention for patients with respiratory failure. Even after 6 hours of MV, diaphragm atrophy and dysfunction (collectively referred to as ventilator-induced diaphragmatic dysfunction, VIDD) occurs in concert with a blunted blood flow and oxygen delivery. The regulation of hypoxia sensitive factors (i.e. hypoxia inducible factor 1α, 2α (HIF-1α,–2α), vascular endothelial growth factor (VEGF)) and angio-neogenetic factors (angiopoietin 1–3, Ang) might contribute to reactive and compensatory alterations in diaphragm muscle. Methods: Male Wistar rats (n = 8) were ventilated for 24 hours or directly sacrificed (n = 8), diaphragm and mixed gastrocnemius muscle tissue was removed. Quantitative real time PCR and western blot analyses were performed to detect changes in angio-neogenetic factors and inflammatory markers. Tissues were stained using Isolectin (IB 4) to determine capillarity and calculate the capillary/fiber ratio. Results: MV resulted in up-regulation of Ang 2 and HIF-1α mRNA in both diaphragm and gastrocnemius, while VEGF mRNA was down-regulated in both tissues. HIF-2α mRNA was reduced in both tissues, while GLUT 4 mRNA was increased in gastrocnemius and reduced in diaphragm samples. Protein levels of VEGF, HIF-1α, -2α and 4 did not change significantly. Additionally, inflammatory cytokine mRNA (Interleukin (IL)-6, IL-1β and TNF α) were elevated in diaphragm tissue. Conclusion: The results demonstrate that 24 hrs of MV and the associated limb disuse induce an up-regulation of angio-neogenetic factors that are connected to HIF-1α. Changes in HIF-1α expression may be due to several interactions occurring during MV. [ABSTRACT FROM AUTHOR]

The article focuses on a study which examined the possible protective effect of proteasome inhibitor, bortezomib, against ventilator-induced diaphragm dysfunction. The study included anesthetized rats that were submitted to controlled mechanical ventilation while receiving bortezomib or saline. It found that bortezomib partially protects the diaphragm from controlled mechanical ventilation-induced diaphragm contractile dysfunction but did not prevent atrophy.

The article discusses a randomized, controlled experiment which investigated the role of the antioxidant, N-acetylcysteine, in protecting the diaphragm from the negative effects of controlled mechanical ventilation. The study hypothesized that the antioxidant will restore the redox balance in the diaphragm of laboratory rats. The experiment showed that N-acetylcysteine prevented diaphragmatic oxidative stress and proteolysis induced by controlled mechanical ventilation. It also abolished contractile dysfunction in the diaphragm.

Background: High dose of corticosteroids has been previously shown to protect against controlled mechanical ventilation (CMV)-induced diaphragmatic dysfunction while inhibiting calpain activation. Because literature suggests that the calpain inhibiting effect of corticosteroid depends on the dose administered, we determined whether lower doses of corticosteroids would also provide protection of the diaphragm during CMV. This may be important for patients undergoing mechanical ventilation and receiving corticosteroids. Methods: Rats were assigned to controls or to 24 hours of CMV while being treated at the start of mechanical ventilation with a single intramuscular administration of either saline, or 5 mg/kg (low MP) or 30 mg/kg (high MP) of methylprednisolone. Results: Diaphragmatic force was decreased after CMV and this was exacerbated in the low MP group while high MP rescued this diaphragmatic dysfunction. Atrophy was more severe in the low MP group than after CMV while no atrophy was observed in the high MP group. A significant and similar increase in calpain activity was observed in both the low MP and CMV groups whereas the high dose prevented calpain activation. Expression of calpastatin, the endogenous inhibitor of calpain, was decreased in the CMV and low MP groups but its level was preserved to controls in the high MP group. Caspase-3 activity increased in all CMV groups but to a lesser extent in the low and high MP groups. The 20S proteasome activity was increased in CMV only. Conclusions: Administration of 30 mg/kg methylprednisolone during CMV protected against CMV-induced diaphragm dysfunction while 5 mg/kg was more deleterious. The protective effect is due mainly to an inhibition of the calpain system through preservation of calpastatin levels and to a lesser extent to a caspase-3 inhibition. [ABSTRACT FROM AUTHOR]

Documentation of community pharmacists' clinical activities, such as the identification and management of drug-related problems (DRPs), is recommended. However, documentation is not systematic in Swiss community pharmacies, and relevant information about DRPs, such as consequences or involved partners, is frequently missing. This study aims to evaluate the interrater and test-retest reliability, appropriateness and acceptability of the Clinical Pharmacy Activities Documented (ClinPhADoc) tool. Ten community pharmacists participated in the study. Interrater reliability coefficients were computed using 24 standardized cases. One month later, test-retest reliability was assessed using 10 standardized cases. To assess the appropriateness, pharmacists were asked to document clinical activities in their own practice using ClinPhADoc. Acceptability was assessed by an online satisfaction survey. Kappa coefficients showing a moderate level of agreement (>0.40) were observed for interrater and test-retest reliability. Pharmacists were able to document 131 clinical activities. The good level of acceptability and brief documentation time (fewer than seven minutes) indicate that ClinPhADoc is well-suited to the community pharmacy setting. To optimize the tool, pharmacists proposed developing an electronic version. These results support the reliability and acceptance of the ClinPhADoc tool. [ABSTRACT FROM AUTHOR]

In chronic obstructive pulmonary disease (COPD), the bronchial epithelium is the first immune barrier that is triggered by cigarette smoke. Although vitamin D (vitD) has proven anti-inflammatory and antimicrobial effects in alveolar macrophages, little is known about the direct role of vitD on cigarette smoke-exposed bronchial epithelial cells. We examined the effects of vitD on a human bronchial epithelial cell line (16HBE) and on air–liquid culture of primary bronchial epithelial cells (PBEC) of COPD patients and controls exposed for 24 h to cigarette smoke extract (CSE). VitD decreased CSE-induced IL-8 secretion by 16HBE cells, but not by PBEC. VitD significantly increased the expression of the antimicrobial peptide cathelicidin in 16HBE and PBEC of both COPD subjects and controls. VitD did not affect epithelial to mesenchymal transition or epithelial MMP-9 expression and was not able to restore impaired wound healing by CSE in 16HBE cells. VitD increased the expression of its own catabolic enzyme CYP24A1 thereby maintaining its negative feedback. In conclusion, vitD supplementation may potentially reduce infectious exacerbations in COPD by the upregulation of cathelicidin in the bronchial epithelium. [ABSTRACT FROM AUTHOR]

Nicotine, an important component of cigarette smoke, is a neurotransmitter that contributes to stress, depression and anxiety in smokers. In rodents, it increases anxiety and reduces exploratory behaviours. However, so far, the measurements of exploratory behaviour in rodents have only been semi-quantitative and lacking in sufficient detail to characterise the temporal effect of smoking cessation. As rodents, such as mice and rats, primarily use whiskers to explore their environment, we studied the effect of 3 months smoking with 1 and 2 weeks smoking cessation on whisker movements in mice, using high-speed video camera footage and image analysis. Both protraction and retraction whisker velocities were increased in smoking mice (p < 0.001) and returned to normal following just one week of smoking cessation. In addition, locomotion speeds were decreased in smoking mice, and returned to normal following smoking cessation. Lung function was also impacted by smoking and remained impaired even following smoking cessation. We suggest that the increased whisker velocities in the smoking mice reflect reduced exploration and impeded tactile performance. The increase in whisker velocity with smoking, and its reduction following smoking cessation, also lends support to acetylcholine being involved in awareness, attention and alertness pathways. It also shows that smoking-induced behavioural changes can be reversed with smoking cessation, which may have implications for human smokers. [ABSTRACT FROM AUTHOR]

Highlights • Enhanced bacterial clearance in vitamin D deficient mice infected with NTHi. • Cigarette smoke inhibited phagocytosis and oxidative burst, but had no effect on bacterial clearance. • Faster resolution of local lung inflammation in vitamin D deficient mice. • Upregulation of CRAMP mRNA as possible mechanism for faster eradication in vitamin D deficient mice. • Misbalance of the protease and anti-protease homeostasis towards proteolysis in NTHi infected vitamin D deficient mice. Abstract Chronic obstructive pulmonary disease (COPD), which is characterized by an excessive inflammatory response of the airways, is often complicated by exacerbations. Vitamin D deficiency has been associated with an increased risk for COPD and may predispose COPD patients to a higher exacerbation rate, particularly during smoking. In the current study, we investigated the effect of vitamin D deficiency and cigarette smoke (CS)-exposure on lung inflammation and bacterial clearance after an acute infection with Nontypeable Haemophilus influenzae (NTHi). Vitamin D deficient or sufficient mice were exposed to nose-only CS or ambient air for 6 weeks and oropharyngeally instilled with 106 NTHi. Residual viable NTHi were measured at different time points post-infection. Mechanisms of bacterial clearance (e.g. phagocytosis, pattern recognition receptors, antimicrobial peptides, surfactant proteins and mucin) and lung remodeling (e.g. metalloproteinases, MMP's) were assessed. Although smoking resulted in reduced phagocytosis capacity of macrophages and neutrophils, bacterial clearance was similar to control mice. By contrast and independent of smoking, bacterial clearance was significantly accelerated in vitamin D deficient mice already from 24 h post-infection (p = 0.0087). This faster and complete eradication was associated with a more rapid resolution of cytokines and neutrophils 72 h post-infection and dominated by an upregulation of cathelicidin-related antimicrobial peptide (CRAMP) mRNA during infection (p = 0.026). However, vitamin D deficiency also resulted in more MMP12 protein in broncho-alveolar lavage and a shift in mRNA expression of MMP12/TIMP1 (p = 0.038) and MMP9/TIMP1 (p = 0.024) ratio towards more protease activity. Overall, vitamin D deficient mice resolved NTHi infection faster with a faster resolution of local lung inflammation, possibly through upregulation of CRAMP. This was associated with a disruption of the protease/anti-protease balance, which may potentially scale towards a higher extracellular matrix breakdown. [ABSTRACT FROM AUTHOR]