Skeletal muscle molecular responses to resistance training and dietary supplementation in COPD.

BACKGROUND: Skeletal muscle dysfunction is a systemic feature of chronic obstructive pulmonary disease (COPD), contributing to morbidity and mortality. Physical training improves muscle mass and function in COPD, but the molecular regulation therein is poorly understood. METHODS: Candidate genes and proteins regulating muscle protein breakdown (ubiquitin proteasome pathway), muscle protein synthesis (phosphatidylinositol 3 kinase/Akt/mammalian target of rapamycin pathway), myogenesis (MyoD, myogenin and myostatin) and transcription (FOXO1, FOXO3 and RUNX1) were determined in quadriceps muscle samples taken at four time points over 8 weeks of knee extensor resistance training (RT) in patients with COPD and healthy controls (HCs). Patients with COPD were randomly allocated to receive protein/carbohydrate or placebo supplements during RT. RESULTS: 59 patients with COPD (mean (SD) age 68.0 (9.3) years, forced expiratory volume in 1 s (FEV1) 46.9 (17.8) % predicted) and 21 HCs (66.1 (4.8) years, 105.0 (21.6) % predicted) were enrolled. RT increased lean mass (~5%) and strength (~20%) in all groups. Absolute work done during RT was lower throughout in patients with COPD compared with HCs. RT resulted in increases (from basal) in catabolic, anabolic, myogenic and transcription factor protein expression at 24 h, 4 weeks and 8 weeks of exercise in HCs. This response was blunted in patients with COPD, except for myogenic signalling, which was similar. Nutritional supplementation did not augment functional or molecular responses to RT. CONCLUSIONS: The potential for muscle rehabilitation in response to RT is preserved in COPD. Except for markers of myogenesis, molecular responses to RT are not tightly coupled to lean mass gains but reflect the lower work done during RT, suggesting some caution when identifying molecular targets for intervention. Increasing post-exercise protein and carbohydrate intake is not a prerequisite for a normal training response in COPD.

Ultrasound assessment of lower limb muscle mass in response to resistance training in COPD.

BACKGROUND: Quantifying the improvements in lower limb or quadriceps muscle mass following resistance training (RT), is an important outcome measure in COPD. Ultrasound is a portable, radiation free imaging technique that can measure the size of superficial muscles belonging to the quadriceps group such as the rectus femoris, but has not been previously used in COPD patients following RT. We compared the responsiveness of ultrasound derived measures of quadriceps mass against dual energy x-ray absorptiometry (DEXA), in patients with COPD and healthy controls following a programme of high intensity knee extensor RT. METHODS: Portable ultrasound was used to assess the size of the dominant quadriceps in 45 COPD patients and 19 healthy controls-before, during, and after 8 weeks of bilateral high intensity isokinetic knee extensor RT. Scanning was performed at the mid-thigh region, and 2 indices of quadriceps mass were measured-rectus femoris cross-sectional area (RFcsa) and quadriceps muscle thickness (Qt). Thigh lean mass (Tdexa) was determined by DEXA. RESULTS: Training resulted in a significant increase in Tdexa, RFcsa and Qt in COPD patients [5.7%, 21.8%, 12.1% respectively] and healthy controls [5.4%, 19.5%, 10.9 respectively]. The effect size for the changes in RFcsa (COPD= 0.77; Healthy=0.83) and Qt (COPD=0.36; Healthy=0.78) were greater than the changes in Tdexa (COPD=0.19; Healthy=0.26) following RT. CONCLUSIONS: Serial ultrasound measurements of the quadriceps can detect changes in muscle mass in response to RT in COPD. The technique has good reproducibility, and may be more sensitive to changes in muscle mass when compared to DEXA. TRIAL REGISTRATION: http://www.controlled-trials.com (Identifier: ISRCTN22764439).

Impact of transcutaneous neuromuscular electrical stimulation or resistance exercise on skeletal muscle mRNA expression in COPD.

Background: Voluntary resistance exercise (RE) training increases muscle mass and strength in patients with chronic obstructive pulmonary disease (COPD). Nonvolitional transcutaneous neuromuscular electrical stimulation (NMES) may be an alternative strategy for reducing ambulatory muscle weakness in patients unable to perform RE training, but little comparative data are available. This study, therefore, investigated changes in muscle mRNA abundance of a number of gene targets in response to a single bout of NMES compared with RE. Methods: Twenty-six patients with stable COPD (15 male; FEV1, 43±18% predicted; age, 64±8 years; fat free mass index, 16.6±1.8 kg/m2) undertook 30 minutes of quadriceps NMES (50 Hz, current at the limit of tolerance) or 5×30 maximal voluntary isokinetic knee extensions. Vastus lateralis muscle biopsies were obtained at rest immediately before and 24 hours after intervention. Expression of 384 targeted mRNA transcripts was assessed by real time TaqMan PCR. Significant change in expression from baseline was determined using the ΔΔCT method with a false discovery rate (FDR) of <5%. Results: NMES and RE altered mRNA abundance of 18 and 68 genes, respectively (FDR <5%), of which 14 genes were common to both interventions and of the same magnitude of fold change. Biological functions of upregulated genes included inflammation, hypertrophy, muscle protein turnover, and muscle growth, whilst downregulated genes included mitochondrial and cell signaling functions. Conclusions: Compared with NMES, RE had a broader impact on mRNA abundance and, therefore, appears to be the superior intervention for maximizing transcriptional responses in the quadriceps of patients with COPD. However, if voluntary RE is not feasible in a clinical setting, NMES by modifying expression of genes known to impact upon muscle mass and strength may have a positive influence on muscle function.

The use of chitosan as a flocculant in mammalian cell culture dramatically improves clarification throughput without adversely impacting monoclonal antibody recovery.

Flocculants have been employed for many years as aides in the clarification of wastewater, chemicals and food. Flocculants aggregate and agglutinate fine particles resulting in their settling from the liquid phase and a reduction in solution turbidity. These materials have not been widely used in the clarification of mammalian cell culture harvest. In this paper we examined chitosan as a flocculent of cells and cell particulates in NS0 culture harvest and the subsequent further clarification of this material by continuous flow centrifugation followed by depth and absolute filtration. Chitosan is an ideal flocculant for biotechnology applications as it is produced from non-mammalian sources (typically arthropod shells) and is also available in a highly purified form that is low in heavy metals, volatile organics and microbial materials. Chitosan is a polymer of deacetylated chitin. The deacetylation imparts limited solubility on insoluble chitin and the amino groups on the polymer result in a polycationic material at acidic and neutral pH that can interact with polyanions, such as DNA and cell culture debris (typically negatively charged). Likely the interaction of chitosan with cell culture particulate forms a germinal center for further interaction and agglomeration of particulates thereby reducing the solubility of these materials resulting in their settling out into the solid phase. Chitosan improved the clarification throughput six to seven folds without a deleterious effect on monoclonal antibody recovery or purity. The procedure for utilizing chitosan is facile, easily implemented, and highly effective in improving material clarity and increasing material throughput.

Ventilatory requirements of quadriceps resistance training in people with COPD and healthy controls.

BACKGROUND: It is proposed that resistance training (RT) does not activate the cardiopulmonary system to the same extent as whole-body exercise. This is important for patients with chronic obstructive pulmonary disease (COPD) who are ventilatory limited. OBJECTIVE: The aim was to assess the ventilatory response to an isokinetic quadriceps RT program in people with COPD and healthy controls. DESIGN: Observational. REGISTRATION NUMBER: ISRCTN22764439. SETTING: Outpatient, university teaching hospital. PARTICIPANTS AND OUTCOME MEASURES: People with COPD (n=14) and healthy controls (n=11) underwent breath-by-breath analysis of their ventilation during an RT session (five sets of 30 maximal knee extensions at 180°/sec). Subjects performed a maximal cycle ergometry test (CET) at baseline. Peak ventilation (VE; L/min) and oxygen consumption (VO2; mL/kg/min) were collected. The same system measured VO2 and VE during the RT session. Parameters are presented as a percentage of the maximal CET. Isokinetic workload, symptom scores, heart rate (HR), and oxygen saturation were documented post-training. RESULTS: People with COPD worked at higher percentages of their maximal capacity than controls (mean range between sets 1-5 for VO2 =49.1%-60.1% [COPD], 45.7%-51.43% [controls] and for VE =57.6%-72.2% [COPD], 49.8%-63.6% [controls]), although this was not statistically significant (P>0.1 in all cases). In absolute terms, the difference between groups was only significant for actual VO2 on set 2 (P<0.05). Controls performed more isokinetic work than patients with COPD (P<0.05). Median Borg symptom scores after RT were the same in both groups (3 breathlessness, 13 exertion), no de-saturation occurred, and both groups were training at ≥65% of their maximum HR. CONCLUSION: No statistically significant differences were found between people with COPD and healthy controls for VO2 and VE achieved during training. The symptoms associated with training were within acceptable limits.

Inflammatory and satellite cells in the quadriceps of patients with COPD and response to resistance training.

BACKGROUND: Quadriceps dysfunction in COPD may be mediated by inflammatory mechanisms or impaired satellite cell function. Resistance training is of proven efficacy in these patients, but data on muscle inflammatory and satellite cell response to resistance exercise in COPD are lacking.We aimed to examine the inflammatory and satellite cell profile of the quadriceps in patients with COPD and healthy control subjects at rest and after acute and chronic resistance exercise. METHODS: Seventeen patients with COPD and 10 healthy control subjects underwent 8 weeks ofbilateral lower-limb, high-intensity resistance training, thrice weekly, on an isokinetic dynamometer.Quadriceps muscle biopsy specimens from the dominant thigh were obtained at baseline,24 h following the fi rst exercise bout, and after 8 weeks 24 h after the last exercise bout. Glycolmethacrylate-embedded muscle biopsy specimens were analyzed using immunohistochemistry to identify neutrophils, macrophages, and satellite cells. RESULTS: Neutrophils were significantly elevated in the quadriceps of patients with COPD at baseline compared with healthy control subjects ( P 5 .03). Inflammatory cells were increased significantly at 24 h in both groups but were similar to baseline values at week 8, with no difference detectable between healthy control subjects and patients with COPD. Satellite cell numbers were comparable between patients and control subjects at baseline, tended to increase at 24 h, and remained elevated at week 8. CONCLUSIONS: Inflammatory cells are elevated in the resting quadriceps of patients with COPD. Acute resistance exercise leads to an inflammatory myositis, which is attenuated with regular training. Satellite cells in patients and control subjects are comparable and are increased in response to exercise. TRIAL REGISTRY: ISRCTN Register ; No.: ISRCTN22764439; URL: www.controlled-trials.com.

Modeling of purification operations in biotechnology: enabling process development, optimization, and scale-up.

Process modeling involves the use of a set of mathematical equations to represent key physical phenomena involved in the process. An appropriately validated model can be used to predict process behavior with limited experimental data, identify critical ranges for process variables, and guide further process development. Although process modeling is extensively used in the chemical process industries, it has not been widely used in purification unit operations in biotechnology. Recent FDA guidelines encourage the use of process modeling during process development, along with multivariate statistical methods, detailed risk assessment, and other quantifiers of uncertainty. This paper will review recent advances in the modeling of key downstream unit operations: chromatography, filtration, and centrifugation. The focus will be on the application of modeling for industrial applications. Relevant papers presented at a session on this topic at the recent American Chemical Society National Meeting in San Francisco will also be reviewed.