Sputum cytology during late-phase responses to inhalation challenge with different allergens.

BACKGROUND: In mouse models of allergic asthma, exposure to different allergens can trigger distinct inflammatory subtypes in the airways. We investigated whether this observation extends to humans. METHODS: We compared the frequency of sputum inflammatory subtypes between mild allergic asthma subjects (n = 129) exposed to different allergens in inhalation challenge tests. These tests were performed using a standardized protocol as part of clinical trials of experimental treatments for asthma, prior to drug randomization. Five allergen types were represented: the house dust mites Dermatophagoides pteronyssinus and Dermatophagoides farinae, ragweed, grass, and cat. RESULTS: Of 118 individuals with a sputum sample collected before allergen challenge (baseline), 45 (38%) had paucigranulocytic, 51 (43%) eosinophilic, 11 (9%) neutrophilic, and 11 (9%) mixed granulocytic sputum. Of note, most individuals with baseline paucigranulocytic sputum developed eosinophilic (48%) or mixed granulocytic (43%) sputum 7 hours after allergen challenge, highlighting the dynamic nature of sputum inflammatory subtype in asthma. Overall, there was no difference in the frequency of sputum inflammatory subtypes following challenge with different allergen types. Similar results were observed at 24 hours after allergen challenge. CONCLUSIONS: Unlike reported in mice, in humans the sputum inflammatory subtype observed after an allergen-induced asthma exacerbation is unlikely to be influenced by the type of allergen used.

OX40L blockade and allergen-induced airway responses in subjects with mild asthma.

BACKGROUND: The OX40/OX40L interaction contributes to an optimal T cell response following allergic stimuli and plays an important role in the maintenance and reactivation of memory T effector cells. OBJECTIVE: We tested whether treatment with an anti-OX40L monoclonal antibody (MAb) would inhibit allergen-induced responses in subjects with asthma. METHODS: Twenty-eight mild, atopic asthmatic subjects were recruited for a double-blind, randomized, placebo-controlled, parallel-group trial (ClinicalTrials.gov identifier NCT00983658) to compare blockade of OX40L using a humanized anti-OX40L MAb to placebo-administered intravenously in 4 doses over 3 months. Allergen inhalation challenges were carried out 56 and 113 days after the first dose of study drug. The primary outcome variable was the late-phase asthmatic response. Other outcomes included the early-phase asthmatic response, airway hyperresponsiveness, serum IgE levels, blood and sputum eosinophils, safety and tolerability. RESULTS: Treatment with anti-OX40L MAb did not attenuate the early- or late-phase asthmatic responses at days 56 or 113 compared with placebo. In the anti-OX40L MAb treatment group, total IgE was reduced 17% from pre-dosing levels, and sputum eosinophils decreased 75% by day 113 (both P = 0.04). There was no effect of anti-OX40L MAb on airway hyperresponsiveness or blood eosinophils. The frequency of AEs was similar in both groups. CONCLUSION AND CLINICAL RELEVANCE: Pharmacological activity of anti-OX40L MAb was observed by decreases in serum total IgE and airway eosinophils at 16 weeks post-dosing, but there was no effect on allergen-induced airway responses. It is possible that the treatment duration or dose of antibody was insufficient to impact the airway responses.

Sputum inflammatory cells and allergen-induced airway responses in allergic asthmatic subjects.

BACKGROUND: Allergen inhalation causes early and late bronchoconstrictor responses, airway hyperresponsiveness and airway inflammation in allergic asthmatics. The role of airway inflammatory cells in causing allergen-induced bronchoconstriction and airway hyperresponsiveness is controversial. The objective of this study was to examine the relationships between allergen-induced increases in airway inflammatory cells, early and late bronchoconstrictor responses and methacholine airway hyperresponsiveness. METHODS: Allergen inhalation challenge was conducted in 50 allergic asthmatics. Changes in the forced expired volume in 1 s (FEV(1%) ) were followed for 7 h, induced sputum was obtained at 7 and 24 h, and the provocative concentration of methacholine causing a 20% fall in FEV(1) (MCh PC(20) ) was measured at 24 h. RESULTS: There was a significant negative correlation between the baseline methacholine PC(20) and baseline sputum eosinophils (r = -0.512, P = 0.0001). Allergen-induced changes in methacholine PC(20) were also significantly negatively correlated to allergen-induced change in sputum eosinophils at 24 h (r = -0.434, P = 0.002), but not to changes in any other inflammatory cells. There were no significant correlations between sputum eosinophils or other inflammatory cells and the allergen-induced early or late asthmatic responses. CONCLUSION: Allergen-induced increases in airway eosinophils in asthmatic dual responders may contribute to allergen-induced changes in methacholine PC(20) , but not the late asthmatic responses.

The objective measurement of breathlessness.

Both direct and indirect psychophysical methods can be used to measure breathlessness. With indirect methods, detection thresholds can be defined. Direct methods are easier to apply and give more information than indirect methods. Direct methods include interval (partition) and ratio scales, which are easy to apply; both have advantages and disadvantages. The selection of scale depends largely on the question addressed. For comparison across individuals or for the measurement of absolute magnitude (however imprecise), simple category scales are adequate and useful. Open magnitude scaling is best used to define the stimulus parameters influencing perceptual magnitude. Comparison across groups or individuals using exponents as an index of perceptual sensitivity should be interpreted with caution. Where possible, alternative methods should be used to validate differences found. The age-old bias against sensory measurement may be in part our inability to understand the sensory mechanisms and have little to do with the validity of the measurements.

Symptom intensity and subjective limitation to exercise in patients with cardiorespiratory disorders.

The aim of the study was to compare (1) the intensity of leg effort and dyspnea during exercise and (2) subjective limitations to performance in normal subjects, patients receiving medication for cardiac disorders, patients with pulmonary impairment, patients with pulmonary impairment who were also receiving cardiac medications, patients experiencing chest pain during exercise, and patients who had a reduced exercise capacity but did not have pulmonary impairment and were not receiving cardiac medication. Five hundred seventy-eight subjects rated the intensity of leg effort, discomfort with breathing (dyspnea), and chest pain every minute (Borg scale) during an incremental exercise task (100 kpm/min each minute) to maximum work capacity on a cycle ergometer and following exercise indicated their subjective limitation by completing a simple questionnaire. Leg effort and dyspnea increased systematically with power output in a positively accelerating manner in all groups; both symptoms were significantly more intense in the impaired groups compared with the normal group at submaximal power outputs. In all groups, there was a significant relationship between symptom intensity at submaximal power outputs and the maximal power output achieved. Leg discomfort in combination with breathing discomfort was the predominant subjective limitation in all groups; chest pain in combination with leg and breathing discomfort was the major subjective limitation in individuals with angina. Activation of the sensory systems during exercise is accompanied by a perception of discomfort associated with the peripheral exercising muscles and discomfort with breathing; both discomfort associated with the exercising muscles and discomfort associated with breathing contribute to exercise limitation to a large degree in normal subjects and patients with cardiorespiratory diseases.

Breathlessness during exercise with and without resistive loading.

The purpose of this study was to quantify the intensity of breathlessness associated with exercise and respiratory resistive loading, with the specific purpose of isolating the quantitative contributions of inspiratory pressure, length, velocity, and frequency of inspiratory muscle shortening and duty cycle to breathlessness. The intensity of inspiratory pressure was quantified by measurement of estimated esophageal pressure (Pes = pressure at the mouth plus lung pressure), the extent of shortening by tidal volume (VT), and the velocity of shortening by inspiratory flow rate (VI). Six normal subjects underwent five incremental (100 kpm X min-1 X min-1) exercise tests on a cycle ergometer to maximum capacity. The first and last test were unloaded and the intervening tests were performed with external added resistances of 33, 57, and 73 cm H2O X l-1 X s in random order. The resistances were selected to provide a range of pressures, tidal volumes, flow rates, and patterns of breathing. At rest and at the end of each minute during exercise the subjects estimated the intensity of breathlessness (psi) by selecting a number ranging from 0 to 10 (Borg rating scale, 0 indicating no appreciable breathlessness and 10 the maximum tolerable sensation). Breathlessness was significantly and independently related to Pes (P less than 0.0001), VI (P less than 0.0001), frequency of breathing (fb) (P less than 0.01), and duty cycle [ratio of inspiratory duration to total breath duration (TI/TT)] (P less than 0.01): psi = 0.11 Pes + 0.61 VI + 1.99 TI/TT + 0.04 fb - 2.60 (r = 0.83). The results suggest that peak pressure (tension), VI (velocity of inspiratory muscle shortening), TI/TT, and fb contribute independently and collectively to breathlessness. The perception of respiratory muscle effort is ideally suited to subserve this sensation. The neurophysiological mechanism purported is a conscious awareness of the intensity of the outgoing motor command by means of corollary discharge within the central nervous system.

Quantification of intensity of sensations during muscular work by normal subjects.

Eleven subjects performed a series of 30-s work bouts on a cycle ergometer at power outputs ranging from 20-120% of the work capacity (Wcap) achieved during an incremental cycle to exhaustion and estimated the intensity of several sensations (leg effort, muscle tension, muscle discomfort, muscle pain, and breathing discomfort) by using Borg's category-ratio scale (range 0-10 units). Leg effort was perceived as "just noticeable" at 31 +/- 15% Wcap, muscle tension was just noticeable at 31 +/- 16% Wcap, muscle discomfort was just noticeable at 47 +/- 21% Wcap, breathing discomfort was just noticeable at 52 +/- 19% Wcap, and muscle pain was just noticeable at 58 +/- 33% Wcap. The intensity of all sensations increased in a positively accelerating manner with increases in power output (P < 0.001). Above 60% Wcap, the intensity of leg effort and muscle tension exceeded the intensity of muscle pain (P < 0.01), and above 100% Wcap the intensity of muscle discomfort also exceeded the intensity of muscle pain (P < 0.01). At 120% Wcap, leg effort, muscle tension, and muscle discomfort were rated between "severe" and "very severe" (6.1 +/- 2.2, 6.4 +/- 2.0, and 5.6 +/- 2.1 Borg units, respectively), whereas muscle pain and breathing discomfort were rated between "moderate" and "somewhat severe" (3.6 +/- 2.1 and 3.3 +/- 1.9 Borg units, respectively). These results suggest that subjects have a perception of muscle pain during muscular work that is distinct from perceptions of leg effort, muscle tension, and muscle discomfort.

Effect of frequency on perceived magnitude of added loads to breathing.

Using open-magnitude scaling, six normal subjects estimated the perceived magnitude of a range of added elastic loads (20-76 cmH2O/l), applied for a sequence of five breaths, at frequencies varying from 5 to 26.4 breaths/min. Two experiments were performed. In the first, frequency was increased by a reduction in expiratory duration (TE), and the duty cycle (ratio of inspiratory duration to total breath duration, TI/TT) ranged between 0.10 and 0.52. The perceived magnitude psi increased significantly with the peak airway pressure (Pm) (P less than 0.0001) but did not reach conventional significance with frequency (fb) (P = 0.15): psi = K0Pm1.23fb0.07 (r = 0.911). However, the sensory magnitude increased significantly as the duty cycle increased (P less than 0.01), but when it was included, the magnitude decreased minimally with frequency (P less than 0.01): psi = K0Pm1.3fb-0.97 TI/TT1.14 (r = 0.92). In the second experiment the duty cycle (TI/TT) was kept constant [(0.43 +/- 0.008 (SE)] and frequency (5-26.4 breaths/min) increased at the expense of shortening both TI and TE. The perceived magnitude of the added elastances decreased with the increase in frequency. However, when the perceived magnitude was corrected for the duration of inspiration, which is known to increase the sensory magnitude, psi = K0Pm1.3TI0.56, the sensory magnitude increased significantly with frequency (P less than 0.001): psi/TI0.56 = K0Pm1.21fb0.28 (r = 0.773). The decrease in inspiratory duration had a greater quantitative effect decreasing sensory magnitude than frequency had on increasing the magnitude. The effect of increasing frequency is complex and depends on the simultaneous intensity, duration of inspiratory pressure, and the duty cycle.

Inspiratory muscle forces and endurance in maximum resistive loading.

The ability of the respiratory muscles to sustain ventilation against increasing inspiratory resistive loads was measured in 10 normal subjects. All subjects reached a maximum rating of perceived respiratory effort and at maximum resistance showed signs of respiratory failure (CO2 retention, O2 desaturation, and rib cage and abdominal paradox). The maximum resistance achieved varied widely (range 73-660 cmH2O X l-1 X s). The increase in O2 uptake (delta Vo2) associated with loading was linearly related to the integrated mouth pressure (IMP): delta Vo2 = 0.028 X IMP + 19 ml/min (r = 0.88, P less than 0.001). Maximum delta Vo2 was 142 ml/min +/- SD 68 ml/min. There were significant (P less than 0.05) relationships between the maximum voluntary inspiratory pressure against an occluded airway (MIP) and both maximum IMP (r = 0.80) and maximum delta Vo2 (r = 0.76). In five subjects, three imposed breathing patterns were used to examine the effect of different patterns of respiratory muscle force deployment. Increasing inspiratory duration (TI) from 1.5 to 3.0 and 6.0 s, at the same frequency of breathing (5.5 breaths/min) reduced peak inspiratory pressure and increased the maximum resistance tolerated (190, 269, and 366 cmH2O X l-1 X s, respectively) and maximum IMP (2043, 2473, and 2913 cmH2O X s X min-1, but the effect on maximum delta Vo2 was less consistent (166, 237, and 180 ml/min). The ventilatory endurance capacity and the maximum O2 uptake of the respiratory muscles are related to the strength of the inspiratory muscles, but are also modified through the pattern of force deployment.

Inspiratory muscles during exercise: a problem of supply and demand.

The capacity of inspiratory muscles to generate esophageal pressure at several lung volumes from functional residual capacity (FRC) to total lung capacity (TLC) and several flow rates from zero to maximal flow was measured in five normal subjects. Static capacity was 126 +/- 14.6 cmH2O at FRC, remained unchanged between 30 and 55% TLC, and decreased to 40 +/- 6.8 cmH2O at TLC. Dynamic capacity declined by a further 5.0 +/- 0.35% from the static pressure at any given lung volume for every liter per second increase in inspiratory flow. The subjects underwent progressive incremental exercise to maximum power and achieved 1,800 +/- 45 kpm/min and maximum O2 uptake of 3,518 +/- 222 ml/min. During exercise peak esophageal pressure increased from 9.4 +/- 1.81 to 38.2 +/- 5.70 cmH2O and end-inspiratory esophageal pressure increased from 7.8 +/- 0.52 to 22.5 +/- 2.03 cmH2O from rest to maximum exercise. Because the estimated capacity available to meet these demands is critically dependent on end-inspiratory lung volume, the changes in lung volume during exercise were measured in three of the subjects using He dilution. End-expiratory volume was 52.3 +/- 2.42% TLC at rest and 38.5 +/- 0.79% TLC at maximum exercise.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of thoracoabdominal breathing patterns on inspiratory effort sensation.

The respiratory sensations evoked by added inspiratory loads are currently thought to be largely mediated by the activity of the inspiratory muscles. Because of the differences in proprioceptors and in afferent and efferent innervations among the inspiratory muscles, we hypothesized that the sensation evoked by a given load would be different when the motor command is directed mainly to rib cage muscles or mainly to the diaphragm. To test this hypothesis, we studied six normal subjects breathing against several inspiratory resistances while emphasizing the use of rib cage muscles, or the diaphragm, or a combination of both. At the end of 10 loaded breaths the subjects rated the perceived magnitude of inspiratory effort on a Borg scale. A linear and unique relationship (r = 0.96 +/- 0.02; P less than 0.001) was found between the sensation and esophageal pressure (Pes) in the three thoracoabdominal breathing patterns. We conclude that the level of Pes, whether generated mainly by the rib cage muscles or the diaphragm, is the main variable related to the sensation of inspiratory effort under external inspiratory loads.

Metabolic and hemodynamic responses of lower limb during exercise in patients with COPD.

Premature lactic acidosis during exercise in patients with chronic obstructive pulmonary disease (COPD) may play a role in exercise intolerance. In this study, we evaluated whether the early exercise-induced lactic acidosis in these individuals can be explained by changes in peripheral O2 delivery (O2). Measurements of leg blood flow by thermodilution and of arterial and femoral venous blood gases, pH, and lactate were obtained during a standard incremental exercise test to capacity in eight patients with severe COPD and in eight age-matched controls. No significant difference was found between the two groups in leg blood flow at rest or during exercise at the same power outputs. Blood lactate concentrations and lactate release from the lower limb were greater in COPD patients at all submaximal exercise levels (all P < 0.05). Leg D02 at a given power output was not significantly different between the two groups, and no significant correlation was found between this parameter and blood lactate concentrations. COPD patients had lower arterial and venous pH at submaximal exercise, and there was a significant positive correlation between venous pH at 40 W and the peak O2 uptake (r = 0.91, P < 0.0001). The correlation between venous pH and peak O2 uptake suggests that early muscle acidosis may be involved in early exercise termination in COPD patients. The early lactate release from the lower limb during exercise could not be accounted for by changes in peripheral O2. The present results point to skeletal muscle dysfunction as being responsible for the early onset of lactic acidosis in COPD.

Mechanisms of exertional dyspnea.

To understand why someone is dyspneic during exercise, we need to follow the advice of Sir Francis Bacon: "No natural phenomenon can be adequately studied in itself alone, but to be understood must be considered as it stands connected with all of nature." In the present context, this implies the careful measurement of events related to metabolism, circulation, and respiration and of the associated sensory events as these systems adapt to the strain and stress of exercise.

Respiratory muscles and dyspnea.

This article reviews the classical basis on which dyspnea is identified and quantified. The sensation or sensations of discomfort experienced during breathing are then viewed using a background of sensory physiology. Exploiting psychophysical techniques, the origin of the sensation of discomfort is viewed within the constraints of the presently-known sensory structures. The contribution of these sensory structures to the quality and quantity of discomfort is suggested, indicating the central role played by the respiratory muscles.

The use of exercise testing and other methods in the investigation of dyspnea.

The sensation of effort is increased when the tension developed by active muscle is increased or when the muscle is weak; similar factors contribute to the sense of respiratory effort that constitute the symptoms of dyspnea. Exercise testing enables systematic loading of the respiratory muscles to be studied; the components of the ventilatory responses to exercise may be quantified in terms of the pattern and timing of breathing and of inspiratory flow and volume. The associated sensation of respiratory effort may then be related to the tension developed by respiratory muscles and to their strength.

An approach to dyspnea in cancer patients.

There are many potential causes of dyspnea in the patient with cancer. Ultimately, a sense of increased respiratory effort is common to all of these diverse situations. An organized approach to dyspnea in the cancer patient is presented based on psychophysical principles, and treatment modalities are suggested.

Copy number aberrations of BCL2 and CDKN2A/B identified by array-CGH in thymic epithelial tumors.

The molecular pathology of thymic epithelial tumors (TETs) is largely unknown. Using array comparative genomic hybridization (CGH), we evaluated 59 TETs and identified recurrent patterns of copy number (CN) aberrations in different histotypes. GISTIC algorithm revealed the presence of 126 significant peaks of CN aberration, which included 13 cancer-related genes. Among these peaks, CN gain of BCL2 and CN loss of CDKN2A/B were the only genes in the respective regions of CN aberration and were associated with poor outcome. TET cell lines were sensitive to siRNA knockdown of the anti-apoptotic molecules BCL2 and MCL1. Gx15-070, a pan-BCL2 inhibitor, induced autophagy-dependent necroptosis in TET cells via a mechanism involving mTOR pathways, and inhibited TET xenograft growth. ABT263, an inhibitor of BCL2/BCL-XL/BCL-W, reduced proliferation in TET cells when administered in combination with sorafenib, a tyrosine kinase inhibitor able to downregulate MCL1. Immunohistochemistry on 132 TETs demonstrated that CN loss of CDKN2A correlated with lack of expression of its related protein p16(INK4) and identified tumors with poor prognosis. The molecular markers BCL2 and CDKN2A may be of potential value in diagnosis and prognosis of TETs. Our study provides the first preclinical evidence that deregulated anti-apoptotic BCL2 family proteins may represent suitable targets for TET treatment.

Effects of inhaled ciclesonide on circulating T-helper type 1/T-helper type 2 cells in atopic asthmatics after allergen challenge.

BACKGROUND: The predominance of T-helper type 2 (Th2) lymphocytes is thought to underlie the pathogenesis of asthma. Allergen inhalation challenge in atopic asthmatic subjects is associated with decreased interferon-gamma (IFN-gamma) positive CD4+ and CD8+ lymphocytes in peripheral blood and induced sputum. OBJECTIVE: This study examined the effects of an inhaled corticosteroid on these previously described allergen-induced changes in circulating Th1 and Th2 lymphocytes. METHODS: Subjects were randomized to 7 days of placebo, 40 or 80 micro g ciclesonide in a crossover study. Airway responses and peripheral blood were measured before and after treatment, and 24 h after allergen challenge. RESULTS: Ciclesonide 40 and 80 micro g significantly attenuated the late response and sputum eosinophils at 8 h post-allergen (P<0.05). Circulating IFN-gamma positive CD4+ lymphocytes decreased after allergen challenge with placebo (P<0.05), and this was inhibited by 40 micro g ciclesonide treatment (P<0.05). There was no effect of allergen inhalation or ciclesonide on IL-4-positive CD4+ lymphocytes or IFN-gamma and IL-4-positive CD8(high) lymphocytes. The allergen-induced change of IFN-gamma/IL-4 ratio on CD4+ cells correlated with the allergen-induced change of peripheral blood eosinophils. CONCLUSIONS: The results of this study suggest that attenuation of allergen-induced airway responses by ciclesonide may be mediated through regulation of IFN-gamma-positive CD4+ cells.