Ciclesonide improves measures of small airway involvement in asthma
Judith Cohen1, W. Rob Douma1, Nick H.T. ten Hacken1, Judith M. Vonk2, Matthijs Oudkerk3, Dirkje S. Postma1
“ONline data supplement”
Methods
Air trapping on expiratory CT scan
Anonymized MDCT data were sent to MeVis (Center for Medical Diagnostic Systems and Visualization, Bremen, Germany) and analyzed by the advanced image analysis software MeVisPULMO3D. Fully automatic segmentation of the trachea and major bronchi was performed using automatically generated seed points and optimized thresholds. In combination with a watershed algorithm, the bronchi segmentation results are used to separate the left from the right lung. In order to detect the lung lobes, the software analyzes the lung vasculature and allows an interactive segmentation based on the regions supplied by the lobar arteries and veins as well as the lobar fissures(E1). The method was validated in vivo on a human lung with respect to intra- and inter-observer reproducibility. The same lung was segmented 5 times by a single person, and once by 5 different persons. Each pair of segmentations was compared by computing the percentage of the overall lung volume that was classified identically. In both studies reproducibility amounted to at least 99.5%.Based on the segmentation, quantitative volumetric and densitometric analyses were performed of total lung, right and left lung separately and for each individual lung lobe. The parameters used in the analysis were volume (mL), mean lung density (MLD; in Hounsfield Units [HU]), 15th percentile density (PD15; HU), and percentage of low attenuation areas (LAA; %). LAA were defined at a cut-off point of -950 HU. Methacholine-induced air trapping on CT was defined at baseline as the absolute change in MLD, PD15 and LAA between the two expiratory scans before and after methacholine. The change in volume between the two expiratory scans before and after Methacholine was also corrected for inspiratory lung volume by using the following equation:
Volume Change=((Inspiration-Expiration) – (Inspiration-Expiration postMCh))*100%
(Inspiration-Expiration)
Combined measurement of FVC and SVC
FVC and SVC were measured in a combined manoeuvre, which is schematically illustrated in Figure E1. Subjects were coached into tidal breathing, after which they were asked to exhale completely to RV level. After a complete expiration, they inhaled slowly to TLC level, which was measured as a SVC manoeuvre. After slow complete inspiration, subjects were asked to perform a forceful complete expiration, during which FEV1 and FVC were measured.
Figure E1 Schematic illustration of the combined measurement of FVC and SVC
FVC: forced expiratory vital capacity
SVC: Slow inspiratory vital capacity
Cytokines measured in epithelial lining fluid (ELF) in peripheral airways
In a subset of 7 subjects, a bronchoscopy was performed under local anesthesia at baseline and after treatment. ELF was sampled by advancing 3 microsample probes (BC-401C, Olympus, Tokyo, Japan) in the lumen of the right main bronchus (central) and 3 probes in the anterior, lateral and posterior segments of right lower lobe (peripheral). A probe consists of a 1.8 mm outer diameter polyethylene sheath and a 1.1 mm inner polyester probe attached to a stainless steel guide wire. Sampling was performed by making contact with the bronchial wall for 10 seconds. Probes contaminated with blood were excluded from analysis. Each probe was sectioned from its guide wire and placed in an empty tube to which 1 mL of saline was added, centrifuged for 10 minutes at 30 rates/minute. Subsequently, the probe was removed from the tube and supernatant was stored at -80°C until use. Interleukin-4 (IL)-4, IL-5, IL-6, IL-8, tumor necrosis factor-alpha (TNF-α), Leukotriene B4 (LTB4) eosinophil cationic protein (ECP)and Thymus and Activation-regulated Chemokine (TARC) were measured in centrally and peripherally-sampled supernatant. IL-4, IL-5, IL-6, IL-8 and TNF-α were measured by Luminex (Linco, Nuclilab BV, Ede, The Netherlands). LTB4 (Amersham LTB4 Biotrak Assay, GE Healthcare Europe, Eindhoven, The Netherlands), ECP (Unicap, Phadia BV, Nieuwegein, The Netherlands) and TARC (Quantikine Immunoassays, R&D Systems, Oxon, United Kingdom) were measured by ELISA’s. The mean of cytokine concentrations was used for analysis if more than 1 probe was available from 1 sampling site (central or peripheral).
RESULTS
Table E1 Methacholine-induced air trapping at baseline
Placebo (n=7) / Ciclesonide (n=9)Total lung volume increase after MCh, % / 14 (2-35) / 24 (9-39)
Mean lung density decrease after MCh, HU / 45 (13-92) / 60 (36-157)
15th percentile density decrease after MCh, HU / 40 (-4-63) / 39 (15-101)
% LAA increase after MCh, % / 3 (-1-7) / 5 (1-9)
HU: Hounsfield Units
LAA: Low Attenuation Areas (areas with density -950 HU)
Table E2 Cytokine concentration in ELF sampled from peripheral airways in 3 subjects
Subject4 (ciclesonide) / Subject 6 (ciclesonide) / Subject 7(placebo)
Baseline / Treatment / Baseline / Treatment / Baseline / Treatment
ECP (ng/mL) / 0 / 0 / 136.4 / 0 / 0 / 0
IL-4 (pg/mL) / 0 / 0 / 168.0 / 0 / 1359.2 / 968.8
IL-5 (pg/mL) / 0 / 0 / 50.4 / 0 / 0 / 7.9
IL-6 (pg/mL) / 383.9 / 0 / 74.6 / 180.6 / 924.1 / 1086.7
IL-8 (pg/mL) / 4.6 / 11.5 / 4.0 / 9.6 / 8.8 / 6.8
LTB4 (pg/mL) / 714.9 / 2614.9 / 219.5 / 358.4 / 1805.4 / 3561.7
TNF-α (pg/mL) / 7.0 / 11.0 / 45.1 / 4.0 / 27.9 / 25.7
REFERENCES
E1. Kuhnigk JM, Dicken V, Zidowitz S, Bornemann L, Kuemmerlen B, Krass S, Peitgen HO, Yuval S, Jend HH, Rau WS, Achenbach T. Informatics in radiology (infoRAD): new tools for computer assistance in thoracic CT. Part 1. Functional analysis of lungs, lung lobes, and bronchopulmonary segments.Radiographics 2005;25:525-536.