Resources of Airway Remodeling in Severe Asthma

Study Design

As part of the Severe Asthma Research Program (SARP) at the National Institutes of Health, a prospective cohort of subjects, following informed consent, underwent detailed testing and MDCT scanning using a standardized protocol that was developed by the SARP. MDCT scan data were analyzed and compared in a total of 123 subjects, as follows: patients with severe asthma, n = 63; patients with mild-to-moderate asthma, n = 35; healthy subjects, n = 25. In a subset of patients who underwent airway biopsies (n = 32), we correlated segmental airway WT and wall area (WA) from MDCT scans with measures of airway remodeling from biopsy specimens, including epithelial and lamina reticularis (LR) thickness, from the corresponding segment. All subjects at the Washington University SARP site were given the option of participating in the airway biopsy substudy. The study was approved by Institutional Review Board at each site and monitored by an independent Data and Safety Monitoring Board.

Human Subjects

The inclusion criteria by group were as follows: for healthy subjects: 18 to 60 years of age; in good overall health; no smoking within past 5 years; < 5 pack-years of smoking; and a provocative concentration of methacholine causing a 20% fall in FEV1 (PC20) of > 16 mg/mL; for patients with mild-to-moderate asthma: 18 to 60 years of age; physician diagnosis of asthma; receiving asthma therapy for > 12 months; daytime asthma symptoms more than two times per week but less than continual and/or nocturnal asthma symptoms more than two times per month but less than nightly; no smoking within the past 5 years and < 5 pack-years of smoking; no concurrent lung disease; and PC20 of < 8 mg/mL or > 15% improvement in FEV1 postbronchodilator therapy; for patients with severe asthma: 18 to 60 years of age; subjects with one or both of two major criteria and two of the minor criteria (American Thoracic Society workshop). The major criteria include (in order to achieve control to a level of mild-to-moderate persistent asthma), the following: (1) continuous or nearly continuous treatment with (ie, > 50% of the previous year) oral corticosteroids; or (2) treatment with high-dose inhaled corticosteroids (beclomethasone [> 1,260 μg/d]; budesonide [> 1,200 μg/d]; flunisolide [> 2,000 μg/d]; fluticasone [> 880 μg/d]; or triamcinolone [> 2,000 μg/d]). The minor criteria include the following: (1) daily treatment with a longterm controller medication in addition to inhaled corticosteroids (eg, long-acting P-agonist, theophylline, or leukotriene antagonist); (2) asthma symptoms requiring short-acting P-agonist use on a daily or nearly daily basis; (3) persistent airflow obstruction (FEV1, <80% predicted and diurnal peak flow variability > 20%); (4) one or more urgent care visits for asthma per year; (5) three or more oral corticosteroid “bursts” per year; (6) prompt deterioration with < 25% reduction in the dosage of oral or inhaled corticosteroids; and (7) a near-fatal asthma event in the past. In these subjects with severe asthma, conditions other than asthma were excluded, exacerbating factors were treated, and the subject did not have a history of poor adherence to treatment. See more articles in the category Asthma.

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MDCT scanning

CT Scan Technique

All subjects underwent an MDCT scan of the chest with 16 or 64 detector rows (Light Speed Ultra 16; GE Healthcare; Milwaukee, WI; or Volume Zoom, Sensation 16 or 64; Siemens; Forchheim, Germany) after maximal bronchodilation with albuterol (540 to 720 μg) to minimize the effect of acute broncho-constriction on airway dimensions. Subjects were administered increasing doses of albuterol until the FEV1 percent predicted difference was < 5% or a maximal dose of albuterol (8 puffs or 720 μg) was reached. Suspended full inspiratory measurements were obtained at the following settings: for the GE scanner: pitch, 1.675 to 1.75 (mm table increment per rotation/detector collimation X number of detectors); rotation time, 0.6 s; 120 kV; reconstructed slice thickness, 0.625 to 1.25 mm; and medium smooth “standard” reconstruction algorithm; for the Siemens scanner: pitch, 1.5; rotation time, 0.5 s; 120 kV; reconstructed slice thickness, 1 mm; medium smooth reconstruction algorithm; and effective, mAs 30 to 57. To obtain isotropic voxels, the slice reconstruction interval was set to equal the in-plane spatial resolution (field of view measured in millimeters per 512 pixels).

MDCT Scan Airway Evaluation Software

MDCT scans were analyzed using automated, quantitative software that was designed to reliably label and segment the first five to six airway generations, and to allow the accurate measurement of airway wall and lumen diameters obtained perpendicular to the long axis of each airway (Pulmonary Workstation, version 0.139; VIDA Diagnostics; Iowa City, IA). Previous studies have validated the lung and airway segmentation methods when compared to manual measurements. Airway measurements for each segment were made at each centerline voxel and were averaged over the middle third of the segment. The specific MDCT scan measurements used included airway WT, percentage of WT (WT%), WA, percentage of WA (WA%), luminal area (LA) and percentage of LA (LA%). The calculations are as follows: WT = average outer diameter — average inner diameter; WT% = (WT/average outer diameter) X 100; WA = total area (TA) — LA; WA% = (WA/TA) X 100; and LA% = (LA/TA) X 100 (Fig 1).

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In the primary analysis, we averaged third-generation airway wall measurements for all automatically segmented and labeled airways in each subject. An average of 18 third-generation airways per subject were measured (9 for each lung). In the secondary analysis of subjects who underwent biopsies, we obtained crosssectional CT scan measurements of each biopsied airway in a plane perpendicular to the airway long-axis at a distance 30% of the segment length distal to the origin of the airway segment. Two independent readers trained in the use of the Pulmonary Workstation were blinded to the subject status or the results of the biopsy measurements. Interrater reliability on a random sample of 50 subjects between these two readers was excellent (intraclass correlation, 0.98). The biopsy can be done in four ways, more can be viewed on this website.

Pulmonary Function Tests

Pulmonary Function Tests

Spirometry, methacholine bronchoprovocation tests, and ple-thysmographic lung volume measurements were performed within 1 to 2 days of the MDCT scan among the SARP sites in accordance with standardized American Thoracic Society crite-ria. Subjects were asked to abstain from the use of long-acting P2 agonists for 12 h and from the use of short-acting p2 agonists for 4 h prior to assessment. Spirometry was performed before and after therapy with a short-acting P2-agonist (4 puffs or 360 μg of albuterol) was delivered by metered-dose inhaler and spacer.

Endobronchial Biopsies

A subset of 32 subjects (15 subjects with severe asthma; 9 subjects with mild-to-moderate asthma; and 8 healthy subjects) underwent bronchoscopy, and 12 to 16 endobronchial biopsy specimens were obtained from the third-generation segmental carinas of the upper lobes. Three readers independently measured the areas of the epithelium and LR of at least three biopsy specimens with intact airway epithelium using software for morpho-metric analysis. The areas of the epithelium and LR were normalized for the length of the basement membrane, yielding an epithelial and LR ratio. The average of these measurements made in triplicate was then used for subsequent analysis.

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Statistical Analysis

All data were analyzed using analysis of variance and x2 tests to compare continuous and categoric variables across groups (SAS, version 9; SAS Institute; Cary, NC). Stepwise multiple linear regression identified variables that had independent significant associations with outcome measures that included FEV1 percent predicted, WA%, and WT%. Variables were retained in these models if they had a significant (p < 0.05) or borderline significant (p < 0.1) association with the outcome measure.

Figure 1
Figure 1. Airway measurements by MDCT scanning. Multiple airway measurements can be performed (using the Pulmonary Workstation program; VIDA Diagnostics). Depicted are the two primary measurements used in the current study. WT is measured over the middle one third of each segment at each centerline voxel. This value is averaged, resulting in a single WT value for each segment. The average outer diameter for the segment is calculated in a similar fashion. The WT% for each segment is calculated by dividing the WT by the average outer diameter. TA and LA are similarly calculated at each centerline voxel and averaged over the middle one third of the segment. WA was calculated by subtracting the LA from the TA. WA% was calculated by dividing the WA by the TA.