INTRODUCTION

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The absence of a maximal dose-response plateau (1), the presence of gas trapping (2), and increases in closing capacity (CC) (3) during bronchoconstriction in asthma suggest that airway closure is an important functional abnormality of asthmatic airways. Airway closure can be assessed from the closing volume (CV) of the single-breath nitrogen curve. It may be increased in clinically stable, asymptomatic asthmatic subjects (4), and may be present at functional residual capacity (FRC) in both the supine and seated positions, leading to arterial hypoxemia secondary to low ventilation/perfusion ratios (5). The degree of airway closure should increase during sleep as the result of a combination of factors, including the decrease in FRC caused by the supine posture, reduced respiratory muscle activity, deep breathing, and a circadian increase in airway tone. Such an increase in closure may result in nocturnal oxygen desaturation.

Studies of airway closure have been done with boluses of inert radioactive gas isotopes inhaled from residual volume (RV), with measurement of the radioactivity of the inhaled bolus made with five pairs of radiation counters placed equidistant from one another along the vertical dimension (height) of the lung. This technique showed that: (1) airway closure was present at low lung volume in normal subjects; (2) airway closure completely explained the distribution of a gas bolus inhaled from RV (6); and (3) CV represented the onset of basal airway closure in normal subjects (7). The distribution of airway closure in relation to lung height was described by the ratio of counts in the two uppermost counters compared with that in the two lowermost counters. This ratio suggested that in normal subjects, airway closure was predominantly basal in distribution, but became more extensive during induced bronchoconstriction, and that a larger vertical gradient of airway closure resulted in a greater height of phase IV of a Xe bolus washout (8). The height of phase IV was subsequently shown to be increased in asymptomatic asthmatic subjects, improving to some degree after isoprenaline inhalation, suggesting that increased bronchomotor tone in asthma also resulted in a more widespread apical-to-basal distribution of airway closure (4).

The use of gamma cameras and radioactive gases allowed a more complete assessment of the radioactivity within the chest. A two-dimensional picture of this radioactivity, called a planar scan, allowed much greater detail but also facilitated an indirect measurement of the lung volume subtended by closed airways at RV, which was shown to be proportional to CC (9, 10). These studies also confirmed the predominantly basal distribution of airway closure in normal subjects, which increased in extent and apical-to-basal distribution with increasing age (10), in accordance with previous findings of increasing CV with age (11). The apical-to-basal gradient of pleural pressure therefore appears to be the main determinant of the magnitude and distribution of airway closure in normal individuals. A study in asthmatic subjects also concluded that airway closure was increased, but that this occurred predominantly in the lung bases (12).

Single-photon emission computed tomography (SPECT) is a mode of gamma camera imaging in which three-dimensional images of radioactivity are generated. The relationship between SPECT and planar imaging is analogous to that between CT scanning and plain radiography. SPECT thus allows the direct measurement of lung volumes affected by airway closure, as well as allowing a detailed analysis of the topographic distribution of airway closure. We have previously confirmed that in normal subjects, the lung volume subtended by closed airways (LVclosed) at low lung volume, as measured with SPECT, correlated with CV and CC (13). In the present study, our aims were to compare, in clinically stable asthmatic subjects and normal subjects: (1) the amount of airway closure; (2) the apical-to-basal distribution of airway closure; and (3) the relationship between LVclosed and CV.

	METHODS

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION REFERENCES

Subjects

Thirteen normal and 23 asthmatic subjects were recruited from the staff and patients of the Royal Prince Alfred Hospital in Sydney. Informed consent was obtained and ethical approval was given by the ethics committee of the Central Sydney Area Health Service. All normal subjects denied any history of chronic illness, respiratory symptoms, or medication use. One subject was a current moderate smoker and two subjects were ex-smokers. Subjects in the normal group were excluded if their CV or CC was greater than the upper limit of predicted normal. This led to the exclusion of only one subject from the study.

All asthmatic subjects reported an asthma diagnosis by a physician, were either taking regular inhaled medications for their asthma, or had reported symptoms and medication use for their asthma within the preceding 12 mo. No subject had experienced any exacerbations requiring an increase in medications in the preceding 3 mo.

Lung Function

Spirometry, lung volumes, and CV were measured in all subjects except for two asthmatic subjects who were lost to follow-up during the study, and for whom only spirometric results were recorded. FEV₁ and FVC were measured with a Cavitron SC-20 electronic spirometer (Cavitron Corp., Anaheim, CA). The greater of two measurements that were within 100 ml of each other was chosen. Lung volumes of subjects in the upright position were measured with a Gould 2800 constant-volume body plethysmograph (Gould Electronics, Dayton, OH). Subdivisions of lung volume were computed from the mean FRC and inspiratory capacity (IC) recorded in three measurements, and from the largest vital capacity (VC). CV was measured through a previously described modification (13) of the single-breath nitrogen test described by Buist and Ross (11). CV was expressed as percent VC. CC was the sum of CV and RV, expressed as percent total lung capacity (TLC).

Asthmatic subjects were also divided into two groups to examine the relationship between abnormal spirometric function and airway closure. Those subjects with an FEV₁/FVC ratio of 0.75 or greater and an FEV₁ % predicted of 80% or greater were defined as the "normal spirometric group"; other asthmatic subjects were designated as the "abnormal spirometric group." All lung function data are expressed at body temperature-ambient pressure-water saturation (BTPS). Predicted values for spirometry were those of Morris and colleagues (14), for lung volumes those of Crapo and coworkers (15), and for the single-breath nitrogen test those of Buist and associates (11).

SPECT Scanning

Simultaneous emission/transmission SPECT was used to measure airway closure by determining the distribution of an inhaled bolus of Technegas. The method used for this has been previously described in detail (13), but will be briefly summarized. Technegas is an ultrafine carbon particle of median diameter 7 to 23 nm (16). It distributes similarly to an inert gas radioisotope (17, 18), but once it is inhaled the particles adhere to alveolar structures without appreciable movement for at least 40 min (18).

Four hundred to seven hundred mBq of (^99mTc)pertechnetate was used to generate Technegas from a Technegas generator according to the manufacturer's instructions (Tetley Manufacturing Ltd., Sydney, Australia). Having previously exhaled to RV from TLC, the subject slowly inhaled a bolus of Technegas, which was approximately 6% of VC, followed by inhalation of air during a slow inspiration back to TLC. The breath was then held for 10 s before resumption of normal breathing. The inhalation was performed in the standing position. The radioactivity within the thorax was measured in a single posteroanterior (PA) projection, and the inhalation was repeated if necessary until the radioactivity exceeded 900 counts/s (13).

SPECT studies were done by rotating a gamma camera around the subjects, who were lying supine and with their arms raised above their heads. Emission counts from the Technegas in the lungs, and transmission counts from an external moving line source, were simultaneously recorded on a computer (PDP-11 with NCV-11C gamma camera interface; Digital Equipment Corporation, Maynard, MA). The resulting data were reconstructed into transaxial slices of Technegas lung activity (emission data) and density maps of the chest (transmission data), which were inherently spatially registered because they were acquired at the same time (Figure 1). There were no gaps between slices for either emission or transmission studies, and absolute volumes could be estimated by counting the number of voxels in a region and multiplying by the known voxel volume of 0.2 ml.

View larger version (73K): [in this window] [in a new window]   Figure 1.   Coronal slices of emission and transmission images from a normal subject (age 27 yr). (a) Reconstructed transmission image. (b) Transmission image after segmentation to define lung outlines (LVtrans). (c) Reconstructed emission image. (d ) Emission image after application of the threshold to determine LVvent. (e) Superimposition of images (b) and (d ), demonstrating lung regions affected by airway closure as grey regions within the lung outlines.

Ventilated lung volume (LVvent) (i.e., regions containing Technegas) was determined from the number of voxels in the emission slices that emitted gamma radiation above a threshold derived to exclude background noise (Figures 1c and 1d). Lung regions in the density maps derived from the transmission study were determined with an automated segmentation algorithm, which separated lung tissue density from the densities of soft tissue and air (Figures 1a and 1b). Lung volume (LVtrans) was then determined from the number of voxels classified by the segmentation program as showing lung tissue density.

According to this technique, airway closure in SPECT scans (LVclosed) was defined as the percent of lung volume that Technegas failed to reach (Figure 1e). Thus, LVclosed = ([LVtrans  LVvent]/ Lvtrans) × 100, and has units %LVtrans.

The apical-to-basal distribution of airway closure was determined by dividing the lung outlines in the transmission scans into three vertical zones of equal height: an apical zone, a middle zone, and a lower zone or base. LVclosed was then determined for each of the three zones.

Data Analyses

Differences in means between groups were examined with Student's t test. The relationships between LVclosed and age, CV, and CC were examined with Pearson's correlation coefficient. Intergroup differences in LVclosed in the three lung zones were examined with analysis of variance (ANOVA) with repeated measures.

	RESULTS

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Lung Function

Normal subjects. Table 1 contains the anthropometric and lung function data for both the asthmatic and normal groups. FEV₁ and FVC of all the normal subjects were greater than 80% predicted, and all FEV₁/FVC ratios were greater than 0.75 except that for one subject (a male subject 56 yr of age and nonsmoker) in whom it was 0.74. There was a strong relationship between age and both CV (r = 0.83, p < 0.01; CV = 0.56 × age  8) and CC (r = 0.88, p < 0.01; CC = 0.60 × age + 13).

Asthmatic subjects. There were 13 asthmatic subjects with normal spirometric results and 10 with abnormal results as defined earlier. The subjects' mean lung function values are shown in Table 1. The mean FEV₁ and FVC % predicted and FEV₁/FVC ratios were significantly lower in asthmatic subjects than in normals (p < 0.05). CV was identified in all but one subject, even though the alveolar plateau for that subject was not curvilinear. CV was above the normal predicted value in two (9%) subjects, and CC was above the predicted value in six (26%) subjects. CC was greater in the group with abnormal spirometry results (138 ± 22.0% predicted) than in the normal subjects (106 ± 9.8% predicted, p < 0.01), but was not greater than that in the group with normal spirometric results (119 ± 18.7% predicted, p > 0.05). There were no significant differences between the normal and asthmatic subjects as groups in mean percent predicted CV or CC. The RV/TLC ratio was higher in the group with abnormal spirometric results (36 ± 6.8%) than in both the normal subjects (25 ± 2.1%, p < 0.01) and the group with normal spirometric results (25 ± 3.2%, p < 0.01); however, there was no difference in the ratio between normal and asthmatic subjects as groups. The geometric mean PD₂₀ was 1.36 (95% CI: 0.733 to 2.54) µmol methacholine.

SPECT Scans

The mean LVclosed was 31 ± 6.0% in asthmatic subjects and 30 ± 7.8% in normals; the difference was not significant. In normal subjects there was a strong correlation between LVclosed and age (r = 0.89, p < 0.01; LVclosed = 1.22 × age  10.6), CV (r = 0.86, p < 0.01; LVclosed = 0.44 × CV  2.2), and CC (r = 0.86, p < 0.01; LVclosed = 0.43 × age + 20.3). In asthmatic subjects there was no significant correlation between LVclosed and age (r = 0.36, p > 0.05), CV (r = 0.10, p > 0.05), or CC (r = 0.30, p > 0.05) in the asthmatic group as a whole or when divided into groups with normal and abnormal spirometric results.

Figure 2 shows the comparison of mean regional LVclosed in normal and asthmatic subjects. In normal subjects, regional LVclosed was greatest in the lung base (p < 0.01). LVclosed in the apical zone was smaller than in the lower zone (p < 0.01), but was not different from that in the middle zone. In asthmatic subjects, LVclosed was greatest in the lower zone, intermediate in the middle zone, and smallest in the apical zone (p < 0.01). Comparison of normal and asthmatic subjects revealed that LVclosed was significantly greater in the upper zones in normal subjects (p < 0.01); however, in the middle or lower zones there were no differences between normal and asthmatic subjects.

View larger version (15K): [in this window] [in a new window]   Figure 2.   LVclosed was greater in the upper lung zones in normal subjects (open circles) than in asthmatic subjects (open squares) (p < 0.01). In normals, LVclosed was greatest in the lung base (p < 0.01), but was similar in the upper and middle zones. In asthmatic subjects, LVclosed was greatest in the lung base, intermediate in the middle zone, and least in the lung apex (p < 0.01). Points represent mean ± 95% CI.

Table 2 shows regional LVclosed in the normal subjects and in the asthmatic subjects with normal and abnormal spirometric function. There were no differences in regional closure in asthmatic subjects with normal spirometric function and those with abnormal spirometric function. LVclosed in the upper zone in the groups of asthmatic subjects with both normal and abnormal spirometric results was significantly smaller than that in normals (p < 0.05).

In normal subjects there was a strong relationship between regional airway closure and age in the middle (r = 0.94, p < 0.01; LVclosed = 1.4 × age  31; Figure 3a) and lower (r = 0.92, p < 0.01; LVclosed = 2.69 × log_e[age  21.1]; Figure 3b) zones. In asthmatic subjects there was a weak relationship between regional airway closure and age in the middle zone only (r = 0.46, p < 0.05; Figure 3a). Separate analysis of the asthmatic groups with normal and abnormal spirometric results showed no significant correlations between regional closure and age in either group. Among asthmatic subjects under 35 yr of age, LVclosed was greater in the lower zone in the group with abnormal spirometric results (71 ± 11.2% LVtrans) than in the normal subjects (36 ± 18.2% LVtrans, p = 0.01) despite similar mean ages for the two groups (27 ± 2.9 yr and 27 ± 1.8 yr, respectively). There was no difference in LVclosed in the lower zone for the asthmatic group with normal spirometric results (56 ± 18.9% LVtrans) and for the normal subjects.

View larger version (12K): [in this window] [in a new window]   Figure 3.   Relationship between airway closure in the middle (a) and lower (b) lung zones with age in normal subjects and two asthmatic spirometric groups. (a) Extent of airway closure in the middle zone correlated strongly with increasing age in normal subjects (solid line) but only weakly in asthmatic subjects (dashed line) as a group. There was no correlation within the asthmatic spirometric groups individually. (b) In the lower zone there was a rapid increase in airway closure with increasing age in normal subjects, so that almost the entire zone was unventilated in those older than 35 yr (solid line). However, there was no correlation in asthmatic subjects.

Figure 4 shows scans from three normal subjects and three asthmatic subjects. Areas affected by airway closure were contiguous and relatively uniform in normal subjects. In asthmatic subjects, regions affected by airway closure were discrete and patchy in distribution, with large, wedge-shaped and peripherally located defects present in some. The affected areas did not favor any particular lung zone in their distribution.

View larger version (70K): [in this window] [in a new window]   Figure 4.   Single coronal slices from three normal subjects (aged 27, 30 and 43 yr, respectively; top row) and three asthmatic subjects (bottom row) showing the distribution of airway closure. Unventilated lung regions are relatively uniform and contiguous in normal subjects. Slices show typical examples of large, peripheral triangular defects in asthmatic subjects that suggest closure of subsegmental airways. Emission and transmission images have been superimposed. Soft tissue is black, unventilated lung regions are grey within the soft-tissue image, and ventilated lung regions are white.

	DISCUSSION

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION REFERENCES

The present study compared airway closure as measured with SPECT and single-breath nitrogen washout in normal and asthmatic subjects. The results of the study showed that: (1) the distribution of airway closure was predominantly basal in both asthmatic and normal subjects; (2) the greatest increase in airway closure with increasing age occurred in the lung base and progressed in a cephalic direction in normal subjects; (3) Airway closure was observed in the lung apex in both normal and asthmatic subjects; (4) despite similar mean values for age, CC, and LVclosed in the asthmatic and normal groups, the distribution of airway closure as measured with LVclosed was different in these groups; and (5) although there were strong correlations between airway closure as measured with SPECT and both CV and CC in normal subjects, no such correlations were present in asthmatics.

Advantages and Limitations to the SPECT Technique

The volume of lung affected by airway closure has not previously been directly measured in asthmatic subjects. The abnormal pattern of airway closure demonstrated in this study is therefore important because the theoretical limitations inherent in measurements of airway closure with planar imaging methods are not applicable to measurements with SPECT. An abnormal pattern of closure represents what is arguably the most important and serious consequence of excessive airway narrowing in asthmatic subjects, and suggests that premature closure of airways, suspected on the basis of an increased CC, is not occurring in the same airways or sites as in normal subjects.

The accuracy of the technique used in our study for detecting ventilation defects was estimated to be approximately 51 ml in phantom experiments. The technique is therefore unlikely to detect widespread and patchy closure of small airways resulting in small nonventilation lung regions of less than 51 ml volume. This may lead to an underestimation of the extent of airway closure in asthmatic subjects, and the technique may therefore detect the closure of central as opposed to peripheral airways, or the closure of many parallel peripheral airways.

The volume of the inhaled Technegas bolus in the SPECT and single-breath nitrogen washout technique is approximately 250 to 300 ml. This volume is necessary in order to distribute sufficient radioactivity within the subject's lungs. However, in lung units distal to closed airways in which gas trapping has occurred, airways may reopen suddenly during the bolus inhalation, leading to a further underestimation of the extent of airway closure in asthmatic subjects with this technique. Airway reopening may be less in normal subjects in whom gas trapping is less at RV.

A threshold technique was used in this study to exclude background counts, so that the ventilated lung volume (i.e., the volume containing Technegas) could be calculated (Figure 1). The use of thresholds is a fundamental technique in image analysis in nuclear medicine, and has been used in previous planar studies of airway closure (9, 10, 12), with thresholds based on 20% of the maximum count made in the study. A limitation of this method is reduced sensitivity in detecting areas of extremely low ventilation relative to other areas with much greater ventilation (and hence greater activity), which results in a higher threshold value. The techniques used in the present study address these limitations to some extent. First, the threshold was derived from the mean value in the largest cross-sectional slice, rather than the maximum value in the study. Using a mean value allows regions of low activity to influence the threshold value, limiting the effect that regions of very high activity have on the threshold. Second, three-dimensional data eliminated the effects of underlying and overlying activity that are inherent in the two-dimensional data of planar scans.

Airway Closure in Normal Subjects

The presence of an appreciable LVclosed in the apical regions of the lung in normal subjects was somewhat surprising. A similar finding was made in one previous study (10), but not in another (9). However, the disparity in the results of these two previous planar imaging studies may have come from differences in image-analysis methodology. Airway closure could be one possible cause of the high values for apical LVclosed, and would suggest that gravity and the vertical gradient of elastic recoil do not fully explain the distribution of airway closure. Variations in mechanical properties of the lungs and bronchomotor tone of the airways and chest wall may be factors influencing regional airway closure. The persistence of a vertical gradient of ventilation and CV in sustained microgravity (19) would support this hypothesis.

The most likely explanation for the higher values of LVclosed in the apical zone, however, is its high regional elastic recoil, resulting in high RV/TLC ratios (20) and consequently much lower ventilation in this region than in other regions. This would result in some areas in the apex with low activity, which may fall below the sensitivity of the SPECT method for detecting abnormalities in ventilation. It may be possible to confirm this by studying subjects in the supine position following inhalation of a Technegas bolus.

The strong correlation of LVclosed with age and CC in normal subjects suggests the presence of a narrow range of closing pressures, so that airway closure is principally determined solely by lung elastic recoil. The rapid increase in regional closure of the lower zone with age (Figure 3b) may be attributed to the shape of the thorax and the dome of the diaphragm, so that the reduction in pulmonary elastic recoil with age had a greater effect on airway closure in the lower zone than in the more proximal regions. The presence of regional mechanical differences responsible for the persistence of inequalities of ventilation and closing volume during sustained microgravity (19) supports this explanation. With regard to explaining the difference between the middle and lower zones, it is unlikely that age-related changes affect closing pressures to any greater extent in the basal airways than in the more apical airways. The linear increase in LVclosed in the middle zone with age, as shown in Figure 3a, appears to reflect the expected increase in the number or size of airways that close with increasing age, owing to changes in elastic recoil.

Airway Closure in Asthmatic Subjects

Closing volume may be difficult to interpret in asthmatic subjects because of the change in VC that occurs during bronchoconstriction (21). Both CC and CV may be underestimated by the single-breath nitrogen washout method as compared with the bolus method in asthmatic subjects (4, 22). This is thought to be due to failure to establish a large vertical nitrogen concentration gradient in the presence of gas trapping. Airway closure in this situation would, however, be detected with inhaled boluses of tracer gases such as ¹³³Xe and Technegas, and may partly account for the absence of a correlation between LVclosed and CC in asthmatic subjects in the present study. The lack of a correlation between LVclosed and age in asthmatic subjects, however, suggests that the differences between CC measured with the single-breath nitrogen and bolus methods are not systematic.

Airway closure in asthmatic airways is determined by a complex mechanical interaction of airway structures, involving: (1) smooth-muscle tone (23); (2) load applied to the airway smooth-muscle as determined by pulmonary elastic recoil, parenchymal interdependence, and accumulation of fluid in the peribronchial space (24, 25), and by the thickness and elastic properties of the airway wall and submucosa (26, 27); (3) mucosal folding; and (4) mucosal liquid surface tension. Airway closure is therefore likely to be patchy and variable with respect to distribution and the volume at which closure occurs (4, 21), and the absence of any significant correlation between LVclosed and age, in addition to lack of correlation with CC, is therefore not surprising.

Airway closure in the present study was greatest in the lower lung zones in both asthmatic and normal subjects, which suggests pulmonary elastic recoil and shear modulus remain very important factors in determining airway closure in asthmatic subjects. However, higher elastic recoil pressures of nondependent lung zones may be required to overcome the influence of factors within the asthmatic airway wall itself that affect closure, as discussed in the previous paragraph. This may explain the weak correlation between LVclosed and age in the middle zone, where elastic recoil is higher, as compared with the lower zone, where a correlation was absent. The effect of these airway abnormalities on airway closure appears to be most pronounced in the lower zone, where elastic recoil pressure is lowest, so that the relationship between age and LVclosed is lost.

Airway closure appears to be more extensive in asthmatic subjects under the age of 35 yr and with abnormal baseline spirometric results than in normal subjects of similar age (Figure 3). In these asthmatic subjects, almost all of the lower zone was affected by airway closure. This would suggest that abnormalities in asthmatic airways may reduce the FEV₁/FVC ratio as well as enhance airway closure. The more extensive airway closure in this lower zone can be seen in subjects with poorer spirometric results, presumably because of the lower elastic recoil in this zone and more severe disease affecting the airways. For similar reasons, differences would be unlikely to be found in this zone in asthmatic and normal subjects over the age of 35 yr because airway closure becomes extensive in normal subjects beyond this age (28).

The lung regions affected by airway closure in the normal subjects in the present study were contiguous and relatively even in distribution. In the asthmatic subjects the distribution was more patchy and uneven, with discrete, wedge-shaped defects observed in some, which suggests that closure may occur in large airways in asthmatic subjects, and that the sites of airway closure may be variable. Closure at low lung volume may result from mucus plugging or increased resting airway smooth-muscle tone, or from structural changes and airway smooth-muscle contraction leading to airway instability. Although it is thought that the cartilage in large airways provides a load against airway smooth-muscle shortening, thus limiting excessive airway narrowing (29), there is some evidence that closure of cartilaginous airways may occur in asthmatic individuals (30) and in animal models (31, 32).

An increased range of closing pressures and sites of airway closure in asthmatic subjects may influence gas exchange even during symptom-free periods. Some airways may close during normal tidal breathing, resulting in areas of low ventilation/ perfusion ratios and causing hypoxemia. This would be in keeping with the finding in previous studies of minor abnormalities in ventilation and ventilation/perfusion matching in clinically stable asthmatic subjects that did not correlate with the subjects' spirometric abnormalities (33, 34).

Summary

SPECT provides important information on airway closure beyond that provided by CC and CV. CV is thought to be the volume at which there is a rapid increase in closure (7), whereas LVclosed is the volume of lung subtended by airways that were closed at RV. In addition to giving measurements of volumes affected by closure the three-dimensional images provided by SPECT (Figure 4) can also demonstrate the patchy distribution of airway closure in asthmatic subjects, which would support the hypothesis of variability in the severity of pathologic changes at different sites in these subjects' airways.

The present study has shown that the predictable pattern of basal airway closure in normals is lost in asthmatic subjects. Although pulmonary elastic recoil influences the distribution of airway closure in asthmatic as it does in normal subjects, there appears to be an increased range of closing pressures in the former group, with the result that airway closure becomes more unpredictable and patchy in distribution.