Journal of Applied Physiology
Vol. 82, No. 4, pp. 1378-1378, April 1997
PULMONARY CIRCULATION AND LUNG FLUID BALANCE

Retention of soluble ^99mTc-DTPA in the human lung: 24-h postdeposition

W. Michael Foster, Pamela T. Stetkiewicz, and Arthur N. Freed

Department of Environmental Health Sciences, School of Hygiene and Public Health, The Johns Hopkins University, Baltimore, Maryland 21205

ABSTRACT

INTRODUCTION

METHODS

RESULTS

DISCUSSION

ACKNOWLEDGEMENTS

FOOTNOTES

REFERENCES

ABSTRACT

Foster, W. Michael, Pamela T. Stetkiewicz, and Arthur N. Freed. Retention of soluble ^99mTc-DTPA in the human lung: 24-h postdeposition. J. Appl. Physiol. 82(4): 1378-1382, 1997.Clearance of low-molecular-weight solutes, e.g., radiolabeled chelate diethylenetriaminepentaacetate (DTPA), across epithelial surfaces of distal airways and the lung parenchyma is a broadly used technique to assess epithelial integrity. It has been generally assumed that clearance of solute follows a simple first-order process and that DTPA clearance through the respiratory epithelium and into blood and lymphatic channels is complete within a few hours. Using -camera imaging and a radiolabeled aerosol of ^99mTc-labeled DTPA, we observed in eight healthy subjects lung retention of radioisotope ~24 h postdeposition of the ^99mTc-DTPA. Residual lung retention at the 24-h end point averaged 6.0 ± 1.8 (SD)% of the amount of radioisotope initially deposited in the lung. This suggests that for normal healthy subjects a small amount of the ^99mTc radioisotope, either in a dissociated or chelated form, is nonpermeable or slowly cleared from respiratory tisssues.

respiratory epithelium; airway; ozone; technetium-99m-labeled diethylenetriaminepentaacetate

INTRODUCTION

AEROSOLIZED LOW-MOLECULAR-WEIGHT solutes have been commonly used to gauge the permeability of the respiratory epithelium (2). The clearance of radiolabeled chelate diethylenetriaminepentaacetate (DTPA) across epithelial surfaces of distal airways and the lung parenchyma can be assessed noninvasively by external detection (single scintillation probe or by two-dimensional scanning of the thorax). Since its introduction in 1979 by Rinderknecht and associates (14), this technique has been applied in numerous experimental and clinical investigations to assess the integrity of the respiratory epithelium (6, 11). A number of -emitting radioisotopes have been utilized for the labeling of DTPA, but, by far, technetium 99m-labeled DTPA (^99mTc-DTPA; mol wt 492) has been the preferred chelate species in both model and clinical studies (11). The clearance rate of small radiolabeled solutes from pulmonary tissues under normal conditions has been found to be linear with clearance half times of <100 min. It is generally assumed that the clearance of solute, e.g., radiolabeled chelate ^99mTc-DTPA, follows a simple first-order process and that solute clearance through the respiratory epithelium and into the blood and lymphatic channels is complete within a few hours (17).

We recently utilized this technique to assess the permeability of the respiratory epithelium of healthy human subjects at baseline and 20 h after laboratory chamber exposures to filtered air or ambient concentrations of an oxidant air pollutant ozone (O₃) (4). As part of these evaluations, we reimaged the subjects by -camera at ~24 h postinhalation of the ^99mTc-DTPA aerosol and we found that a residual amount of the radioisotope could be counted within the lung field.

The purpose of this communication is to acknowledge our finding that a residual amount of radiolabel is retained in lung tissues 24 h postinhalation. Thus, for normal healthy subjects, regions of the respiratory epithelium may be non- or slowly permeable to ^99mTc-DTPA. This finding has an impact on physiological and clinical studies in which the chelate ^99mTc-DTPA is used to characterize epithelial integrity. The amount of radioisotope retained in respiratory tissues may need to be considered as a correction factor when clearance is reevaluated within a 24-h period.

METHODS

As participants in a research study, eight healthy subjects (7 men and 1 woman) had their pulmonary clearance of ^99mTC-DTPA evaluated. All of the subjects were nonsmokers, had no history of lung disease, and were not receiving medications for any other disease. The subjects had a mean age of 26 ± 2 (SD) yr and were free of respiratory infection at the time of the baseline study. The forced vital capacity, forced expiratory volume at 1 s, and midmaximal expiratory flow rate of the group averaged >92 ± 11% of predicted. Consent was obtained from the subjects before admission to the study, and the research was approved by the University Review Board.

The lung clearance of small solute was measured by using freshly prepared ^99mTc-labeled DTPA (Medi-Physics, Arlington Hts, IL). A submicronic ^99mTc-DTPA aerosol (0.95 µm count median diameter and geometric SD = 1.8) was generated by jet nebulization, and the subjects, seated in an erect position, inhaled tidal breaths of aerosol at ambient pressures by mouth. ^99mTc-DTPA sampled from the nebulizer reservoir postnebulization was assayed for unbound ^99mTc by using silica gel media and thin-layer chromatography to verify labeling procedures (1, 11). Aerosol was delivered early in the inspiratory cycle of tidal breaths to enhance transport of the aerosol into the lung periphery (7). A visual indicator of the inspiratory flow rate assisted subjects in maintenance of flow <0.4 l/s during the aerosol inhalations (mode Elektro-2, Respiratory Care Center, Finland); a range of three to four aerosol breaths was required to achieve significant deposition of radioisotope in the lung field.

Immediately after inhalation and deposition of ^99mTc-DTPA aerosol, the initial distribution and retention of aerosol were measured in subjects in a seated and erect position by imaging the thorax with a posteriorly positioned -camera (Maxi Camera, General Electric Medical Systems, Pittsburgh, PA). The camera was set with a 18% window around the peak energy of ^99mTc and was shielded by a parallel-hole collimator. Clearance of ^99mTc-DTPA from the lung was then monitored for at least a 100-min period; subjects returned to the facility ~24 h postinhalation of ^99mTc-DTPA and were reimaged for the presence of residual radioactivity within the lung. Images were stored by computer (Sopha Med, Columbia, MD) for subsequent analysis.

The eight subjects were restudied as volunteers in an experimental protocol to evaluate the effects of O₃ exposure on respiratory epithelial permeability. Thus, on 2 additional study days, the subjects were exposed for 130 min in an experimental chamber to an ambient profile of O₃ concentrations or filtered air (activity during the exposure periods alternated between 10 min of rest and light treadmill exercise). The order of exposures was randomized, and washout time between exposures was 7-14 days. After completion of the exposure period, the subjects left the laboratory and returned 18-20 h later to have lung clearance of ^99mTc-DTPA reassessed. After each of these evaluations of ^99mTc-DTPA clearance, subjects again returned to the laboratory ~24 h postinhalation of the labeled aerosol to image the lung for residual retention of ^99mTc.

Analysis of radioimages. On an initial screening day, after history taking and spirometric assessment of pulmonary function, subjects who qualified for the study had a ¹³³Xe ventilation scan performed. The ventilation scan was acquired to evaluate regional volume and identify lung regions for subsequent analysis of ^99mTc-DTPA deposition and retention. With noseclips in place, the subjects rebreathed ¹³³Xe gas by mouth (Pulmonex, Atom Products, Shirley, NY) to achieve a steady-state count rate (defined as the point at which there was no further increment in counts, i.e., a plateau in the count rate for the thorax had occurred), and a lung image was acquired and stored by computer. The steady-state image stored on a video screen enabled regions of interest to be selected by cursor manipulation and drawn to cover 1) the entire right lung field, 2) a central lung zone surrounding the hilus, and 3) a peripheral lung zone that included an outer envelope of the right lung. The regional area of the central and peripheral zones encompassed on average 16 and 36%, respectively, of the area covered by the right lung field. For a number of the subjects, ^99mTc-DTPA radioactivity immediately after inhalation was within extrapulmonary areas (digestive tract) and made it difficult to clearly define the left lung base; therefore, the left lung was not included in the analysis.

^99mTc-DTPA deposition was quantitated by using a technique modified from Foster et al. (3). To characterize the distribution of ^99mTc-DTPA aerosol deposited in the lung, a deposition index for the right lung was calculated as the ratio of ^99mTc-DTPA radioactivity in the central (C) and peripheral (P) lung zones (described above) divided by the ratio of ¹³³Xe radioactivity (steady state) in the central and peripheral lung zones as follows Therefore, an index value near unity signifies a homogeneous distribution of deposited ^99mTc-DTPA aerosol equivalent to the distribution of regional lung volume (at functional residual capacity) of central and peripheral regions, and with less penetration of aerosol and increased bronchial deposition the index was >1.00 (3, 7).

The amount of residual ^99mTc activity retained in the lung 24 h postinhalation of the ^99mTc-DTPA aerosol was assessed for the entire right lung region after background subtraction and decay correction. This value was expressed as a percentage of the amount of ^99mTc activity deposited initially at inhalation.

Statistical analysis. Means ± SE were calculated with standard statistical methods, and comparison of lung retentions between studies (baseline vs. treatments with filtered air or O₃) was accomplished by paired analysis using Student's t-test.

RESULTS

For the eight subjects evaluated under baseline conditions, the initial distributions of ^99mTc-DTPA aerosol deposited centrally and peripherally in the right lung field are presented in Table 1. The ratio of aerosol (^99mTc-DTPA) to ventilation (¹³³Xe) counts for central-to-peripheral lung zones is frequently used as a deposition index to reference the homogeneity of aerosol deposition (7). For the eight subjects evaluated, the deposition index had a mean value of 1.19. 

Table 1. Baseline study: lung distribution of 99mTc-DTPA aerosol deposition and retention at 24 h postinhalation Subject No. Deposition, %  Distribution Index (C/P) 24-h Lung Retention, %  Central Peripheral 1 23 31 0.95 4.0 2 20 35 0.84 6.4 3 19 30 0.97 5.3 4 19 23 1.55 7.0 5 20 22 1.14 8.7 6 23 20 1.41 6.5 7 24 19 1.35 7.2 8 23 17 1.29 3.2 Mean ± SD 21 ± 2  25 ± 6  1.19 ± 0.25  6.0 ± 1.8 C/P, index of distribution of 99mTc-labeled diethylenetriaminepentaacetate (DTPA) deposited in central (C) and peripheral (P) zones of right lung. Lung zones based on 133Xe ventilation image (at steady state) and index are calculated as ratio of CDTPA/PDTPA radioactivity to CXe/PXe, radioactivity (see METHODS).

Also listed in Table 1 are the amounts of ^99mTc counts retained in the right lung field ~24 h postdeposition of the ^99mTc-DTPA aerosol. The percentage of counts retained was small (range: 3.2-8.8% of the aerosol counts deposited initially); however, for each subject, an image was definable, and counts within the lung region were above background. The lung was not analyzed regionally, but on visual examination of the images the radiolabel appeared diffusely distributed throughout the lung field. Figure 1 demonstrates one subject's radioimage of the lung field acquired at this time point, and included for comparison is the initial scan of deposited ^99mTc-DTPA aerosol, acquired immediately after inhalation.

The mean lung retentions (at 24 h postinhalation) observed during the additional ^99mTc-DTPA clearance studies after the experimental exposures to filtered air and O₃ are represented in Fig. 2, and included for comparison is the mean of the lung retentions observed during the baseline ^99mTc-DTPA clearance studies. The administration of the ^99mTc-DTPA aerosol and imaging techniques were identical to those used to acquire the baseline data. Although the subjects were predisposed to treatment (exposure to O₃ or filtered air) at a time point 18-20 h before their DTPA clearance study, the lung retentions observed 24 h postinhalation of ^99mTc-DTPA were again small but exceeded background counts for each subject (and were comparable to the %lung retentions observed in the baseline studies).

DISCUSSION

Our experimental results suggest that a residual amount of the radiolabeled DTPA did not clear the lung in a 24-h period after deposition. To our knowledge, this observation has not been previously noted, although the measurement of pulmonary clearance of ^99mTc-DTPA by external detection has been applied for almost 20 years as an index of epithelial leakiness. A number of explanations are possible for this observation: 1) after deposition, dissociation of the ^99mTc label from the DTPA chelate and adherence of the label to intracellular or extracellular elements prevents clearance; 2) after clearance through the respiratory epithelium and passage into the pulmonary circulation, a redistribution within pulmonary tissues occurs; and 3) phagocytosis and retention of ^99mTc-DTPA within parenchymal cells and/or the lymphatic system slow clearance. Increases in lung volume and its attended effects on surface area also influence the lung clearance of ^99mTc-DTPA in humans (9), but changes in lung volume, in excess of changes that normally occur during tidal breathing, were absent from our protocols (4).

Nolop and co-workers (10) demonstrated that the bond between DTPA and ^99mTc was labile to oxidative dissociation and could lead to the formation of ^99mTcO₄. Placed on the tracheal epithelium of the dog, ^99mTcO₄ behaves as a pseudohalide, in addition to its primary pathway for passive transport through paracellular diffusion, a proportion is actively transported across epithelial cells via chloride channels (8). However, recent studies with DTPA labeled with indium, in comparison with ^99mTc-DTPA in humans, found little difference in clearance kinetics between the two labeled species of DTPA. Analyses of urine for ^99mTcO₄ did not support a significant dissociation of ^99mTc-DTPA in the normal or diseased lung (12).

In a study by Stather et al. (16), the retention in humans of intravenous vs. inhaled DTPA was conducted. After intravenous injection, DTPA retention in the blood could be described by three exponential components with half times of ~1.4, 14.3, and 95 min; by 24 h, >99% of the DTPA had been excreted in the urine and <0.5% remained in the plasma. After inhalation, DTPA retention in the lungs could be represented by a single component with a half time of ~75 min.

Therefore, neither dissociation of ^99mTc from DTPA and subsequent retention of ^99mTcO₄ within pulmonary cells nor redistribution of ^99mTc-DTPA to respiratory tissues after transfer into blood would appear to be explanations for lung retention of residual ^99mTc radioactivity.

Although the pathway(s) for removal of ^99mTc-DTPA is not fully defined, evaluations using the sheep model and lung lymph flow support the concept that ^99mTc-DTPA is normally cleared into the bloodstream. For example, after injury by infusing air into the circulation, increased amounts of ^99mTc-DTPA are removed by lymphatic drainage. However, even with this condition, <1% of the ^99mTc-DTPA lung clearance is by the lymphatic route (13). In the dog model, occlusion of the pulmonary artery delayed ^99mTc-DTPA lung clearance, and thus this circulation has been interpreted to be the primary clearance route, as occlusion of the bronchial artery had little influence on clearance (15).

Supplementing the retention data observed after baseline clearance of ^99mTc-DTPA, the same eight subjects, as participants in an experimental study to compare exposure to O₃ (ambient levels) and filtered air, had clearance of ^99mTc-DTPA evaluated on two additional occasions. The lung retention results at 24 h postinhalation of aerosol in these additional studies were similar to 24-h retentions observed during the baseline study. There was a tendency for the lung retention values 24 h postinhalation to be lower when subjects were pretreated with O₃, but this trend was not significant. Thus, in our laboratory, residual radiolabel is found that apparently does not clear the pulmonary system within the initial 24-h period postinhalation. This occurred with ^99mTc-DTPA chelate as the permeable solute and appeared to be a reproducible observation in the normal lung.

In an effort to assess whether the residual radioactivity was tightly adhered to pulmonary tissues, we delivered to an anesthetized dog model a ^99mTc-DTPA aerosol with inhalation techniques identical to those used in our human subjects; although, in the animals, the aerosol was delivered via an endotracheal tube. In all three dogs evaluated in this manner, activity was measurable 24 h postinhalation of the ^99mTc-DTPA aerosol, but in only two of the dogs was the amount significantly above background. After imaging of the thorax at the 24-h time point, distal lung units of the dogs were lavaged by bronchoscopic technique (5) to assess whether radiolabel was removable and/or redistributed by the lavage procedures. The dogs were reimaged after lavage. Lung retention data for these three dogs at ~24 h postinhalation of the ^99mTc-DTPA aerosol are presented in Table 2. Although only a single lobe of the dog lung was lavaged, activity was not found in the lavage fluid nor were any differences noted in the retention of the ^99mTc radioactivity, compared with the lung images acquired prelavage. Thus it appears that residual retention of radiolabel can also occur for the dog lung, as in the human lung, and that the radioactivity is not recoverable by bronchoalveolar lavage.

Table 2. Lung retention in dog lung 24 h postinhalation of 99mTc-DTPA aerosol Dog No. Right Lung Retention, %  24 h 24 h postlavage 1 6.9 6.7 2 0.7 0.9 3 3.3 2.9 Right lung retention, %initially deposited radioisotope retained in right lung at 24 h, pre- and postlavage.

In summary, the results presented in this communication do not nullify lung clearance of ^99mTc-DTPA chelate as a valid technique for assessing respiratory epithelial integrity. However, the results suggest caution in assuming that the respiratory epithelium is uniformly permeable to small-molecular-weight solutes, e.g., ^99mTc-DTPA (mol wt 492), and that chelate removal from the lungs by pulmonary blood flow is complete within a 24-h period. The influence of destructive pulmonary disease on residual retention or delayed clearance of ^99mTc-DTPA may need to be evaluated.

ACKNOWLEDGEMENTS

The authors thank Kristen Macri and Theresa Myers for their superb technical assistance.

FOOTNOTES

Address for reprint requests: W. M. Foster, no. 7006 Hygiene Bldg., 615 North Wolfe St., Baltimore, MD 21205 (E-mail: mfoster{at}welchlink.welch.jhu.edu).

Received 1 August 1996; accepted in final form 26 November 1996.

REFERENCES