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A Technique for Isolated Airway Segment Lavage
A Technique for Isolated Airway Segment Lavage* W. L. Eschenbacher M.D., F.C.C.P.; and T. R. Gravelyn, M.D.
A novel technique has been developed for the isolation and lavaging of a central airway segment in humans. The procedure uses a double-balloon airway-sampling catheter that is placed into the airways with 6beroptic bronchoscopic guidance. A protocol using this technique was attempted in 32 subjects (18 normal subjects apd 14 subjects with mild asthma), and 27 of the subjects were able to complete all phases of the protocol, in<:luding bronchoalveolar lavage
and lavage of an isolated airway segment. There was no significant complication in any subject. Cellular composition of the lavages demonstrated differences in cell differential and total cell counts for the two different sampled regions (central airway vs alveolar spaces~ We believe that this technique of lavage of an isolated airway segment is safe and has potential usefulness in many areas of pulmonary research.
from the airways alone would be important Sampling in many areas of pulmonary research. Currently,
subjects with asthma at baseline conditions and after a local challenge.
bronchoalveolar lavage is used to obtain material from the lung for cellular, protein, and mediator analyses; however, it is believed that the material sampled in bronchoalveolar lavage comes predominantly from the alveolar spaces. 1 The ability to sample directly from the airways alone would allow the recovery of material that would have particular relevance in certain diseases such as asthma. Although narrowing of the airways in asthma occurs throughout the tracheobronchial tree, many authors believe that the more central airways are more predominantly involved in the acute bronchoconstriction to both allergic and nonallergic stimuli. u Thus, obtaining material from the more central airways, rather than the alveoli, appears to be more relevant in identifying mediators and cells that are involved in the acute bronchoconstrictive response to stimuli in asthma. The ability to differentiate central from peripheral material may also be important in other research such as surfactant, protease inhibitors, or other pulmonary protein regional differences, or in studying aerosol deposition or pulmonary injury due to inhaled agents. With these needs in mind, we have developed a technique that allows the isolation and lavage of an upper airway segment in human subjects with minimal contamination from distal pulmonary units. This lavage of an isolated airway segment involves the use of a double-balloon catheter that can be placed into the airways using fiberoptic bronchoscopy fur guidance. Initially, we have used this technique for evaluating the fluid obtained from the airways of normal subjects and *From the Division of Pulmonary and Critical Care Medicine, University of Michigan Medical Center, Ann Arbor. Manuscript received October 6; revision accepted December 6. Reprint requests: Dr. Eschenbacher, BIH272, Universify Hospital, 1500 East Medical Center Drive, Ann Arbor 48104
MATERIALS AND METHODS
Airway-Sampling Catheter Based upon the design criteria of one of the authors (W L. E.), a commercial manufacturer (American Edwards Corp.) produced the catheters that were used in the development of this technique. The catheters are modifications of the pulmonary arterial catheter that is used fur hemodynamic evaluation in the pulmonary circulation. Each catheter made fur airway sampling was llO em in length and 2 mm in diameter and was made of plasticized polyvinylchloride. 1Wo latex balloons are located at the distal end of the catheter separated by 2 to 4 em (depending upon the particular design). Each balloon can be in8ated with up to 2.5 ml of air or 8uid (other models have been made with balloons with 1.5-ml capacity). In the interballoon segment, several ports are present that open to one channel fur infusion and to another channel fur withdrawal of 8uid. Thus, there are four channels throughout the length of the catheter: two fur the in8ations of the balloons and two fur the introduction and withdrawal of 8uid. A diagram of the catheter and its position when placed into the lungs is shown in Figure 1.
Protocol The placement of the airway catheter into the subjects was perfOrmed while observing the precautions outlined by the National Heart, Lung, and Blood Institutes Workshop on Fiberoptic Bronchoscopy and Bronchoalveolar Lavage in Asthmatics. • Each subject signed a consent form that had been approved by the human research committee at the University of Michigan Medical Center. The subjects with asthma were considered to have mild asthma and could stop their inhaled and oral medication fur 24 hours befOre the study. No subject had an upper respiratory infection within fuur weeks befOre the study. The subjects were carefully monitored throughout the protocol and fur several hours after the protocol. On the day of the study, we first determined baseline specific airway resistance (SRaw) fur each subject using a constant-volume whole-body plethysmograph (Airco 3000 System; Ohio Medical Products). For these measurements, we used the technique described by DuBois et al.1 We then gave each subject 1 mg of atropine sulfate by intramuscular injection 30 minutes before the placement of the bronchoscope or catheter into the airways. During this 30minute period, we gave routine nasal, pharyngeal, laryngeal, and CHEST I 92 I 1 I JULY, 1987
105
FIGURE 1. Double-balloon airway-sampling catheter and its position in left main-stem bronchus during isolated airway segment lavage. Bronchoalveolar lavage (BAL) is perfOrmed in right middle lobe. upper-airway anesthesia with topical solutions of 2 percent and 4 percent lidocaine. At the end of the 30 minutes, we repeated the measurement of SRaw. We then placed the subject in a supine position, attached a pulse-oximeter (N-100 Pulse Oximeter; Nellcor Inc) to their index finger, gave supplemental oxygen by mouth (4 to 5 Umin), and inserted a fiberoptic bronchoscope (Olympus BF-2TR), through one naris, past the vocal cords, and into the airways. We used some additional topical lidocaine within the airways and wedged the bronchoscope into a subsegmental orifice in the right middle lobe. We then perfOrmed bronchoalveolar lavage in a standard fashion 1 using three to five 30-ml aliquots of physiologic saline solution. After the bronchoalveolar lavage, we withdrew the bronchoscope back above the vocal cords. At this time, we inserted the airwaysampling catheter into the other naris and advanced it past the vocal cords using the bronchoscope for visual guidance. We positioned the distal end of the catheter into the left main-stem bronchus so that the distal balloon was proximal to the bifurcation of the left upper and left lower lobes. The proximal balloon would then be di$tal to the main carina in the left main-stem bronchus. Again, the position of the catheter in place is shown in Figure 1. We then inflated the distal balloon and checked the adequacy of the seal visually through the bronchoscope. We then inflated the proximal balloon, creating an isolated airway segment. Next we lavaged the isolated segment with different fluids. By calculation of the potential volume of the isolated airway segment (using standard anatomic bronchial dimensions and the length of the interballoon segment) and from experience in perfOrming these lavages of isolated segments, we found that 2-ml aliquots of fluid would almost completely fill the airway segment without spilling outside the segment. We introduced 2-ml aliquots of physiologic saline solution into the segment, checking again for the adequacy of the balloon seals both proximally and distally, and then withdrew the saline solution through the other channel using a 3-ml syringe and hand suction only. We recorded the recovery from each separate aliquot introduced but combined all the aliquots from a
108
particular series into a common container. In all studies, we first perfOrmed four to ten lavages with 2-ml aliquots of physiologic saline for baseline values. We would then use one of two different fluids for the second series of lavages, either distilled water or hypertonic sodium chloride solution (3.6 percent sodium chloride or approximately 900/mOsm). Again, we used fOur to ten lavages with 2-ml aliquots for this second series. Finally, for the third series, we again used four to ten 2-ml lavages of physiologic saline solution. After completion of this final series oflavages, we deBated the balloons and removed the catheter and the bronchoscope. We then repeated the measurement of SRaw for the subject. Subjects were then followed for the next two to fOur hours for symptoms, but no further measurements of SRaw were made. On the average, it took three to five mintues for the insertion ofthe bronchoscope, five to eight minutes for the bronchoalveolar lavage, five to ten minutes for the insertion of the catheter, and three to five minutes for each lavage of the isolated airway segment. Overall, the entire procedure after anesthesia lasted 30 to 45 mintues. The fluid obtained from the bronchoalveolar lavage and the isolated airway segment lavages was then centrifuged at 1,000 g for five minutes. The supernatant was stored at -70"C for future mediator analyses. The cell pellets were resuspended in Hanks balanced salt solution and were washed twice. A portion of each suspension was then spun in a cytocentrifuge (Cytospin 2; Shandon Southern Instruments) and stained using Papanicolaou$ stain for differential cell count. Three hundred cells were counted for each sample. Another portion of the cell suspension was used for total cell count using a hemacytometer.
REsur.:rs We have perfunned this protocol on 32 nonsmoking volunteers (18 nonnal subjects and 14 subjects with mild asthma). The subjects with asthma all had positive methacholine challenges with provocative concentra-
Table 1-Changea in SBmD after Atropine Sulfote and after Bronchoacopy* SRaw, L x em H10/Usect Time
Nonnal Subjects
Asthmatic Subjects
Baseline 30 min after atropinet Afterbronchoscopy
4.8± 1.3 3.2± 1.0 3.4±1.0
7.3±4.3 4.2±2.6 6.2±3.1
*Bronchoscopy includes bronchoalveolar lavage and isolated airway segment lavage. tValues shown are means± SO. tAtropine sulfate (1 mg given by intramuscular injection).
tions of methacholine that resulted in a 100 percent increase in baseline SRaw ofless than 1 m~ml. Of the 32 subjects studied, only five subjects (two normal subjects and three subjects with mild asthma) were unable to complete all phases of the protocol (bronchoalveolar lavage and three isolated airway segment lavages). One of the normal subjects and two of the subjects with asthma noted a sensation of shortness of breath with the inflation of the airway catheter's balloons. We deflated the balloons and withdrew the catheter and the bronchoscope and repeated the measurement of SRaw. In all three subjects, the SRaw measured immediately after the catheter removal was within 20 percent of their baseline SRaw measured at the beginning of the study. We were not able to pass the catheter through the vocal cords of the other normal subject who did not complete the protocol. This was not a significant problem in any of the other 31 subjects. The remaining subject with asthma also noted an increase in shortness of breath with the balloon inflation; however, in her case, audible wheezing was also heard. We elected not to obtain a subsequent measurement of SRaw immediately after removing the catheter and bronchoscope but instead treated her with two puffs from an inhaler of albuterol (salbutamol). Within ten minutes, the SRaw had returned to baseline. The changes in SRaw fur the remaining 16 normal subjects and the 11 subjects with mild asthma are shown in 'Thble 1. Except fur some bronchodilation after the atropine sulfate, there was no overall change
in SRaw in either group. In addition, no individual subject had greater than a 45 percent increase in SRaw after the bronchoscopy, when compared to the baseline value. The oximetric measurements during the procedure revealed less than a 2 percent decrease in oxygen saturation during bronchoalveolar lavage but a fall in saturation from 99 percent to 93 to 95 percent during isolated airway segment lavage despite the use of supplemental oxygen. In one subject the saturation fell transiently to 88 percent during isolated airway segment lavage. The temporal course of the decrease in oxygen saturation remained consistentin that it began to fall 1.5 to 2.5 minutes after balloon inflation and reached its nadir at 3.5 to 4.5 minutes after inflation. After balloon deflation the saturation would return to normal during the fOllowing one to two minutes. The volume of fluid recovered in the bronchoalveolar lavage averaged 45 to 67 percent of the fluid introduced. There was no significant difference in the recovery from bronchoalveolar lavage between the normal subjects and the subjects with asthma. The volume recovered in isolated airway segment lavage varied more than fur bronchoalveolar lavage, ranging from 20 percent to close to 100 percent. Viewing through the bronchoscope while isolated segment lavage was perfOrmed, we observed fluid occasionally leaking out of the segment, but we did not see fluid entering into the isolated segment from distal pulmonary units. The cellular compositions fur the bronchoalveolar lavage and the isolated airway segment lavage are given in Table 2. The cellular composition fur only the first or baseline isolated airway segment lavage is shown. The results from bronchoalveolar lavage are somewhat comparable to previously reported values fur normal nonsmoking subjects and subjects with asthma, 1.u although the percentage of macrophages is slightly higher in our subjects, and the percentage oflymphocytes is slightly lower. There were no differences in cellular composition of bronchoalveolar lavage between our two groups of subjects. For the results of isolated airway segment lavage, there was an increase in the percentage of ciliated epithelial cells and neu-
Table 2-Cellular Compoaition tf Bronchoalveolar lAooge and laolated AinDGy Segment lAooge* Bronchoalveolar Lavage
Isolated Airway Segment Lavage
Data
Nonnal Subjects
Asthmatic Subjects
Normal Subjects
Asthmatic Subjects
Macrophages, percent Lymphocytes, percent Neutrophils, percent Eosinophils, percent Ciliated Epithelial Cells, percent
95.4± 1.9 4.6±1.9 0 0 0
95.3±1.9 3.6±1.3 0.4± 1.1 0.6±1.0 0
41.1±25.1 5.3±2.1 3.9±5.8 0 49.4±25.8
37.5±10.2 6.2±3.8 7.0±8.9 0.7±0.8 48.2±16.2
*Values are means± SO. CHEST I 92 I 1 I JULY, 1987
107
trophils when compared to bronchoalveolar lavage. Again, there were no real differences between the two groups of subjects. The approximate total number of cells recovered from bronchoalveolar lavage was 6 to 10 X 1()6 cells and from all three isolated airway segment lavages was 0.2 to 1.0 X 1()6 cells. DISCUSSION
A technique has been developed fur the lavage of an isolated airway segment in human subjects. The technique involves the use of a specially designed doubleballoon catheter that can be placed into the airways under fiberoptic bronchoscopic guidance. The technique is relatively safe, as evidenced by trials in normal subjects and subjects with mild asthma, provided precautions are fOllowed in the selection and monitoring of subjects. Although the technique appears to sample from the airway segment alone, it is possible that "contamination" from areas of the lung outside the segment could occur. We did not observe this during our studies. It is also possible that differences in the material obtained from the isolated airway segment may be inherently different when compared to material from the alveolar spaces (such as the differences in cellular composition noted in our study); but the difference in the airway segment material could also be the result of the presence of the balloons in the airway. Also, since a mild decrease in systemic oxygen saturation during isolated airway segment lavage may be considered a problem in the interpretation of results from future studies, we will be developing a catheter with a fifth channel that opens distally to the distal balloon through which oxygen could be supplied to the occluded lung. Previous attempts at lavaging separate regions of the lung, including limited central-airway lavage, have been reported. Wiggins et al10 tried to separately obtain tracheal secretions, bronchial lavage, and bronchoalveolar lavage. Their technique fur bronchial lavage consisted of the instillation of 5 to 10 ml of sterile physiologic saline solution into either the right or left main-stem bronchus through a bronchoscope and then aspiration of the fluid back through the bronchoscope. Others have previously attempted bronchial lavage by instilling "small quantities" of fluid into the airways using rigid bronchoscopes and catheters. 1u 2 A slightly different approach was tried by Lam et al, 13 who used a specialized irrigation catheter that would allow simultaneous infusion and suctioning of fluid to create a vortex effect around the catheter tip. None of these previous methods attempted to isolate an airway segment. Their usefulness over a more extended period of time of sampling would also be limited. In a more closely related technique, Sielczak et al14 have used a nasotracheal tube in sheep fur creating an isolated 108
tracheal segment that can be lavaged by a catheter affixed to the side of the tube above the inflated cuff; however, this technique would require nasotracheal or endotracheal intubation. Our technique does have potential application fur many areas of pulmonary research. It is currently being used to evaluate the quantity, pattern, and temporal course of mediator release in the airways of normal subjects and subjects with asthma to nonallergic stimuli. It could not only provide infOrmation regarding regional cellular compositional differences but also functional differences in the cells obtained from different regions of the lung. The technique may also be helpful in evaluating other regional differences in lung-produced proteins, immunoglobulins, enzymes, proteases, protease inhibitors, or phospholipids. It could even be useful fur evaluating aerosol deposition to determine central vs more peripheral deposition of different materials. These studies of aerosol deposition could be perfOrmed by giving the subjects inhalations of aerosols with markers (radioactive or otherwise) and then introducing the bronchoscope and sampling catheter to obtain material from the central airways (isolated airway segment lavage) and the peripheral airways (bronchoalveolar lavage). Evaluation of the separate lavage fluids fur the markers could then help determine regional deposition of the aerosol. Finally, response of the lung to inflammatory agents may show regional differences that could be identified by regional sampling. In summary, isolated airway segment lavage can be perfOrmed simply and safely in humans and may have application in pulmonary research in both human and animal models. ACKNOWLEDGMENTS: We thank Ms. Karen Ciffin fur technical assistance and Ms. Rose Welcome fur assistance in the preparation of this report. REFERENCES
1 Reynolds HY, Chretien J. Respiratory tract fluids: analysis of content and contemporary use in understanding lung diseases. DM 1984; 30:6-103 2 Antic R, Macklem PT. The influence of clinical factors on site of airway obstruction in asthma. Am Rev Respir Dis 1976; 114: 851-59 3 Mildon A, Leroux M, Hutcheon M, Zamel N. The site of airways obstruction in exercise-induced asthma. Am Rev Respir Dis 1974; 110:409-14 4 Chan-Yeung M, Abboud R, Tsao MS, Maclean L. Effect of helium on maximal expiratory flow in patients with asthma befOre and during induced bronchoconstriction. Am Rev Respir Dis 1976; 113:433-43 5 Laube BL, Swift DL, Wagner HN, Norman PS, Adams GK. The effect of bronchial obstruction on central airway deposition of a saline aerosol in patients with asthma. Am Rev Respir Dis 1986; 133:740-43 6 Bernstein L, Boushey HA, Chemiack RM, FinkJN, Fulmer JD, Goetze! EJ, et al. Summary and recommendations of a workshop on the investigative use of fiberoptic bronchoscopy on bron-
choalveolar lavage in asthmatics. Am Rev Respir Dis 1985; 132:180-82 7 DuBois AB, Botelho SY, Comroe JH. Anew method for measuring airway resistance in man using a body plethysmograph: values in normal subjects and in patients with respiratory disease. J Clio Invest 1956; 35:327-35 8 Daniele RP, Elias JA, Epstein PE, Rossman MD. Bronchoalveolar lavage: role in the pathogenesis, diagnosis, and management of interstitial lung disease. Ann Intern Med 1985; 102:93-
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9 Metzger WJ, Richerson HB, Worden K, Monick M, Hunninghake GW Bronchoalveolar lavage of allergic asthmatic patients following allergen bronchoprovocation. Chest 1986; 89:477-83 10 Wiggins J, Hill SL, Stockley RA. Lung secretion sol-phase proteins: comparison of sputum with secretions obtained by direct
sampling. Thorax 1983; 38:102-07 11 Falk GA, Okinaka AJ, Siskin GW Immunoglobulins in the bronchial washings of patients with chronic obstructive pulmonary disease. Am Rev Respir Dis 1972; 105:14-21 12 Mandell MA, Dvorak KJ, Worman LW, DeCosse JJ. Immunoglobulin content in bronchial washings of patients with benign and malignant pulmonary disease. N Eng! J Med 1976; 295:694-98 13 LamS, LeRiche J, Chan-Yeung M, Kijek K, Phillips D. Bronchial lavage: comparison of two methods (abstract). Am Rev Respir Dis 1985; 131:A9 14 Sielczak MW, Denas SM, Abraham WM. Airway cell changes in tracheal lavage of sheep after ozone exposure. J Toxicol Environ Health 1983; 11:545-53
Doppler Echocardiography Seminar The Advanced Echocardiography Seminar, sponsored by the Center for Medical Ultrasound, Bowman Gray School of Medicine, will be held September 10-12 at the Innisbrook Resort Conference Center, Thrpon Springs, Florida. For information, contact the Registrar, Ultrasound Center, Bowman Gray School of Medicine, 300 South Hawthorne Road, Winston-Salem, NC 27103 (919:748-4505).
CHEST I 92 I 1 I JULY, 1987
109
A novel technique has been developed for the isolation and lavaging of a central airway segment in humans. The procedure uses a double-balloon airway-sampling catheter that is placed into the airways with 6beroptic bronchoscopic guidance. A protocol using this technique was attempted in 32 subjects (18 normal subjects apd 14 subjects with mild asthma), and 27 of the subjects were able to complete all phases of the protocol, in<:luding bronchoalveolar lavage
and lavage of an isolated airway segment. There was no significant complication in any subject. Cellular composition of the lavages demonstrated differences in cell differential and total cell counts for the two different sampled regions (central airway vs alveolar spaces~ We believe that this technique of lavage of an isolated airway segment is safe and has potential usefulness in many areas of pulmonary research.
from the airways alone would be important Sampling in many areas of pulmonary research. Currently,
subjects with asthma at baseline conditions and after a local challenge.
bronchoalveolar lavage is used to obtain material from the lung for cellular, protein, and mediator analyses; however, it is believed that the material sampled in bronchoalveolar lavage comes predominantly from the alveolar spaces. 1 The ability to sample directly from the airways alone would allow the recovery of material that would have particular relevance in certain diseases such as asthma. Although narrowing of the airways in asthma occurs throughout the tracheobronchial tree, many authors believe that the more central airways are more predominantly involved in the acute bronchoconstriction to both allergic and nonallergic stimuli. u Thus, obtaining material from the more central airways, rather than the alveoli, appears to be more relevant in identifying mediators and cells that are involved in the acute bronchoconstrictive response to stimuli in asthma. The ability to differentiate central from peripheral material may also be important in other research such as surfactant, protease inhibitors, or other pulmonary protein regional differences, or in studying aerosol deposition or pulmonary injury due to inhaled agents. With these needs in mind, we have developed a technique that allows the isolation and lavage of an upper airway segment in human subjects with minimal contamination from distal pulmonary units. This lavage of an isolated airway segment involves the use of a double-balloon catheter that can be placed into the airways using fiberoptic bronchoscopy fur guidance. Initially, we have used this technique for evaluating the fluid obtained from the airways of normal subjects and *From the Division of Pulmonary and Critical Care Medicine, University of Michigan Medical Center, Ann Arbor. Manuscript received October 6; revision accepted December 6. Reprint requests: Dr. Eschenbacher, BIH272, Universify Hospital, 1500 East Medical Center Drive, Ann Arbor 48104
MATERIALS AND METHODS
Airway-Sampling Catheter Based upon the design criteria of one of the authors (W L. E.), a commercial manufacturer (American Edwards Corp.) produced the catheters that were used in the development of this technique. The catheters are modifications of the pulmonary arterial catheter that is used fur hemodynamic evaluation in the pulmonary circulation. Each catheter made fur airway sampling was llO em in length and 2 mm in diameter and was made of plasticized polyvinylchloride. 1Wo latex balloons are located at the distal end of the catheter separated by 2 to 4 em (depending upon the particular design). Each balloon can be in8ated with up to 2.5 ml of air or 8uid (other models have been made with balloons with 1.5-ml capacity). In the interballoon segment, several ports are present that open to one channel fur infusion and to another channel fur withdrawal of 8uid. Thus, there are four channels throughout the length of the catheter: two fur the in8ations of the balloons and two fur the introduction and withdrawal of 8uid. A diagram of the catheter and its position when placed into the lungs is shown in Figure 1.
Protocol The placement of the airway catheter into the subjects was perfOrmed while observing the precautions outlined by the National Heart, Lung, and Blood Institutes Workshop on Fiberoptic Bronchoscopy and Bronchoalveolar Lavage in Asthmatics. • Each subject signed a consent form that had been approved by the human research committee at the University of Michigan Medical Center. The subjects with asthma were considered to have mild asthma and could stop their inhaled and oral medication fur 24 hours befOre the study. No subject had an upper respiratory infection within fuur weeks befOre the study. The subjects were carefully monitored throughout the protocol and fur several hours after the protocol. On the day of the study, we first determined baseline specific airway resistance (SRaw) fur each subject using a constant-volume whole-body plethysmograph (Airco 3000 System; Ohio Medical Products). For these measurements, we used the technique described by DuBois et al.1 We then gave each subject 1 mg of atropine sulfate by intramuscular injection 30 minutes before the placement of the bronchoscope or catheter into the airways. During this 30minute period, we gave routine nasal, pharyngeal, laryngeal, and CHEST I 92 I 1 I JULY, 1987
105
FIGURE 1. Double-balloon airway-sampling catheter and its position in left main-stem bronchus during isolated airway segment lavage. Bronchoalveolar lavage (BAL) is perfOrmed in right middle lobe. upper-airway anesthesia with topical solutions of 2 percent and 4 percent lidocaine. At the end of the 30 minutes, we repeated the measurement of SRaw. We then placed the subject in a supine position, attached a pulse-oximeter (N-100 Pulse Oximeter; Nellcor Inc) to their index finger, gave supplemental oxygen by mouth (4 to 5 Umin), and inserted a fiberoptic bronchoscope (Olympus BF-2TR), through one naris, past the vocal cords, and into the airways. We used some additional topical lidocaine within the airways and wedged the bronchoscope into a subsegmental orifice in the right middle lobe. We then perfOrmed bronchoalveolar lavage in a standard fashion 1 using three to five 30-ml aliquots of physiologic saline solution. After the bronchoalveolar lavage, we withdrew the bronchoscope back above the vocal cords. At this time, we inserted the airwaysampling catheter into the other naris and advanced it past the vocal cords using the bronchoscope for visual guidance. We positioned the distal end of the catheter into the left main-stem bronchus so that the distal balloon was proximal to the bifurcation of the left upper and left lower lobes. The proximal balloon would then be di$tal to the main carina in the left main-stem bronchus. Again, the position of the catheter in place is shown in Figure 1. We then inflated the distal balloon and checked the adequacy of the seal visually through the bronchoscope. We then inflated the proximal balloon, creating an isolated airway segment. Next we lavaged the isolated segment with different fluids. By calculation of the potential volume of the isolated airway segment (using standard anatomic bronchial dimensions and the length of the interballoon segment) and from experience in perfOrming these lavages of isolated segments, we found that 2-ml aliquots of fluid would almost completely fill the airway segment without spilling outside the segment. We introduced 2-ml aliquots of physiologic saline solution into the segment, checking again for the adequacy of the balloon seals both proximally and distally, and then withdrew the saline solution through the other channel using a 3-ml syringe and hand suction only. We recorded the recovery from each separate aliquot introduced but combined all the aliquots from a
108
particular series into a common container. In all studies, we first perfOrmed four to ten lavages with 2-ml aliquots of physiologic saline for baseline values. We would then use one of two different fluids for the second series of lavages, either distilled water or hypertonic sodium chloride solution (3.6 percent sodium chloride or approximately 900/mOsm). Again, we used fOur to ten lavages with 2-ml aliquots for this second series. Finally, for the third series, we again used four to ten 2-ml lavages of physiologic saline solution. After completion of this final series oflavages, we deBated the balloons and removed the catheter and the bronchoscope. We then repeated the measurement of SRaw for the subject. Subjects were then followed for the next two to fOur hours for symptoms, but no further measurements of SRaw were made. On the average, it took three to five mintues for the insertion ofthe bronchoscope, five to eight minutes for the bronchoalveolar lavage, five to ten minutes for the insertion of the catheter, and three to five minutes for each lavage of the isolated airway segment. Overall, the entire procedure after anesthesia lasted 30 to 45 mintues. The fluid obtained from the bronchoalveolar lavage and the isolated airway segment lavages was then centrifuged at 1,000 g for five minutes. The supernatant was stored at -70"C for future mediator analyses. The cell pellets were resuspended in Hanks balanced salt solution and were washed twice. A portion of each suspension was then spun in a cytocentrifuge (Cytospin 2; Shandon Southern Instruments) and stained using Papanicolaou$ stain for differential cell count. Three hundred cells were counted for each sample. Another portion of the cell suspension was used for total cell count using a hemacytometer.
REsur.:rs We have perfunned this protocol on 32 nonsmoking volunteers (18 nonnal subjects and 14 subjects with mild asthma). The subjects with asthma all had positive methacholine challenges with provocative concentra-
Table 1-Changea in SBmD after Atropine Sulfote and after Bronchoacopy* SRaw, L x em H10/Usect Time
Nonnal Subjects
Asthmatic Subjects
Baseline 30 min after atropinet Afterbronchoscopy
4.8± 1.3 3.2± 1.0 3.4±1.0
7.3±4.3 4.2±2.6 6.2±3.1
*Bronchoscopy includes bronchoalveolar lavage and isolated airway segment lavage. tValues shown are means± SO. tAtropine sulfate (1 mg given by intramuscular injection).
tions of methacholine that resulted in a 100 percent increase in baseline SRaw ofless than 1 m~ml. Of the 32 subjects studied, only five subjects (two normal subjects and three subjects with mild asthma) were unable to complete all phases of the protocol (bronchoalveolar lavage and three isolated airway segment lavages). One of the normal subjects and two of the subjects with asthma noted a sensation of shortness of breath with the inflation of the airway catheter's balloons. We deflated the balloons and withdrew the catheter and the bronchoscope and repeated the measurement of SRaw. In all three subjects, the SRaw measured immediately after the catheter removal was within 20 percent of their baseline SRaw measured at the beginning of the study. We were not able to pass the catheter through the vocal cords of the other normal subject who did not complete the protocol. This was not a significant problem in any of the other 31 subjects. The remaining subject with asthma also noted an increase in shortness of breath with the balloon inflation; however, in her case, audible wheezing was also heard. We elected not to obtain a subsequent measurement of SRaw immediately after removing the catheter and bronchoscope but instead treated her with two puffs from an inhaler of albuterol (salbutamol). Within ten minutes, the SRaw had returned to baseline. The changes in SRaw fur the remaining 16 normal subjects and the 11 subjects with mild asthma are shown in 'Thble 1. Except fur some bronchodilation after the atropine sulfate, there was no overall change
in SRaw in either group. In addition, no individual subject had greater than a 45 percent increase in SRaw after the bronchoscopy, when compared to the baseline value. The oximetric measurements during the procedure revealed less than a 2 percent decrease in oxygen saturation during bronchoalveolar lavage but a fall in saturation from 99 percent to 93 to 95 percent during isolated airway segment lavage despite the use of supplemental oxygen. In one subject the saturation fell transiently to 88 percent during isolated airway segment lavage. The temporal course of the decrease in oxygen saturation remained consistentin that it began to fall 1.5 to 2.5 minutes after balloon inflation and reached its nadir at 3.5 to 4.5 minutes after inflation. After balloon deflation the saturation would return to normal during the fOllowing one to two minutes. The volume of fluid recovered in the bronchoalveolar lavage averaged 45 to 67 percent of the fluid introduced. There was no significant difference in the recovery from bronchoalveolar lavage between the normal subjects and the subjects with asthma. The volume recovered in isolated airway segment lavage varied more than fur bronchoalveolar lavage, ranging from 20 percent to close to 100 percent. Viewing through the bronchoscope while isolated segment lavage was perfOrmed, we observed fluid occasionally leaking out of the segment, but we did not see fluid entering into the isolated segment from distal pulmonary units. The cellular compositions fur the bronchoalveolar lavage and the isolated airway segment lavage are given in Table 2. The cellular composition fur only the first or baseline isolated airway segment lavage is shown. The results from bronchoalveolar lavage are somewhat comparable to previously reported values fur normal nonsmoking subjects and subjects with asthma, 1.u although the percentage of macrophages is slightly higher in our subjects, and the percentage oflymphocytes is slightly lower. There were no differences in cellular composition of bronchoalveolar lavage between our two groups of subjects. For the results of isolated airway segment lavage, there was an increase in the percentage of ciliated epithelial cells and neu-
Table 2-Cellular Compoaition tf Bronchoalveolar lAooge and laolated AinDGy Segment lAooge* Bronchoalveolar Lavage
Isolated Airway Segment Lavage
Data
Nonnal Subjects
Asthmatic Subjects
Normal Subjects
Asthmatic Subjects
Macrophages, percent Lymphocytes, percent Neutrophils, percent Eosinophils, percent Ciliated Epithelial Cells, percent
95.4± 1.9 4.6±1.9 0 0 0
95.3±1.9 3.6±1.3 0.4± 1.1 0.6±1.0 0
41.1±25.1 5.3±2.1 3.9±5.8 0 49.4±25.8
37.5±10.2 6.2±3.8 7.0±8.9 0.7±0.8 48.2±16.2
*Values are means± SO. CHEST I 92 I 1 I JULY, 1987
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trophils when compared to bronchoalveolar lavage. Again, there were no real differences between the two groups of subjects. The approximate total number of cells recovered from bronchoalveolar lavage was 6 to 10 X 1()6 cells and from all three isolated airway segment lavages was 0.2 to 1.0 X 1()6 cells. DISCUSSION
A technique has been developed fur the lavage of an isolated airway segment in human subjects. The technique involves the use of a specially designed doubleballoon catheter that can be placed into the airways under fiberoptic bronchoscopic guidance. The technique is relatively safe, as evidenced by trials in normal subjects and subjects with mild asthma, provided precautions are fOllowed in the selection and monitoring of subjects. Although the technique appears to sample from the airway segment alone, it is possible that "contamination" from areas of the lung outside the segment could occur. We did not observe this during our studies. It is also possible that differences in the material obtained from the isolated airway segment may be inherently different when compared to material from the alveolar spaces (such as the differences in cellular composition noted in our study); but the difference in the airway segment material could also be the result of the presence of the balloons in the airway. Also, since a mild decrease in systemic oxygen saturation during isolated airway segment lavage may be considered a problem in the interpretation of results from future studies, we will be developing a catheter with a fifth channel that opens distally to the distal balloon through which oxygen could be supplied to the occluded lung. Previous attempts at lavaging separate regions of the lung, including limited central-airway lavage, have been reported. Wiggins et al10 tried to separately obtain tracheal secretions, bronchial lavage, and bronchoalveolar lavage. Their technique fur bronchial lavage consisted of the instillation of 5 to 10 ml of sterile physiologic saline solution into either the right or left main-stem bronchus through a bronchoscope and then aspiration of the fluid back through the bronchoscope. Others have previously attempted bronchial lavage by instilling "small quantities" of fluid into the airways using rigid bronchoscopes and catheters. 1u 2 A slightly different approach was tried by Lam et al, 13 who used a specialized irrigation catheter that would allow simultaneous infusion and suctioning of fluid to create a vortex effect around the catheter tip. None of these previous methods attempted to isolate an airway segment. Their usefulness over a more extended period of time of sampling would also be limited. In a more closely related technique, Sielczak et al14 have used a nasotracheal tube in sheep fur creating an isolated 108
tracheal segment that can be lavaged by a catheter affixed to the side of the tube above the inflated cuff; however, this technique would require nasotracheal or endotracheal intubation. Our technique does have potential application fur many areas of pulmonary research. It is currently being used to evaluate the quantity, pattern, and temporal course of mediator release in the airways of normal subjects and subjects with asthma to nonallergic stimuli. It could not only provide infOrmation regarding regional cellular compositional differences but also functional differences in the cells obtained from different regions of the lung. The technique may also be helpful in evaluating other regional differences in lung-produced proteins, immunoglobulins, enzymes, proteases, protease inhibitors, or phospholipids. It could even be useful fur evaluating aerosol deposition to determine central vs more peripheral deposition of different materials. These studies of aerosol deposition could be perfOrmed by giving the subjects inhalations of aerosols with markers (radioactive or otherwise) and then introducing the bronchoscope and sampling catheter to obtain material from the central airways (isolated airway segment lavage) and the peripheral airways (bronchoalveolar lavage). Evaluation of the separate lavage fluids fur the markers could then help determine regional deposition of the aerosol. Finally, response of the lung to inflammatory agents may show regional differences that could be identified by regional sampling. In summary, isolated airway segment lavage can be perfOrmed simply and safely in humans and may have application in pulmonary research in both human and animal models. ACKNOWLEDGMENTS: We thank Ms. Karen Ciffin fur technical assistance and Ms. Rose Welcome fur assistance in the preparation of this report. REFERENCES
1 Reynolds HY, Chretien J. Respiratory tract fluids: analysis of content and contemporary use in understanding lung diseases. DM 1984; 30:6-103 2 Antic R, Macklem PT. The influence of clinical factors on site of airway obstruction in asthma. Am Rev Respir Dis 1976; 114: 851-59 3 Mildon A, Leroux M, Hutcheon M, Zamel N. The site of airways obstruction in exercise-induced asthma. Am Rev Respir Dis 1974; 110:409-14 4 Chan-Yeung M, Abboud R, Tsao MS, Maclean L. Effect of helium on maximal expiratory flow in patients with asthma befOre and during induced bronchoconstriction. Am Rev Respir Dis 1976; 113:433-43 5 Laube BL, Swift DL, Wagner HN, Norman PS, Adams GK. The effect of bronchial obstruction on central airway deposition of a saline aerosol in patients with asthma. Am Rev Respir Dis 1986; 133:740-43 6 Bernstein L, Boushey HA, Chemiack RM, FinkJN, Fulmer JD, Goetze! EJ, et al. Summary and recommendations of a workshop on the investigative use of fiberoptic bronchoscopy on bron-
choalveolar lavage in asthmatics. Am Rev Respir Dis 1985; 132:180-82 7 DuBois AB, Botelho SY, Comroe JH. Anew method for measuring airway resistance in man using a body plethysmograph: values in normal subjects and in patients with respiratory disease. J Clio Invest 1956; 35:327-35 8 Daniele RP, Elias JA, Epstein PE, Rossman MD. Bronchoalveolar lavage: role in the pathogenesis, diagnosis, and management of interstitial lung disease. Ann Intern Med 1985; 102:93-
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9 Metzger WJ, Richerson HB, Worden K, Monick M, Hunninghake GW Bronchoalveolar lavage of allergic asthmatic patients following allergen bronchoprovocation. Chest 1986; 89:477-83 10 Wiggins J, Hill SL, Stockley RA. Lung secretion sol-phase proteins: comparison of sputum with secretions obtained by direct
sampling. Thorax 1983; 38:102-07 11 Falk GA, Okinaka AJ, Siskin GW Immunoglobulins in the bronchial washings of patients with chronic obstructive pulmonary disease. Am Rev Respir Dis 1972; 105:14-21 12 Mandell MA, Dvorak KJ, Worman LW, DeCosse JJ. Immunoglobulin content in bronchial washings of patients with benign and malignant pulmonary disease. N Eng! J Med 1976; 295:694-98 13 LamS, LeRiche J, Chan-Yeung M, Kijek K, Phillips D. Bronchial lavage: comparison of two methods (abstract). Am Rev Respir Dis 1985; 131:A9 14 Sielczak MW, Denas SM, Abraham WM. Airway cell changes in tracheal lavage of sheep after ozone exposure. J Toxicol Environ Health 1983; 11:545-53
Doppler Echocardiography Seminar The Advanced Echocardiography Seminar, sponsored by the Center for Medical Ultrasound, Bowman Gray School of Medicine, will be held September 10-12 at the Innisbrook Resort Conference Center, Thrpon Springs, Florida. For information, contact the Registrar, Ultrasound Center, Bowman Gray School of Medicine, 300 South Hawthorne Road, Winston-Salem, NC 27103 (919:748-4505).
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