Bronchial Responsiveness Is Not Increased by Bronchoalveolar and Bronchial Lavage Performed after Allergen Challenge 1- 3
PIERO GIANIORIO, MARCO BONAVIA, EMANUELE CRIMI, SABINA LANTERO, PAOLO CRIMI, GIOVANNI A. ROSSI, and VITO BRUSASCO
Introduction
Bronchoalveolar lavage has been increasingly used for studying the inflammatory cells and mediators present in the asthmatic lung both during baseline conditions (1-3) and after inhalation challenges with allergens(4, 5) or occupational agents (6, 7). A limitation to the use of bronchoalveolar lavage in asthmatic patients is represented by the risk that this procedure may lead to deterioration of lung function and increase bronchial responsiveness. During baseline conditions, bronchial responsiveness to methacholine was found to be unchanged after bronchoalveolar lavage in patients with mild airway hyperresponsiveness (8), but it was increased in patients with a high initial hyperresponsiveness (9). After allergen inhalation, bronchial responsiveness may be increased in some asthmatic patients (10). Whether bronchoalveolar lavage performed after an allergen inhalation challenge may induce or aggravate airway hyperresponsiveness has not been evaluated. The present study was designed to investigate if in allergicpatients with different patterns of response to inhaled allergen a fiberoptic bronchoscopy with bronchoalveolar and bronchial lavage performed 4 or 24 h after allergen bronchial challenge may deteriorate lung function and increase bronchial responsiveness. Methods Patients Twenty-three outpatients, whose characteristics are summarized in table 1, were studied. They had a history of perennial rhinitis and/or bronchial asthma with allergic sensitization to Dermatophagoidespteronyssinus as documented by skin prick test and radio-allergosorbent test. At the time of study all patients were asymptomatic, with a FEV 1 larger than 70070 of predicted. None of the patients were receiving antiasthmatic treatment other than
SUMMARY Nonspecific bronchial responsiveness was studied In 23 allergic patients with a history of rhinitis and/or bronchial asthma who underwent flberoptlc bronchoscopy with bronchoalvaolar and bronchial lavage (BAI.-BL) 4 h (Group A) or 24 h (Group B) after an allergen Inhalation challenge. In all patients, the dose of methacholine causing an FEY1 filII of 15% (P01 5) was determined at baseline, 24 h before allergen challenge. Methacholine bronchial challenge wes repeated 1 h before BAI.BL In patients of both groups and again 12 to 14 h after BAI.-BL In Group A and 24 h after BAI.-BL in Group B. In patients of Group A, the values of methacholine POu after BAL-BL were not significantly different from those determined before BAI.-BL. This wes also the case In patients In whom bronchial responsiveness weslncreased after allergen challenge. In patients of Group B, methacholine POlS was significantly decreased after allergen challenge, and this decrease was correlated with the occurrence and the severity of the late asthmatic reaction. Evan In patients who showed dual asthmatic reactions and an Increased responsiveness after allergen challenge, methacholine P0 15 was not further decreased after BAL-BL. These dlta support the safety of a procedure combinIng bronchial allergen challenge with BAL-BL, which can be used for studies on the p~thophyslology of bronchial asthma. AM REV RESPIR DIS 1991; 143:105-108
13rstimulants.These, if any,werediscontinued at least 12 h before entering the study.
trol between 3 and 8 h after the end of inhalation challenge.
Bronchial Challenges Methacholine aerosols were delivered by an ampul-dosimeter device (MEFAR, Brescia, Italy) (11). The challenge was started from a dose of 0.02 mg. The same ampul was used for each patient. The best of three FEV 1 maneuvers measured immediately after inhalation of each methacholine dose was taken to construct dose-response curves. The dose of methacholine wastwofold-incremented until FEV 1 was decreased below 80070 of control (inhalation of saline), or up to a maximal dose of 5 mg. The dose causing a 15070 fall of FEV 1 (PD 1s ) was calculated by interpolation of the dose-response curves. For allergen bronchial challenge, solutions of Dermatophagoides pteronyssinus reconstituted from predosed (arbitrary units, AU) dried allergen (Pharmacia, Uppsala, Sweden) were used. FEV 1 was measured 15 min after inhalation of allergen solutions. After control inhalation of saline, the allergen bronchial challenge was started from a dose of 4 AU, with twofold increments until the FEV 1 fell below 80070 of control values, or up to a maximal dose of 500 AU. Thereafter, FEV 1 was monitored every hour for 8 h to detect the occurrence of late asthmatic reactions, defined as an FEV 1 fall exceeding 15070 of con-
Bronchoalveolar and Bronchial Lavage Patients weregiven atropine (0.5 mg) and diazepam (10mg) intramuscularly 5 min before the introduction of the bronchofiberoscope (Olympus BF, type PlO; Olympus Corp. of America, New Hyde Park, NY). The bronchoscope was passed through the nose after local anesthesia of the nostrils (lidocaine solution 2070 and adrenaline solution 0.1/1000, 1 ml each side). Bronchoalveolar lavage was performed by injecting into the middle lobe
(Received in original form February 23, 1990 and in revised form July 13, 1990) 1 From the Istituto di Medicina delloSport, Cattedra di Fisiopatologia Respiratoria, and the Instituto di Igiene e Medicina Preventiva, Cattedra di Medicina di Comunita, Universita di Genova, and the Prima Divisione di Pneumologia, Ospedale S. Martino, Genova, Italy. 1 Supported in part by a grant from The Italian Ministry of Public Education by Grant No. 87.00560.56 from the Consiglio Nazionale delle Richerche. 3 Correspondence and requests for reprints should be addressed to Vito Brusasco, M.D., Istituto di Medicina dello Sport, Viale Benedetto XV, 10. 16132, Genova, Italy.
105
GIANIORIO, BONAVIA, CRIMI, LANTERO, CRIMI, ROSSI, AND BRUSASCO
106
the end of the bronchoscopic procedures, with a contact telephone number that they could call at any time.
TABLE 1 PATIENTS' CHARACTERISTICS, BRONCHODILATOR USAGE, AND ADVERSE EFFECTS OF BAL-BL Bronchodilator Patient No. Group A 1 2 3 4 5 6 7 8 9 Mean SO Group B 10 11 12 13 14 15 16 17 18 ,19 20 21 22 23 Mean SO
Age (yr)
25 19 24 18 18 19 19 19 27 21 3 28 19 26 26 19 38 17 18 17 21 18 19 21 20 22 6
Sex
Post-BPT
Post-BAL-BL
Adverse Effects
79 100 107 98 100 98 76 95 111 96 12
No No No No No No No No No
No No No No Yes Yes Yes No No
None None None None None None Fever, malaise None None
89 111 99 98 129 74 109 98
Yes Yes Yes No No Yes No Yes Yes No Yes Yes Yes Yes
No No Yes No No Yes No No Yes No No Yes No Yes
None Sore throat None None None None None None None None None None None Cough
Baseline FEV 1 (% pred)
M
M M M M M M M M
M
F M M M M M M M M M M M M
Definition of abbreviations: BPT
n
106 108 113 103 109 101 14
= allergen bronchial provocation
test; BAL-BL
Statistical Analysis The values of methacholine PD 15 were transformed in logarithmic form for statistical analysis and reported as geometric mean. In order to control the per-experiment error rate, the significance of changes among observation times was assessed by F statistics. If p values from this preliminary analysis were less than 0.05, separate paired t tests were performed. Pearson's correlation coefficient was also used; p values less than 5010 were considered to be statistically significant. Results
= bronchoalveolar
and bronchial lavage.
bronchus 100 ml of sterile saline, fractionated in five aliquots of 20 ml each, which were aspirated at a negative pressure of 50 to 120 mm Hg. Bronchial lavage was performed by injecting into the orifice of the lower lobe bronchus 40 ml of saline, fractionated in two aliquots of 20 ml each, which were immediately suctioned at a pressure of about 50 to 120 mm Hg.
Experimental Protocols All patients underwent a methacholine bronchial challenge in the morning of the first day of study, and an allergen challenge in the morning of the following day. Bronchoalveolar and bronchial lavage (BAL-BL) was performed 4 h after the end of the allergen inhalation challenge in nine patients (Group A), and 24 h after the allergen challenge in the remaining 14patients (Group B). Methacholine challenge was repeated 1 h before and again 12 to 14 h after BAL-BL in Group A, and 1 h before and again 24 h after BAL-BL in Group B. A synopsis of the experimental protocols used in the two groups is given in table 2. The time intervals between procedures was chosen so that methacholine bronchial challenge was always done at the same time in the morning. At the end of the methacholine challenge performed 1 h before BAL-BL, patients inhaled 200 ug salbutamol to reverse the induced bronchospasm. A single dose of
salbutamol (200 ug by metered aerosol) was also given to the patients who experienced symptoms of asthma after allergen bronchial challenge or BAL-BL (table 1). Patients were allowed to inhale additional doses of salbutamol if nocturnal symptoms of asthma developed. Patient 6 showed a severelate asthmatic reaction after bronchial challenge and was given steroid intramuscularly (bethamethasone 4 mg) immediately before BAL-BL. All patients left the hospital 8 h after the end of the bronchial challenge or 1 h after
TABLE 2 SYNOPSIS OF EXPERIMENTAL PROTOCOLS Time between Procedures
Procedures
Group A
Group B
(h)
(h)
24
24
3
24
12-14
24
Methacholine challenge BPT Methacholine challenge BAL-BL Methacholine challenge For definition of abbreviations, see table 1.
Group A The results of allergen inhalation challenge are given in table 3, and changes of methacholine sensitivity and baseline FEV 1 after allergen challenge and BALBL are shown in figure 1. Methacholine PD 15 (geometric mean) was 0.214mg before allergen challenge, 0.170 mg 3 h after allergen challenge, and 0.322 mg 12 to 14 h after BAL-BL. The corresponding baseline values of FEV 1 (mean ± SD) were 4.12 ± 0.55 L, 4.05 ± 0.58 L, and 3.98 ± 0.71 L, respectively. None of these differences was statistically significant. No significant correlations resulted between changes in methacholine PD 1 5 after BAL-BL and either baseline or prelavage methacholine PD 15 • There was a wide variability of changes in PD 15 after allergen challenge, which was likely due to different inflammatory responses of individual patients. In four patients, methacholine PD 15 was reduced by 500/0 or more after allergen challenge. One of these patients (Patient 7) took several doses of salbutamol overnight after BALBL. This patient was the only one in whom PD 15 to methacholine was found to be further decreased after BAL-BL compared with the postchallenge value. Group B The results of allergen inhalation challenge are given in table 3, and changes of methacholine sensitivity and baseline FEV 1 after allergen challenge and BALBL are shown in figure 2. 1\venty-four hours after allergen challenge, both methacholine PD 15 and baseline FEV 1 were reduced from 0.136mg to 0.096 mg (p = 0.(05) and from 4.39 ± 0.85 to 3.89 ± 0.94 L (p = 0.0008), respectively. The changes in methacholine PD 15 and baseline FEV 1 were significantly correlated (r = 0.58, p = 0.03 and r = 0.74, p = 0.0025, respectively) with the late maxi-
107
BRONCHOALVEOLAR LAVAGE AND ALLERGEN CHALLENGE IN ASTHMA
TABLE 3 RESULTS OF BASELINE METHACHOLINE CHALLENGE ANO ALLERGEN CHALLENGE
Patient No.
Methacholine POlS (mg)
Group A 1 2 3 4 5 6 7 8 9 Mean SO Group B 10 11 12 13 14 15 16 17 18 19 20 21 22 23 Mean SO
Maximal FEVl Fall EAR· (% of control)
31 200 2,016 337 61 105 205 96 1,760 214 (geometric)
14 21 12 8 27 22 44 23 11 20 11
15 131 24 345 5,000 34 693 70 31 5,000 104 55 43 137 135 (geometric)
32 37 24 15 5 20 36 28 24 14 30 29 33 20 24 9
Maximal FEV1 Fall LARt (% of control)
30 25 17
6 2 53
3 31 30
o 30
2 31
2 18 16
• Maximal fall of FEV, within 1 h after allergen inhalation challenge. Maximum fall of FEV, between 3 and 8 h after allergen inhalation challenge.
t
mal fall of FEV 1. This indicates that the significant changes observed after allergen challenge were due to eight patients who developed a late asthmatic response. Twenty-four hours after BAL-BL, methacholine PD 15 was 0.152 mg and baseline FEV 1 was 4.19 ± 0.94 L. These values were significantly higher than after allergen challenge (p = 0.04 and p = 0.006, respectively); however, FEV 1 values were still significantly lower than before allergen challenge (p = 0.004). There wereno significant correlations be-
tween changes in methacholine PD 15 after BAL-BL and either baseline or prelavagemethacholine PD 1S • Three patients (Patients 15, 18, and 23) took overnight doses of salbutamol both after bronchial challenge and after BAL-BL, one patient (Patient 21) used salbutamol only after bronchial challenge. Discussion
In this study, we found that BAL-BL performed 4 or 24 h after allergen bronchial challenge neither deteriorated lung func-
Group A.
Group A.
•
=== ;:::::
4
§ 2
0..1...---
8.for. BPT
Aft... BAL-8L Aft.r BPT
_
8.ro ... 8PT
Aft... 8AL-BL Aft.,. BPT
Fig. 1. ~hanges in bronchial sensitivity (PD,s) to methacholine and baseline FE~ 3 h after allergen bronchial provocation test (BPT)and 12to 14 h after bronchoalveolar and bronchial lavage (BAL-BL). None of the differences was statistically significant.
tion nor increased airway responsiveness even in patients in whom airway responsiveness was increased as a consequence of the previous allergen inhalation. In primates, it has been reported that bronchoalveolar lavage may elicit an inflammatory response in the lung (12). Airway inflammation is a key feature of bronchial asthma and is thought to be a main mechanism in the pathogenesis of bronchial hyperreactivity (13).Therefore, although in normal humans and in nonasthmatic patients (9), bronchoalveolar lavage does not appear to cause an increase in bronchial reactivity to methacholine, one may expect that this procedure in asthmatic patients may produce or worsen airway hyperresponsiveness. However, Kirby and coworkers (8) found no changes in bronchial sensitivity to methacholine measured 24 h after BAL in patients with mild asthma. Bycontrast, in a group of asthmatic patients with various degrees of airway responsiveness, Kelly and colleagues (9) found that sensitivity to methacholine was increased 24 h after BAL, and this increase was positively correlated with the baseline responsiveness. Experimental exposure to allergen has been shown to cause an increase of nonspecific bronchial responsiveness that, in patients with dual asthmatic reaction, may persist for several days (10). On the basis of these data, one might expect that if bronchoalveolar lavage is performed after allergen inhalation challenge any allergen-induced bronchial hyperresponsiveness might be eventually enhanced. The only study in which the effect of the combined procedure of bronchial provocation and BAL was considered is that of Metzger and coworkers (14). They found a marginal deterioration of lung function after locally instilled allergen followed by BAL. In the present study, we found no evidence of increased methacholine sensitivity after BAL-BL performed either early (Group A) or late (Group B) after allergen bronchial challenge. This was also true in patients with an elevated degree of baseline or prelavage bronchial responsiveness. This finding does not confirm what was previously reported by Kelly and colleagues (9). More importantly, we also found that BAL-BL did not further enhance bronchial responsiveness when this had been previously increased as a result of an allergen inhalation challenge. In a fewpatients, symptoms of asthma occurred early after BAL-BL, but they werein most cases easily controlled by a single dose of f3r
108
GIANIORIO, BONAVIA, CRIMI, LANTERO, CRIMI, ROSSI, AND BRUSASCO
Group B
Group B 10
..
---
~-~ - --
" - -::= ~-I= ::,=/~---...
.01
~
.001
Before BPT
Afte,. BAL-BL After aPT
~
2
~
0.1...-.-----------B.fo,.e BPT
Afte,. BAL-BL After aPT
Fig. 2. Changes in POts to methacholine and baseline FE'/, 24 h after BPT and 24 h after BAL. Continuous lines indicate patients with late asthmatic reaction, and broken lines indicate patients with isolated early asthmatic reaction or without asthmatic reaction. POtsafter BPT versus POtsbefore BPT: p = 0.005; POts after BAL versus POts after BPT: p = 0.04. FE'/, after BPT versus FE'/, before BPT: p = 0.0008; FE'/, after BAL versus FE'/, after BPT: p = 0.006; FE'/, after SAL versus FE'/, before BPT: p = 0.004.
stimulant and did not result in a sustained increase of bronchial responsiveness. We cannot know which patients of Group A would have experienced a late asthmatic response to allergen since the medications givenbefore and after BALBL may have prevented it. However, we can reasonably think that the four patients in whom airwayresponsiveness was increased after the allergen challenge were likely to develop a late asthmatic reaction. In three of these patients, the increase in airway responsiveness observed after allergen challenge was reversed after BAL-BL. In this group, methacholine sensitivity was determined 12to 14h after BAL-BL;therefore, it cannot be excluded that the tendency for methacholine sensitivity to be decreased after BAL-BL was in part due to a persistent effect of the atropine given intramuscularly as a premedication for bronchoscopy. Even though premedication drugs might have somehow affected the results in this group, the lack of worsening of airway responsiveness after BAL-BL performed 4 h after allergen inhalation challenge seems of practical interest. In patients of Group B, we found that methacholine sensitivity was increased after allergen challenge and, consistent with a previous report (10), this increase was positively correlated with the severity of the late asthmatic reaction. Cartier and coworkers (10) found that allergeninduced airway hyperresponsiveness in most patients with definite late reactions lasted 48 h or more. In our study, an increased methacholine sensitivity was observed 24 h after allergen challenge in
four of eight patients who developed a late asthmatic reaction, but this change was completely reversed24 h after BALBL. In these patients, thus, allergen-induced hyperresponsiveness apparently lasted less than 48 h. We do not know the reason for this finding, whichapparently contrasts with those of Cartier and coworkers. It may be possible that antigenic stimulation was less strong in our patients, but we cannot exclude that premedication given for bronchoscopy may have interrupted the pathophysiologic events that lead to persistent increase in bronchial responsiveness after exposure to allergen. In three patients of this group, methacholine sensitivity determined after BAL-BLmight have been affected by the I3rstimulant treatment received during the previous 12 h. However, these patients took a similar number of puffs of the same drug during the night after bronchial challenge; therefore, the results of methacholine challenges after bronchial challenge and BAL-BL should be comparable. The only deterioration in lung function observed at the end of all procedures in this group was a marginal (less than 5070) decrease of mean FEV 1 compared with baseline. In the patients of this group, methacholine sensitivity was determined 24 h after BAL-BL; it is, therefore, unlikely that results may have been affected by the premedication given for bronchoscopy since intramuscularly administered atropine is almost completely eliminated within 12 h (15). In conclusion, our results confirm and extend the observations reported by Kirby and coworkers (8) that BAL-BL does
not alter airway responsiveness in asthmatic patients. In addition, we have shown that the combination of allergen inhalation challenge and BAL-BL seems to be a safe procedure sinceBAL-BLdoes not appear to further increase allergeninduced airway hyperresponsiveness. References 1. Godard P, Chaintreuil J, Damon M, et 01. Functional assessment of alveolar macrophages; comparisons of cells from asthmatics and normal subjects. J Allergy Clin Immunol 1982; 70:88-93. 2. Crimi E, Scordamaglia P, Crimi P, Zupo S, Barocci S. Total and specific IgE in serum, bronchial lavage and bronchoalveolar lavage of asthmatic patients. Allergy 1983; 38:553-9. 3. Tomioka M, Ida S, Shindoh Y, Ishihara T, Takishima T. Mast cells in bronchoalveolar lumen of patients with bronchial asthma. Am Rev Respir Dis.1984; 129:1000-5. 4. De Monchy GR, Kauffman HF, VengeP, et 01. Bronchoalveolar eosinophils during allergeninduced late-phase asthmatic reactions. Am Rev Respir Dis 1985; 131:373-6. 5. Metzger WJ, Richerson HB, Worden K, Monick M, Hunningake GW. Bronchoalveolar lavage of allergic asthmatic patients following allergen bronchoprovocation. Chest 1986; 89:477-83. 6. Fabbri LM, Boschetto P, Zocca E, et 01. Bronchoalveolarneutrophilia during late asthmatic reactions induced by toluene diisocyanate. Am Rev Respir Dis 1987; 136:36-42. 7. Lam S, Leriche J, Philips D, Chan-Yeung M. Cellular and protein changes in bronchoalveolar lavage fluid after late asthmatic reactions in patients with red cedar asthma. J Allergy Clin Immunol 1987; 80:44-50. 8. Kirby JG, O'Byrne PM, Hargreave FE. Bronchoalveolar lavage does not alter airway responsivenessin asthmatic subjects. Am Rev Respir Dis 1987; 135:554-6. 9. KellyC, Hendrick D, Walters H. The effect of bronchoalveolar lavage on bronchial responsiveness in patients with airflow obstruction. Chest 1988; 93:325-8. 10. Cartier A, Thomson NC, Frith PA, Roberts R, HargreaveFE. Allergen-induced increasein bronchial responsiveness to histamine: relationship to the late asthmatic response and change in airway caliber. J Allergy Clin Immunol 1982; 70:170-7. 11. Crimi E, Brusasco V, Losurdo E, Crimi P. Predictive accuracy of late asthmatic reaction to Dermatophagoides pteronyssinus. J Allergy Clin Immunol 1986; 78:908-13. 12. Cohen AB, Batra GK. Bronchoscopyand lung lavage induced bilateral pulmonary neutrophil influx and blood leukocytosis in dogs and monkeys. Am Rev Respir Dis 1980; 122:239-47. 13. Hargreave FE, Ramsdale EH, Kirby JG, O'Byrne PM. Asthma and the role of inflammation. Eur J Respir Dis 1986;69(Suppl 147:16-21). 14. Metzger M, Zavala D, Richerson HB, et al. Local allergen challenge and bronchoalveolar lavage of allergic asthmatic lungs. Am Rev Respir Dis 1987; 135:433-40. 15. Weiner N. Atropine, scopolamine, and related antimuscarinic drugs. In: Gilman AG, Goodman LS, Gilman A, eds. The pharmacological basis of therapeutics. 6th ed. New York: Macmillan, 1980; 120-37.
PIERO GIANIORIO, MARCO BONAVIA, EMANUELE CRIMI, SABINA LANTERO, PAOLO CRIMI, GIOVANNI A. ROSSI, and VITO BRUSASCO
Introduction
Bronchoalveolar lavage has been increasingly used for studying the inflammatory cells and mediators present in the asthmatic lung both during baseline conditions (1-3) and after inhalation challenges with allergens(4, 5) or occupational agents (6, 7). A limitation to the use of bronchoalveolar lavage in asthmatic patients is represented by the risk that this procedure may lead to deterioration of lung function and increase bronchial responsiveness. During baseline conditions, bronchial responsiveness to methacholine was found to be unchanged after bronchoalveolar lavage in patients with mild airway hyperresponsiveness (8), but it was increased in patients with a high initial hyperresponsiveness (9). After allergen inhalation, bronchial responsiveness may be increased in some asthmatic patients (10). Whether bronchoalveolar lavage performed after an allergen inhalation challenge may induce or aggravate airway hyperresponsiveness has not been evaluated. The present study was designed to investigate if in allergicpatients with different patterns of response to inhaled allergen a fiberoptic bronchoscopy with bronchoalveolar and bronchial lavage performed 4 or 24 h after allergen bronchial challenge may deteriorate lung function and increase bronchial responsiveness. Methods Patients Twenty-three outpatients, whose characteristics are summarized in table 1, were studied. They had a history of perennial rhinitis and/or bronchial asthma with allergic sensitization to Dermatophagoidespteronyssinus as documented by skin prick test and radio-allergosorbent test. At the time of study all patients were asymptomatic, with a FEV 1 larger than 70070 of predicted. None of the patients were receiving antiasthmatic treatment other than
SUMMARY Nonspecific bronchial responsiveness was studied In 23 allergic patients with a history of rhinitis and/or bronchial asthma who underwent flberoptlc bronchoscopy with bronchoalvaolar and bronchial lavage (BAI.-BL) 4 h (Group A) or 24 h (Group B) after an allergen Inhalation challenge. In all patients, the dose of methacholine causing an FEY1 filII of 15% (P01 5) was determined at baseline, 24 h before allergen challenge. Methacholine bronchial challenge wes repeated 1 h before BAI.BL In patients of both groups and again 12 to 14 h after BAI.-BL In Group A and 24 h after BAI.-BL in Group B. In patients of Group A, the values of methacholine POu after BAL-BL were not significantly different from those determined before BAI.-BL. This wes also the case In patients In whom bronchial responsiveness weslncreased after allergen challenge. In patients of Group B, methacholine POlS was significantly decreased after allergen challenge, and this decrease was correlated with the occurrence and the severity of the late asthmatic reaction. Evan In patients who showed dual asthmatic reactions and an Increased responsiveness after allergen challenge, methacholine P0 15 was not further decreased after BAL-BL. These dlta support the safety of a procedure combinIng bronchial allergen challenge with BAL-BL, which can be used for studies on the p~thophyslology of bronchial asthma. AM REV RESPIR DIS 1991; 143:105-108
13rstimulants.These, if any,werediscontinued at least 12 h before entering the study.
trol between 3 and 8 h after the end of inhalation challenge.
Bronchial Challenges Methacholine aerosols were delivered by an ampul-dosimeter device (MEFAR, Brescia, Italy) (11). The challenge was started from a dose of 0.02 mg. The same ampul was used for each patient. The best of three FEV 1 maneuvers measured immediately after inhalation of each methacholine dose was taken to construct dose-response curves. The dose of methacholine wastwofold-incremented until FEV 1 was decreased below 80070 of control (inhalation of saline), or up to a maximal dose of 5 mg. The dose causing a 15070 fall of FEV 1 (PD 1s ) was calculated by interpolation of the dose-response curves. For allergen bronchial challenge, solutions of Dermatophagoides pteronyssinus reconstituted from predosed (arbitrary units, AU) dried allergen (Pharmacia, Uppsala, Sweden) were used. FEV 1 was measured 15 min after inhalation of allergen solutions. After control inhalation of saline, the allergen bronchial challenge was started from a dose of 4 AU, with twofold increments until the FEV 1 fell below 80070 of control values, or up to a maximal dose of 500 AU. Thereafter, FEV 1 was monitored every hour for 8 h to detect the occurrence of late asthmatic reactions, defined as an FEV 1 fall exceeding 15070 of con-
Bronchoalveolar and Bronchial Lavage Patients weregiven atropine (0.5 mg) and diazepam (10mg) intramuscularly 5 min before the introduction of the bronchofiberoscope (Olympus BF, type PlO; Olympus Corp. of America, New Hyde Park, NY). The bronchoscope was passed through the nose after local anesthesia of the nostrils (lidocaine solution 2070 and adrenaline solution 0.1/1000, 1 ml each side). Bronchoalveolar lavage was performed by injecting into the middle lobe
(Received in original form February 23, 1990 and in revised form July 13, 1990) 1 From the Istituto di Medicina delloSport, Cattedra di Fisiopatologia Respiratoria, and the Instituto di Igiene e Medicina Preventiva, Cattedra di Medicina di Comunita, Universita di Genova, and the Prima Divisione di Pneumologia, Ospedale S. Martino, Genova, Italy. 1 Supported in part by a grant from The Italian Ministry of Public Education by Grant No. 87.00560.56 from the Consiglio Nazionale delle Richerche. 3 Correspondence and requests for reprints should be addressed to Vito Brusasco, M.D., Istituto di Medicina dello Sport, Viale Benedetto XV, 10. 16132, Genova, Italy.
105
GIANIORIO, BONAVIA, CRIMI, LANTERO, CRIMI, ROSSI, AND BRUSASCO
106
the end of the bronchoscopic procedures, with a contact telephone number that they could call at any time.
TABLE 1 PATIENTS' CHARACTERISTICS, BRONCHODILATOR USAGE, AND ADVERSE EFFECTS OF BAL-BL Bronchodilator Patient No. Group A 1 2 3 4 5 6 7 8 9 Mean SO Group B 10 11 12 13 14 15 16 17 18 ,19 20 21 22 23 Mean SO
Age (yr)
25 19 24 18 18 19 19 19 27 21 3 28 19 26 26 19 38 17 18 17 21 18 19 21 20 22 6
Sex
Post-BPT
Post-BAL-BL
Adverse Effects
79 100 107 98 100 98 76 95 111 96 12
No No No No No No No No No
No No No No Yes Yes Yes No No
None None None None None None Fever, malaise None None
89 111 99 98 129 74 109 98
Yes Yes Yes No No Yes No Yes Yes No Yes Yes Yes Yes
No No Yes No No Yes No No Yes No No Yes No Yes
None Sore throat None None None None None None None None None None None Cough
Baseline FEV 1 (% pred)
M
M M M M M M M M
M
F M M M M M M M M M M M M
Definition of abbreviations: BPT
n
106 108 113 103 109 101 14
= allergen bronchial provocation
test; BAL-BL
Statistical Analysis The values of methacholine PD 15 were transformed in logarithmic form for statistical analysis and reported as geometric mean. In order to control the per-experiment error rate, the significance of changes among observation times was assessed by F statistics. If p values from this preliminary analysis were less than 0.05, separate paired t tests were performed. Pearson's correlation coefficient was also used; p values less than 5010 were considered to be statistically significant. Results
= bronchoalveolar
and bronchial lavage.
bronchus 100 ml of sterile saline, fractionated in five aliquots of 20 ml each, which were aspirated at a negative pressure of 50 to 120 mm Hg. Bronchial lavage was performed by injecting into the orifice of the lower lobe bronchus 40 ml of saline, fractionated in two aliquots of 20 ml each, which were immediately suctioned at a pressure of about 50 to 120 mm Hg.
Experimental Protocols All patients underwent a methacholine bronchial challenge in the morning of the first day of study, and an allergen challenge in the morning of the following day. Bronchoalveolar and bronchial lavage (BAL-BL) was performed 4 h after the end of the allergen inhalation challenge in nine patients (Group A), and 24 h after the allergen challenge in the remaining 14patients (Group B). Methacholine challenge was repeated 1 h before and again 12 to 14 h after BAL-BL in Group A, and 1 h before and again 24 h after BAL-BL in Group B. A synopsis of the experimental protocols used in the two groups is given in table 2. The time intervals between procedures was chosen so that methacholine bronchial challenge was always done at the same time in the morning. At the end of the methacholine challenge performed 1 h before BAL-BL, patients inhaled 200 ug salbutamol to reverse the induced bronchospasm. A single dose of
salbutamol (200 ug by metered aerosol) was also given to the patients who experienced symptoms of asthma after allergen bronchial challenge or BAL-BL (table 1). Patients were allowed to inhale additional doses of salbutamol if nocturnal symptoms of asthma developed. Patient 6 showed a severelate asthmatic reaction after bronchial challenge and was given steroid intramuscularly (bethamethasone 4 mg) immediately before BAL-BL. All patients left the hospital 8 h after the end of the bronchial challenge or 1 h after
TABLE 2 SYNOPSIS OF EXPERIMENTAL PROTOCOLS Time between Procedures
Procedures
Group A
Group B
(h)
(h)
24
24
3
24
12-14
24
Methacholine challenge BPT Methacholine challenge BAL-BL Methacholine challenge For definition of abbreviations, see table 1.
Group A The results of allergen inhalation challenge are given in table 3, and changes of methacholine sensitivity and baseline FEV 1 after allergen challenge and BALBL are shown in figure 1. Methacholine PD 15 (geometric mean) was 0.214mg before allergen challenge, 0.170 mg 3 h after allergen challenge, and 0.322 mg 12 to 14 h after BAL-BL. The corresponding baseline values of FEV 1 (mean ± SD) were 4.12 ± 0.55 L, 4.05 ± 0.58 L, and 3.98 ± 0.71 L, respectively. None of these differences was statistically significant. No significant correlations resulted between changes in methacholine PD 1 5 after BAL-BL and either baseline or prelavage methacholine PD 15 • There was a wide variability of changes in PD 15 after allergen challenge, which was likely due to different inflammatory responses of individual patients. In four patients, methacholine PD 15 was reduced by 500/0 or more after allergen challenge. One of these patients (Patient 7) took several doses of salbutamol overnight after BALBL. This patient was the only one in whom PD 15 to methacholine was found to be further decreased after BAL-BL compared with the postchallenge value. Group B The results of allergen inhalation challenge are given in table 3, and changes of methacholine sensitivity and baseline FEV 1 after allergen challenge and BALBL are shown in figure 2. 1\venty-four hours after allergen challenge, both methacholine PD 15 and baseline FEV 1 were reduced from 0.136mg to 0.096 mg (p = 0.(05) and from 4.39 ± 0.85 to 3.89 ± 0.94 L (p = 0.0008), respectively. The changes in methacholine PD 15 and baseline FEV 1 were significantly correlated (r = 0.58, p = 0.03 and r = 0.74, p = 0.0025, respectively) with the late maxi-
107
BRONCHOALVEOLAR LAVAGE AND ALLERGEN CHALLENGE IN ASTHMA
TABLE 3 RESULTS OF BASELINE METHACHOLINE CHALLENGE ANO ALLERGEN CHALLENGE
Patient No.
Methacholine POlS (mg)
Group A 1 2 3 4 5 6 7 8 9 Mean SO Group B 10 11 12 13 14 15 16 17 18 19 20 21 22 23 Mean SO
Maximal FEVl Fall EAR· (% of control)
31 200 2,016 337 61 105 205 96 1,760 214 (geometric)
14 21 12 8 27 22 44 23 11 20 11
15 131 24 345 5,000 34 693 70 31 5,000 104 55 43 137 135 (geometric)
32 37 24 15 5 20 36 28 24 14 30 29 33 20 24 9
Maximal FEV1 Fall LARt (% of control)
30 25 17
6 2 53
3 31 30
o 30
2 31
2 18 16
• Maximal fall of FEV, within 1 h after allergen inhalation challenge. Maximum fall of FEV, between 3 and 8 h after allergen inhalation challenge.
t
mal fall of FEV 1. This indicates that the significant changes observed after allergen challenge were due to eight patients who developed a late asthmatic response. Twenty-four hours after BAL-BL, methacholine PD 15 was 0.152 mg and baseline FEV 1 was 4.19 ± 0.94 L. These values were significantly higher than after allergen challenge (p = 0.04 and p = 0.006, respectively); however, FEV 1 values were still significantly lower than before allergen challenge (p = 0.004). There wereno significant correlations be-
tween changes in methacholine PD 15 after BAL-BL and either baseline or prelavagemethacholine PD 1S • Three patients (Patients 15, 18, and 23) took overnight doses of salbutamol both after bronchial challenge and after BAL-BL, one patient (Patient 21) used salbutamol only after bronchial challenge. Discussion
In this study, we found that BAL-BL performed 4 or 24 h after allergen bronchial challenge neither deteriorated lung func-
Group A.
Group A.
•
=== ;:::::
4
§ 2
0..1...---
8.for. BPT
Aft... BAL-8L Aft.r BPT
_
8.ro ... 8PT
Aft... 8AL-BL Aft.,. BPT
Fig. 1. ~hanges in bronchial sensitivity (PD,s) to methacholine and baseline FE~ 3 h after allergen bronchial provocation test (BPT)and 12to 14 h after bronchoalveolar and bronchial lavage (BAL-BL). None of the differences was statistically significant.
tion nor increased airway responsiveness even in patients in whom airway responsiveness was increased as a consequence of the previous allergen inhalation. In primates, it has been reported that bronchoalveolar lavage may elicit an inflammatory response in the lung (12). Airway inflammation is a key feature of bronchial asthma and is thought to be a main mechanism in the pathogenesis of bronchial hyperreactivity (13).Therefore, although in normal humans and in nonasthmatic patients (9), bronchoalveolar lavage does not appear to cause an increase in bronchial reactivity to methacholine, one may expect that this procedure in asthmatic patients may produce or worsen airway hyperresponsiveness. However, Kirby and coworkers (8) found no changes in bronchial sensitivity to methacholine measured 24 h after BAL in patients with mild asthma. Bycontrast, in a group of asthmatic patients with various degrees of airway responsiveness, Kelly and colleagues (9) found that sensitivity to methacholine was increased 24 h after BAL, and this increase was positively correlated with the baseline responsiveness. Experimental exposure to allergen has been shown to cause an increase of nonspecific bronchial responsiveness that, in patients with dual asthmatic reaction, may persist for several days (10). On the basis of these data, one might expect that if bronchoalveolar lavage is performed after allergen inhalation challenge any allergen-induced bronchial hyperresponsiveness might be eventually enhanced. The only study in which the effect of the combined procedure of bronchial provocation and BAL was considered is that of Metzger and coworkers (14). They found a marginal deterioration of lung function after locally instilled allergen followed by BAL. In the present study, we found no evidence of increased methacholine sensitivity after BAL-BL performed either early (Group A) or late (Group B) after allergen bronchial challenge. This was also true in patients with an elevated degree of baseline or prelavage bronchial responsiveness. This finding does not confirm what was previously reported by Kelly and colleagues (9). More importantly, we also found that BAL-BL did not further enhance bronchial responsiveness when this had been previously increased as a result of an allergen inhalation challenge. In a fewpatients, symptoms of asthma occurred early after BAL-BL, but they werein most cases easily controlled by a single dose of f3r
108
GIANIORIO, BONAVIA, CRIMI, LANTERO, CRIMI, ROSSI, AND BRUSASCO
Group B
Group B 10
..
---
~-~ - --
" - -::= ~-I= ::,=/~---...
.01
~
.001
Before BPT
Afte,. BAL-BL After aPT
~
2
~
0.1...-.-----------B.fo,.e BPT
Afte,. BAL-BL After aPT
Fig. 2. Changes in POts to methacholine and baseline FE'/, 24 h after BPT and 24 h after BAL. Continuous lines indicate patients with late asthmatic reaction, and broken lines indicate patients with isolated early asthmatic reaction or without asthmatic reaction. POtsafter BPT versus POtsbefore BPT: p = 0.005; POts after BAL versus POts after BPT: p = 0.04. FE'/, after BPT versus FE'/, before BPT: p = 0.0008; FE'/, after BAL versus FE'/, after BPT: p = 0.006; FE'/, after SAL versus FE'/, before BPT: p = 0.004.
stimulant and did not result in a sustained increase of bronchial responsiveness. We cannot know which patients of Group A would have experienced a late asthmatic response to allergen since the medications givenbefore and after BALBL may have prevented it. However, we can reasonably think that the four patients in whom airwayresponsiveness was increased after the allergen challenge were likely to develop a late asthmatic reaction. In three of these patients, the increase in airway responsiveness observed after allergen challenge was reversed after BAL-BL. In this group, methacholine sensitivity was determined 12to 14h after BAL-BL;therefore, it cannot be excluded that the tendency for methacholine sensitivity to be decreased after BAL-BL was in part due to a persistent effect of the atropine given intramuscularly as a premedication for bronchoscopy. Even though premedication drugs might have somehow affected the results in this group, the lack of worsening of airway responsiveness after BAL-BL performed 4 h after allergen inhalation challenge seems of practical interest. In patients of Group B, we found that methacholine sensitivity was increased after allergen challenge and, consistent with a previous report (10), this increase was positively correlated with the severity of the late asthmatic reaction. Cartier and coworkers (10) found that allergeninduced airway hyperresponsiveness in most patients with definite late reactions lasted 48 h or more. In our study, an increased methacholine sensitivity was observed 24 h after allergen challenge in
four of eight patients who developed a late asthmatic reaction, but this change was completely reversed24 h after BALBL. In these patients, thus, allergen-induced hyperresponsiveness apparently lasted less than 48 h. We do not know the reason for this finding, whichapparently contrasts with those of Cartier and coworkers. It may be possible that antigenic stimulation was less strong in our patients, but we cannot exclude that premedication given for bronchoscopy may have interrupted the pathophysiologic events that lead to persistent increase in bronchial responsiveness after exposure to allergen. In three patients of this group, methacholine sensitivity determined after BAL-BLmight have been affected by the I3rstimulant treatment received during the previous 12 h. However, these patients took a similar number of puffs of the same drug during the night after bronchial challenge; therefore, the results of methacholine challenges after bronchial challenge and BAL-BL should be comparable. The only deterioration in lung function observed at the end of all procedures in this group was a marginal (less than 5070) decrease of mean FEV 1 compared with baseline. In the patients of this group, methacholine sensitivity was determined 24 h after BAL-BL; it is, therefore, unlikely that results may have been affected by the premedication given for bronchoscopy since intramuscularly administered atropine is almost completely eliminated within 12 h (15). In conclusion, our results confirm and extend the observations reported by Kirby and coworkers (8) that BAL-BL does
not alter airway responsiveness in asthmatic patients. In addition, we have shown that the combination of allergen inhalation challenge and BAL-BL seems to be a safe procedure sinceBAL-BLdoes not appear to further increase allergeninduced airway hyperresponsiveness. References 1. Godard P, Chaintreuil J, Damon M, et 01. Functional assessment of alveolar macrophages; comparisons of cells from asthmatics and normal subjects. J Allergy Clin Immunol 1982; 70:88-93. 2. Crimi E, Scordamaglia P, Crimi P, Zupo S, Barocci S. Total and specific IgE in serum, bronchial lavage and bronchoalveolar lavage of asthmatic patients. Allergy 1983; 38:553-9. 3. Tomioka M, Ida S, Shindoh Y, Ishihara T, Takishima T. Mast cells in bronchoalveolar lumen of patients with bronchial asthma. Am Rev Respir Dis.1984; 129:1000-5. 4. De Monchy GR, Kauffman HF, VengeP, et 01. Bronchoalveolar eosinophils during allergeninduced late-phase asthmatic reactions. Am Rev Respir Dis 1985; 131:373-6. 5. Metzger WJ, Richerson HB, Worden K, Monick M, Hunningake GW. Bronchoalveolar lavage of allergic asthmatic patients following allergen bronchoprovocation. Chest 1986; 89:477-83. 6. Fabbri LM, Boschetto P, Zocca E, et 01. Bronchoalveolarneutrophilia during late asthmatic reactions induced by toluene diisocyanate. Am Rev Respir Dis 1987; 136:36-42. 7. Lam S, Leriche J, Philips D, Chan-Yeung M. Cellular and protein changes in bronchoalveolar lavage fluid after late asthmatic reactions in patients with red cedar asthma. J Allergy Clin Immunol 1987; 80:44-50. 8. Kirby JG, O'Byrne PM, Hargreave FE. Bronchoalveolar lavage does not alter airway responsivenessin asthmatic subjects. Am Rev Respir Dis 1987; 135:554-6. 9. KellyC, Hendrick D, Walters H. The effect of bronchoalveolar lavage on bronchial responsiveness in patients with airflow obstruction. Chest 1988; 93:325-8. 10. Cartier A, Thomson NC, Frith PA, Roberts R, HargreaveFE. Allergen-induced increasein bronchial responsiveness to histamine: relationship to the late asthmatic response and change in airway caliber. J Allergy Clin Immunol 1982; 70:170-7. 11. Crimi E, Brusasco V, Losurdo E, Crimi P. Predictive accuracy of late asthmatic reaction to Dermatophagoides pteronyssinus. J Allergy Clin Immunol 1986; 78:908-13. 12. Cohen AB, Batra GK. Bronchoscopyand lung lavage induced bilateral pulmonary neutrophil influx and blood leukocytosis in dogs and monkeys. Am Rev Respir Dis 1980; 122:239-47. 13. Hargreave FE, Ramsdale EH, Kirby JG, O'Byrne PM. Asthma and the role of inflammation. Eur J Respir Dis 1986;69(Suppl 147:16-21). 14. Metzger M, Zavala D, Richerson HB, et al. Local allergen challenge and bronchoalveolar lavage of allergic asthmatic lungs. Am Rev Respir Dis 1987; 135:433-40. 15. Weiner N. Atropine, scopolamine, and related antimuscarinic drugs. In: Gilman AG, Goodman LS, Gilman A, eds. The pharmacological basis of therapeutics. 6th ed. New York: Macmillan, 1980; 120-37.
