Airway Inflammation and Occurrence of Delayed Bronchoconstriction in Exercise-induced Asthma 1- 3

EMANUELE CRIMI, ANTONELLA BALBO, MANLIO MILANESE, ANTONIO MIADONNA, GIOVANNI A. ROSSI, and VITO BRUSASCO

Introduction

In asthmatic subjects, experimental exposure to allergen or occupational sensitizers causes an acute bronchospasm associated with recruitment of inflammatory cells in the airways. In a variable number of patients, a bronchospastic response may also be triggered by exercise. It is widely accepted that the initiating stimulus for exercise-induced asthma (EIA) is represented by thermal changes occurring in the airways (1-3). Whether a relationship exists between airway inflammation and EIA is controversial (4, 5). Another controversial issue in EIA is whether a late-phase asthmatic response to exercise may occur (6-8). In allergic asthma, a link between the latephase asthmatic response and the degree of airway inflammation has been clearly documented (9, 10). Yet no evidence has been provided that the late-phase response after exercise reported by some investigators (6-8) was associated with inflammatory changes in the airways. In this study, we investigated whether changes in the number of inflammatory cells and in the levels of mediators occur in the airways after EIA. We observed that exerciseis associated with an increase of mast cell degranulation and an influx of eosinophils in the airway lumen. Furthermore, we wondered if a late-phase response occurred after exercise and whether this was specifically associated with the inflammatory changes in the airways. The results show that a delayed bronchoconstriction may develop after exercise, but this seems to be the result of fluctuations in lung function occurring in patients with prominent airway inflammation. Methods Patients Nine asthmatic patients whose characteristics are shown in table 1 participated in the study. To be included in the study, patients had to meet the following criteria: to have a recent history of EIA, to have been free of

SUMMARY We studied nine asthmatic patients with a history of exercise-induced asthma (EIA) in order to Investigate whether inflammatory changes in the airways occur after exercise and are eventually associated with the development of a late-phase asthmatic response. On two separate study days, bronchoalveolar lavage (BAl), bronchial lavage (Bl), and bronchial biopsy (BB) were performed 3 h after an exercise or a methacholine (MCh) challenge. On two other separate occasions, FEV1 was monitored for 12 h after Identical exercise and MCh challenges not followed by BAl, Bl, and BB. We found a greater percentage of eoslnophlls In BAl after exercise versus MCh challenge (p < 0.05). In five patients, BAl eoslnophils after exercise were ~ 2% of total Inflammatory and immunoeffector cells. In three of these patients an FEV1 fall > 20% of control was recorded 5 to 12 h after exercise. However, two of these patients had 2% or more eoslnophlls In BAl and similar late falls of FEV1 after MCh challenge. The percentage of degranulating mast cells in BB was higher (p < 0.05) after EIA than after MCh, but no significant differences were found In Bl histamine and leukotrienes. We conclude that (1) exercise may enhance mast cell degranulation and eosinophilic Inflammation of the airways, and (2) a delayed bronchoconstrlction after exercise Is not specific to EIA but is more likely the result of fluctuations in lung function associated with airway AM REV RESPIR DIS 1992; 146:507-512 inflammation.

severe exacerbations of asthma for at least 1 month, and not to be currently undergoing treatment with antiasthmatic drugs other than ~2 stimulants; these, if any, were avoided for 12h before studies. Measurements of pulmonary function were made by using a turbine spirometer (MicroMedical Ltd., Rochester, UK). Studies were carried out provided that the FEV 1 was not < 65070 of the predicted value (11). Patients attended the laboratory on 4 separate days. On the first 2 consecutive days (screening days), patients underwent methacholine (MCh) and exercise challenges. On both occasions, FEV 1 was monitored hourly for 12 h after the initial asthmatic response. The results of bronchial challenges obtained on these screening days are shown in table 1. Approximately 1 wk after this initial screening, patients attended the laboratory on 2 study days to undergo bronchoalveolar lavage (BAL), bronchial lavage (BL), and bronchial biopsy (BB) 3 h after MCh or exercise challenge. The order 0 f bronchial challenges was randomized, and 1 wk was allowed between the 2 study days. All patients were volunteers who gavewritten consent after receiving complete information about the nature and the aim of the study. Bronchial Challenges MCh. Aerosols of MCh chloride in saline were delivered by an ampul-dosimeter device (MEFAR, Brescia, Italy) with the same ampul being used for each patient. Three tech-

nically acceptable forced expiratory maneuvers were obtained at baseline and 1 min after each inhalation dose. The challenge was started from a dose of 20 ug and continued with twofold increments until FEV 1 fell below 80070 of control (inhalation of saline), or up to a maximum dose of 5,000 ug, Doseresponse curves wereconstructed by using the largest FEV 1 value obtained at each step, and the dose causing the FEV 1 to fall by 15070 (PD 1s) was calculated by interpolation. Exercise. Exercise was performed on a cycloergometer for 6 min with the work load adjusted to increase the heart rate to 85070 of the maximum predicted for age (12). During exercise, patients breathed through a mouthpiece connected with a twoway low-resistance valve. The inspired air was conditioned to have

(Received in original form July 29, 1991, and in revised form February 3, 1992) 1 From the Dipartimento eli Scienze Motorie, Cattedra di Fisiopatologia Respiratoria, Universita di Genova, the Istituto G. Gaslini, Divisione di Pneumologia, Genova, and the Dipartimento di Medicina Interna, Malattie Infettive ed Immunopatologia, Universita di Milano, Milano, Italy. 2 Supported in part by Grant No. 87.00560.56 from the Consiglio Nazionale delle Ricerche, Rome, Italy. 3 Correspondence and requests for reprints should be addressed to Emanuele Crimi, M.D., Medicina dello Sport, Ospedale S. Martino, Viale Benedetto XV, 10,16132, Genova, Italy.

507

508

CRIMI, BALBO, MILANESE, MIADONNA, ROSSI, AND BRUSASCO

TABLE 1 PATIENT DEMOGRAPHIC CHARACTERISTICS AND PULMONARY FUNCTION DATA ON SCREENING DAYS' Maximal FEV, Fall (%) Patient No.

FEV, (%)

Age

After MCh

After Exercise

(yr)

Sex

Before MCh

Before Exercise

MCh PD,. (JIg)

1 2 3 4 5 6 7 8 9

27 28 25 25 24 21 21 22 20

M F M M M M M M M

83 82 76 83 70 86 102 88

90 90 78 80 70 82 95 65 90

87 37 131 90 27 30 107 20 30

31 16 19 31 34 23 23 23 29

5 40 2 0 35 3 2 4 7

16 22 34 22 50 36 13 10 17

9 25 3 0 45 2 5 0 25

Mean SE

23 1

82 3

82 3

50 (Geometric)

25 2

11 5

24 4

13 5

72

Early

Late

Early

Late

Definition of abbreviation : FEV, (0/0) = FEV, as percent of predicted.

• None of the differences between the MCh-challenge day and exercise-challenge day were statistically significan t.

a relative humidity of ""0070 and a temperature at the mouth of - 5 ± 2 0 C. The expired air was measured with a gasometer. FEY! measurements were taken before exercise and 1, 5, 15, 30, 60, 120, and 180 min after exercise.The maximum postexerciseFEY1 fall was retained for analysis.

BAL-BL-BB Studies Procedure. Patients receivedatropine (0.5 mg intramuscularly) and diazepam (10 rng intramuscularly) as premedication for bronchoscopy. From previous studies it was apparent that this premedication inhibits neither eosinophil influ x in the airways (10, 13)

nor mast cell degranulation (14). Nasal anes thesia was obtained with 2% lidocaine and 0.01% adrenaline solutions (I ml each nostril). A fiberoptic bronchoscope (Olympus BF, type PIO; Olympus Corp. of America, New Hyde Park, NY) was passed through the nose and, after local anesthesia of pharynx and airways, was wedged into a subsegmental branch of the right middle lobe. BAL was performed by infusing five 20-ml aliquots of sterile saline that were suctioned at a negative pressure of 50 to 120 mm Hg. BL was performed to obtain more concentrated samples for mediator analysis. For this purpose two 20-ml aliquots of sterile saline were in-

jected into the orifice of the left lower lobe bronchus and immediatelysuctioned at a pressure of 50 to 120mm Hg. BB was taken, after BAL and BL, from the carina of the main stem bronch i and from the carina of the right upper bronchus. Inhaled salbutamol (200 ug) was given to patients who experienced respiratory symptoms after bronchoscopy. Patients left the hosp ital 1 h after completion of the procedure with a contact telephone number that they could call at any time. Processing ofBAL and BL fluids. The recovered fluids were filtered through two layers of sterile gauze and cells were separated from the supernatant by centr ifugation at 250 x g for 5 min. The cell pellet was washed once and resuspended in Hanks' balanced salt solution without Ca H or Mg" at a concentration of 106 cells/ml. A small sample of the cell suspension was centrifuged by using a cytocentrifuge (Cytospin; Shandon Southern Instruments, Sewickley, PA) spinning approximately 100,000 cells at 500 rpm for 5 min onto a glass slide. BAL cells were air-dried and stained with Diff-Quik staining (Merz & Dade A. G., Dudigen, Switzerland). Cell differentials weredetermined by counting at least 300 cells per slide, using a light microscope (Carl Zeiss, Oberkochen, Germany). The supernatants of BL were frozen at -70 0 C before mediator anal ysis. Histamine in the BL fluid was determined in duplicate by an automated fluorimetric technique accord ing to the method of Siraganian (15). The presence of histamine wasconfirmed by a reduction of the fluorescence of the samples after treatment with diamine oxidase. Immunoreactive leukotrienes LTC.,

Fig. 1. Examples of intact mast cell observed at xl ,OOO after MCh challenge (left) and degranulating mast cell observed at x500 after exercise (right).

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AIRWAY INFLAMMATION IN EXERCISE-INDUCED ASTHMA

TABLE 2 PULMONARY FUNCTION (FEV 1) DATA ON STUDY DAYS* MCh-Challenge Day Patient No.

Exercise-challenge Day

Before MCh (0/0 predicted)

After MCh (maximum % fall)

Before Bronchoscopy (0/0 predicted)

Before Exercise (0/0 predicted)

After Exercise (maximum % fall)

Before Bronchoscopy (0/0 predicted)

1 2 3 4 5 6 7 8 9

83 86 76 83 65 83 99 65 95

28 18 28 17 20 25 25 14 31

84 89 78 82 71 83 101 65 85

79 90 77 85 69 73 95 76 89

38 15 37 31 17 15 20 30 49

81 89 79 89 77 86 103 69 84

Mean SE

82 4

23 2

82 3

81 3

28 4

84 3

* None of the differences between the MCh-challenge day and exercise-challenge day were statistically significant.

LTD4 , and LTE 4 in BL fluids were assayed Statistical Analysis the screening days (p >0.37 and p >0.62, by a competitive radioimmunologic method Data normally distributed were analyzed by respectively). Furthermore, on neither of (16, 17) using commercially available kits '" Student's paired t test and are presented as the study days was the FEV 1 value deter(Amersham, Littlechalfont, UK). Cross re- . mean ± SEM; data that were not normally mined immediately before bronchoscoactivity of rabbit anti-Ll'C, antiserum was distributed were analyzed by Wilcoxon's py significantly different from baseline 64070 with both LTD4 and LTE4 , according signed rank test, and are presen ted as the me- (p > 0.82 and p > 0.25, respectively), into the manufacturing firm. The antibodydian with upper and lower quartiles. Values dicating a complete recovery of pulmoantigen complexes were separated from free were considered to be statistically significant nary function after both MCh and EIA. antigen by absorption of free tracer onto acat p < 0.05. tivated charcoal. A supernatant containing BAL Cell Data the antigen-antibody complexes was obtained Results by centrifugation and then decanted into a Inflammatory cell differentials observed Pulmonary Function polyethylene vial. Thereafter, 10 ml of a liqin BAL after MCh and EIA are presentuid scintillation cocktail (Histagel; Packard On the screening days (table 1), the early ed in table 3. The percentage of eoInstrument Co., Downers Grove, IL) was addairway responses to MCh and exercise sinophils was significantly higher (p < ed, and radioactivity was measured in a betawere similar. Three patients exhibited a 0.05) after EIA than after MCh. Five pacounter (Tri-Carb 4000; Packard). Results are substantial decrease of FEV 1 between 5 tients had 20/0 or more eosinophils in expressed in LTC4 equivalents. Albumin in and 12 h after the early response to exer- BAL after EIA; three of them were those BL fluid was measured with a radial immucise. Twoof these patients exhibited simi- who had a late fall of FEV 1 after EIA nodiffusion technique using ultralow-Ievel lar late falls of FEV 1 also after MCh. on the screening day. Two of these paplates (Behring; Scoppito, L'Aquila, Italy). On the study days (table 2), FEV 1 was tients had 2 % or more eosinophils in Processing ofBB. The specimens were immediately fixed in 10070 buffered formalin sosimilar before MCh and exercise chal- BAL also after MCh; these patients were lution at room temperature, then embedded lenge (p > 0.17). The maximum falls of those who had also a late fall of FEV 1 in paraffin and cut at 5 J.1m with a rotative FEV 1 caused by MCh and exercise were after MCh on the screening day. The permicrotome. Toluidine blue stain was used for also similar (p > 0.17) and not signifi- centage of macro phages was significantly mast cells and hematoxylin-eosin for other cantly different from those observed on lower (p < 0.05) after EIA than after parenchymal and inflammatory cells. BB were analyzed by two independent observers who were unaware of the results of bronchial TABLE 3 challenges. BB not correctly oriented were disINFLAMMATORY CELL DIFFERENTIALS IN BAL 3 H AFTER MCh AND EIA* carded. The following morphologic characteristics were evaluated: abnormalities in the Eosinophils Lymphocytes Neutrophils Total Macrop hages ratio of cylindrical to goblet cells, thickening (%) (%) (%) (%) (x 10"/ml) Patient or other abnormalities of the basement memMCh MCh EIA EIA MCh EIA EIA EIA MCh No. MCh brane, and angiectasis of the lamina propria. Each of these morphologic abnormalities was 1 96.5 0.5 0.5 0.5 6.4 5.4 94 5 3 0 scored from 0 to 3, with 0 meaning absence 15 76.5 2 7 0 1.5 3 4.5 2 8.0 95 1 85.5 4 12 0 1.5 0 of abnormalities. Cells lying within 200 J.1m 2.4 0.75 96 3 80.0 13 1 1 14.5 0.5 5 1.75 85 3.5 from the epithelial basement membrane were 4 2.5 78.0 2 3 19 0 0.5 2.5 4.0 95 counted and identified by means of an eye- 5 4 85.5 91 8 10 1 0.5 0 5.2 5.3 6 piece graticule at x 500 over five fields. En0.5 95.5 97 2 1 1 0 11.6 14.1 3 dothelial cells, pericytes, and Schwann cells 7 92.0 1 1 87.5 7.5 4 1 7.1 6 8 22..8 were not included in this count. Mast cells 9 2.5 1 96 82.0 2.5 2.1 3 13 0 1.5 were also observed at x 1,000to 2,000 to bet2.5 1 4.5 95 85.5 4 0.5 1 Median 5.2 7 ter detect the presence of extracellular gran1 2.1 91 80.0 0.5 0 0 Lower quartile 2.5 3 6 ules. Mast cells showing one or more 4 1 92.0 1 1.5 5.4 96 7.5 13 8 metachromatic granules around it (figure 1) Upper quartile were said to be degranulating, * The differences in macrophages and eosinophils were statistically significant (p < 0.05).

510

CRIMI, BALBO, MILANESE, MIADONNA, ROSSI, AND BRUSASCO

TABLE 4 INFLAMMATORY CELLS AND MORPHOLOGIC ABNORMALITY SCORE IN BRONCHIAL BIOPSY 3 H AFTER MCh AND EIA *

Total (x 102/mm 2 )

Lymphocytes (x 102/mm 2 )

Eosinophils (1/mm 2 )

Mast Cells (1/mm 2 )

MCh

EIA

MCh

EIA

MCh

EIA

MCh

EIA

MCh

EIA

MCh

EIA

1 2 3 4 5 6 7 8

19.5 5.5 20.0 5.5 11.0 13.5 13.5 8.5

9.0 8.5 22.5 8.5 17.5 8.5 8.5 8.0

9.7 2.2 16.0 0.5 2.2 6.7 6.7 7.2

0.4 5.1 6.7 0.8 5.2 2.5 5.9 7.2

26 3 3 10 50 10 19 3

59 5 43 16 48 14 2 14

2 2 2 5 10 11 10 0

6 2 2 3 13 16 6 0

9.5 3.2 3.9 4.8 8.2 6.5 6.5 1.2

7.9 3.3 15.3 7.5 11.6 5.7 2.5 0.6

5 4 4 1 4 3 4 0

2 1 2 1 2 4 7 1

Median Lower quartile Upper quartile

12.2 7.0 16.5

8.5 8.5 13.2

6.7 2.2 8.6

5.1 1.6 6.3

9.6 3.2 22.4

15.2 9.6 45.6

3.5 2 10

5.6 3.5 7.4

6.6 2.9 9.7

4 2 4

2 1 3

MCh. Neither the number of total inflammatory cells nor the percentages of lymphocytes and neutrophils was significantly different after MCh and EIA. The numbers of epithelial cells per milliliter of BAL fluid recovered after MCh and EIA also were not significantly different (medians were 254 and 227, respectively; p > 0.5).

BB Data The numbers of inflammatory cells per unit area of bronchial mucosa observed after MCh and EIA are presented in table 4. No significant differences were found in the numbers of lymphocytes, eosinophils, and mast cells. The number of other cells, including plasma cells, neutrophils, fibroblasts, histiocytes, and unclassified cells, was not significantly different after MCh (range 15 to 88070) than after EIA (range 29 to 88070). When the ratio of degranulating to intact mast

4.5 2 9.5

not suitable for analysis.·

cells was evaluated (table 5), it was found that the percentage of degranulating mast cells was significantly higher after EIA than after MCh (p < 0.05). The morphologic abnormality score was similar after MCh and EIA.

BL Mediators As shown in table 5, we did not observe significant differences in either histamine or LTC (LTC4 equivalent) levels after MCh and EIA. The concentration of histamine was below the detection limit of the assay in four and that of LT in eight of 12 samples available. The concentrations of albumin per milliliter of BAL fluid recovered after MCh and EIA were not significantly different. Discussion

In this study we have found that EIA was associated with an increased proportion of degranulating mast cells in BB and

TABLE 5 PERCENTAGE OF DEGRANULATING MAST CELLS IN BS AND MEDIATOR LEVELS IN BL 3 H AFTER MCh AND EIA

1 2 3 4 5 6 7

8 Mean

SE

Morpholog ic Abnormalities (score)

Patient No.

• No difference was statistically significant. In Patient 9 bronchial biopsy was

Patient No.

Other Cells (x 102/mm 2 )

Degranulating Mast Cells (0/0)

BL Fluid Recovered (ml)

MCh

EIA

MCh

EIA

MCh

EIA

0 0 0 0 20 90 100 0

50 100 100 100 85 63 100 0

20 4.5 6 6 10 NA 3.5 NA

16 13 6 5 21 NA 12 NA

0.25 0.5 0.6 1.5 < 0.2 NA 0.5 NA

26 15

75 13

8.3 2.5

12 2.4

0.6 0.2

BL Histamine (ng/ml)

BL Peptide Leukotriene LTC4 Equivalent (ng/ml) MCh

EIA

< 0.2

0.27

1.5 1.25 1.5 < 0.2 NA < 0.2 NA

< 0.125 0.16 0.13 < 0.125 NA 0.27 NA

< 0.125 < 0.125 < 0.125 < 0.125 < 0.125

0.8 0.1

0.18 0.02

Definition of abbreviation: NA = not available. * The percentage of degranulating mast cells observed after EIA was significantly higher (p difference was significant. In Patient 9 neither 88 nor BL were available.

< 0.05) than after

NA

< 0.125 NA

< 0.125 0 MCh. No other

with an increased percentage of eosinophils in BAL. Furthermore, the results of this study suggest that the occurrence of a delayed bronchoconstriction after exercise challenge is a nonspecific phenomenon likely due to the instability of bronchial tone in patients with chronic bronchial inflammation. Changes in bronchial blood flow or in the osmolarity of the bronchial mucosa that accompany airway cooling and rewarming have been proposed as mechanisms leading to airway narrowing during EIA (1-3, 18). Whether recruitment of inflammatory cells in the airway may also play a role in the pathogenesis of EIA has not been established. An increased neutrophil chemotactic factor activity was found in serum after exercise in patients with EIA, and this increase was prevented by pretreatment with the mast cell membrane stabilizer cromolyn sodium (19), suggesting a role for mast cell degranulation. However, histamine was found to be either increased (20, 21) or unchanged (22, 23) in blood during EIA. Studies using BAL also gave inconsistent results. Broide and coworkers (24) neither found increased concentrations of the most important mast cell-derived mediators nor increased percentages of inflammatory cells during EIA. After isocapnic hyperpnea, which is considered to be a stimulus very similar to exercise (2, 21), Pliss and associates (25) found significant increases of eosinophils, epithelial cells, and leukotrienes, thus suggesting a signi ficant role for airway inflammation. Several factors may account for some discrepancies between studies in which BAL is performed after an experimentally induced bronchoconstriction. First,

511

AIRWAY INFLAMMATION IN EXERCISE-INDUCED ASTHMA

changes in airway caliber at the time of deed, the finding of a larger proportion BAL may cause differences in the distri- of degranulating mast cells in BB after bution of the lavage fluid, which may EIA than after MCh challenge suggests thus reflect different portions of the lung. that EIA is associated with mediators Second, the time of sampling after chal- released from mast cells:it is possible that lenge may affect the local level of medi- 3 h after exercise most of the mediators ators as they are synthesized, secreted, released during EIA were metabolized, and degradated at different rates. Simi- thus reducing their concentration below larly, the number of inflammatory cells the detection limits of the available in BAL may depend on time because they methods. The mechanism of recruitment are moving from one compartment to an- of eosinophils in the airways after EIA other, e.g., from blood to bronchial is not clear, but seems to suggest that exmucosa and to airway lumen during the ercise in these subjects may enhance airinflammatory process. In both Broide way inflammation. We think it unlikely and coworkers' (24) and Pliss and associ- that the increase of eosinophils after EIA ates' (25) studies, BAL was performed might have been a nonspecific phenomeduring EIA and compared with a BAL non resulting from acute disruption of performed during baseline conditions. In epithelial tight junction (26), thus proour study BAL, BL, and BB procedures voking an increased recovery of loosened were done when EIA had completely inflammatory cells. Under these circumresolved, and results were compared with stances, in fact, all inflammatory cell those obtained when a nonspecifically in- - types should have been found increased duced bronchospasm had also complete- in BAL after EIA and a larger number ly resolved. In this way, we attempted to of epithelial cells should have been reminimize any functional effect of bron- covered in BAL after EIA than in BAL chospasm on BAL and BL fluid compo- after MCh challenge. In a high proportion of allergic asthsition. We chose to perform BAL 3 h after exercise or MCh challenge because, matic subjects a late-phase bronchospasin a previous study (13), we observed a tic reaction occurs after inhalation of alsignificant increase of airway responsive- lergen (27). This late-phase response has ness 3 h after allergen challenge in pa- been shown to be associated with an eartients with late-phase asthmatic response, ly influx of eosinophils in the airway lusuggesting that an increased airway in- men and an increase of airway responflammation was already present at that siveness (9, 10). The existence of a latetime. Indeed, BB and BAL performed phase response after exercisein asthmatic thereafter showed a significant mast cell subjects is questionable (6, 7, 8, 28), and degranulation in bronchial mucosa (14) an increase of airway responsiveness folaccompanied, in patients with late-phase lowing EIA could not be demonstrated response, by an influx of eosinophils (10). (29, 30) even when a late-phase asthmatWe allowed a similar time between exer- ic response was reported (31). Moreover, cise challenge and BAL, BL, and BB there are no studies in which a possible studies in order to gather data that could link between airway inflammation and be compared with those after allergen the late-phase response after exercise was challenge with respect to mast cell de- investigated. We found that three of five granulation and recruitment of inflam- patients with 2070 or more eosinophils in matory cells in the airways. BAL after EIA had a marked delayed fall We have found that the percentage of of FEV 1 after exercise. However, two of eosinophils in BAL after exercise was them showed a similar delayed FEV 1 fall greater than after MCh challenge. How- and similar percentages of eosinophils in ever, when expressed as absolute num- BAL after MCh. This suggests that the bers the difference between eosinophils occurrence of a delayed bronchospasm in BAL after EIA and after MCh did not after exercise was not specific to EIA but reach statistical significance. This was more likely resulted from an instability likely related to a difference in the num- of lung function that may be present in ber of total inflammatory cells recovered patients with more pronounced eosinofrom BAL, which tended to be greater philic inflammation of airways. Indeed, after MCh than after EIA. In addition, unpublished data from our laboratory we have found a larger proportion of show that patients with 2070 or more eodegranulating mast cells in BB after EIA sinophils in BAL at baseline are those than after MCh. These results are in good with the highest degree of bronchial hyagreement with those of Pliss and associ- persensitivity, requiring more frequent ates (25), even if we could not find a con- medication. In this study we did not find sistent increase of mediators in BL. In- a relationship between the increase in eo-

sinophils in BAL and the development of the late-phase response similar to that observed after allergen inhalation. It is possible that the influx of eosinophils observed after EIA is not followed by their activation, which may be necessary for initiating the cascade of inflammatory events leading to the late-phase response. In conclusion, in the present study we have shown that EIA is associated with mast cell degranulation and eosinophil influx in the airway lumen. Furthermore, in some patients with EIA and chronic airway inflammation a delayed bronchoconstriction may be observed, but this does not seem to be a specific consequence of exercise. References 1. Chen WY, Horton DJ. Heat and water loss from the airways and exercise-induced asthma. Respiration 1977; 34:305-13. 2. Deal EC, McFadden ER, Ingram RH, Jaeger J J. Hyperpnea and heat flux: initial reaction sequence in exercise-induced asthma. J Appl Physiol 1979; 43:476-83. 3. McFadden ER, Lenner KA, Strohl KP. Postexertional airway rewarming and thermally induced asthma. J Clin Invest 1986; 78:18-25. 4. Deal EC, Wasserman S, Soter N, Ingram RH, McFadden ER. Evaluation of role played by mediators of immediate hypersensitivity in exerciseinduced asthma. J Clin Invest 1980; 65:659-65. 5. Lee TH, Nagakura T, Cromwell 0, Brown M, Causon R, Kay AB. Neutrophil chemotactic activity and histamine in atopic and nonatopic subjects after exercise-induced asthma. Am Rev Respir Dis 1984; 129:409-12. 6. Lee TH, Nagakura T, Papageorgiou N, Iikura Y, Kay AB. Exercise-induced late asthmatic reactions with neutrophil chemotactic. N Engl J Med 1983; 308:1502-5. 7. Foresi A, Mattoli S, Corbo G M, VergaA, Sommaruga A, Ciappi G. Late bronchial response and increase in methacholine hyperresponsiveness after exercise and distilled water challenge in atopic subjects with asthma with dual asthmatic response to allergen inhalation. J Allergy Clin Immunol1986; 78:1130-9. 8. Zawadsky DK, Lenner KA, McFadden ER. Reexamination of the late asthmatic response to exercise. Am Rev Respir Dis 1988; 137:837-41. 9. De Monchy JGR, Kauffman HF, Venge P, et of. Bronchoalveolar eosinophilia during allergeninduced late asthmatic reaction. Am Rev Respir Dis 1985; 131:373-6. 10. Rossi GA, Crimi E, Lantero S, et of. Late-phase asthmatic reaction to inhaled allergen is associated with early recrui tment of eosinophils in the airways. Am Rev Respir Dis 1991; 143:379-83. 11. Quanjer P. Standardized lung function testing. Report of wor king party on standardization of lung function tests-European community for coal and steel. ECCS. Bull Eur Physiopathol Respir 1983; 19 (Suppl 5:1-94). 12. Jones NL, Makrides L, Hitchcoch C, Chypchar T, McCartney N. Normal standards for incremental progressive cycleergometer test. Am Rev Respir Dis 1985; 131:700-8. 13. Brusasco V, Crimi E, Gianiorio P, Lantero S, Rossi GA. Allergen-induced increase in airway re-

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Airway inflammation and occurrence of delayed bronchoconstriction in exercise-induced asthma.

We studied nine asthmatic patients with a history of exercise-induced asthma (EIA) in order to investigate whether inflammatory changes in the airways...
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