![Inflammatory Response in 7 Mg N02·M· a Larger Proportion of the AM Were 3 BALF After Exposure to 7 Mg N02·M· [16]](http://data.docslib.org/img/e0a995e8b152e94fa9a138e3729b72f8-1.webp)
Eur Reaplr J 1991, 3, 332-339
Inflammatory cell response in bronchoalveolar lavage fluid after nitrogen dioxide exposure of healthy subjects: a dose-response study
T. Sandstrom•t, N. Stjernber~J", A. Eklund .. , M-C. Ledin·, L.. Bjermert, B. Kolmodin-Hedman·, K. Lmdstrom·, L. Rosenhallt, T. Angstromtt
Inflammatory cell response in bronchoalveolar lavage fluid after nitrogen • National Institute of Occupational Health, dioxide exposure of healthy subjects: a dose response study. T. Sandstrom, Medical Division, Umel, Sweden. N. Stjernberg, A. Eklund, M-C. Ledin, L. Bjermer, B. Kolmodin-Hedman, K. Lindstrom, L. Rosenha/1, T. Angstr6m. ' Dept of Thoracic Medicine, Karolinska Hospital, ABSTRACT: The combinatJon of environmental chamber exposure and Stockhoim, Sweden. broncboalveolar lavage (BAL) was used to study the effects of the • • Dept of Thoracic Medicine, Karolinska Hospital, ). common air pollutant nitrogen dioxide (N01 Eighteen healthy Stockholm, Sweden. nonsmokers were exposed to N01 during 20 mln in an exposure chamber during llgbt bicycle ergometer work. All subjects were examined with "Cytology Laboratory, University Hospital, UmeA, BAL at least 3 wks before exposure, as a reference. The subjects were Sweden. re-examined wltb BAL, in groups of eight, 24 h after exposure to 4, 7 and 10 mg N0 ·m·' (2.25, 4.0 and S.S ppm), respectively. An Correspondence: T. SandstrOm, National Institute of 1 inflammatory cell response was found after exposure to all concentra Occupational Health, Medical Division, Box 6104, tions. An Increase In the number of lymphocytes in BAL fluid was S-900 04 Umel, Sweden. observed after 7 and 10 mg·m~ (p<0.05 and 0.02, respectively). An Keywords: Air pollution; bronchoalveolar lavage; Increase In the number of mast cells, that appears to be dose-dependent, lymphocyte; lysozyme; macrophage; mast cell. was found after exposure to all concentrations. The proportion of lysozyme posltJve alveolar macropbages was elevated after exposure to Received: February 8, 1990; accepted after revision 7 mg·m·'. The Inflammatory mediators fibronectln, hyaluronan, August 23, 1990. angiotensin converting enzyme (ACE) and beta -mfcroglobulln were 1 unchanged by exposure. Due to the findings of Inflammatory cell changes This study was supported by grants from the Swedish far below the peak exposure limits for work places In Industrialized Heart Lung Foundation, the University of UmeA and countries, 9-18 mg·m·', the safety of these limits Is questioned. the Norrlands Gas AB Fund. Studies are In progress In our laboratory using BAL to evaluate the effects of repeated N01 exposure. Eur Respir 1., 1991, 4, 332-339.
Nitrogen dioxide (NOJ is a common air pollutant both occur after high N02 levels or long-term exposure [10, in community air in urban areas and in the indoor 11]. In animal studies increased numbers of neutrophils environment in industries using combustion processes. and macrophages [12-14], and in some investigations Due to its poor solubility it is to a large extent deposited also lymphocytes and mast cells [10, 15], have been in the peripheral air spaces [1, 2] where it acts on cells found in histological specimens and bronchoalveolar according to its oxidant and free radical properties [3]. lavage fluid (BALF).
Inhalation of relatively low concentrations of N02 may In man bronchoalveolar lavage (BAL) has only increase airway resistance in healthy subjects and recently been employed in investigating the effects of repeated exposure may give enhanced susceptibility to N02 inhalation. Recently, we showed an altered airway infections [4, 5]. Reduced diffusion capacity composition of cells in BALF as the time-course was for carbon monoxide has been reported after a investigated for the inflammatory cell response induced 3 3 single exposure to 9 mg N02·m· [6]. Exposure to by 7 mg N02·m· healthy subjects [16]. N02 exposure high concentrations may result in chronic obstructive has also been shown to cause an altered antiprotease lung disease, emphysema or bronchiolitis obliterans activity, alpha2-macroglobulin increase in BALF and [7]. impaired virus inactivation by human alveolar Morphological studies in animals have shown that macrophages (AM) [17-19). the most sensitive part of the lung to N02 exposure is This investigation was undertaken to examine the the transitional zone between the terminal bronchiole dose-response relationship between N02 exposure in man and the alveolar duct [8, 9]. Alveolar effects mainly during light bicycle ergometer work and changes in the N0 EPFECI'S INVESTIGATED WITH BRONCHOALVEOLAR LAVAGE 2 333
BALF content of cells with regard to their concentra Bronchoalveolar lavage tion and function. This was measured as phagocytic ability and release of soluble components considered as All bronchoscopies were performed by the same a reflection of cellular activity. investigator. Atropine was given s.c. prior to the examination and lidocaine 200 mg was used for topical anaesthesia. No sedative or other complementary Subjects and methods medication was given. The flexible fibreoptic bronchoscope (Olympus BF 1T or BF 1T10, Japan) was Subjects inserted through the mouth with the subject in the supine position. After careful wedging of the broncho Eighteen healthy nonsmoking male volunteers with a scope tip in a middle lobe bronchus, sterile phosphate mean age of 26 yrs (range 22-32 yrs) participated. buffered saline pH 7.3 (PBS-A) at 37°C was infused in None of them had experience symptoms of airway four aliquots of 60 ml and gently suctioned back after infection for at least six weeks prior to the study or each infusion to a siliconized container placed in ice had a history of asthma. Informed consent was obtained water. and the study was approved by the local Ethics Committee. Lavage fluid analysis
Design of the study Cell counts. The chilled BALF was filtered through a nylon filter (pore diameter 100 J.tm, Syntab Product AB, Malmo, Sweden) and centrifuged at 400 g for 15 min. Flexible fibreoptic bronchoscopy with BAL was The cell pellet was resuspended in balanced salt solu performed in all subjects in order to obtained reference 6 BALF. In the following exposure series, each subject tion to a concentration of 10 cells per ml. The total number of cells in the lavage fluid was counted in a was exposed to at least one N0 concentrations. Six of 2 Biirker chamber. Cytocentrifugal specimens with Sx104 the subjects were exposed to two N0 concentrations, 2 non-epithelial cells per slide were prepared using a with a time interval of at least 3 wks. The environmental Cytospin 2* (Shandon Southern Instruments Inc., chamber exposure was performed with 4, 7 and 10 Sewickly, PA, USA) 100 rpm (96 G) for 15 min. Slides mg·m·3 (2.25, 4.0 and 5.5 ppm) for 20 min. Eight were stained according to May-Griinwald-Giemsa for subjects were studied at each concentration, according standard cell differential counts and two hundred cells to a standardized protocol [20]. During the last 15 min per slide were counted. Mast cells were counted on 10 of the exposure the subjects were working on a bicycle visual fields at 16x magnification on slides stained with ergometer with a work load of 75 W. A second BAL acid toluidine blue and counterstained with Mayer's acid was performed 24 h after the end of N0 exposure, which 2 haematoxylin [21]. Lysozyme positive AM were was at least 4 wks after the first lavage. Immediately demonstrated with Lysozyme* antibody using an before and after exposure, and before the post-exposure immunoperoxidase technique (Dakopatts A/S, BAL, forced expiratory volume in one second (FEV ) 1 Copenhagen, Denmark). Two hundred alveolar and forced vital capacity (FVC) were recorded using a macrophages (AM) were counted. The ratio helper Vitalograph* spirometer (Vitalograph Ltd, Buckingham, inducer/cytotoxic-suppressor (CD8•/CD4•) T-cells was UK). counted on 200 lymphocytes using light microscopy using the Simultest T Helper/Suppressor Test11 (Becton Dickinsson AB, Stockholm, Sweden). Nitrogen dioxide exposure Non-cellular components. Albumin was measured with The exposure chamber measured 3.20 x 2.00 x 2.20 rocket electroimmunoassay according to LAuRELL [22]. 3 1 m with an air volume of 14.1 m • It was built with Concentrations were expressed in mg·l· • anodized aluminium, with windows in one wall. Fibronectin was analysed by a double-sandwich Ambient air was drawn continuously through the enzyme-linked immunosorbent assay (ELISA) developed 1 chamber at 150 m3·h· , resulting in one air exchange by Bu.sCHKE et al. [23]. The aliquots of BAL fluid were approximately every 6 min. During the exposures, the defrozen at 37°C. Briefly, microtitre plates (NUNC, chamber air temperature was kept at 21 °C and the Denmark) were coated with rabbit-antihuman fibronectin relative humidity at 45%. The desired N02 concen antibodies (Dakopatts, Denmark) in phosphate-saline trations in the exposure chamber were achieved by buffer, pH 7.2. After addition of unconcentrated BAL adding a gas stream from a gas tube containing 1% fluid samples the plates were incubated at room N02 gas into the chamber air inlet. The gas flow temperature for 2 h. Horse radish peroxidase-labelled was maintained evenly by a Mass Flowmeter, Brooks antihuman fibronectin (Dakopatts) was added as second 5850 Tr, Brooks Instruments, Vendeen, The Netherlands. antibody and the plates were incubated for 1 h. The The chamber air was continuously analysed with a amount of bound peroxidase, which is proportional direct indicating equipment (Nitrogen oxides analyzer to the amount of fibronectin in the sample, was 8440 B, Monitor Labs, San Diego, Ca, USA). measured by analysing the enzymatic activity on 334 T. SANDSTROM ET AL.
orthophenylendiamine. Serum fibronectin of nephelom (Pharmacia Diagnostics AB, Sweden). The detection etric quality from Behringer-Hoechst (Frankfurt am limit was 40 f,.lg·J·1• Concentrations were expressed in Main, GFR) was used as standard. Concentrations of f,.lg·l-1 and the detection limit was 40 f,.lg·l· 1• The fibronectin were expressed in f,.lg·l·1• The detection limit intra- and interassay variability was 6.2% and 7.0%, was 10 f,.lg·l·1• Intra- and interassay variability were 3.7% respectively. and 6.4%, respectively. Macrophage phagocytosis was measured as percent Angiotension-converting enzyme (ACE) activity was age engulfment positive cells, using a glass surface analysed in principle according to RYAN et al. [24). It adherence method as described previously [27). The was measured in unconcentrated fluid with a commer method was modified according to the use of cial radioassay (Ventrex Lab Inc; Portland, Maine, USA) bronchoalveolar lavage cells. Briefly, 2x10' cells in 200 using 3H-benzoylphenylalanyl-alanyl-proline as sub f..ll medium containing 10% pooled human AB+ serum strate. Aliquots of the subcellular fractions containing were allowed to adhere to a glass surface for 30 min in 1-4 f,.lg protein were mixed with the substrate in cell culture conditions. After rinsing the non-adherent hydroxyethylpiperazine ethanesulphonic acid (HEPES) cells away, yeast cells labelled with FITC and opsonized NaCl buffer pH 7.4 and incubated for 15 min at 37°C. with human serum where added to the slides (2.5x107 The amount of ACE was measured after extraction of yeast cells in PBS buffer solution, pH 7.4). After 30 the acidified reaction mixtures with Ventrex scintilla min the phagocytosis was stopped by dipping the slides tion fluid. The analysis was also performed in the into ice-cold PBS containing 1 mM edetic acid (EDTA). presence of the ACE inhibitor captopril and the differ The fluorescence of non-ingested yeast cells were ence in enzyme activity was calculated. The enzyme quenched by dripping toluidine blue in saline (1 activity (U) was expressed as that quantity of enzyme mg·m·1, pH 4.7) onto the slides. Yeast cell adherence that hydrolyses the substrate at an initial rate of 1% per (attachment) to a macrophage was defined as visible min at 37°C. contact between a brown yeast cell and a macrophage. Hyaluronan was analysed in duplicate in uncon Engulfment was defined as the presence of fluorescent centrated BALF by a radiometric assay using the yeast cells within a macrophage. Pharmacia Research Kit according to principles outlined previously [25, 26]. Concentrations were expressed in f,.lg·l· 1• The detection limit was 10 f,.lg·l·1• Intra- and Statistics interassay coefficients of variation were 4.0% and 6.4%, respectively. Values are given as medians and interquartile ranges
Beta2 -microglobulin was measured in duplicate, in (Ql-Q3) since the observed values were not normally unconcentrated fluid with a radioimmunoassay distributed. Wilcoxon's non-parametric signed rank test
Table 1. - Cell numbers and alveolar macrophage phagocytic capacity in BALF 24 h after controlled chamber 3 exposure with 4, 7 and 10 mg N02·m·
Total cells Lymphocytes Mast cells AM LZM+AM AM 7 7 x107·t·1 107·/'1 % x1Q4·[·1 % x10 ·/'1 % x10 ·1"' %of % macrophages engulfment positive cells
Before exposure 7.4 0.4 6 1.9 0.05 6.8 92 0.8 8 88 (n=18) 6.4-12 0.3-{).7 5-9 0-4.8 {}-{).03 5.6-11.5 89-92 0.3-1.5 6-18 83-92
24 h after 9.6 1.0 10 12.5• o.n• 8.4 88 1.0 10 87 3 7.1-13.9 83-90 0.7-1.7 83-92 4 mg N02·m· 8.2-15.2 0.5-1.7 5-13 5-17.2 0.07-0.11 8-12 (n=8)
24 h after 7.9 1.4• 16• 11.7• 0.14••• 6.4 79• 0.8 14• 87 3 7 mg N02·m· 6.7-9.0 0.7-2.0 12-24 6.9-13.6 0.11-0.22 5.(}-7.6 68-85 0.7-1.1 11-18 85-89 (n=8)
24 h after 9.6 1.4 .. 12• 19.7• 0.17 .. 7.8 85• 1.0 12 87 N0 3 8-20 12.2-27.8 5.2-14 0.7-1.9 8-18 83-89 10 mg 2·m· 5.6-15.4 0.7-2.1 0.07-0.27 78-90 (n=8)
Values are expressed as medians with interquartile ragnes. •: p<0.05; ..: p<0.02; •••: p
N02 concentrations (p<0.05, p<0.01 and p<0.02, 2 respectively). The response increased by the dose as seen in figure 1. The relative changes in cell ppm numbers after exposure are given in figure 1, calculated 0 as ratio after/before exposure. Total cell number, 0 2 4 I 10 mg·m·3 neutrophils, eosinophils, epithelial cells, and T-helper N02 concentration inducer/supressor-cytotoxic cell ratio were unaffected Fig. 1. - The relative changes in total cell counts for mast cells, lymphocytes and alveolar macrophages 24 h after exposure to 4, 7 after all exposure concentrations (data not shown). The phagocytic activity of AM in vitro was 88% and 10 mg N02·m·' (2.25, 4.0 and 5.5 ppm). The values are given as median ratio of cell counts after/before exposure:tsEM. •: p Table 2. - Soluble components in BALF before and 24 h after N02 exposure Before N02 concentrations exposure 4 mg·m·' 7 mg·m·' 10 mg·m·' n=18 n=8 n=8 n=8 Fibronectin Median 74 76 70 74 ~o~g·[·l QI-QJ 59-101 55--80 58-108 55-112 Hyaluronan Median 15 17 18 14 J.lg•/'1 Ql-03 11-25 14-24 13-26 12-27 13 -micro- Median 138 182 176 141 globulin Ql-Q) 105-165 148-214 166-215 129-175 ~g·tl ACE Median 9066' 4885" 9999 15544 u Qt-QJ 5783-17136 4464-5706 6064-18379 10636-17695 Albumin Median 46 48 48 54 mg·L·t Qt-QJ 31-50 33-55 42-55 47-63 : tt; 0 1-03 first-third quartile; ': n=16; n=6. BALF: bronchoalveolar lavage fluid; ACE: angiotensin converting enzyme. 336 T. SANDSTROM ET AL. Non-cellular components. The albumin, hyaluronan, presently unclear whether these mast cells are active in the immune response or not. Support for an active role fibronectin, beta2-microglobulin concentrations and ACE activity are shown in table 2. No significant changes has previously been presented after observation of were detected after exposure. antihistamine inhibition of increased airway resistance after N02 exposure (36]. Furthermore, N02 induced degranulation of mast cells has been observed in a Discussion morphological study in rodents [15]. Considering the present and previous investigations, In the present study the N02 concentration range, 4- we can conclude that short-term exposure to 3, moderately high N0 concentrations in man does not 10 mg N02·m· chosen to reflect conditions encoun 2 tered mainly in industrial work places. We found an induced increase in the number of neutrophils, as altered composition of the population of cells recovered reflected in BALF. This is surprising considering that in the lavage fluid all through the investigated neutrophilia has been a consistent finding in rodents concentration interval, i.e. even far below the peak after N02 exposure [12, 13] and increased neutrophil 3 exposure limits for N02 in work places, 9-8 mg·m· chemotactic activity has been demonstrated after in (5-10 ppm) in different countries. The increase in vitro challenge with N02 of human AM [37]. Varying cell numbers appears to be dose-dependent for mast susceptibility in different species, differences in cells, but cannot be determined conclusively for exposure duration and concentration, and various lymphocytes. The pattern for AM stained positive for conditions in in vitro test situations may account for lysozyme is not conclusive in the present material. The the diverging results. latter may be a consequence of the relatively low number In addition to neutrophilia, increase in AM numbers of subjects investigated at each concentration. Involve has been the most pronounced inflammatory cell ment of all three cell types in the toxicological response to NO in BALF and lung tissue in animal reaction in the human lung to N02 is further supported studies [12, 13]. ~e were unable to detect any significant by a parallel study in our laboratory investigating increase in the total AM number, but after exposure to 3 the time-kinetics of the inflammatory response in 7 mg N02·m· a larger proportion of the AM were 3 BALF after exposure to 7 mg N02·m· [16]. In the lysozyme positive. This increase in the intracellular earliest N0 study using BALF analyses in man. concentration of lysozyme has previously been shown 2 Frampton and eo-workers [19] reported no changes in to occur after a variety of stimuli in man including S02 3 cell numbers after 1.1 mg·m· • MosHENIN and GEE [17] exposure [29, 30, 38, 39]. It is consistent with recent were also unable to detect any significant changes in results from a time-kinetic study, where the increased 3 the cell populations recovered in BALF after 7 mg·m· • proportion of lysozyme positive AM persisted 72 h after However, in that study the N02 exposed individuals were exposure [16]. The findings could be caused by an cod to a small separate control group. Consequently, enhanced production of lysozyme in the AM or be due interindividual differences in cell numbers may well have to an inhibited secretory process after exposure. influenced the results. It should, furthermore, be noted Increased susceptibility to viral infections following that mast cell and lysozyme stains were not applied in NO exposure has been proposed in epidemiological any of these studies. studies [4, 5]. This corresponds with in in vitro tests In the present study an increase in the lymphocyte with animal [40, 41] and human AM [19] demonstrat population, expressed as total lymphocyte number and ing N02 induced decrease in phagocytosis and inacti percentage of recovered cells was a significant response vation of microorganisms. FRAMPTON and eo-workers [19] to N02 exposure. However, no change in the ration of found this to occur after an N02 concentration as low as T helper-inducer/suppressor-cytotoxic cells was seen, 1.1 mg·m·3• In this study we investigated whether in vivo in agreement with previous BAL studies with N02 and exposure to N02 would impair phagocytosis in vitro by human AM, but were unable to detect any significant S02 in man [16, 29, 30]. The concentrations of betaz-microglobulin, a potential marker ofT-cell activa effect. This could have different explanations. Firstly, tion, was also unaffected after the exposures. Thus, the human AM phagocytosis may not be affected by N02 expansion of the lymphocyte population does not seem exposure in vivo, at least not in the investigated con to be accompanied by a simultaneously enhanced activity centration interval. Secondly, impaired phagocytosis of the cells. possibly occurs only after repeated exposures. Thirdly, Mast cells constitute 2% of the bronchial epithelial the in vitro test situation per se may have stimulated cells in human lung [31] and have previously been the macrophages so that an impairment in phagocytosis associated mainly with immunoglobulin (IgE) mediated present in vivo was masked in vitro. allergic reactions and asthma [32, 33]. Recent studies Other signs of altered AM activity were also lacking have focused attention on other properties of this cell, as the concentrations of ACE and fibronectin in the which appears to be involved in conditions with alveolitis lavage fluid were normal. ACE has been demonstrated tissue damage and development of fibrosis [34, 35]. to be release by AM in an activated state [42, 43] and The observed increase in the number of mast cells is an increased concentration in BALF has been reported consistent with our ~arlier findings after exposure to in conditions with alveolitis [44 ]. Fibronectin is also both N0 and S0 [16, 29, 30]. It remains to be known to be produced by AM in various lung disorders 2 2 established what significance this has and its is e.g. in association with fibrosing processes [26, 45]. 337 N02 EFFECfS INVESTIGATED WITH BRONCHOALVEOLAR LAVAGE It has, furthermore, been shown to be a sensitive marker nitrogen dioxide toxicity. Am Rev Respir Dis, 1978, 118, of irradiation induced damage to the respiratory 1061-1090. epithelium [46]. The discrepancy in the findings of an 4. Speizer FE, Ferris B, Bishop YMM, Spengler J. - Res increased portion of lysozyme positive AM and piratory disease rates and pulmonary function in children associated with N0 exposure. Am Rev Respir Dis, 1980, 121, unaffected phagocytic capacity and mediator 2 3-10. production does not support Lhe hypothesis that positive 5. Shy CM, Creason JP, Pearlman Me, McClain KE, Benson lysozyme staining of AM reflects a state of activation. FB, Young MM. -The Chattanooga school children study: Hyaluronan is a glycosaminoglucan with a pronounced effects of community exposure to nitrogen dioxide II. ability to immobilize water [47]. It has been demon Incidence of acute respiratory illness. J Air Pollut Control strated to correlated with reduced alveolar-capillary Assoc, 1970, 20, 582-588. diffusion capacity for carbon monoxide in alveolitis 6. von Nieding 0, Krekeler H, Fuchs R, Wagner M, Koppenhagen K. -Studies of the acute effects of N0 on lung [48, 49]. 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Int Arch Allergy Appllmmunol, 1987, 82, 298-301. 36. von Nieding G, Wagner M, Krekler H, Loellgen H, Fries Reponse des cellules inflammatoires du liquide de lavage W, Beuthan A. - Controlled studies of human exposure to broncho-alveolaire apres exposition de sujets sains au bioxyde NO~ • single and combined action of o, and S02 lnt Arch d'azote. T. Sandstrom, N. Stjernberg, A. Eklund, M-C. Ledin, Occup Environ Health, 1979, 3, 19:l-210. L. Bjermer, B. Kolmodin-Hedman, K Lindstrom, L. Rosenhall, 37. Bart F, Aerts F, Deroubaix C, Wallaert B, Voisin C. - T. Angstrom. In vitro short-term exposure to 0.2 ppm nitrogen dioxide, RESUME: Les effets de polluant tr~s commun de !'air ambient, le N0 , ont ete etudies par la combinaison de evidence for an increased generation of superoxide anion and 2 neutrophil chemotactic activity. Am Rev Respir Dis, 1988, 137, !'exposition dans une chambre environnementale et le lavage A168. broncho-alveolaire (BAL). Dix-huit sujets sains non-fumeurs 38. Bjermer L, Back 0 , Roos G, Thunell M. - Mast cells furent exposes a N02 pendant 20 minutes dans une chambre and lysozome positive macrophages in bronchoalveolar lav d'exposition au cours d'un travail leger a la bicyclette age from patients with sarcoidosis. Acta Med Scand, 1986, ergometrique. Tout les sujets ont ete examines par lavage au 220, 161-166. moins trois semaines avant !'exposition, a titre de reference. 39. Leakes ES, Myrvik QN.- Changes in the morphology Les sujets ont ete reexamines par lavage broncho-alveolaire and lysozyme content of free alveolar cells after intravenous dans des groupes de 8x24 heures apr~s exposition a 4, 7 et N0 EFFECTS INVESTIGATED WlTif BRONCHOALVEOLAR LAVAGE 2 339 10 mg N02·m·' (2.25, 4.0 et 5.5 ppm), respectivement. Une l'hyaluronan, !'enzyme de conversion de l'angiotensine (ACE) reponse cellulaire inCJammatoire a ete observee apr~s et la betafmicroglobuline, restent inchanges apres exposition. exposition A toutes les concentrations. Une augmentation du Vu nos observations concemant les modifications des cellules nombre de lymphocytes dans le liquide de BAL a ete observee inflammatories observees en dessous des limites tolerees pour apr~s 7 et 10 mg·m·) (p<0.05 et 0.02, respectivement). Une les pointes d'exposition dans les lieux de travail des pays augmentation du nombre de mastocytes, qui s'avere industrialises (9-18 mg·m·3), l'ont met en question la securite dose-dependante, est observee apres exposition ~ toutes les de ces limites. Des etudes sont poursuivies dans notre concentrations. La proportion de m.acrophages alveolaires laboratoire au moyen du BAL, pour evaluer les effets de lysozome positifs augmente apres exposition A 7 mg·m·3• Les !'exposition repetee au NO . mediateurs inflammatoires que sont le fibronectine, Eur Respir J., 1991, 4, 33}-339.