Bronchoprotective effects of atrial natriuretic peptide against propranolol-induced bronchoconstriction after allergic reaction in guinea pigs

OBJECTIVE: To study the effects of intravenous atrial natriuretic peptide (ANP) on antigen-induced bronchoconstriction, propranolol-induced bronchoconstriction (PIB) after antigen challenge, and histamine-induced bronchoconstriction in guinea pigs. METHODS: Allergic bronchoconstriction was evoked by inhalation of ovalbumin (OA) and PIB was caused when 10 mg/mL of propranolol was inhaled 20 min after OA challenge in passively sensitized and artificially ventilated guinea pigs. 25, 50, 100 and 200 microg/mL of histamine were inhaled for 20 s at 5-min intervals in non-sensitized guinea pigs. RESULTS: Pretreatment with ANP in doses of 0.1 and 1.0 nmol/kg injected intravenously 15 min after antigen challenge reduced PIB in a dose-dependent manner, and 5 min before antigen challenge significantly attenuated PIB but not antigen-induced bronchoconstriction. Intravenous ANP significantly reduced bronchial responses to increasing concentrations of inhaled histamine in a dose-dependent manner. CONCLUSION: These results suggest that ANP possesses protective effects against propranolol-induced and histamine-induced bronchoconstriction, albeit by a non-specific mechanism in guinea pig in vivo.     

Inhibition of antigen-induced late asthmatic response and bronchial hyperresponsiveness by cyclosporin A

We have previously demonstrated that Cyclosporin A (CyA), a T lymphocyte-selective immunosuppressive agent, reduced the delayed-phase bronchial eosinophil infiltration after antigen challenge in a guinea pig model of asthma. In the present study, we studied the effects of CyA on antigen-induced late asthmatic response (LAR) and bronchial hyperresponsiveness following LAR. Guinea pigs immunized by repeated exposure to aerosolized ovalbumin (OA) were intravenously given metopirone, a cortisol synthesis inhibitor, 24 hours before and 30 minutes before antigen challenge, and to prevent death from immediate severe bronchoconstriction, chlorpheniramine maleate was also injected. After antigen challenge with high dose of OA, LAR occurred in twelve of fifteen animals (80%) and the bronchial responsiveness to acetylcholine was significantly increased. However, when guinea pigs were treated with CyA from the beginning of immunization period, the development of LAR was completely inhibited, although similar magnitude of immediate bronchoconstriction was observed, and a subsequent increase in bronchial responsiveness was partially but significantly blocked. Since CyA has been shown to suppress activation of guinea pig T lymphocytes and their production of lymphokines, these results suggest that T cell factor(s) may be important for the elicitation of LAR and the antigen-induced bronchial hyperresponsiveness.     

Significant changes of bronchial responsiveness to methacholine after early asthmatic reaction to toluene diisocyanate (TDI) in a TDI-sensitive asthmatic worker

Current asthma is often diagnostically excluded by the presence of normal bronchial responsiveness. We report on a TDI-induced occupational asthma patient with normal bronchial responsiveness. He had suffered from shortness of breath during and after TDI exposure for several months. His initial methacholine bronchial challenge test showed a negative response. The bronchoprovacation test with TDI showed an isolated immediate bronchoconstriction. The following methacholine bronchial challenge tests revealed that the bronchial hyperresponsiveness developed seven hours after the TDI challenge (methacholine PC20:5.1 mg/ml), progressed up until 24 hours, and returned to normal on the seventh day. This case provides evidence that the response of the airway to TDI may not always be accompanied by bronchial hyperresponsiveness to methacholine. Screening programs utilizing methacholine challenges may not always identify TDI-sensitized asthmatic workers.     

Inhibitory effect of FK-506 on the development of late asthmatic response and on the increased bronchial responsiveness]

We examined the effects of FK-506, a potent immunosuppressive agent, on the development of late asthmatic response (LAR) and on the increased bronchial responsiveness to acetylcholine following LAR in guinea pig model of asthma. Guinea pigs sensitized by repeated inhalation of ovalbumin (OA) were intravenously given metopiron 24 hours before and 30 minutes before antigen challenge and to prevent death from immediate severe bronchoconstriction, chlorpheniramine maleate was also injected. When we defined LAR as the responses with a two-fold increase in respiratory resistance during the late phase of antigen challenge, twelve out of fifteen control animals demonstrated apparent LAR. However, when guinea pigs were treated with FK-506 from the beginning of immunization period, the development of LAR was completely inhibited, although similar magnitude of immediate bronchoconstriction was observed, and a subsequent increase in bronchial responsiveness was significantly blocked. We also measured bronchial responsiveness to acetylcholine before, 24 and 72 hours after antigen challenge. FK-506-treated animals inhibited an increase in bronchial responsiveness to acetylcholine. These results suggest that the involvement of cell-mediated immunity may be important in the development of LAR and an increase in bronchial responsiveness.     

Changes in sputum cell counts after exposure to occupational agents: what do they mean?

BACKGROUND: Exposure to occupational agents can induce eosinophilic inflammation in subjects with occupational asthma (OA). It might also induce nonspecific changes in airway inflammation in subjects without OA. OBJECTIVES: We sought to investigate the changes in airway inflammation induced by exposure to occupational agents in subjects with and without OA and to determine which changes in sputum eosinophil numbers and bronchial responsiveness to methacholine should be regarded as clinically significant for predicting a 20% fall in FEV(1). METHODS: We performed specific inhalation challenges (SICs) in 3 groups of subjects: subjects reporting a history consistent with OA with a positive SIC response (n = 17); subjects reporting a history consistent with OA with a negative SIC response (n = 14); and asthmatic subjects without any history of OA (n = 10). Induced sputum and methacholine challenges were performed at the end of the control day and again at the end of the last day of exposure; the last day of exposure was always performed in the laboratory. RESULTS: There was an increase in median sputum eosinophil and neutrophil numbers in subjects with positive SIC responses. Cell counts remained unchanged after exposure in asthmatic subjects without OA. A combination of a greater than 0.26 10(6)/mL increase in sputum eosinophil numbers and a decrease in the concentration of methacholine inducing a 20% fall in FEV(1) of at least 1.8-fold compared with baseline values predicted a 20% fall in FEV(1) in 96% (95% CI, 70%-99%) of patients. CONCLUSION: Exposure to occupational agents per se does not induce airway inflammation. Changes in both sputum eosinophil counts and methacholine responsiveness are satisfactory predictors of a significant bronchial responsiveness to occupational agents.     

In vivo airway eosinophil accumulation induced by polymyxin-B reduces bronchial responsiveness in guinea pigs

BACKGROUND: Chronic desquamative eosinophilic bronchitis and bronchial hyperresponsiveness have been considered essential for bronchial asthma. However, it has not been studied whether airway eosinophils enhance or inhibit bronchial responsiveness in vivo. OBJECTIVE: This study was conducted to elucidate the influence of airway eosinophil accumulation on bronchial responsiveness in vivo. MATERIALS AND METHODS: Guinea pigs were transnasally treated with 75 microg/kg of polymyxin-B or vehicle twice a week for a total of 3 weeks. Guinea pigs were surgically cannulated and artificially ventilated 24 h after the last administration of polymyxin-B or vehicle. Ten minutes after the installation of artificial ventilation, ascending doses of methacholine, acetylcholine or histamine were inhaled for 20 s at intervals of 5 min. Subsequent study was conducted 20 min after treatment of 60 mg/kg of indomethacin in the same manner. Final study was conducted in naive guinea pigs after single inhalation of 75 microg/mL of polymyxin B. RESULTS: The proportion of eosinophils in bronchoalveolar lavage fluid significantly increased in guinea pigs treated with polymyxin-B compared with vehicle. Bronchial responsiveness to inhaled methacholine, acetylcholine and histamine was significantly decreased by the polymyxin-B treatment. This protective effect induced by polymyxin B was abolished by pretreatment of indomethacin. A significant increase in bronchial responsiveness was observed after a single inhalation of polymyxin B. CONCLUSION: These results suggest that in vivo airway eosinophils may reduce non-specific bronchial responsiveness through inhibitory or bronchoprotective prostanoids.     

Allergen-induced late bronchoconstriction and airway hyperresponsiveness in guinea pigs

Allergen-induced bronchoconstrictive responses were examined in a guinea pig model actively sensitized by the inhalation of aerosolized ovalbumin (OA), which showed reproducible late bronchial responses (LBR). OA-challenge was performed under cover of mepyramine through the inhaled route in spontaneous breathing without anesthesia. The respiratory resistance (Rrs) was measured by the oscillation method for 96 h after OA challenge. All of the 22 guinea pigs displayed immediate bronchial responses (IBR), followed by 2 or 3 phase LBR that peaked at 6-8 h and 24 h after OA challenge. Examination of bronchoalveolar lavage fluid (BALF) revealed a significant increase in neutrophils at 1 h (p less than 0.01) and eosinophils at 7 h and 96 h (each p less than 0.01) after OA challenge. OA-challenge induced airway hyperresponsiveness to histamine (p less than 0.01) at 96 h that was associated with a significant increase in eosinophils (p less than 0.01) in BALF compared with lavages at 7 h. Intravenous administration of 1% OA induced a significant increase of leukotriene (LT)B4, C4, D4 in tracheal lavage fluids at IBR phase in each control (p less than 0.01). The increases of Rrs in LBR were inhibited almost completely by the pretreatment of KC-404, which is reported to have antagonistic action against LTC4/D4. These results suggest that allergen-induced late broncho-constrictions accompanied by airway hyperresponsiveness in guinea pigs are associated with the extensive infiltration into the airway lumen of inflammatory cells, and are concerned with the release of LTs. We also suggest that this LBR guinea pig model is useful in studying the mechanisms of the occurrence of LAR in humans, and in evaluating the action of anti-allergic drugs in LAR.     

Bronchial hyperresponsiveness induced by chronic treatment with albuterol: Role of sensory nerves

BACKGROUND: It has recently been suggested that regular treatment with racemic beta(2)-adrenoceptor agonists might result in bronchial hyperresponsiveness (BHR) to a range of spasmogens, and this might be due to adverse effects of the distomer. OBJECTIVE: We sought to determine whether BHR induced by means of continuous exposure to racemic and S-albuterol was mediated by sensory nerves. METHODS: Naive or ovalbumin-sensitized guinea pigs were treated for 10 days with RS-, R-, or S-albuterol (1 mg.kg(-1).d(-1)) through subcutaneously implanted minipumps. Lung function was then determined in response to a number of spasmogens and assessed on the basis of an increase in total airway resistance. A separate group of animals were chronically treated with capsaicin (80 mg/kg) before the albuterol treatment. RESULTS: Treatment with RS- or S-albuterol increased airway responsiveness to bradykinin, leukotriene C(4), and capsaicin in naive guinea pigs (P <.05) and to histamine and ovalbumin in immunized guinea pigs (P <.05). Chronic treatment with capsaicin prevented the development of RS- and S-albuterol-induced BHR in these models. The bronchodilator efficacy of acute intravenously administered RS-albuterol was unaffected in RS-, R-, or S-albuterol-treated guinea pigs compared with in vehicle-treated animals. CONCLUSION: We have provided evidence demonstrating that continuous exposure to RS- and S-albuterol increases bronchial responsiveness to a range of stimuli, an effect not attributed to beta-adrenoceptor occupancy or desensitization. Furthermore, capsaicin-sensitive sensory nerves mediate the development of BHR, at least in part.     

Effects of aerosol administration of a thromboxane receptor antagonist (S-1452) on experimental asthma in guinea pigs

The importance of thromboxane A₂(TXA₂), one of the arachidonate metabolites, in the pathogenesis of bronchial asthma has been emphasized recently. Because aerosolized administration of antiasthmatic drugs is effective and safe, this study examined the effect of aerosolized TXA-₂ receptor antagonist (S-1542) on allergic bronchoconstriction in passively sensitized and mechanically ventilated guinea pigs. Under the cover of antihistamine, antigen-induced bronchoconstriction was markedly inhibited by pretreatment with aerosolized S-1452 inhalation in a dose-dependent manner. Although aerosolized S-1452 itself provoked weak bronchoconstriction for its partial agonist effect, bronchial responsiveness to inhaled histamine did not change 10 min after S-1452 inhalation. These results indicate that aerosolized S-1452 may be useful in treating bronchial asthma.     

Neurokinin-1 receptor antagonist inhibits short-term sulfuric-acid-induced airway hyperresponsiveness in sensitized guinea pigs

BACKGROUND: Tachykinins are involved in the development of bronchial inflammation and airway hyperresponsiveness (AHR); however, the role of the neurokinin-1 (NK(1)) receptor in acid-aerosol-induced bronchial impairment in asthmatic patients remains controversial. METHODS: To investigate the effects on the NK(1) receptor antagonist FK888 the neurokinin-2 (NK(2)) receptor antagonist SR48968 on sulfuric-acid (H(2)SO(4))-induced AHR in guinea pigs, specific airways resistance (sRaw) and airways responsiveness to methacholine (MCh) were measured before and after 6 h of exposure to H(2)SO(4) aerosol (pH 1.7, 82 mg/m(3)) in ovalbumin-sensitized guinea pigs. RESULTS: Airway responsiveness to MCh significantly increased (p<0. 05) after the exposure, however sRaw did not. Treatment with FK888 significantly inhibited (p<0.05) H(2)SO(4)-induced AHR in a dose-dependent manner, as did SR48968. CONCLUSIONS: These results suggest that not only NK(2) but also NK(1) receptors might have important roles in the development of acid-aerosol-induced AHR.     

Comparison between repeated antigen and acetylcholine-induced bronchoconstriction in development of airway wall thickness in a chronic guinea pig model of asthma]

Asthma is associated with a chronic remodeling of the airways, but it remains to be elucidated whether repeated bronchoconstriction (BC) influences any structural attributes of the lungs. To investigate the role of repeated BC on the morphology of the airways, we chronically exposed guinea pigs to an aerosol of acetylcholine (ACH) over a period of 6 months. The guinea pigs were challenged with either ACH or saline daily for 10 days and thereafter were challenged once a week. To compare repeated BC with a chronic model of allergic inflammation, an additional group of animals were sensitized to aerosolized ovalbumin (OA) and subsequently exposed with OA. The airway wall area and basement membrane (BM) thickness were measured by standard morphometric techniques. Both airway wall area and BM thickness were significantly increased in OA exposed guinea pigs compared with both saline control and AHC-exposed guinea pigs. Our results suggest that persistent bronchoconstriction and/or allergic airway inflammation in asthmatics may contribute to the thickness of the airway wall observed in this disorder. However, repeated acute bronchoconstriction events may not contribute to the increase in the airway wall or BM thickness.     

Sodium cromoglycate attenuates pulmonary inflammation without influencing bronchial responsiveness in healthy subjects exposed to organic dust

BACKGROUND: Inhalation of organic dust from a pig house induces airway inflammation and increases bronchial responsiveness to methacholine in healthy subjects. OBJECTIVE: To study whether sodium cromoglycate influences the airway inflammatory reaction and the increase in airway responsiveness induced by inhalation of organic dust. METHODS: Bronchoalveolar and nasal lavages, and bronchial methacholine challanges were performed and blood samples were drawn in 32 healthy subjects before and after exposure to dust in a pig farm. Sodium cromoglycate was inhaled (20 mg, twice a day) and administered intranasally (5.2 mg, twice a day) by 16 and a corresponding placebo was given to the other 16 healthy controls for two weeks prior to exposure. RESULTS: Exposure induced a significant increase in inflammatory cells and soluble components (pro-inflammatory cytokines, inflammatory mediators) in bronchoalveolar and nasal lavage fluid in both groups. The increase in neutrophils, interleukin (IL)-6 and tumour necrosis factor (TNF)-alpha as well as myeloperoxidase and soluble intracellular adhesion molecule (ICAM)-1 in bronchoalveolar lavage (BAL) fluid was significantly reduced by treatment with sodium cromoglycate. Although sodium cromoglycate inhalation largely influenced a variety of inflammatory indices in bronchoalveolar lavage fluid it had no effect on the increase in bronchial responsiveness to methacholine. CONCLUSION: Sodium cromoglycate alters the airway inflammatory response to inhaled organic dust without influencing the dust-induced increase in bronchial responsiveness to methacholine.     

Repeated aeroallergen challenge induces lung dysfunction but not bronchial hyperresponsiveness in conscious guinea pigs

Adult male Hartley-strain guinea pigs were sensitized by 10 min exposure to aerosolized 1% ovalbumin (OA; 10 mg/ml in normal saline containing 4% heat-killedB. pertussis vaccine and 0.02% antifoam B emulsion). One week after sensitization, animals were placed in an exposure chamber and challenged (nebulized OA 0.5%) until each animal showed labored breathing. Maximal exposure time was 10 min. Diphenhydramine (20 mg/kg, i.p.) was given 1 h before each OA challenge to protect the animals from bronchospasmic death. Antigen challenge was repeated twice a week for 2 weeks. The specific airway resistance (sR _aw) changes in response to increasing concentrations of aerosolized acetylcholine (Ach) were determined. The data obtained in this study demonstrated that repeated antigen challenge produced a significant bronchial tone i.e. an increase in sR _aw and a decline in specific airway conductance (sG _aw) and failed to induce bronchial hyperreactivity to aerosolized acetylcholine (Ach) in conscious guinea pigs.     

Impaired beta-adrenoceptor function, increased leukocyte respiratory burst, and bronchial hyperresponsiveness.

Inflammatory processes have potential importance in the pathogenesis of bronchial hyperresponsiveness and asthma. Because beta-adrenoceptor function may be impaired in asthma, we studied regulation of the leukocyte respiratory burst using blood samples from subjects with bronchial hyperresponsiveness to methacholine. Leukocytes from hyperresponsive subjects were less responsive to the beta-agonist isoproterenol than were leukocytes from healthy control subjects. The magnitude of the respiratory burst was increased in cells from hyperresponsive subjects and correlated with the degree of methacholine responsiveness. These results demonstrate that peripheral leukocytes reflect a functional impairment in beta-adrenergic responsiveness that parallels airway hyperresponsiveness. Because untreated subjects demonstrated a reduction in beta-adrenergic response, the impairment in beta-adrenoceptor function was not a result of drug therapy and may be associated with the pathogenesis of asthma.     

Maximal forced expiratory maneuver to measure airway obstruction in allergen challenged mice

Our purpose was to develop a method of using a maximal forced expiratory maneuver (MFEM) for the study of bronchoconstriction and bronchial hyperreactivity (BHR) induced in mice by ovalbumin (OA) inhalation challenge. Eight mice (group I) were sensitized and then provocated with OA. Pulmonary function testing (PFT) at baseline and after varying doses of acetylcholine challenge was performed. Eight weight-matched normal mice served as controls (group II). Pulmonary functions include MFEM, dynamic respiratory system compliance (Crs) and respiratory system resistance (Rrs). The results showed that mice treated with OA had worse PFTs than normal controls, characterized by lower MFEF 50%, FEV0.1 and Crs but higher Rrs. The OA-sensitized mice also had more severe bronchoconstriction in response to acetylcholine, characterized by greater decreases in MFEF 50%, FEV0.1 and Crs but a higher Rrs than the controls. There was a good correlation between PD20MFEF50%Ach and PD20FEV0.1Ach with PD20CrsAch and PD20RrsAch. In conclusion, the MFEM can be used to evaluate airway obstruction and BHR induced in mice by allergen challenge.     

Peptide leukotrienes mediate acetaldehyde-induced bronchial hyper-responsiveness in guinea-pigs

Background We previously reported that inhaled acetaldehyde, a metabolite of ethanol atid a maiti factor in alcohol-induced asthma, causes bronchial hyper-responsiveness (BHR) in asthmatics. However, the mechanisms are unclear. Objective The purpose of this study was to investigate a role of a peptide leukotriene (LT) in acetaldehyde-induced BHR. Methods Effects of LT antagonists, ONO-1078 (O.l-l.Omg/kg) and ICI-198, 615 (0.03–0.3 mg/kg), on acetaldehyde-induced bronchoconstriction and BHR to inhaled methacho-line were examined using a modified Konzett-Rössler method in guinea pigs. Results Aeetaldehyde at 0.8mg/ml, which failed to induce significant changes in Pao (pressure at the airway opening), enhanced an increase in Pao induced by subsequent inhalations of ascending doses (50–200 μg/ml) of methacholine. suggesting a potentiating effect of acetaldehyde oti bronchial responsiveness. Although ONO-1078 had no inhibitory effect on bronchoconstriction caused by ascending doses (5.0–20 mg/ml) of acetaldehyde, ONO-1078 and ICI-198, 615 reduced the acetaldehyde-induced BHR. Conclusion Acetaldehyde causes BHR via LT release in guinea-pigs.     

Bronchoconstriction due to isocapnic cold air inhalation minimally influences bronchial hyperresponsiveness to methacholine in asthmatic subjects

The aim of this study was to investigate if bronchial hyperresponsiveness to methacholine could be influenced by a previous bronchoconstriction due to isocapnic inhalation of cold air. Twelve adult asthmatic subjects in a clinical steady state were seen on four different days in a randomized way according to three different sequences. After assessment of spirometry, bronchial responsiveness to inhaled methacholine was determined on each occasion by the provocative concentration causing a fall of 20% in FEV1 (PC20). On two occasions, the methacholine test was preceded by the inhalation of dry cold air which caused significant (greater than 20% change in FEV1) bronchoconstriction. The methacholine test was performed after functional recovery. There was a significant (t = 2.53; p less than 0.05) but minimal (mean changes of 0.65 single two-fold concentration difference) reduction in PC20 after cold air inhalation. It is concluded that cold air-induced bronchoconstriction causes significant but minimal changes in bronchial responsiveness to methacholine in asthmatic subjects.     

Effect of ethanol on airway caliber and nonspecific bronchial responsiveness in patients with alcohol-induced asthma

No study has investigated the effects of ethanol on bronchial responsiveness in patients with alcohol-induced asthma, although acetaldehyde, which is a metabolite of ethanol and is thought to be a main factor in alcohol-induced asthma, causes both bronchoconstriction and bronchial hyperresponsiveness. The purpose of this study was to investigate the direct action of ethanol on the airway in patients with alcohol-induced asthma. First, we investigated the bronchial response to inhalation of ascending doses (5, 10, and 20%) of ethanol in nine patients with alcohol-induced asthma. Then, the bronchial responsiveness to methacholine was measured in 14 patients who were pretreated with saline or 20% ethanol in a double-blind, randomized, placebo-controlled, crossover fashion. Ascending doses of inhaled ethanol caused no significant changes in FEV₁. The methacholine concentrations producing a 20% fall in FEV₁ (PC₂₀-MCh) after 20% ethanol (0.769 mg/ml, GSEM 1.514) were significantly ( P = 0.0357) higher than those after saline (0.493 mg/ml, GSEM 1.368). This indicates that ethanol has a reducing effect on nonspecific bronchial responsiveness in patients with alcohol-induced asthma; this paper is the first report on the effects of ethanol on bronchial responsiveness.     

Role of sensory neuropeptides in post-allergic propranolol-induced bronchoconstriction in guinea pigs in vivo

Background Administration of propranolol can provoke bronchoconstriction in asthmatic patients. We hypothesized that such bronchoconstriction may result from the inflammatory mediators released by an allergic reaction. We recently developed a guinea pig model for propranolol-induced bronchoconstriction (PIB). Neuropeptides which are released from C-fibre nerve endings have been postulated to induce bronchial hyperresponsiveness through neurogenic inflammation. Objective The purpose of this study was to examine whether sensory neuropeptides are involved in the development of PIB after allergic reaction. Methods Propranolol at a concentration of 10mg/ml was inhaled 20min after antigen challenge in passively sensitized, anaesthetized and artificially ventilated guinea pigs. The animals were treated intravenously with a NK₁ and NK₂ dual antagonist, FK224, in a dose of 1 or 10 mg/kg or vehicle or a selective NK₁ antagonist, FK888, in a dose of 1 or 10mg/kg or vehicle 10min before or 15min after antigen challenge. Results Propranolol inhaled 20 min after antigen challenge caused bronchoconstriction. FK224 or FK888 administered 15 min after antigen challenge as well as 10 min before antigen challenge did not reduce the PIB. Conclusion We conclude that neuropeptides such as neurokinin A and substance P do not directly contribute to the development of PIB after allergic reaction.     

Ozone-induced airway hyperresponsiveness in guinea pigs

We studied the time related changes of airway hyperresponsiveness induced by ozone inhalation (2.9 ppm, 30 min) in guinea pigs. In unanesthetized and spontaneously breathing guinea pigs, the respiratory resistance was measured by a forced oscillation technique. The respiratory resistance and respiratory frequency were unchanged until 24 hours after ozone inhalation. In mechanically ventilated guinea pigs, airway responsiveness to inhaled methacholine was determined using a modification of the Konzett-R?ssler technique, and after methacholine challenge bronchoalveolar lavage (BAL) was performed for cell yield. At 1 hour and 3 hours after ozone inhalation, airway responsiveness was increased significantly, but returned to the control level at 6 hours. In the BAL fluid, there was a significant increase in neutrophils at 3 hours after ozone inhalation and thereafter. In the separated groups, before air or ozone inhalation, human serum albumin (HSA) was administered intravenously, and BAL was performed 1 hour after inhalation. In the ozone inhalation group, the concentration of HSA in BALF was increased significantly compared to the air inhalation group. These results suggest that airway hyperresponsiveness induced by ozone inhalation may occur before the influx of neutrophils into the airways and may depend on some structural changes such as submucosal and mucosal edema induced by the enhancement of capillary permeability.