Lung function and bronchial responsiveness after bronchoalveolar lavage and bronchial biopsy performed without premedication in stable asthmatic subjects.

We evaluated tolerance, safety, and effects on lung function and bronchial responsiveness of BAL (4 x 50 ml) combined with BB (three to five specimens) performed without premedication in 13 mild and stable asthmatics and eight healthy volunteers. All subjects tolerated bronchoscopy procedures well and without serious side effects. During procedures, no supplemental oxygen was administered and no ECG abnormalities were noted. The PEFR was measured before and immediately after bronchoscopy and at 5-min intervals up until recovery. The maximal percentage fall in PEFR after bronchoscopy was significantly greater in asthmatics (23.1 +/- 13.9 percent) compared to normal subjects (7.8 +/- 8.2 percent, p less than 0.01). Changes in PEFR returned to baseline values within 120 min in all asthmatics. The tcPO2 was recorded at baseline, during and after bronchoscopy. In both groups, a significant change in tcPO2 was measured during the infusion of BAL aliquots, and persisted throughout the procedure. A significant difference in asthmatics compared to healthy subjects was evident during BB and at the end of the procedure (p less than 0.05). In asthmatics, M challenge was performed on three different days over a three-week period prior to bronchoscopy, and was repeated at intervals of 2, 6, and 24 h following procedure. The PC20 M values measured before bronchoscopy were found to have a very high reproducibility (intraclass correlation coefficient = 0.93). The PC20 values measured during experiment times after bronchoscopy were not significantly different from baseline values. These data demonstrate that in mild and stable asthmatics, BAL combined with BB can be safely performed following administration of only local anesthesia. In carefully selected asthmatic subjects, transient bronchoconstriction and a lowering of oxygen tension can be induced by BAL and BB, whereas changes in bronchial responsiveness are more unlikely to occur.     

Early changes in T lymphocytes recovered by bronchoalveolar lavage after local allergen challenge of asthmatic airways.

To assess the role of T lymphocytes in the initiation of the allergic asthmatic response we have investigated T-cells subsets and their activation markers in bronchoalveolar lavage (BAL) fluid recovered 10 min after local challenge of the bronchial mucosa with allergen or saline. Endobronchial challenge was performed in 13 mildly atopic asthmatic patients (FEV1% predicted range, 78.2 to 116.5) and 10 normal volunteers. In all of the asthmatics but in none of the normal subjects allergen but not saline exposure resulted in visible bronchoconstriction. Analysis of BAL by flow cytometry showed no differences in the overall number of T cells (CD3+) and their CD4+ and CD8+ subsets per milliliter of BAL between the groups of normal subjects and asthmatics. However, within 10 min of allergen challenge, in the asthmatics but not in the normal subjects, there occurred a significant loss of CD3+ cells (p less than 0.01) comprising mostly CD4+ (p less than 0.05) but also CD8+ cells, with a consequent decrease in the CD4:CD8 ratio. At this early time point no differences in the extent of expression of the T-cell activation markers, IL-2 receptor, and HLA-DR were found. These results provide evidence to support a role of T lymphocytes early in the allergen-induced inflammatory response in asthma.     

Interleukin 16 and T-cell chemoattractant activity in bronchoalveolar lavage 24 hours after allergen challenge in asthma

IL-16 has been shown to be one of the earliest CD4(+) cell chemoattractants present in BAL 4-6 h after antigen challenge but little is known about its persistence and biological activity after 6 h. We determined the concentration of IL-16 using ELISA and the T-cell chemoattractant activity using a modified Boyden chamber assay in unconcentrated BAL fluid from 13 patients with mild asthma and 9 nonatopic control subjects at baseline and 24 h after segmental allergen or saline challenge. Furthermore, the percentage of IL-16-producing T cells was determined in the different samples of BAL fluid using a flow cytometric intracellular cytokine assay. Although no substantial levels of IL-16 protein were detectable in BAL fluid from control subjects and patients with asthma at baseline and after saline challenge, IL-16 concentrations were significantly elevated in patients with asthma after allergen challenge (median, 97 pg/ml; range, 38-362 pg/ml; p < 0.01). Furthermore, there was an increased T-cell chemoattractant activity after allergen challenge in patients with asthma (p < 0.01), which could be blocked by preincubation with anti-IL-16 antibodies and which correlated significantly with the IL-16 protein levels (R = 0.90, p < 0.01) and with the level of Fas ligand expression on BAL CD4(+) cells (R = 0. 80, p < 0.05). A high percentage (mean 70-90%) of CD4(+) and CD8(+) cells stained positively for IL-16 in both patients with asthma and control subjects without differences after allergen or saline challenge. These data demonstrate that the increased chemotactic activity for T cells in patients with asthma is mainly attributable to IL-16. Although T cells by themselves are able to produce IL-16, other cells, such as epithelial cells, have to be considered as further sources for this cytokine in patients with asthma.     

Alterations in bronchoalveolar lavage fluid but not in lung function and bronchial responsiveness in swine confinement workers.

Testing of lung function and bronchial reactivity, bronchoalveolar lavage (BAL), and a skin prick test with a standard panel and six "swine" extracts obtained from swine and swine environment were performed in 20 randomly selected nonsmoking swine confinement workers. In addition, blood samples for detection of antibodies by the diffusion in gel-enzyme-linked immunosorbent assay (DIG-ELISA) technique and precipitating antibodies were drawn. Air samples for measurement of dust and endotoxin levels were collected. All the farmers regarded themselves as healthy. The results were compared with reference groups consisting of urban nonsmoking subjects who had not been exposed to pig farming environment. The pig farmers had normal lung function and the bronchial reactivity was not different from the reference group. In the BAL fluid of the farmers, the concentration of total cells and granulocytes was increased while the concentrations of lymphocytes and macrophages were normal. The BAL fluid concentrations of albumin, fibronectin, and hyaluronan were elevated in the farmers. Skin prick tests with swine extracts were negative in all farmers. Antibodies (assessed by DIG-ELISA) against swine dander, swine dust, and pig feed were increased and precipitating antibodies against swine dander were found in 14, against pig food in five, and against swine confinement dust in three of the 20 pig farmers. The concentration of airborne total dust was 7.4 mg/cu mm and the endotoxin concentration was 37 (22 to 60) ng/cu mm during tending the pigs and increased, during feeding, to 13.8 mg/cu mm and 315 (194 to 716) ng/cu mm, respectively. There was no correlation between exposure and lung function or lavage findings. In conclusion, randomly selected pig farmers had signs of airway inflammatory reaction and activation of the immune system without alteration in lung function and bronchial reactivity.     

Elevated levels of leukotriene C4 in bronchoalveolar lavage fluid from atopic asthmatics after endobronchial allergen challenge

Sulfidopeptide leukotrienes have been implicated in the pathogenesis of asthma because of their ability to induce bronchospasm, airways hyperreactivity, and increased mucus production. In the present study, the leukotrienes (LT) C4, D4, and E4 were measured in bronchoalveolar lavage fluid (BALF) before and 5 min after endobronchial allergen challenge in four subject groups: nonatopic nonasthmatic, nonatopic asthmatic, atopic nonasthmatic, and atopic asthmatic. As determined by high performance liquid chromatography (HPLC), after allergen challenge, the predominant sulfidopeptide leukotriene found in BALF from atopic asthmatics was LTC4. Smaller amounts of LTD4 and LTE4 were detectable. The baseline level of leukotrienes in the atopic asthmatics was 64 +/- 18 pg/ml, with measurable levels being found in nine of 11 samples. Atopic nonasthmatics had measurable levels in only one of seven baseline samples, whereas five of six nonatopic subjects had undetectable levels. Allergen challenge in atopic asthmatics resulted in significant increases in LTC4 over prechallenge levels (64 +/- 18 to 616 +/- 193 pg/ml) (p less than 0.01) and over levels in the three control groups after challenge (p = 0.0297). The atopic nonasthmatic group also had detectable leukotriene levels after allergen challenge (88 +/- 32 pg/ml), whereas leukotrienes remained undetectable in five of the six nonatopic samples. For comparison, histamine and the prostanoids prostaglandin D2 (PGD2) and thromboxane B2 (TxB2) were also measured in BALF. The levels of all three of these mediators increased in BALF from atopic asthmatics after allergen challenge. After allergen challenge, the best correlation was found between the levels in BALF for the prostanoids PGD2 and TxB2 (r = 0.88).(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of bronchoalveolar lavage on bronchial responsiveness in patients with airflow obstruction

This study assessed the effect of bronchoalveolar lavage (BAL) on nonspecific bronchial responsiveness in 31 patients. Of these, 20 had airflow obstruction; 11 control subjects had normal pulmonary function. Bronchial responsiveness to methacholine, expressed as the dose of inhaled methacholine required to provoke a 20 percent fall in forced expiratory volume in one second (PD20 FEV1), was measured before and after BAL. We found no evidence for the induction of responsiveness by BAL in 11 control subjects with negative methacholine tests prior to the procedure. There were small but significant falls in FEV1 following BAL in both the control group and in patients with airflow obstruction. Thus, BAL does not appear to induce nonspecific bronchial hyperresponsiveness in subjects without airflow obstruction, nor does it affect airway responsiveness in emphysema patients. Among asthmatics, bronchial responsiveness can be increased as a result of BAL; this increase was greatest in patients who were most responsive initially.     

Salmeterol does not alter increased bronchial responsiveness caused by organic dust exposure

BACKGROUND: Exposure in a swine house induces airway inflammation and increases bronchial responsiveness to methacholine in healthy subjects. STUDY OBJECTIVES: The aim was to investigate whether a long-acting beta2-agonist, salmeterol, alters the increased bronchial responsiveness induced in healthy subjects following exposure to organic dust in a swine barn. DESIGN AND SUBJECTS: The study includes three separate parts. In the first part (part 1), healthy subjects inhaled salmeterol (50 microg bid, n = 8) or placebo (n = 8) over 2 weeks. In part 2, healthy subjects inhaled one single dose of salmeterol (100 microg, n = 6) or placebo (n = 6) 1 h prior to exposure in a swine barn, which was followed by a bronchial methacholine challenge. In part 3, eight healthy individuals inhaled placebo or salmeterol (100 microg), 2 h or 8 h prior to a bronchial methacholine provocation, without being exposed in the swine barn. RESULTS: Exposure caused an increase of bronchial responsiveness to methacholine by 3.2 doubling concentration steps (25 to 75th percentiles, 2.8 to 4.1) and 2.6 doubling concentration steps (25 to 75th percentiles, 1.4 to 3.7) in the placebo and salmeterol groups (2 weeks), respectively, with no significant differences between the groups (p = 0.3; part 1). Similar results were obtained when salmeterol was administered as a single dose (part 2) prior to exposure. However, salmeterol significantly attenuated the bronchial responsiveness to methacholine by 1.2 doubling concentration steps (0.8 to 1.7) 8 h after inhalation (part 3). CONCLUSIONS: Salmeterol inhalation did not protect against the increased bronchial responsiveness induced in healthy subjects following exposure to organic dust when administered for 2 weeks or as a single dose prior to exposure. This lack of protection cannot be explained by homologous beta2-adrenoceptor desensitization. We hypothesize that exposure to organic material may alter the airway response to beta2-agonists.     

Extracellular matrix metalloproteinase inducer is increased in smokers' bronchoalveolar lavage fluid

Extracellular matrix metalloproteinase inducer (EMMPRIN), also called basigin, is present in the lung during development, but its expression in normal adult lung is minimal. Increases of EMMPRIN have been found in various forms of experimental lung injury. To determine whether EMMPRIN might be involved in alveolar injury/repair associated with smoking, we developed an ELISA for EMMPRIN and applied it to bronchoalveolar lavage fluids from never-smokers (n = 7), former smokers (n = 16), and current smokers (n = 58). The smoker groups included subjects with emphysema, as determined by high-resolution chest computed tomography. EMMPRIN levels were significantly elevated in current and former smokers (315 +/- 20 and 175 +/- 15 pg/ml SEM, respectively, compared with 31 +/- 7 pg/ml in never-smokers), but the EMMPRIN levels of smokers with emphysema were not different from smokers without emphysema. Immunohistochemistry of smokers' lung tissue showed EMMPRIN in bronchiolar epithelium and alveolar macrophages, but EMMPRIN mRNA in alveolar macrophages was not different between current and never-smokers. Matrix metalloproteinase-1 was also detectable in the bronchoalveolar lavage fluid from some smokers but not in never-smokers. These findings indicate that smoking is associated with increased intrapulmonary EMMPRIN. Whether EMMPRIN is involved in smoking-induced lung pathology remains to be determined.     

Altered apoptosis in bronchoalveolar lavage lymphocytes after allergen exposure of atopic asthmatic subjects.

The increased number of lymphocytes in airways during an asthmatic response is believed to be the result of increased recruitment of these cells. However, it is possible that a decreased apoptotic rate could also contribute to the increased number. The aim of the present study was to investigate whether allergen airway provocation influences the apoptotic phenotype of lung and peripheral blood lymphocytes (PBL) in subjects with atopic asthma. Bronchoalveolar lavage (BAL) lymphocytes and PBL from 12 asthmatic subjects previously challenged with allergen (n = 7) or saline (n = 5) were exposed to the apoptotic stimulus tributyltin (TBT) in vitro and assayed for apoptosis. Airway allergen provocation resulted in decreased sensitivity of BAL lymphocytes to TBT-induced apoptosis, with 42.2% (range 33.9-62.5%) apoptotic cells before challenge versus 23.5% (range 15.3-42.4%) after challenge, while PBL were unaffected. The increased apoptosis resistance correlated with higher numbers of Bcl-2-expressing lymphocytes. Interestingly, baseline caspase-3-like activity was significantly elevated in viable BAL lymphocytes compared with viable PBL, and was unaltered by allergen exposure. In conclusion, allergen inhalation renders bronchoalveolar lavage lymphocytes more resistant to apoptosis while peripheral blood lymphocytes were not influenced at all, indicating that the apoptotic phenotype of airway lymphocytes may play a role in asthmatic inflammation.     

Local allergen challenge and bronchoalveolar lavage of allergic asthmatic lungs. Description of the model and local airway inflammation

The local mechanisms that result in the cellular inflammation and bronchial airway hyperreactivity that characterize allergic bronchial asthma are poorly defined. In order to study these processes, we developed a method for local allergen challenge using a fiberoptic bronchoscope and direct observation and bronchoalveolar lavage (BAL) to assess the airway responses to allergen. In these studies, 11 allergic asthmatics (all of whom had previously demonstrated a late-phase asthmatic response to aeroallergen challenge) and 6 healthy, asymptomatic subjects volunteered to undergo bronchoalveolar lavage after local airway challenge via a bronchoscope wedged into subsegmental airways. These studies revealed that asthmatic airways respond to allergen with an immediate pallor followed by reactive hyperemia, edema, and bronchial narrowing. This site and a control site were relavaged at 48 or 96 h after the immediate response. Neutrophils and eosinophils increased significantly at 48 h after challenge, as did helper T-lymphocytes. Characteristically, at 96 h, neutrophil counts returned to normal values, whereas eosinophiles and helper T-cells remained elevated. Peroxidase-staining cells were also elevated at 48 h after local allergen challenge. Electron microscopy revealed degranulation of mast cells and eosinophils, both immediately and later (48 and 96 h) after local allergen challenge. Macrophages were highly activated and had phagocytized, partially intact granules from both eosinophils and mast cells. There was a significant correlation (p less than 0.001) between the concentration of allergen required to produce a visible airway response and a positive end-point skin titration in the asthmatic subjects.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cathepsin D activity is increased in alveolar macrophages and bronchoalveolar lavage fluid of smokers

Cathepsin D activity was determined in alveolar macrophages (AM) and cell-free bronchoalveolar lavage fluid (BALF) from volunteers who were current cigarette smokers and compared with that found in lifetime nonsmokers. Enzyme activity was determined with a highly sensitive and specific substrate [D-Phe-Ser(0-CH2-C6H5)-Phe-Phe-Ala-Ala-pAB]. Specific activity was more than three times higher in AM from smokers than in cells from nonsmokers (37,880 +/- 2,090 versus 10,300 +/- 1,200; p less than 0.001) and approximately seven times higher in BALF from smokers than from nonsmokers (3,620 +/- 490 versus 515 +/- 165; p less than 0.001). This study demonstrated that cigarette smoke is a potent inducer of cathepsin D activity in AM in vivo. Because cathepsin D is capable of degrading a variety of proteins, the finding of high concentrations of the enzyme in AM and BALF from smokers, along with previous observations of elevated cathepsin B activity, suggests that lysosomal enzymes may cause or contribute to structural lung damage associated with cigarette smoking.     

Intestinal helminth infestation is associated with increased bronchial responsiveness in children

Non-atopic asthma is the predominant phenotype in non-affluent parts of Latin America. We recently reported that infestation with Ascaris lumbricoides increased the risk of non-atopic asthma in less affluent areas of Brazil but the mechanism is unclear. The present study was conducted to determine whether helminth infestation is associated with heightened bronchial responsiveness (BHR), a common finding in asthma. A random sample of 50 asthmatic and 50 non-asthmatic controls (mean age 10.1 years) were selected from a larger cohort (n = 1,011) without knowledge of their helminth infestation status. Three stool samples were collected from each child on different days and each sample was analyzed by the Kato-Katz method for quantitative determination of helminth eggs. Bronchial provocation tests were performed with inhaled 4.5% hypertonic saline using the ISAAC Phase II standardized protocol. There was no difference between the prevalence of positive BHR in the asthmatics (20.4%) compared with the controls (14.6%) (P = 1.0). Helminth infestation was detected in 24.0% of children, with A. lumbricoides being the most common. Children with high load infestation (>or=100 eggs/g) were five times more likely to have BHR than children with low load or no infestation. Despite the small sample size the results of the present study suggest that the link between high load helminth infestation and non-atopic asthma may be mediated via heightened bronchial responsiveness, possibly due to an inflammatory response to the pulmonary phase of the helminth life cycle.     

Chronic inflammation is associated with an increased proportion of goblet cells recovered by bronchial lavage

To evaluate the possibility that bronchoalveolar lavage could provide sufficient respiratory epithelial cells to quantify changes in epithelial cell types associated with chronic inflammation, we examined the epithelial cells obtained in the first infused (20 ml) aliquots that were processed separately from later aliquots, a process known to enrich for bronchial contents. Epithelial cells, including ciliated cells, goblet cells, and fragments of desquamated epithelium, were easily identified after preparation by cytocentrifugation and staining with a modified Giemsa stain. Quantification of the columnar cell types revealed that those with chronic bronchitis and asymptomatic smokers have increased goblet cells as a percentage of the total columnar epithelial cells (chronic bronchitics 36 +/- 2 percent, asymptomatic smokers 22 +/- 2 percent) compared with normal subjects (9 +/- 1 percent, p less than 0.001, ANOVA). Significantly, the goblet cell percentage was strongly correlated with other measures of bronchitis and measures of airflow obstruction such as the bronchitis index, a visually derived score at bronchoscopy of airway inflammation (r = 0.72, p less than 0.001), the percent neutrophils in the first infused aliquots (r = 0.44, p less than 0.05), and the FEV1 percent (r = -0.74, p less than 0.001). Thus, bronchoalveolar lavage is capable of providing sufficient bronchial epithelial cells for analysis, and the changes seen in the spectrum of columnar epithelial cells may reflect important underlying pathologic changes.     

Vitronectin in bronchoalveolar lavage fluid is increased in patients with interstitial lung disease.

Vitronectin, also known as S-protein, is a 75,000-dalton serum glycoprotein that has a variety of functions, including the capacity to interact with the terminal components of the complement cascade, the coagulation system, and cell surfaces. By virtue of its ability to interact with cells, vitronectin is capable of mediating cell-spreading and adhesion and may also influence cell differentiation and cell growth. To investigate the possibility that vitronectin might contribute to the pathogenesis of interstitial lung disease, vitronectin was measured in bronchoalveolar lavage fluid from patients with sarcoidosis, idiopathic pulmonary fibrosis, and, for comparison, normal volunteers. Vitronectin was detected in lavage fluid and serum of all study subjects. Increased lavage fluid concentrations were found in patients with interstitial lung disease when compared with normal subjects (p less than 0.005), and glucocorticoid-treated patients with interstitial lung disease had lower vitronectin levels than did untreated patients. Furthermore, on SDS-PAGE and Western blot analysis lavage fluid vitronectin comigrated with serum vitronectin, suggesting similar molecular size. Thus, vitronectin is a normal constituent of the epithelial lining fluid, and lavage fluid vitronectin is similar to serum vitronectin. The increase of vitronectin concentrations in the epithelial lining fluid of patients with interstitial lung disease suggests that vitronectin may contribute to the pathogenesis of interstitial lung disease.     

Dissimilarity in methacholine and adenosine 5'-monophosphate responsiveness 3 and 24 h after allergen challenge

Bronchial hyperresponsiveness (BHR) to methacholine and adenosine 5'-monophosphate (AMP) was studied in 15 allergic asthmatic patients before and 3 and 24 h after allergen challenge with house dust mite (HDM). Subjects attended the clinic on 3 consecutive days. On the first day a control solution was inhaled, and methacholine or AMP challenge was performed 3 h later. The next day HDM was inhaled, and 3 and 24 h later methacholine or AMP challenge was performed again. There were no significant difference in FEV1 baseline value between any of the study days. PD20 HDM, percentage decrease in FEV1, and AUC for both the EAR and LAR were not significantly different in the methacholine and AMP studies. After HDM challenge, PC20 methacholine decreased significantly from a geometric mean (+/- SEM) starting value of 1.39 +/- 0.63 mg/ml to 0.30 +/- 0.78 mg/ml (p less than 0.001) at 3 h and to 0.22 +/- 0.75 mg/ml (p less than 0.001) at 24 h. The magnitude of the decrease in PC20 methacholine at 3 h correlated with the severity of the late asthmatic reaction (LAR) as measured by the percentage fall in FEV1 and area under the curve (AUC) (r = -0.60 and r = 0.55; p less than 0.05). A significant decrease was observed in the PC20 AMP at 3 h, from a geometric mean value of 12.2 +/- 0.96 mg/ml after challenge with the control solution to 4.47 +/- 0.99 mg/ml (p less than 0.05) after HDM challenge.(ABSTRACT TRUNCATED AT 250 WORDS)     

Urinary leukotriene E4 levels after allergen and exercise challenge in bronchial asthma.

Urinary leukotriene E4 (LTE4) concentrations were measured in six asthmatic subjects after treadmill exercise, and in five asthmatic subjects after allergen challenge. Exercise and allergen challenge produced a 42 +/- 18% (mean +/- SD) and 22 +/- 8% fall in FEV1, respectively. The baseline concentration of urinary LTE4 in subjects challenged with exercise was 64 (27 to 150) pg/mg creatinine (geometric mean and 95% confidence interval), and in those challenged with allergen it was 36 (23 to 59) pg/mg creatinine. Urinary LTE4 concentrations did not change significantly in the 24 h after exercise. In contrast, there was a mean 4-fold increase in urinary LTE4 during the 3 h after allergen challenge.     

Cell profile and elastase activity in diffuse panbronchiolitis investigated by bronchoalveolar and bronchial lavage.

To examine the mechanism of tissue damage which causes bronchiolectasis in diffuse panbronchiolitis (DPB), the cellular components, elastase and its main inhibitor, alpha 1-protease inhibitor (alpha 1-PI) were measured in bronchoalveolar and bronchial lavage fluid (BALF and BLF) from 14 DPB patients. A predominant increase in the neutrophil count was observed in DPB. Elastase activity in BALF and BLF was about 1,000-fold higher in the DPB group than in the control group. An inhibitor study and a positive correlation between elastase activity and the neutrophil count in both lavage fluids from the DPB group indicated that the activity was mainly that of neutrophil elastase. Western blot analysis of alpha 1-PI showed that most of the alpha 1-PI in the lavage fluids from DPB group was degraded. These results indicated that neutrophil infiltration increases the level of elastase in the DPB lesions; this increase seems to be closely related to tissue damage.     

Guinea pig ozone-induced airway hyperreactivity is associated with increased N-acetyl-beta-D-glucosaminidase activity in bronchoalveolar lavage fluid

High level ozone exposure is known to cause acute, neutrophil-independent airway hyperreactivity in the guinea pig. The precise biochemical mechanisms involved remain unclear. Because of its potential pathophysiologic importance, we examined whether a lysosomal hydrolase, N-acetyl-beta-D-glucosaminidase (NAGA) was released from the airways in vivo and from bronchoalveolar cells, specifically macrophages. Muscarinic reactivity was determined by measuring specific airway resistance (sRaw) in response to increasing doses of aerosolized acetylcholine in guinea pigs that were either exposed to air or to ozone (3.0 ppm, 2 h). The ozone-exposed animals showed substantial muscarinic hyperreactivity 30 min after exposure. In addition, both total and percent released NAGA in bronchoalveolar lavage fluid obtained immediately after reactivity testing were significantly greater in the ozone-exposed group. It was also found that substantially more NAGA was released from mixed bronchoalveolar lavage cells in response to 20 microM A23187. Moreover, bronchoalveolar macrophages of ozone-exposed animals secreted more NAGA upon stimulation in vitro by either 20 microM A23187 or 200 micrograms/ml opsonized zymosan. We conclude that ozone-induced airway hyperreactivity in guinea pigs is associated with the presence of increased NAGA activity in bronchoalveolar fluid. Our data suggest that bronchoalveolar macrophages may, at least in part, be responsible for release of this enzyme into the airways after ozone exposure.     

Fractional processing of sequential bronchoalveolar lavage to separate bronchial and alveolar samples

Bronchoalveolar lavage has been widely used to sample the lower respiratory tract. Most of the material recovered with this technique represents alveolar contents. A number of modifications have been suggested in order to obtain samples relatively enriched for bronchial material. In order to be able to use a standard technique for bronchoalveolar lavage to sample both airways and "routine" alveolar material, a simple modification of the technique as described by Reynolds and Newball was used: five sequential 20-ml aliquots were infused into the lower respiratory tract, and each aliquot was immediately aspirated. The return from the first aliquot was processed separately from the return from the subsequent four aliquots. These last four aliquots were pooled. Analysis of the first aliquot revealed it to be enriched for ciliated epithelial cells when compared with the subsequent aliquots. There were also differences in inflammatory cell composition with the bronchial sample containing relatively more neutrophils and relatively less lymphocytes. Aspiration during transoral bronchoscopy was documented by quantifying salivary amylase in the bronchial and alveolar lavage fluids. It was estimated, however, that the aspiration was not of quantitative significance in the vast majority of subjects studied. Finally, with the technique of fractional processing of bronchoalveolar lavage samples, it was possible to compare the protein concentrations in bronchial and alveolar lavages. Most prominent among the differences was a marked relative enrichment in the bronchial samples for immunoglobulin A. The technique of fractional processing of bronchoalveolar lavage samples provides a simple means to obtain samples enriched for bronchial and alveolar components. This should facilitate analysis of lower respiratory tract specimens in airway disease.