Interleukin-17A and vascular remodelling in severe asthma; lack of evidence for a direct role.

BACKGROUND: Bronchial vascular remodelling may contribute to the severity of airway narrowing through mucosal congestion. Interleukin (IL)-17A is associated with the most severe asthmatic phenotype but whether it might contribute to vascular remodelling is uncertain. OBJECTIVE: To assess vascular remodelling in severe asthma and whether IL-17A directly or indirectly may cause endothelial cell activation and angiogenesis. METHODS: Bronchial vascularization was quantified in asthmatic subjects, COPD and healthy subjects together with the number of IL-17A+ cells as well as the concentration of angiogenic factors in the sputum. The effect of IL-17A on in vitro angiogenesis, cell migration and endothelial permeability was assessed directly on primary human lung microvascular endothelial cells (HMVEC-L) or indirectly with conditioned medium derived from normal bronchial epithelial cells (NHBEC), fibroblasts (NHBF) and airway smooth muscle cells (ASMC) after IL-17A stimulation. RESULTS: Severe asthmatics have increased vascularity compared to the other groups, which correlates positively with the concentrations of angiogenic factors in sputum. Interestingly, we demonstrated that increased bronchial vascularity correlates positively with the number of subepithelial IL-17A+ cells. However IL-17A had no direct effect on HMVEC-L function but it enhanced endothelial tube formation and cell migration through the production of angiogenic factors by NHBE and ASMC. CONCLUSIONS & CLINICAL RELEVANCE: Our results shed light on the role of IL-17A in vascular remodelling, most likely through stimulating the synthesis of other angiogenic factors. Knowledge of these pathways may aid in the identification of new therapeutic targets.

Exacerbation risk in severe asthma is stratified by inflammatory phenotype using longitudinal measures of sputum eosinophils.

BACKGROUND: Airway inflammatory phenotyping is increasingly applied to subjects with asthma. However, its relationship to clinical outcomes in difficult asthma is incompletely elucidated. OBJECTIVE: The goal of our study was to determine the relationship between exacerbation rates and phenotypes of difficult asthma based on the longitudinal measures of sputum eosinophils and neutrophils. METHODS: Subjects in the longitudinal observational study from two tertiary care centres that completed 1 year of observation and provided at least three sputum samples were classified by inflammatory phenotypes using previously established thresholds. Kaplan-Meier curves and univariable and multivariable Cox proportional hazard models were used to determine the association between inflammatory phenotypes and exacerbation rate. RESULTS: During the study, 115 exacerbations occurred in 73 severe asthmatic subjects. Subjects with the persistently eosinophilic phenotype had a significantly shorter time to first exacerbation and greater risk of exacerbation over a 1-year period than those with the non-eosinophilic phenotype based on the univariable and multivariable Cox proportional hazard model (hazard ratio [HR], 3.24; 95% confidence interval [CI], 1.35-7.72; adjusted HR, 3.90; 95% CI, 1.34-11.36). No significant differences in time to first exacerbation or exacerbation risk over a 1-year period were observed among the neutrophilic phenotypes. CONCLUSIONS: The persistent eosinophilic phenotype is associated with increased exacerbation risk compared with the non-eosinophilic phenotype in severe asthma. No differences in time to first exacerbation or exacerbation risk over a 1-year period were detected among neutrophilic phenotypes.

Phenotyping of difficult asthma using longitudinal physiological and biomarker measurements reveals significant differences in stability between clusters.

BACKGROUND: Although the heterogeneous nature of asthma has prompted asthma phenotyping with physiological or biomarker data, these studies have been mostly cross-sectional. Longitudinal studies that assess the stability of phenotypes based on a combination of physiological, clinical and biomarker data are currently lacking. Our objective was to assess the longitudinal stability of clusters derived from repeated measures of airway and physiological data over a 1-year period in moderate and severe asthmatics. METHODS: A total of 125 subjects, 48 with moderate asthma (MA) and 77 with severe asthma (SA) were evaluated every 3 months and monthly, respectively, over a 1-year period. At each 3-month time point, subjects were grouped into 4 asthma clusters (A, B, C, D) based on a combination of clinical (duration of asthma), physiological (FEV1 and BMI) and biomarker (sputum eosinophil count) variables, using k-means clustering. RESULTS: Majority of subjects in clusters A and C had severe asthma (93 % of subjects in cluster A and 79.5 % of subjects in cluster C at baseline). Overall, a total of 59 subjects (47 %) had stable cluster membership, remaining in clusters with the same subjects at each evaluation time. Cluster A was the least stable (21 % stability) and cluster B was the most stable cluster (71 % stability). Cluster stability was not influenced by changes in the dosage of inhaled corticosteroids. CONCLUSION: Asthma phenotyping based on clinical, physiologic and biomarker data identified clusters with significant differences in longitudinal stability over a 1-year period. This finding indicates that the majority of patients within stable clusters can be phenotyped with reasonable accuracy after a single measurement of lung function and sputum eosinophilia, while patients in unstable clusters will require more frequent evaluation of these variables to be properly characterized.

Depletion of OX-8 lymphocytes from the blood and airways using monoclonal antibodies enhances the late airway response in rats.

Recent evidence supports a role for T lymphocytes in allergic airway responses. We hypothesized that reducing blood T suppressor cells (Ts) might increase the late airway response (LR). Sprague-Dawley (SD) rats were sensitized with ovalbumin (OA). On days 8, 10, and 12, post-sensitization test SD (n = 14) received monoclonal antibody intravenously (OX-8; 1 mg) specific to rat Ts. Controls received saline (n = 7) or mouse ascites IgG (n = 7). On day 14, animals were challenged with OA aerosol (5% wt/vol) for 5 min, lung resistance was recorded for 8 h (n = 18) and bronchoalveolar lavage was performed. The LR was determined from the area under the lung resistance vs time curve from 75 to 480 min after challenge. In the remaining 10 rats, airway lymphocyte subsets were measured 8 h after OA aerosol challenge in minced and digested lungs. A decrease in percentage of blood and airway Ts, respectively, in test animals was observed vs controls (blood: 6.27 +/- 0.84 vs 32.95 +/- 1.94, P < 0.001); (airway: 5.05 +/- 0.66 vs 24.5 +/- 3.05, P < 0.02). Blood and airway helper T lymphocytes did not differ between test and control animals. The LR was significantly increased in test (22.89 +/- 3.92) vs controls (4.22 +/- 2.18, P < 0.001). Bronchoalveolar lavage macrophages, neutrophils and lymphocytes, and serum OA-specific IgE were also significantly elevated (P < 0.05) in test animals. We conclude that Ts play an important role in attenuating the LR in SD rats.

Structural changes and airway remodelling in occupational asthma at a mean interval of 14 years after cessation of exposure.

BACKGROUND: Occupational asthma (OA) may cause alterations of airways with inflammation and remodelling after cessation of exposure. Although the long-term clinical, functional and induced sputum sequelae have been examined in workers removed from exposure, the long-term pathological outcomes are unknown. OBJECTIVE: We aimed to investigate whether airway inflammation and remodelling were present in bronchial biopsies of subjects with prior OA but without evidence of persisting asthma at a mean interval of 14 years after cessation of exposure. METHODS: Ten clinically and functionally asymptomatic subjects with a prior diagnosis of OA were recruited and underwent bronchoscopy, bronchoalveolar lavage and bronchial biopsy. Comparisons were made with biopsies from normal control subjects. Epithelial detachment, epithelial metaplasia, mucous gland and airway smooth muscle (ASM) areas as well as the distance between the epithelium and ASM were measured by image analysis. The amount of collagen present was assessed by van Gieson staining. The numbers of TGF-beta1- and eosinophil cationic protein (ECP)-positive cells were evaluated by specific immunostaining. RESULTS: Statistically significant increases were found in the numbers of TGF-beta1- and ECP-positive cells and in the amount of subepithelial fibrosis present in the biopsies of subjects with prior OA compared with control biopsies. The distance between the epithelium and ASM was significantly reduced in the OA group. Increases in epithelial metaplasia, ASM mass, mucous gland numbers, collagen deposition and eosinophilia in the OA group were not statistically significant. There was no evidence of ongoing inflammation in the group with prior OA as assessed by the number of T lymphocytes present. CONCLUSION: Some aspects of airway inflammation and remodelling persist in subjects with prior OA long after cessation of exposure even in the absence of clinical, sputum and functional abnormalities. These findings are relevant to the assessment of long-term sequelae in subjects with OA when reviewed after cessation of exposure.

A comparison of responses to albuterol delivered by two aerosol devices.

Nineteen outpatients with stable obstructive pulmonary disease (mean forced expiratory volume in one second [FEV1], 1.00 + 0.10 L) were evaluated for airway response to albuterol (salbutamol) administered by metered-dose inhaler and Bosch ultrasonic nebulizer (BUSN). Albuterol administered by metered-dose inhaler but not by nebulizer caused a significant increase in FEV1 and the mean forced expiratory flow over the middle half of the forced vital capacity (FEF25-75%) (p less than 0.02). Absolute increase from baseline of FEV1 and FEF25-75% was significantly greater for metered-dose inhaler (0.21 +/- 0.05 L; 0.32 +/- 0.13 L/sec) compared to ultrasonic nebulizer (0.07 +/- 0.03 L; 0.03 +/- 0.04 L/sec) (p less than 0.02). In 11 subjects (mean FEV1, 1.08 + 0.14 L), the placebo effect of inhalation of the diluent from the metered-dose inhaler (Freon) and the ultrasonic nebulizer (isotonic saline solution) was determined. Freon produced the mean increase of 1.5 percent, whereas the ultrasonic aerosol of isotonic saline solution resulted in a mean decrease of 8 percent in FEV1. Therefore, the inferior response to albuterol administered by ultrasonic nebulizer was at least in part due to the superimposed broncho-constriction occurring with ultrasonically administered saline solution. The metered-dose inhaler was more effective than the ultrasonic nebulizer for administration of albuterol in stable obstructive pulmonary disease, and the latter device is not recommended. A specific ultrasonic nebulizer should be prescribed only if its superiority to a metered-dose inhaler can be objectively documented.

Increased expression of the chemoattractant cytokines eotaxin, monocyte chemotactic protein-4, and interleukin-16 in induced sputum in asthmatic patients.

BACKGROUND: Induced sputum from asthmatic patients has been recently used to assess inflammatory cells. We have previously reported an increased expression of Th-2-type cytokines in induced sputum of asthmatic patients. C-C chemokines, particularly eotaxin and monocyte chemotactic protein (MCP)-4, are associated with eosinophilic infiltration. Interleukin (IL)-16 is associated with chemotactic activity for CD4+ cells. Chemokine expression in BAL and bronchial biopsy specimens has been demonstrated in asthmatic airways, but not in induced sputum. METHODS: We examined whether eotaxin, MCP-4, and IL-16 expression could be detected in induced sputum of asthmatic patients (n = 10), and whether the expression was increased compared to normal control subjects (n = 9). Eotaxin, MCP-4, and IL-16 immunoreactivity were determined by immunocytochemistry. In addition, inflammatory cells were investigated using markers for T cells (CD3), eosinophils (major basic protein [MBP]), macrophages (CD68), neutrophils (elastase), and epithelial cells (cytokeratin). RESULTS: Our results showed that there was a significant difference in the percentages of MBP-positive and epithelial cells between asthmatic patients and normal control subjects (p < 0.05). However, there was no difference between these two groups in the percentage of CD3-, elastase-, and CD68-positive cells. Immunoreactivity for eotaxin, MCP-4, and IL-16 was expressed in the induced sputum of all asthmatic patients, and expression of these chemotactic cytokines was significantly greater than in control subjects (p < 0.001, p < 0.005, and p < 0.001, respectively). CONCLUSIONS: This study showed that induced sputum could be used to detect chemokines in patients with bronchial asthma, and that the upregulation of chemotactic cytokines in the airways can be seen using noninvasive techniques.

Inhaled budesonide inhibits OVA-induced airway narrowing, inflammation, and cys-LT synthesis in BN rats.

The objective of the present investigation was to examine the effects of an inhaled glucocorticoid, budesonide, on antigen-induced production of cysteinyl leukotrienes (cys-LTs) and pulmonary inflammatory cell infiltration in the Brown Norway rat, an animal model of asthma. Two weeks after sensitization to ovalbumin, rats were treated with budesonide (2.5 mg/kg) 18 and 1 h before challenge with antigen. Budesonide abolished the late response to ovalbumin (P<0.02) and strongly inhibited the in vivo synthesis of N-acetyl-leukotriene E(4), an indicator of cys-LT synthesis, during this period (P<0.005). Both total bronchoalveolar lavage (BAL) cells (P<0.01) and BAL macrophages (P<0.005) were markedly reduced to approximately 25% of their control levels after treatment with budesonide. It can be concluded that inhibition of the antigen-induced late response in Brown Norway rats by budesonide is associated with reductions in both BAL macrophages and cys-LT synthesis. It is possible that the effect of budesonide on cys-LT synthesis is related to its effects on pulmonary macrophages.

Homeokinesis and short-term variability of human airway caliber.

We hypothesized that short-term variation in airway caliber could be quantified by frequency distributions of respiratory impedance (Zrs) measured at high frequency. We measured Zrs at 6 Hz by forced oscillations during quiet breathing for 15 min in 10 seated asthmatic patients and 6 normal subjects in upright and supine positions before and after methacholine (MCh). We plotted frequency distributions of Zrs and calculated means, skewness, kurtosis, and significance of differences between normal and log-normal frequency distributions. The data were close to, but usually significantly different from, a log-normal frequency distribution. Mean lnZrs in upright and supine positions was significantly less in normal subjects than in asthmatic patients, but not after MCh and MCh in the supine position. The lnZrs SD (a measure of variation), in the upright position and after MCh was significantly less in normal subjects than in asthmatic patients, but not in normal subjects in the supine position and after MCh in the supine position. We conclude that 1) the configuration of the normal tracheobronchial tree is continuously changing and that this change is exaggerated in asthma, 2) in normal lungs, control of airway caliber is homeokinetic, maintaining variation within acceptable limits, 3) normal airway smooth muscle (ASM) when activated and unloaded closely mimics asthmatic ASM, 4) in asthma, generalized airway narrowing results primarily from ASM activation, whereas ASM unloading by increasing shortening velocity allows faster caliber fluctuations, 5) activation moves ASM farther from thermodynamic equilibrium, and 6) asthma may be a low-entropy disease exhibiting not only generalized airway narrowing but also an increased appearance of statistically unlikely airway configurations.

Increased plasma catecholamines in patients with Friedreich's ataxia.

We studied free plasma catecholamines in 23 patients with Friedreich's ataxia, having a mean age of 22 +/- 9.6 (SD) years. Conjugated catecholamines were also studied in 10 patients. Mean plasma norepinephrine and epinephrine were significantly higher than controls both in the supine and standing positions. In total 15 out of 23 patients (65%) had increase free and/or conjugated plasma catecholamines. The increased in plasma catecholamines was more marked in patients with severe neuromotor impairment. Among the patients with left ventricular concentric hypertrophy (wall thickness greater than 12 mm), only 3 had no demonstrable sympathetic hyperfunction. Since the high local concentrations of norepinephrine at the site of release from sympathetic nerve terminals may serve as a trigger for the hypertrophic response of the myocardial cell, it is suggested that early pharmacological intervention could prevent or limit the cardiomyopathic process or its clinical consequences.

Fuzzy logic and mechanical ventilation.

Closed-loop control can achieve appropriate ventilation of the lungs in a healthy person by involving the continuous interaction of three components: sensors, controllers, and effectors. The sensors are the chemo-mechanoreceptors, which continuously monitor key bodily functions affected by ventilation. This information is relayed to the controllers, the respiratory centers in the brain, allowing them to determine how actual ventilation compares with that needed by the body. Finally, the controllers direct the effectors, the muscles of respiration, to adjust the ventilation accordingly. When a patient is in respiratory failure, the effector's role is taken over by a mechanical ventilator. The issue that is considered in this article is how the physician might be taken out of the feedback loop.

Differences in proteoglycan deposition in the airways of moderate and severe asthmatics.

Excess deposition of proteoglycans (PGs) has been described in the subepithelial layer of the asthmatic airway wall. However, less is known about deposition in the airway smooth muscle (ASM) layer, and whether the pattern of deposition is altered depending upon disease severity. Endobronchial biopsies were performed in patients with severe or moderate asthma (defined using American Thoracic Society criteria) and in control subjects. Biopsies were immunostained for the PGs biglycan, lumican, versican and decorin. PG deposition was measured in the subepithelial and ASM layers, the former by calculating the area of positive staining, and the latter by determining the percentage area stained using point counting. Immunostaining for PGs was prominent in biopsies from both moderate and severe asthmatics, compared with control subjects. While there was no difference in the amount of PG in the subepithelial layer between the two asthmatic groups, the percentage area of biglycan and lumican staining in the ASM layer was significantly greater in moderate versus severe asthmatics. Differences in the deposition of proteoglycans within the airway smooth muscle layer of moderate versus severe asthmatics potentially impact on the functional behaviour of the airway smooth muscle in these two groups of patients.

Evidence for expression of eosinophil-associated IL-12 messenger RNA and immunoreactivity in bronchial asthma.

BACKGROUND: Eosinophils are a source of cytokines within the airways of asthmatic individuals that may exert an important immunoregulatory influence. OBJECTIVES: We examined IL-12 messenger (m)RNA and protein expression in eosinophils from peripheral blood and bronchoalveolar lavage (BAL) fluid obtained from subjects with atopic asthma (n = 7), patients with chronic bronchitis (n = 5), and nonatopic healthy control subjects (n = 7). To further define this IL-12(+) population of eosinophils for the expression of other cytokines, we colocalized IL-12 and IL-5 within the peripheral blood eosinophils. METHODS: To detect IL-12 mRNA and protein expression, we used in situ hybridization and immunocytochemistry techniques. The double-immunocytochemistry technique was used to localize IL-12 protein to BAL eosinophils and to colocalize IL-5 and IL-12 in peripheral blood eosinophils. RESULTS: IL-12 mRNA and immunoreactive protein were localized to peripheral blood eosinophils. BAL fluid-derived eosinophils from asthmatic subjects were also reactive to IL-12. The percentage of peripheral blood eosinophils expressing mRNA for IL-12 was significantly lower in asthmatic subjects compared with that found in eosinophils obtained from patients with chronic bronchitis (P<.001) and control patients (P <.05). Colocalization studies demonstrated that the percentages of IL-12(+) eosinophils that are also IL-5(+) were 72% in asthmatic subjects and only 11% in control subjects (P<.001). CONCLUSION: These results suggest that eosinophils are a potential source of IL-12. Eosinophil-derived IL-12 may contribute and modulate the local allergic inflammatory responses.

Inflammatory cell populations in the airways and parenchyma after antigen challenge in the rat.

We examined the effects of sensitization and antigen challenge on the cellular populations retrieved from the large airways (LA) (generations zero to 5 approximately) and small airways and parenchyma (S/P) of the rat. Male Brown-Norway rats, 7 to 8 wk of age, were either actively sensitized to ovalbumin (n = 24) or sham-sensitized to saline (n = 9), and, 14 days later, they were anesthetized with urethane, intubated endotracheally, and challenged. Aerosols of ovalbumin (5% wt/vol in saline for 5 min) or saline were administered to 12 and six rats, respectively, and measurements of pulmonary resistance (RL) were made for 8 h. The early airway response (ER) was calculated as the highest value of RL in the first hour after challenge, and the late response (LR) was calculated as an increase in RL to greater than 200% of the baseline value in the 4- to 8-h period after challenge. Rats were killed by exsanguination, LA were separated from S/P, and cells were isolated after tissue mincing and digestion with collagenase. Total and differential cell counts and lymphocyte subsets were determined. Antigen challenge significantly increased the cellular yield (mostly neutrophils) from the LA and S/P. Animals with a LR had a lower total cellular yield from the LA and S/P than did animals without a LR. The animals with a LR also had a lower yield of eosinophils and lymphocytes from the S/P than did animals challenged with saline alone. Cellular yields were not lower in the animals with an isolated ER after antigen challenge.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of dexamethasone on airway inflammation and responsiveness after antigen challenge of the rat.

The purpose of this study was to examine the effects of dexamethasone on airway responsiveness and lung inflammation of rats at 8 h, 32 h, and 7 d after allergen challenge. Brown-Norway male rats, 7 to 8 wk old, were actively sensitized to ovalbumin (OA) and challenged 14 d later. The rats were divided into a control group (n = 31) and a test group (n = 34) that received dexamethasone (DEXA) (0.3 mg/kg intraperitoneally) 14 h and 2 h before saline or OA challenge. For challenge, rats were anesthetized with pentobarbital and intubated endotracheally. Aerosols of OA (5% wt/vol in saline) were administered for 5 min. Responsiveness to inhaled aerosols of methacholine and the total and differential leukocyte counts in the large airways (generations 0 to 5), small airways, and parenchyma isolated by tissue mincing and digestion were measured at 8 h, 32 h, and 7 d after OA challenge. The cellular influx into the airways and parenchyma was highest at 8 h and decreased progressively over 7 d. DEXA significantly inhibited the cellular influx after allergen challenge. At 8 h, cellular return from the large airways was 3.61 +/- 0.5 x 10(6) (controls) versus 1.0 +/- 0.2 x 10(6) (DEXA), and from the small airways and parenchyma was 31.7 +/- 2.8 x 10(6) (controls) versus 21.9 +/- 2.9 x 10(6) (DEXA) (p < 0.05). The differences were attributable mostly to decreases in neutrophils in DEXA-treated animals. In control animals, neutrophil yield fell between 8 and 32 h, whereas eosinophils and 32 h, whereas eosinophil and lymphocyte counts increased.(ABSTRACT TRUNCATED AT 250 WORDS)

Extracellular ATP stimulates elastase secretion from human neutrophils and increases lung resistance and secretion from normal rat airways after intratracheal instillation.

The effect of ATP on intracellular Ca2+ levels and elastase secretion in isolated normal human peripheral blood neutrophils was investigated as was its in vivo effect on lung resistance and mucous secretion. ATP (10(-5) M) increased [Ca2+]i from 61 +/- 3 to 165 +/- 15 nM in nonactivated neutrophils; elastase secretion was increased by 40% from nonactivated neutrophils but was unaffected in fMLP (10(-5) M) activated cells. Instillation of ATP (10(-5) and 10(-3) M) into the airways of brown Norway rats increased both lung resistance and secretion. These findings suggest that aerosolization of ATP into the cystic fibrosis-affected bronchial tree might be hazardous in terms of enhancement of parenchymal damage, which would result from neutrophil elastase release, and in terms of impaired respiratory lung function.

Comparison of cellular composition of induced sputum analyzed with Wright staining and immunocytochemistry.

We sought to compare cell counts based on cellular morphology and obtained after Wright staining with cell counts obtained after immunocytochemistry in induced sputum. Counts of eosinophils, neutrophils, macrophages, lymphocytes, and epithelial cells identified after Wright staining were compared with the counts obtained after immunocytochemistry through use of mAbs in 25 samples. Agreement between Wright staining and immunocytochemistry was poor for lymphocytes and epithelial cells. Rather than Wright staining, immunocytochemistry should be used to accurately identify lymphocytes and epithelial cells.

CD8 depletion-induced late airway response is characterized by eosinophilia, increased eotaxin, and decreased IFN-gamma expression in rats.

There is an emerging body of knowledge defining the role of CD8(+) cells in the pathogenesis of allergic asthma. We have previously demonstrated in sensitized Sprague-Dawley (SD) rats that depletion of CD8(+) cells caused an increase in the late airway response (LAR) and cellular infiltration after antigen challenge. To better delineate the mechanism of CD8(+) cell involvement in the development of the LAR and airway inflammation, we investigated the pattern of chemokine and cytokine production after antigen challenge. SD rats were sensitized to ovalbumin (OA) and subsequently treated with anti-CD8 (OX-8) monoclonal antibody (mAb) for the depletion of CD8(+) cells or with control mouse anti-rat IgG(1) mAb as a control procedure. After OA challenge, CD8- depleted SD rats developed an increased LAR when compared with control rats (area under the curve: 16.65 +/- 6.6 in CD8- depleted rats versus 5.39 +/- 2.0 in control animals; p < 0.05). Compared with the control animals, the increase in the LAR was accompanied by a significantly increased eosinophilic infiltration of the airways and was associated with increased eotaxin expression (both messenger RNA [mRNA] and protein) in the CD8-depleted group. There were no differences between the groups in RANTES or monocyte chemoattractant protein-1 (MCP-1) expression. In addition, we found a significantly lower interferon gamma (IFN-gamma) mRNA expression in the CD8-depleted rats, without any effects on interleukin (IL)-4 and IL-5 mRNA expression when measured either by semiquantitative reverse transcriptase/polymerase chain reaction (RT-PCR) or by in situ hybridization for the number of cells expressing these cytokines. Taken together, these results suggest that CD8(+) cells from sensitized SD rats exhibit the functional capacity to suppress the LAR, possibly through downregulation of eotaxin expression and increased expression of IFN-gamma mRNA.

Leukotrienes in bile during the early and the late airway responses after allergen challenge of sensitized rats.

The Brown Norway rat produces high levels of IgE in response to active immunization and develops both early and late airway constrictor responses after subsequent allergen challenge. We have used this model of allergic asthma to investigate the temporal relationship between the in vivo synthesis of peptidoleukotrienes (peptido-LTs) and the late response. Brown Norway rats that had been sensitized by injection of ovalbumin 2 to 3 wk prior to the commencement of the experiment were subjected to bile duct cannulation and tracheal intubation. The rats were challenged 2 h later by intratracheal instillation of ovalbumin. Lung resistance was measured before and at frequent intervals after antigen challenge. Biliary peptido-LTs (LTC4, LTD4, LTE4, and N-acetyl-LTE4) were measured by a combination of high pressure liquid chromatography and radioimmunoassay in bile samples collected for a period of 1 h before instillation of ovalbumin, and between zero and 1 h, 1 and 4 h, 4 and 6 h, and 6 and 8 h, subsequently. All of the 10 rats subjected to antigen challenge developed early responses. Of these, six also developed late responses, whereas two died about 1 h after challenge. The levels of peptido-LTs excreted in bile between 4 and 8 h after antigen challenge (corresponding in time to the late responses) were about four times higher in the ovalbumin-instilled rats that developed late responses (n = 6) than in the ovalbumin-sensitized control rats that had been subjected to instillation of saline (n = 6; p < 0.02).(ABSTRACT TRUNCATED AT 250 WORDS)