Fosinophils may be important in the pathophysiology of exercise-induced bronchoconstriction (EIB) and the response to therapy in patients with chronic asthma (read more). In vitro studies have demonstrated that eosinophils generate and release cysteinyl leu-kotrienes when subjected to a hyperosmolar stimulus, which is an important condition that provokes EIB. Cysteinyl leukotrienes are potent bronchocon-strictor mediators that are implicated in EIB. Furthermore, the presence and severity of EIB is significantly correlated to eosinophil levels measured in the blood and sputum of asthmatic patients. Beyond these observations, the relationship between eosinophilic inflammation and EIB has not been fully explored.
In the present study, the relationship among sputum eosinophil levels, EIB, and the response to ICS therapy was examined. We previously reported on the heterogeneity of the response in EIB to four different doses of ciclesonide, as suggested by an increase in the coefficient of variability of the fall in FEV1 with exercise during treatment compared to baseline. From the same data, we now report on the secondary outcomes, examining the correlation between sputum eosinophil levels and EIB; and whether sputum eosinophil percentages predict the response of EIB to inhaled corticosteroid (ICS) therapy. We hypothesized that airway eosinophils contribute to the mechanisms underlying EIB and the therapeutic effect of ICS therapy. Therefore, high sputum eosinophil levels predict a greater severity in EIB and a response to ICS therapy that is mediated through the suppression of eosinophilic inflammation. In contrast, low eosinophil count is associated with relative ICS insensitivity, which cannot be overcome by increasing the dose or duration of ICS therapy.
Materials and Methods
Asthmatic subjects between 12 and 30 years of age and a baseline FEV1 of > 70% predicted were enrolled into the study. All subjects demonstrated a maximum percentage fall in FEV1 from baseline after exercise challenge of > 15% (þV1 fall) at screening and after a 1-week run-in period. Exclusion criteria included unstable asthma (< 6 weeks), regular medication use (except salbutamol), comorbidity, and current or ex-smokers with a history of > 10 pack-years of smoking. Subjects were randomized to the following two parallel, double-blind, crossover arms, with each arm comparing two doses of ciclesonide: 40 vs 160 Î¼g once daily and 80 vs 320 Î¼g once daily for 3 weeks, with a washout period of 3 to 8 weeks. The data were pooled to give two dose levels of ICS therapy (low-dose therapy, 40 and 80 Î¼g; high-dose therapy, 160 and 320 Î¼g), based on previous findings that low doses (equivalent to budesonide, < 100 Î¼g daily) are ineffective in attenuating chronic sputum eosinophilia; therefore, we anticipate ICS therapy having a limited effect on attenuating EIB. Each period comprised four visits, as follows: baseline; and weekly visits with an exercise challenge and sputum induction. The last dose of ciclesonide was administered 12 h prior to visits. Compliance was assessed by the weighing of canisters. The study was approved by the hospital research ethics board and informed consent was obtained from participants.
Spirometry was performed according to American Thoracic Society standards and reference values taken from Hankinson et al. Exercise challenge was performed on a treadmill for 8 min with continuous heart-rate monitoring and the patient breathing dry air (temperature, 21Â°C; relative humidity, < 10%). The treadmill was at a 10% incline, and the speed was adjusted to achieve 90% of the predicted maximum heart rate (210 â€” age) for the last 4 min of the test. After, the speed was reduced, and subjects walked while breathing dry air for another 2 min. FEV1 was measured at baseline, and at 2, 4, 6, 8, 10, 15, 20, and 30 min postexercise. The severity of EIB was expressed as the þV1 fall postchallenge (ie, preexercise FEV1 â€” lowest FEV1 postexercise/preexercise FEV1 X 100). Area under the curve for FEV1 fall up to 30 min postexercise (AUC0â€”30) was calculated using the trapezoidal rule.
Sputum induction and processing were performed as previously described with selected cell plugs treated with 0.1% dithiothreitol (Sputolysin; Calbio-chem Corp; La Jolla, CA) and Dulbecco phosphate-buffered saline solution (Life Technologies Inc; Gaithersburg, MD). This sample was filtered and centrifuged, and the supernatant was separated. Total cell count (TCC) was determined using a Neubauer hemocytometer chamber (Hausser Scientific; Horsham, PA) and cytospins prepared on slides. Differential cell counts were performed manually (400 nonsquamous cells and metachromatic cells of a total of 1,500 cells). Results are expressed as the percentage of the TCC.
This was a pilot study with 24 subjects as no data were available on the variability of the difference in EIB attenuation caused by ICS therapy. Subjects were divided into noneosinophilic and eosinophilic groups using a baseline sputum eosinophil cutoff of 5%. Sputum cell percentages were log-transformed. To compare the dose-time response of the þV1 fall, AUC0 30, and sputum cell percentage, a multiplicative analysis-of-variance model was used with the following factors: dose; visit; treatment sequence, and eosinophilic group, A mixed-effect regression model was used to examine the rate (slope) of improvement in þV1 fall over time by dose. The significance of eosinophil percentage as a predictor of severity and the response to ICS therapy was tested by its interactions with baseline þV1 fall and the rate of change in þV1 fall over time. A p value (two-sided) of < 0.05 was accepted as significant. All analyses were performed using a statistical software package (SPSS, 12.0; SPSS, Inc; Chicago, IL).