Analysis of induced sputum has provided significant insight into the inflammatory response in chronic respiratory diseases such as asthma. The thick, tenacious nature of cystic fibrosis (CF) sputum presents certain challenges to such evaluation. We describe the development of a methodology to assess CF sputum, and the within-week reproducibility (to limit the possibility of any change in clinical status) of cellular and inflammatory markers.
Methods: Seventeen young adults [9 males, 8 females, mean age 24 (5), median (quartile range) years, percentage of predicted FEV(1) = 64.0 (18.0%)] with CF underwent sputum inductions on the Monday and Thursday of the same week. Patients were pretreated with 400 microg salbutamol and subsequently inhaled 5% saline via a breath enhanced nebulizer. Every 3-min nebulization was interrupted to allow for expectoration of sputum into a polypropylene pot. Sputum samples were dispersed with a solution of dithiothreitol (DTT) and deoxyribonuclease to allow for analysis of total cell count (TCC) and percentage neutrophils (%Neut). Measurement of tumor necrosis factor-alpha (TNF-alpha), interleukin-8 (IL-8), and neutrophil elastase was performed on samples dispersed with DTT alone.
Results: There were no significant differences between the measurements taken in the same week. Values for Day 1 versus Day 2 were as follows: TCC 20.8 (6.4) vs. 17.6 (2.5) x 10(6) cells/ml; %Neut: 94.1 (0.0) vs. 95.4 (0.5) %; TNF-alpha 7.5 (26.0) vs. 21.0 (44.0) pg/ml; IL-8 610.0 (422.0) vs. 524.0 (587.0) ng/ml and neutrophil elastase 110.0 (19.75) vs. 49.75 (60.75) microM. High intraclass correlation coefficients (ICC) for TCC, %Neut, TNF-alpha IL-8 and neutrophil elastase were found (ICC = 0.76, 0.82, 0.93, 0.82, 0.74, respectively).
Conclusions: The method developed here for the analysis of CF sputum shows good reproducibility and can be used to evaluate therapeutic interventions in patients with cystic fibrosis.