Granulocyte markers in induced sputum in patients with respiratory disorders and healthy persons obtained by two sputum-processing methods

Induced sputum is increasingly used to detect and monitor airway inflammati on in respiratory diseases. However, the processing of sputum has been rath er laborious for clinical practice. The aim of this study was to improve the practicality of induced-sputum stu dies by simplifying sample processing. Eosinophil cationic protein (ECP), e osinophil peroxidase (EPO), neutrophil lipocalin (HNL) and myeloperoxidase (MPO) were used as biochemical markers of airway inflammation and the resul ts of the study method were compared with a previously validated (reference ) method. Induced sputum was obtained from 42 healthy controls, 10 subjects with acut e respiratory infection, eight patients with chronic obstructive pulmonary disease (COPD) and 17 asthmatics. The sputum sample was divided into two pa rts and treated either: (i) by the reference method (released markers), whe re sputum was homongenized with dithiotreitol and centrifuged to yield cell -free supernatant and a cell pellet, or (ii)by the study method (total mark ers), where the cells were lysed after homogenization so that cell-associat ed markers were released and solubilized. For comparison, the four biochemi cal markers were measured in sputum supernatant and in sputum lysstle. The differential cell count was performed from the cell pellet in the reference method. Repeatability was assessed in a group of 16 subjects. The effect o f reagents and the recovery of assays were also evaluated. Released and total markers correlated well (ECP r(s) = 0.80, P < 0.0001; EP O r(s) = 0.49, P < 0.0001; HNL r(s) = 0.87, P < 0.0001; MPO r(s) = 0.71, P < 0.0001). Incubation with dithiotreitol and lysis reagent had no negative influence on marker assays. The within-subject variability of total ECP, MP O and HNL in both methods was small in two measurements taken 1 week apart. The study method, measuring total inflammatory markers, gave comparable res ults to the reference method, measuring released markers. In the study meth od the sputum processing was simplified, which may improve its applicabilit y.