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Original Article

Fluorocytometric analysis of induced sputum cells in an asthmatic population

J. Domínguez Ortega, F. León+#, JC Martínez Alonso, A. Alonso Llamazares*,
E. Roldán+, T. Robledo*, M. Mesa*, A. Bootello+, C. Martínez-Cócera*

Allergy Unit. Hospital Virgen de la Concha. Zamora, Spain Lab. of Clinical Investigation/NIAID/NIH. Bethesda, USA,
* Department of Allergy. H. Clínico San Carlos
+ Department of Immunology. H. Ramón y Cajal. Madrid. Spain

J Invest Allergol Clin Immunol 2004; Vol. 14(2): 108-113



Background: Bronchial mucosal inflammation is the major pathogenic process in asthma. In the latest years, induced sputum (IS) examination has become an important non-invasive method of assessing airway inflammation. Flow cytometry has been recently applied to the study of IS though it is not exempt of methodological difficulties.

The aim of the present study was to further study if the fluorocytometric analysis of IS could represent a reliable tool to assess the presence of bronchial activated lymphocytes in stable mild asthmatic patients.

Induced sputa from controls and asthmatic patients were processed in isotonic 3mM dithiothreitol (DTT), a mucolytic agent required for cell dispersion. The individualized cells were then stained with monoclonal antibodies for three-colour flow-cytometric analysis. Total IgE and ECP were measured in serum and in the sputum fluid

Results: The cellularity of asthmatic sputa is enriched in eosinophils (mean, 26.63%) with respect to controls, but not in lymphocytes. However, lymphocytes from asthmatics show increased surface expression of activation markers (CD25 in T cells, CD23 in B cells). Surprisingly, no differences were observed in the detected levels of CD54 on
IS lymphocytes and eosinophils between asthmatics and non-asthmatics. Furthermore, there was a significantly higher concentration of ECP and total IgE in the sputum from the asthmatic group.

Conclusion: Fluorocytometric analysis of induced sputum is a reliable non-invasive method for the study of bronchial immune cells. It could provide complementary information on activated cells in the bronchial mucosa even in non-smokers, mild and stable asthmatics and it is reasonable to speculate that it will be useful in monitoring the effect of the treatment in these patients.

Key words: Induced sputum, Asthma, ECP, Adhesion molecules, CD23, CD25, Lymphocytes, Eosinophils