Differential cell counting
To differentiate the various cell populations in the BALF, cytocentrifuged cell preparations (Shandon Cytospin 3, 400 RPM, 10 min) were used, with ~5x104 cells per slide. Dried cytospots were subjected to a Shandon Kwik-Diff® Fixative (Thermo Fischer Scientific, USA), followed by Hemacolour® eosin and thiazine differential stains (Merck, USA) as outlined in the manufacturer’s instructions. Cell types (i.e. macrophages, neutrophils and lymphocytes) were identified according to standard morphological criteria, using the above-mentioned microscope, with at least 500 cells counted per slide.