BAL and Lung Tissue Collection

Lung tissue and BAL samples were obtained as previously described [34] (link), [35] (link). Briefly, mice were anesthetized and a small-caliber tubing was inserted into the trachea. Two successive volumes of 1 ml of PBS were instilled, aspirated and pooled. BAL samples were centrifuged at 4000 rpm, and supernatants were stored at −80°C until evaluation. Cells in 100 µl aliquots were counted by trypan blue staining. A total of 100,000 viable BAL cells were centrifuged in Cytospin 3 (Thermo Shandon Ltd, Runcorn, UK). Cell differentiation was determined by Diff-quik (Fisher Scientific Co., Newark, DE). The lung was perfused with cold PBS. The whole lung was either excised for RNA and protein analyses or inflated with neutral buffered formalin for histology.