Collection of Bronchoalveolar Lavage Fluid (BALF) and Total Protein Analysis

At the time of lavage, the mice were anesthetized with an i.p. injection of 1% pentobarbital sodium (50 mg/kg). The chest cavity was exposed, and then the trachea was intubated. PBS (1 mL) was slowly instilled into the lungs and withdrawn. The collected solutions were centrifuged at 1000 × g for 10 min at 4°C. The supernatant was harvested and stored at −80°C for further analysis. The total protein concentration of cell-free bronchoalveolar lavage fluid (BALF) was determined by the BCA method.