Il 13

Rg3 and dexamethasone (Dex) were obtained from Sigma-Aldrich (St. Louis, MO, USA). Interleukin-1 beta (IL-1β) was obtained from R&D system Inc. (Minneapolis, MN, USA). Rabbit anti-phospho-NF-κB p65, anti-NF-κB p65, and anti-COX-2 were purchased from Cell Signaling Tech (Beverley, MA, USA). Mouse anti-β-actin, goat anti-rabbit IgG-HRP, and goat anti-mouse IgG-HRP were purchased from Santa Cruz Biotechnology (Dallas, TX, USA). Assay Media Sterile was purchased from MatTek Corp. (Ashland, MA, USA). Human Cytokine 6-plex Flat Bottom Express (TNF-α, eotaxin, IL-4, IL-5, IL-9, and IL-13) and protein assay reagent were purchased from Bio-Rad (Hercules, CA, USA). Protease inhibitor cocktail was obtained from Roche Diagnostics (Mannheim, Germany).

Mice were euthanized using CO₂ and their lungs were surgically exposed. The trachea was cannulated and the lungs were lavaged with 5 aliquots of 800 µL of 0.9% saline (VWR International, West Chester, PA). Bronchoalveolar lavage (BAL) fluid supernatant was stored at −80°C until further use. BAL fluid total protein was quantified with a total protein assay kit using Bicinchoninic Acid (Thermo Scientific, Waltham, MA) and Granzyme B was measured using a specific ELISA kit (R&D systems, Minneapolis, MN). Cytokine analysis was performed on the BAL fluid using mouse 23-plex and 9-plex cytokine assays that included detection of IFN-γ, IL-4, IL-5, IL-13, IL-17, IL-10, IL-15, and CCL5 (Bio-Rad Laboratories, Hercules, CA).