Schematic timelines of the formation of the air pouch (A) and the time course experiment (B). A. Three ml of filtered air are injected under the dorsal skin. The resulting air pouch is reinflated on day 3 with an additional injection of 2 ml. A newly formed thin membrane lines the pouch lumen by day 6. B. Two mg endotoxin-free MSU crystals are injected into the pouch on day 6. At the indicated time points (x-axis), the inside of the pouch is lavaged with PBS to determine the number of leukocytes, and the pouch membrane is then isolated from the surrounding tissues by blunt dissection and processed for gene expression analysis. Four mice were used at t = 1h and t = 4h, and 3 mice at each of the other time points.