Materials and Methods

Lung Slices and Treatments Three female mice (C57Bl/6 or BALB/c; Charles River Laboratories, Kingston, NY) in each experimental group were anesthetized with sodium pentobarbital (100mg/kg, intraperitoneally). The thoracic cavity was opened, the dorsal aorta was cut, and the lungs were perfused free of blood with phosphate-buffered saline. After excision of the lungs, the lobes were separated and slices were cut «1 mm) with a scalpel on a glass plate.

Slices were equilibrated by three washes for 5 min each at room temperature in Krebs-Henseleit buffer (pH 7.2)(29) containing 4.5 g glucose (Sigma Chemical Co., St. Louis, MO) per liter (Krebs/glucose buffer). After the equilibration period, three to four slices (10 to 15 mg tissue) were distributed in 24-well culture plates in 1.0 ml Krebs/glucose and shaken (30). The samples were pre-incubated at 37 C for 15 min in 1.0 ml buffer with or without 2.5 mM 3-AB (Sigma) to inhibit PAP. This concentration of 3-AB is selective for polymerase inhibition based on the results of other investigators (31, 32).