All of the animal experiments were approved by the institutional animal research committee of the University of Washington. Mice were housed in a pathogen-free environment according to University of Washington animal use guidelines. Male C57BL/6 mice ("B6") and mice carrying a spontaneous mutation in the Fas gene that impairs Fas signaling (B6. MRL-Fas lpr /J, "lpr") were obtained from the Jackson Laboratories and studied at 7-13 weeks of age. Briefly, the mice were anesthetized with 5% inhaled isoflurane and then intubated endotracheally with a 20-gauge angiocatheter. Placement of the catheter in the trachea was verified by visualizing the movement of a 100 μl bubble of water in response to respiratory efforts. After confirming intubation, the trachea was instilled with either E. coli LPS, 15 ng/kg or PBS. The instillate was suspended in 2.5% colloidal carbon to allow later confirmation of the extent and distribution of the instillation macro and microscopically. After the installations some of the mice were extubated, returned to their cages, and allowed free access to food and water; other mice where kept intubated and subjected to mechanical ventilation with the following settings: tidal volume (Tv) 10 ml/kg; respiratory rate (RR) 150 breaths/minute; fraction of inspired oxygen (FiO 2 ) 0.21; and positive end-expiratory pressure (PEEP) of 3 cm H 2 O. The heart rate, airway pressures, rectal temperatures and EtCO 2 were monitored continuously using a computerized monitoring system (Chart 4, AD Instruments, Colorado Springs, CO). The RR was adjusted to maintain the EtCO 2 between 30 -40 mmHg. The body temperature was maintained between 37 and 38°C with external heating. The mice were hydrated with a continuous intraperitoneal infusion of lactated ringer solution at 500 μl/hour. Muscle relaxation was attained with pancuronium bromide, 1 μg/g i.p followed by 0.5 μg/g i.p. every hour. After four hours of mechanical ventilation the mice were euthanized with 0.30 ml/kg i.p. of Beuthanasia-D (Schering-Plough Animal Health, Union, NJ). The thorax was rapidly opened and the mouse was exsanguinated by direct cardiac puncture. The left lung was removed and flash-frozen in liquid nitrogen. The right lung was lavaged with 0.6 mM EDTA in PBS; an aliquot of the bronchoalveolar lavage fluid (BALF) was removed for cell counts and differentials, the remaining fluid was spun at 1200 x g, and the supernatants stored at -80°C in individual aliquots. Following the BAL, the right lung was fixed in 4% buffered paraformaldehyde at an inflation pressure of 15 cm H 2 O for histological evaluation.