Objective: Pseudomonas aeruginosa-inuced lung injury is characterized not only by the alteration in lung fluid movement but also by apoptosis of lung epithelial and endothelial cells. We studied whether inhibition of apoptosis using a broad spectrum caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp fluoromethylketone (Z-VAD.fmk), would affect lung fluid balance in rat P. aeruginosa pneumonia. Methods: Z-VAD.fmk (3 mg/kg) was administered intravenously simultaneously with P. aeruginosa intratracheal instillation (0.5 ml/kg, 2×109 CFU/ml). Apoptosis was evaluated with the TUNEL technique, cytoplasmic oligonucleosome assay, and caspase 3 activation. To evaluate lung permeability, extravascular plasma equivalent (EPE) and lung wet to dry weight ratio (W/D) were measured 4 h after intratracheal instillation of P. aeruginosa. Results: We found an increase of lung apoptosis 4 h after P. aeruginosa instillation: cytoplasmic oligonucleosome assay increased from 3.17±0.78 to 26.82±4.67 ODx1000/mg of proteins/ml, Z-VAD.fmk administration decreased this parameter to 10.3±2.98 ODx1000/mg of proteins/ml. Caspase 3 levels followed the same pattern. Apoptosis involved both epithelial cells and endothelial cells. Endothelial permeability was increased after Pseudomonas instillation: W/D increased from 3.75±0.28 in the Co group to 4.42±0.23 in the Pn group; EPE was also higher in the Pn group compared with the Co group (0.125±0.04 and 0.002±0.01 ml, respectively). Both of these parameters were improved after Z-VAD.fmk administration; W/D decreased to 3.36±0.25 and EPE to 0.02±0.02 ml. Conclusion: Apoptosis occurs in the early phase of P. aeruginosa pneumonia. Administration of Z-VAD.fmk significantly decreases DNA fragmentation and caspase 3 levels. This is associated with an improvement of endothelial permeability and lung fluid balance. © Springer-Verlag 2004.