Effects of a Neutrophil Elastase Inhibitor (ONO-5046) on Acute Pulmonary Injury Induced by Tumor Necrosis Factor Alpha (TNF α ) and Activated Neutrophils in Isolated Perfused Rabbit Lungs

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Abstract
The aim of this study was to examine the effect of ONO-5046, a neutrophil elastase (NE) inhibitor, on a model of acute lung injury induced by tumor necrosis factor α (TNF α ) and phorbol myristate acetate (PMA)-activated neutrophils in isolated perfused rabbit lungs. 120 min after TNF α (4,000 JRU/ml) was injected into the pulmonary artery (PA), 5 × 107 PMA-stimulated neutrophils were infused into the PA together with 125I-rabbit serum albumin (RSA). In the ONO-5046-treated group (ONO), ONO-5046 (20 mg/kg/h) was continuously infused during the experimental period from 30 min prior to neutrophil administration. Saline, the ONO-5046 vehicle, was infused instead of ONO-5046 in the positive control group (ALD) and nonactivated neutrophils were infused without TNF α in the negative control group (Cont). PA pressure was monitored over a 240 min period, and bronchoalveolar lavage (BAL) was performed at the end of the experiment. Lung tissues were examined immunohistochemically for the expression of thrombomodulin (TM). The levels of TM in the perfusate were also measured by ELISA and the radioactivities in the BAL fluid, lung tissue and perfusate were determined to calculate the permeability index (PI) as an indicator of alveolar septal or vascular endothelial damage. The rabbit lungs infused with ONO-5046 showed slower and less increases in PA pressure compared with ALD group. The PI was significantly higher in ALD group (PIBAL = 0.028 ± 0.014, PILUNG = 0.04 ± 0.003) than Cont (PIBAL = 0.002 ± 0.001, PILUNG = 0.015 ± 0.003) and ONO group (PIBAL = 0.004 ± 0.003, PILUNG = 0.028 ± 0.003 (p < 0.05). ALD group had higher TM levels in the perfusate and showed decreased expression of TM on the vascular endothelium compared to Cont and ONO group, suggesting that there was shedding of TM on endothelium and ONO-5046 attenuated a shedding of TM. In conclusion, ONO-5046 attenuated acute lung injury by inhibiting the alveolar epithelial and vascular endothelial injury triggered by activated neutrophils. NE appears to play an important role in the neutrophil-induced increase of pulmonary epithelial and microvascular permeability observed in acute lung injury.