The influence of bronchoalveolar and circulating tumor necrosis factor-alpha on apoptosis in different models of lung injury

Abstract

Cytokines are involved in a variety of lung diseases, but their pathogenetic role in programmed cell death is still controversial. This study tests whether the activation of an ‘extrinsic’ pathway to cell death is mediated by bronchoalveolar and circulating tumor necrosis factor (TNF)-alpha. Nonlinear male rats weighing 200-230 g were used in all the experiments. For modeling of acid aspiration-induced acute lung injury anesthetized rats underwent tracheostomy and insertion of a fine-bore cannula into the anterior segment of the left lung. This was followed by the instillation of either 1.0 mL/kg HCI, pH 1.2 (n = 12) or 1.0 mL/kg saline in control rats (n = 12). All animals were studied at 6-hour after acid aspiration. For modeling of hepatopulmonary syndrome anesthetized rats underwent common bile duct ligation (CBDL) (n = 12). Sham rats underwent mobilization of the common bile duct without ligation (n = 12). All the animals were studied at 31-day after CBDL or sham operation. Bronchoalveolar lavage (BAL) and blood serum were analyzed for TNF- levels in pg/ml using commercially available ELISA kits. The level of apoptosis was analyzed with the help of Annexin V-FITS and propidium iodide using Beckman Coulter flow cytometer. By 6 h after acid aspiration TNF-alpha values were significantly higher than in control group (in the blood serum: 17,06±1,91 pg/ml vs 11,54±0,59 pg/ml, p<0,001, in the BAL: 16,39±0,80 vs 1,73±0,13, p<0,001). It was also a significant increase in the number of early apoptotic cells present (in the blood serum: 0,63±0,14 vs 0,45±0,03, p<0,001, in the BAL: 2,41±0,15 vs 0,61±0,05, p<0,001). At 31-day after CBDL TNF-alpha values were also significantly higher than in control group (in the blood serum: 46,36±2,33 pg/ml vs 10,35±1,90 pg/ml, p<0,001, in the BAL: 11,50±0,77 vs 2,06±0,44, p<0,001). It was also a significant increase in the number of early apoptotic cells present (in the blood serum: 2,02±0,35 % vs 0,60±0,09 %, p<0,01, in the BAL: 2,77+0,45 % vs 0,47±0,06 %, p<0,01). In acid aspiration-induced acute lung injury and hepatopulmonary syndrome the high level of TNF-a is positively correlated with the increasing level of early apoptosis. In the future there will be investigated TNF-a receptors: TNF-RI and TNF-RII.