Effect of NADPH oxidase inhibition on cardiopulmonary bypass-induced lung injury.

Cardiopulmonary bypass (CPB) causes acute lung injury. Reactive oxygen species (ROS) from NADPH oxidase may contribute to this injury. To determine the role of NADPH oxidase, we pretreated pigs with structurally dissimilar NADPH oxidase inhibitors. Low-dose apocynin (4-hydroxy-3-methoxy-acetophenone; 200 mg/kg, n = 6), high-dose apocynin (400 mg/kg, n = 6), or diphenyleneiodonium (DPI; 8 mg/kg) was compared with diluent (n = 8). An additional group was treated with indomethacin (10 mg/kg, n = 3). CPB was performed for 2 h with deflated lungs, complete pulmonary artery occlusion, and bronchial artery ligation to maximize lung injury. Parameters of pulmonary function were evaluated for 25 min following CPB. Blood chemiluminescence indicated neutrophil ROS production. Electron paramagnetic resonance determined the effect of apocynin and DPI on in vitro pulmonary endothelial ROS production following hypoxia-reoxygenation. Both apocynin and DPI attenuated blood chemiluminescence and post-CPB hypoxemia. At 25 min post-CPB with Fi(O(2)) = 1, arterial Po(2) (Pa(o(2))) averaged 52 +/- 5, 162 +/- 54, 335 +/- 88, and 329 +/- 119 mmHg in control, low-dose apocynin, high-dose apocynin, and DPI-treated groups, respectively (P < 0.01). Indomethacin had no effect. Pa(O(2)) correlated with blood chemiluminescence measured after drug administration before CPB (R = -0.60, P < 0.005). Neither apocynin nor DPI prevented the increased tracheal pressure, plasma cytokine concentrations (tumor necrosis factor-alpha and IL-6), extravascular lung water, and pulmonary vascular protein permeability observed in control pigs. NADPH oxidase inhibition, but not xanthine oxidase inhibition, significantly blocked endothelial ROS generation following hypoxia-reoxygenation (P < 0.05). NADPH oxidase-derived ROS contribute to the severe hypoxemia but not to the increased cytokine generation and pulmonary vascular protein permeability, which occur following CPB.

Effect of bronchial artery blood flow on cardiopulmonary bypass-induced lung injury.

Cardiovascular surgery requiring cardiopulmonary bypass (CPB) is frequently complicated by postoperative lung injury. Bronchial artery (BA) blood flow has been hypothesized to attenuate this injury. The purpose of the present study was to determine the effect of BA blood flow on CPB-induced lung injury in anesthetized pigs. In eight pigs (BA ligated) the BA was ligated, whereas in six pigs (BA patent) the BA was identified but left intact. Warm (37 degrees C) CPB was then performed in all pigs with complete occlusion of the pulmonary artery and deflated lungs to maximize lung injury. BA ligation significantly exacerbated nearly all aspects of pulmonary function beginning at 5 min post-CPB. At 25 min, BA-ligated pigs had a lower arterial Po(2) at a fraction of inspired oxygen of 1.0 (52 +/- 5 vs. 312 +/- 58 mmHg) and greater peak tracheal pressure (39 +/- 6 vs. 15 +/- 4 mmHg), pulmonary vascular resistance (11 +/- 1 vs. 6 +/- 1 mmHg x l(-1) x min), plasma TNF-alpha (1.2 +/- 0.60 vs. 0.59 +/- 0.092 ng/ml), extravascular lung water (11.7 +/- 1.2 vs. 7.7 +/- 0.5 ml/g blood-free dry weight), and pulmonary vascular protein permeability, as assessed by a decreased reflection coefficient for albumin (sigma(alb); 0.53 +/- 0.1 vs. 0.82 +/- 0.05). There was a negative correlation (R = 0.95, P < 0.001) between sigma(alb) and the 25-min plasma TNF-alpha concentration. These results suggest that a severe decrease in BA blood flow during and after warm CPB causes increased pulmonary vascular permeability, edema formation, cytokine production, and severe arterial hypoxemia secondary to intrapulmonary shunt.

Effect of cGMP on lung microvascular endothelial barrier dysfunction following hydrogen peroxide.

The authors determined the effect of cyclic guanosine 3',5'-monophosphate (cGMP) on hydrogen peroxide (H(2)O(2))-induced barrier dysfunction in bovine lung microvascular endothelial cell (BLMVEC) monolayers and compared the results to bovine pulmonary artery endothelial cells (BPAECs). In BLMVECs, H(2)O(2) (250 microM) caused a 31.9% +/- 4.8% decrease in transendothelial electrical resistance (TER) associated with increased actin stress fiber formation, intercellular gaps, and intracellular calcium concentration ([Ca(2+)](i)). The cGMP analogue 8-(p-chlorophenylthio)-cGMP (8p-CPT-cGMP; 30 or 50 microM) prevented the H(2)O(2)-induced decrease in TER (p <.001) as well as the cytoskeletal rearrangement and intercellular gap formation. 8-pCPT-cGMP (50 microM) attenuated the peak (418.8 +/- 42.1 versus 665.2 +/- 38.0 nmol/L; p <.001) and eliminated the sustained increase in [Ca(2+)](i) (193.5 +/- 21.3 versus 418.8 +/- 42.1 nmol/L; p <.001) caused by H(2)O(2). 8-pCPT-cGMP also increased TER (14.2% +/- 2.2%; p <.05) and decreased [Ca(2+)](i) (201.2 +/- 12.5 vs. 214.4 +/- 12.1 nmol/L; p <.03) before H(2)O(2). In BPAECs, 8p-CPT-cGMP significantly attenuated H(2)O(2)-induced increases in permeability and [Ca(2+)](i) but less effectively than in BLMVECs. These results suggest that in BLMVECs, cGMP countered the adverse effects of H(2)O(2) on barrier function by preventing actin cytoskeletal rearrangement and attenuating the increase in [Ca(2+)](i).

Inhibition of inwardly rectifying K(+) channels by cGMP in pulmonary vascular endothelial cells.

Endothelial barrier dysfunction is typically triggered by increased intracellular Ca(2+) concentration. Membrane-permeable analogs of guanosine 3',5'-cyclic monophosphate (cGMP) prevent disruption of endothelial cell integrity. Because membrane potential (E(m)), which influences the electrochemical gradient for Ca(2+) influx, is regulated by K(+) channels, we investigated the effect of 8-bromo-cGMP on E(m) and inwardly rectifying K(+) (K(IR)) currents in bovine pulmonary artery and microvascular endothelial cells (BPAEC and BMVEC), using whole cell patch-clamp techniques. Both cell types exhibited inward currents at potentials negative to -50 mV that were abolished by application of 10 microM Ba(2+), consistent with K(IR) current. Ba(2+) also depolarized both cell types. 8-Bromo-cGMP (10(-3) M) depolarized BPAEC and BMVEC and inhibited K(IR) current. Pretreatment with Rp-8-cPCT-cGMPS or KT-5823, protein kinase G (PKG) antagonists, did not prevent current inhibition by 8-bromo-cGMP. These data suggest that 8-bromo-cGMP induces depolarization in BPAEC and BMVEC due, in part, to PKG-independent inhibition of K(IR) current. The depolarization could be a protective mechanism that prevents endothelial cell barrier dysfunction by reducing the driving force for Ca(2+) entry.