Inhaled nitric oxide reduces ischemia-reperfusion injury in rat lungs from non–heart-beating donors

If lungs could be retrieved from non–heart-beating donors, the critical shortage of lungs for transplantation could be alleviated. However, lungs subjected to warm ischemia develop edema when reperfused. We hypothesized that ventilation of rat lungs from non–heart-beating donors with nitric oxide during the period of warm ischemia alone, with reperfusion, or both might reduce ischemia-reperfusion injury.An isolated perfused rat lung model measured the filtration coefficient and accumulation of lung water by the wet/dry weight ratio. Donor rats were euthanized, and then lungs were retrieved immediately after death or 2 or 3 hours postmortem. Lungs retrieved postmortem were either not ventilated or ventilated with 100% oxygen alone or 40 ppm nitric oxide in oxygen. In the circuit, lungs were ventilated with alveolar gas with or without 40 ppm nitric oxide.Nitric oxide administration to the non–heart-beating donor or in the perfusion circuit reduced filtration coefficient and wet/dry weight ratio. Lungs retrieved 2 hours postmortem ventilated with nitric oxide or treated with nitric oxide on reperfusion had filtration coefficients and wet/dry weight ratios similar to those of lungs retrieved immediately after death. Nitric oxide was most beneficial when administered both during warm ischemia and at reperfusion in lungs retrieved 3 hours postmortem. Nitric oxide administration in the circuit was associated with increased lung levels of lung cyclic guanosine monophosphate, determined by enzyme-linked immunosorbent assay.Administration of nitric oxide to non–heart-beating donors during warm ischemia and with reperfusion might facilitate transplantation of lungs from non–heart-beating donors by reducing ischemia-reperfusion injury and capillary leak

Isoproterenol Reduces Ischemia-Reperfusion Lung Injury Despite ␤-Blockade 1

Background. If lungs could be retrieved from nonheart-beating donors (NHBDs), the shortage of lungs for transplantation could be alleviated. The use of lungs from NHBDs is associated with a mandatory warm ischemic interval, which results in ischemiareperfusion injury upon reperfusion. In an earlier study, rat lungs retrieved 2-h postmortem from NHBDs had reduced capillary leak measured by filtration coefficient (Kfc) when reperfused with isoproterenol (iso), associated with an increase in lung tissue levels of cyclic AMP (cAMP). The objective was to determine if this decrease in Kfc was because of ␤-stimulation, or would persist despite ␤-blockade. Materials and methods. Donor rats were treated intraperitoneally with ␤-blockade (propranolol or pindolol) or carrier, sacrificed, and lungs were retrieved immediately or 2 h postmortem. The lungs were reperfused with or without iso and the ␤-blockers in the reperfusate. Outcome measures were Kfc, wet:dry weight ratio (W/D), lung levels of adenine nucleotides and cAMP. Results. Lungs retrieved immediately after death had normal Kfc and W/D. After 2 h of ischemia, Kfc and W/D were markedly elevated in controls (no drug) and lungs reperfused with ␤-blockers alone. Isoproterenolreperfusion decreased Kfc and W/D significantly (P < 0.01) even in the presence of ␤-blockade. Lung cAMP levels were increased only with iso in the absence of ␤-blockade. Conclusions. The attenuation of ischemia-reperfusion injury because of iso occurs even in the presence of ␤-blockade, and may not be a result of ␤-stimulated increased cAMP. © 2005 Elsevier Inc. All rights reserved. Key Words: lung ischemia-reperfusion injury; filtration coefficient; non-heart-beating lung donor; ␤-blockade; ␤-adrenoreceptor; cyclic AMP. INTRODUCTION Lung transplantation is a successful therapy that palliates many patients with a variety of types of end stage lung disease, but its widespread use is limited by a lack of suitable donors [1]. If lungs could be retrieved from non-heart-beating donors (NHBDs), the critical shortage of lungs for transplantation could be alleviated. This shortage of donor lungs has prompted our laboratory to investigate the use of NHBDs in experimental models of lung transplantation Using an isolated perfused rat lung model (IPRLM), we have investigated the impact of increasing postmortem ischemic time on the early phase of ischemia-reperfusion injury (IRI) independent of circulating leukocytes, assessed by measuring the filtration coefficient (Kfc) using a blood free reperfusate. We demonstrated that rat lungs retrieved 1 h post-mortem from NHBDs had substantially increased Kfc and wet:dry weight ratio (W/D), ameliorated by ventilation of the cadaver with oxygen, and that rat lungs retrieved 2 h postmortem from NHBDs had very elevated Kfc and W/D ratio irrespective of cadaver ventilation Isoproterenol is a potent ␤-adrenoreceptor agonist that activates adenylate cyclase to increase intracellular cAMP. Propranolol (pro) is a non-selective ␤-blocker that acts as a competitive inhibitor of iso MATERIALS AND METHODS Challenge Test Separate experiments established the appropriate dose of pro (Sigma Chemical Co., St. Louis, MO) and pin (ICN Biomedical Inc., Aurora, OH) to achieve ␤-blockade in the rat. Male Sprague-Dawley rats were anesthetized by i.p. injection of pentobarbital sodium (60 mg/kg) (Abbott Laboratories, Chicago, IL), and the trachea was cannulated. Then, ␤-blocked rats were treated with different doses of pro (2 mg/ml saline) or pin (0.4 mg/ml saline) i.p. The control rat (no ␤-blocker or iso-only group) received a similar volume of saline. After waiting for 15 min to allow time for absorption and attainment of therapeutic serum levels, the degree of ␤-blockade was demonstrated by i.v. administration of 0.1 mg/kg iso (Sigma Chemical Co.) administered via the penile vein in a concentration of 0.1 mg/ml saline Isolated Perfused Rat Lung Model The isolated perfused rat lung model (IPRLM) was first used to measure Kfc in dogs Briefly, 60 donor rats weighing 250 to 400 g were anesthetized by i.p. injection of pentobarbital sodium (60 mg/kg) (Abbott Laboratories), and the trachea was cannulated. Then, rats were treated with the ␤-blockers (4 mg/kg pro or 0.8 mg/kg pin) or a similar volume of saline i.p. After 15 min, a small laparotomy incision was made and 600U of heparin (Elkins-Sinn, Cherry Hill, NJ) was injected i.t. under direct vision. The rats were sacrificed with an i.t. injection of pentobarbital sodium (100 mg/kg). Cardiac arrest was documented by observation of cardiac motion transmitted through the diaphragm and by palpation. The heart-lung block was left in situ in an effort to simulate the NHBD's clinical situation as closely as possible. The heart, lungs and mediastinal structures were retrieved en bloc via median sternotomy immediately after death or 2 h postmortem from non-ventilated NHBDs. Subsequently, the main pulmonary artery and the left atrium were cannulated for ventilation and perfusion in the IPRLM as described in detail in an earlier publication Pressure transducers (Cope Laboratories, Lakewood, CO) were positioned at the hila of the lungs, zeroed to atmospheric pressure, and calibrated with a mercury manometer. Pulmonary arterial (Ppa) and venous (Ppv) pressures were measured continuously. In addition, pulmonary airway pressure (Paw) was continuously monitored by positioning a T-tube on the inspiratory limb of the respiration plumbing. All pressure measurements and changes in weight gain were collected by Lab View SCXI Pressure Weight Acquisition System (National Instruments, Austin TX) on a Gateway personal computer. Pulmonary capillary pressure (Ppc) was estimated by the doubleocclusion technique described by Townsle