The Protective Effect Of N-acetylcysteine In Donation After Cardiac Death (DCD) Lung

Lung transplantation is the most effective treatment for end-stage lung disease, but itis facing with the lack of donors. In recent years, Donation after Cardiac Death（DCD）,also termed Non-Heart-Beating Donor(NHBD) before, has increased everyyear and achieved a good clinical results. DCD can be divided into controlled（cDCD）anduncontrolled donor（uDCD）.In the process of pulmonary transplantation with DCDlung,the graft experienced an severe ischemia-reperfusion injury (IRI), which can leadto early primary graft dysfunction (PGD).Although the pathogenesis of IRI is complex, ithas been confirmed that the generation of oxygen free radical and activation of neutrophilwere involved in this injury process.Recently,a novel technique (Ex vivo lung perfusion, EVLP) was presented. EVLP helpsto reduce pulmonary edema, attenuate IRI, and finally reduce the incidence rates of PGD,improve the utilization of DCD lungs. EVLP has been used as a platform to evaluate andpretreat donor lung.In addition, recent studies have proved that N-acetylcysteine(NAC) can inhibitneutrophil generation，thus inhibited NF-kB which was induced by ROS/RNS，avoidlipid peroxidation and cell apoptosis. In vivo，NAC can activate guanylate cyclase,elevate plasma cyclic guanosine monophosphate(cGMP) level, increase the biologicaleffects of NO, inhibit interleukins and platelet activating factors（PAF），and attenuateearly IRI, whether NAC could protect lung from DCD is still unknown. In the first part of this study, we established a rabbit DCD lung model anddeveloped an Ex Vivo Lung Perfusion (EVLP) device. The device was evaluated. In thesecond part of this study, we explored whether NAC could provide protective effect forDCD lung.The results showed the rabbit DCD lung model and EVLP has good reliability andrepeatability. It can be used not only obtain respiratory parameters(peak airway pressureand arterial blood gas)，but also evaluate pre-transplantation pulmonary function.EVLP can be used as a good platform for the study of lung protection. NAC cansignificantly increase the lung GSH level，inhibit the active oxygen free radicals,suppress the release of inflammatory substances, decrease the level of MPO,MDA, andIL-1，and reduce airway resistance. It has a significant protective effect on DCD lung.In summary，this rabbit Ex Vivo Lung Perfusion Model is stable and has agood reproducibility. NAC can attenuate IRI, and might be an alternative approach toprotect lung from DCD. Part I: Establishment of Rabbit Ex Vivo Lung Perfusion(EVLP) Device.Objective: To evaluate Ex Vivo Lung Perfusion (EVLP) we developed on DCD lung.Methods: Six Male New Zealand white rabbits (body weight2-2.5kg) were anesthetizedand undergoing thoractomy. A drainage tube for bloodletting was inserted into thecardiac apex，blood withdrawn (i.e., heart arrest were induced by exsanguinations) andstored in a special blood-collecting bag. After one hour “no touch” period and left in theroom temperature (DCD lung animal model established), donor lung were flushed withcold Perfadex solution (antegrade and retrograde successively), then harvested andimmersed immediately in cold Perfadex solution for3h (cold static storage). Heart-lungwas hanged on the Ex Vivo Lung Perfusion (EVLP) we developed and start to perfusewith the harvested rabbit blood which was deoxygenated in the oxygenator with a gasmixture of N2and CO2.The donor lung ventilated, the deoxygenated blood was perfused tothe lung via main pulmonary, oxygenated in the lung, flow out from the left atrium, andretune to the oxygenator. A peristaltic pump was used to drive the blood circulate theEVLP device throughout the ex vivo perfusion.Results: Except for one animal had a pulmonary artery tear and leakage during thepulmonary catheter insert, and lead to the experiment failed, the all other animals finishedthe experiment. No significant change in water content, respiratory tract secretion, peakairway pressure were found, grossly infarction focuses were not found either. Thestructure of lungs under the light microscopic observation shows some Alveolar ruptureand neutrophil infiltration. Conclusion: This rabbit Ex Vivo Lung Perfusion device we developed is stable and has agood reproducibility. Part II：Protective Effect of N-acetylcysteine in Donation afterCardiac Death (DCD) LungObjective: To explore whether N-acetylcysteine could protect the lung from Donationafter Cardiac Death (DCD).Methods: Eighteen male New Zealand white rabbits (body weight2-2.5kg) wererandomly assigned to three groups (n=6for each group). Group I: Pure Perfadex was usedfor donor’s lung flush and for cold statistic storage. Group II: NAC enriched Perfadexwere used for donor lung flush, but pure Perfadex for cold statistic storage. Group III:NAC enriched Perfadex were used both for donor lung flush and cold statistic storage.After storage, DCD lung was continuously perfused on EVLP for90min. Graft function,including Partial pressure of oxygen (Pa02) and peak airway pressure (PawP) wereassessed, and by histologic examination, and MDA, MPO, IL-1β and GSH of lung tissuewere measured. DCD lung animal model and EVLP were finished in the same way asdescribed in part I.Results: lung water content, MDA, MPO, IL-1β levels in control group weresignificantly higher than that in experimental groups(P <0.05), while, PaO2and GSHlevel in the control group were lower than in experimental groups (P<0.05).Nodifferences were found between all three groups with respect to levels of protein (P>0.05).Pa02decline in all three groups60min after the beginning of perfusion, but it was stilldecline much more in control than in the two experiment groups(P<0.05). ThoughPawP remain stable from the beginning of perfusion until60min after the beginning ofperfusion in three groups, it start to increase in all three groups in the later perfusionperiod,with the control group increase significantly higher than in experimental groups by the end of perfusion (P <0.05). Observation under light microscope showed moresignificant damage change (such as rupture of alveoli, inflammatory cell infiltration,etc) in control group than in experiment groups. Specifically, we observed that there wereno significant differences in MDA, MPO, IL-1β and GSH (P>0.05) between twoexperimental groups.Conclusion: NAC could reduce graft inflammatory reaction and abate oxygen freeradicals injury, leads to better oxygenation, as well as better pulmonary compliance,attenuate early ischemia reperfusion injury, and protect DCD lung against IRI. It impliesthat NAC added to Perfadex might be an alternatively approach for DCD lungpreservation.