Research Of Heme Oxygenase-1 On Lung Transplantation

Part I The improvements in establishment of rat orthotopicleft lung transplantation modelObjective To establish a simple, valid rat orthotopic left lung transplantation model with the improved operation technique. Methods Male rats were randomly divided into donor and recipient groups, SD rats as donors and Wister rats as Recipients. Transplants were performed by using inferior vena flushing , harvesting in situ and the improved cuff anastomosis techniques. Results Successful rate was 88%, the chest CT scan show no atelectasis and blood gas analysis manifest good respiratory function. Conclusions The improved operation technique offer a simple , valid ,cheap and useful method , it can establish rat orthotopic left lung transplantation model successfully. Part II Heme oxygenase-1 and donor lungs ischemia/ reperfusioninjury. Objective Heme oxygenase-1(HO-1) has been found to be a cytoprotective protein. This study is to investigate the expression and significance of heme oxygenase-1 in donor lung ischemia/reperfusion injury. Methods On the base of the rat orthotopic lung transplantation model regulating endogenous heme oxygenase-1 in lung I/R injury. Cobalt protoporphyrin is the HO-1 inducer, and Zinc protoporphyrin is the H0-1 inhibitor. They both can regulate endogenous heme oxygenase-1 expression.Three groups of rats were examined. At 24 hours before the harvest, the donors in group 1, which is the control group, received 0.9% saline, the donors in group 2 were pretreated with CoPP, the donors in group 3 received ZnPP.Obtained the grafted lungs after lungs at 1, 2, 4, 8, 12h after reperfusion. The grafted lungs were resected into two parts, one part was put in liquid nitrogen for freezing preservation, the other part was fixated with 10% formaldehyde. We detected expression of HO-1 in donor lung tissue with immunohistochemistry technique, and we measured HO-1mRNA activity in those tissue by using reverse transcription-polymerase chain raction (RT-PCR) after donors received CoPP or ZnPP. At the same time, in suit TUNEL was used to survey the apoptotic in lung. Results Apoptotic index(AI) increased after at 1 hour after reperfusion, reached the peak at 4 hours after reperfusion, and then decreased gradually after 4 hours. Lung ischemia/reperfusion can induce HO-1 expression, which increased stepwise along with reperfusion, and reached to the peak at 8 hours after reperfusion. Administrating CoPP and ZnPP at 24 hours before the operation can effectively modulate the expression of HO-1 protein in lung tissue. As the HO-1 inducer, CoPP can induce HO-1 its up-regulation, HO-1 protein can degrade apoptosis incidence rate. Conclusions Apoptosis participate in postgraft lung I/R injury, it changes dynamically with reperfusion. Apoptosis generates during the initial stage after lung transplantation. Pretreating the donor lungs with CoPP before operation can lead HO-1 expression up-regulation. HO—1 hyperexpression can inhibit pneumonocyte apoptosis, so that it can relieve donor lung I/R injuryPart III Heme oxygenase-1 and acute lung allograft rejectionObjective To investigate the effectiveness and mechanism of HO-1 protein in acute rejection of lung transplantation. Methods We regulated endogenousin rat heme oxygenase-1 model by using CoPP as the HO-1 inducer and using ZnPP as the HO-1 inhibitor. Three groups were studied, including control group, CoPP group and ZnPP group. The donors were pretreated with 0.9% saline, CoPP, ZnPP 24 hours before harvest respectively. Inspecting the respiration of the grafted lung ever day, obtaining the grafted lungs at day 3,6,9, 12 after transplantation. The grade of acute rejection was estimated lungs by HE staining of histological sections. The expression of HO-1 protein in lung tissue were measured by immunohistochemistry technique. Western blot was used to assess the relative expression of HO-1 protein quantitatively. Results The expression of the HO-1 protein stronger in allograft lungs than in isograft lungs, and it increased in grafts with the acute rejecton in rat rejecton in rat. The HO-1 expression primarily in alveolar brochus periphery, alveolar epithelial cells and infiltrating inflammatory cells. Compared with control group and ZnPP group, CoPP treated group didn' t show graft survival significantly. (The median survival time of control group was 12.6 days, which was 14.7 days in CoPP group and 10.2 days in ZnPP group, P>0. 05) Conclusions HO-1 protein participated in pathological process of post-graft acute rejection, it increased gradually with acute rejection aggravationly. HO-1 protein may be considered monitor index of acute rejection after lung transplantation. Preoperative treatment with CoPP can induce up-regulation of HO-1 expression.The over expression of HO-1 didn't inhibit acute pneumal graft rejection significantly, and it couldn't prolong allograft survival time obviously.