Experimental Study On The Change Of NF-κB And Cytokine During Isolated Reperfusion In Rat Donor Lung

Objective In this study, we established a isolated lung reperfusion model with extrocopreal circle apparatus and extracorporeal membrane oxygenation filled with nitrogen to study the preservation of donor lung , to observe the changes of airway pressure, the gas blood exchange analysis, the lung wet-to-dry weight, pulmonary histology, the gene expressions of intercellular adhesion molecule-1 (ICAM-1) and cytokine-induced neutrophil chemoattractant (CINC), the DNA binding activity of nuclear factor-kB (NF-kB) and the level of IkBa during reperfusion in rat donor lung in order to explore the change of them on reperfusion injury in rat lung, offer the experimental and theoretical evidence for the preservation of donor lung in clinical lung transplantation.Methods 36 male Sprague-Dawley rats were randomly divided into 3 goup:5min during reperfusion group,30 min group and 60 min group,each group was divided 2 subgroup again:donor group and recieptor group, each subgroup contains 6 rats. Donor lung undergoing ventilation with 4℃ LPD from left ventricle and six hours ischemia, the lung artery was perfused by the venous blood made by membrane oxygenation filled with nitrogen, and the isolated lung was ventilated by the ventilator. The changes of airway pressure and arterial blood gas analysis were studied at 5min, 15min, 30min, 60min of reperfusion. the lung wet-to-dry weight ratio (W/D) were obtained at 5min, 30min, 60min of reperfusion, the mRNA expressions of ICAM-1 and CINC were detected by reverse-transcription polymerase chain reaction (RT-PCR) , the level of IkBa was determined by the method of western blot and the DNA binding activity of NF-B in rat lung was measured by electrophoretic mobility shift assay (EMSA ) at thesame time .Results (1) The change of PaC^: PaC*2 decreased gradually and was obviously during reperfusion, the mean PaQ were 346+18.16, 327 + 13.40, 301 + 12.14 and 217 + 24.05mmHg at 5min, 15min,30min and 60min, respectively .There was significant difference in differnet time groups(P<0.05). (2) The airway peak pressure: It increased gradually and was obviously during reperfusion,the mean pressure were 12.17+1.47> 12. 67+1.75, 14.17+1.47and 16.33 +1.97cmH2O at Omin, 15min,30min and 60min, respectively .There was significant difference in differnet time groups(P<0.05).(3) W/D: During reperfusion the W/D increased progressively, the mean W/D were 2.08 + 0.52, 2. 79±0.94 and 3.83 + 0.12at Omin, 30min and 60min, respectively. there was significant difference in the lung W/D among groups(P<0.05). (4) Histological examination: the lung injury was aggravated gradually after reperfusion and there was marked pulmonary capillary congestion, interstitial edema and leukocyte infiltration, intra-alveolar hemorrhage in the light microscopy.(5)The gene expressions of ICAM-1 and CINC in lung tissue: Compared among groups, marked increase in gene expressions of ICAM-1 and CINC was observed during the reperfusion,there was significant difference (P<0.05). (6) The DNA binding activity of NF-kB in rat lung: The DNA binding activity of NF-kB increased rapidly as early as 3 Omin of reperfusion and the increment persisted to 60min of reperfusion (P<0.05), The level of IkBa decreased obviously during reperfusion(P<0.05).(7) Significant positive correlation was observed between the expression of ICAM-1 mRNA or CINC mRNA and NF-kB DNA binding activity during reperfusion, IkBa and NF-kB DNA binding activity was significant negative correlation,respectively (P<0.01).Conclusions (1) This model provides a simple, reliable model for determining the fuction of the donor lung. It can reflect the injury ofreperfusion.(2) Nuclear factor-B, activation of which can regulate the expression of ICAM-1 mRNA and CINC mRNA, plays a important role during reperfusion in rat donor lung.