Fischer S, Maclean A A, Liu M, Cardella J A, Slutsky A S, Suga M, Moreira J F, Keshavjee S
Thoracic Surgery Research Laboratory, Toronto General Hospital Research Institute, University of Toronto, Toronto, Ontario, Canada.
Am J Respir Crit Care Med. 2000 Nov;162(5):1932-9. doi: 10.1164/ajrccm.162.5.9910064.
Ischemia-reperfusion (IR) injury is a major cause of organ dysfunction following lung transplantation. We have recently described increased apoptosis in transplanted human lungs after graft reperfusion. However, a direct correlation between ischemic time, cell death, and posttransplant lung function has not yet been demonstrated. We hypothesized that an increased ischemic period would lead to an increase in cell death, and that the degree and type of cell death would correlate with lung function. To investigate this, we preserved rat lungs at 4 degrees C for 20 min and 6, 12, 18, and 24 h, and then transplanted the lungs and reperfused them for 2 h. Cell viability was determined with a triple staining technique combining trypan blue, terminal deoxynucleotidyl transferase-uridine nucleotide end-labeling, and propidium iodide nuclear staining. Percentages of apoptotic and necrotic cells were calculated from total cell numbers. Following 20 min and 6 and 12 h of cold preservation, less than 2% of graft cells were dead, whereas after 18 and 24 h of cold preservation, 11% and 27% of cells were dead (p < 0.05), the majority of which were necrotic. After transplantation and reperfusion, the mode of cell death changed significantly. In the 6- and 12-h groups, approximately 30% of cells were apoptotic and < 2% were necrotic, whereas in the 18- and 24-h groups, 21% and 29% of cells, respectively, were necrotic and less than 1% were apoptotic. Lung function (Pa(O(2))) decreased significantly (p < 0.05) with increasing preservation time. The percentage of necrotic cells was inversely correlated with posttransplant graft function (p < 0.0001). The study demonstrates a significant association among cold preservation time, extent and mode of cell death, and posttransplant lung function, and suggests new potential strategies to prevent and treat IR injury.
缺血再灌注（IR）损伤是肺移植后器官功能障碍的主要原因。我们最近报道了移植的人肺在移植肺再灌注后凋亡增加。然而，缺血时间、细胞死亡与移植后肺功能之间的直接相关性尚未得到证实。我们假设缺血时间延长会导致细胞死亡增加，并且细胞死亡的程度和类型与肺功能相关。为了研究这一点，我们将大鼠肺在4℃保存20分钟以及6、12、18和24小时，然后进行肺移植并再灌注2小时。采用台盼蓝、末端脱氧核苷酸转移酶介导的dUTP缺口末端标记和碘化丙啶核染色相结合的三重染色技术测定细胞活力。根据总细胞数计算凋亡细胞和坏死细胞的百分比。在冷保存20分钟以及6和12小时后，少于2%的移植细胞死亡，而在冷保存18和24小时后，11%和27%的细胞死亡（p<0.05），其中大多数为坏死细胞。移植和再灌注后，细胞死亡模式发生了显著变化。在6小时和12小时组中，约30%的细胞凋亡，<2%的细胞坏死，而在18小时和24小时组中，分别有21%和29%的细胞坏死且<1%的细胞凋亡。肺功能（Pa（O₂））随着保存时间的延长而显著降低（p<0.05）。坏死细胞百分比与移植后移植物功能呈负相关（p<0.0001）。该研究表明冷保存时间、细胞死亡程度和模式与移植后肺功能之间存在显著关联，并提出了预防和治疗IR损伤的新潜在策略。
