METHODS: Experimental, randomized pilot trial of parallel groups and final blind analysis using a swine model of LT. Donor animals (n=8) were submitted to organ procurement. Lungs were subjected to 6h (n=4) or 12h (n=4) aerobic hypothermic preservation. The left lung was transplanted and re-perfused for 4h. Lung biopsies were obtained at (i) the beginning of CIT, (ii) the end of CIT, (iii) 30min after reperfusion, and (iv) 4h after reperfusion. Lung-grafts were histologically assessed by microscopic lung injury score and wet-to-dry ratio. Inflammatory response was measured by determination of inflammatory cytokines. Caspase-3 activity was determined as apoptosis marker.
RESULTS: We observed no differences on lung injury score or wet-to-dry ratio any given time between lungs subjected to 6h-CIT or 12h-CIT. IL-1β and IL6 showed an upward trend during reperfusion in both groups. TNF-α was peaked within 30min of reperfusion. IFN-γ was hardly detected. Caspase-3 immunoexpression was graded semiquantitatively by the percentage of stained cells. Twenty percent of apoptotic cells were observed 30min after reperfusion.
CONCLUSIONS: We observed that 6 and 12h of CIT were equivalent in terms of microscopic lung injury, inflammatory profile and apoptosis in a LT swine model. The extent of lung injury measured by microscopic lung injury score, proinflammatory cytokines and caspase-3 determination was mild.
方法:实验,平行组的随机试点试验和使用LT猪模型的最终盲分析。将供体动物(n=8)提交到器官采购。对肺进行6h(n=4)或12h(n=4)有氧低温保存。左肺移植并再灌注4h。在(i)CIT开始时获得肺活检,(Ii)CIT的结尾,(iii)再灌注后30min,和(iv)再灌注后4h。通过微观肺损伤评分和干湿比对肺移植物进行组织学评估。通过测定炎性细胞因子来测量炎症反应。测定Caspase-3活性作为凋亡标志物。
结果:我们观察到接受6h-CIT或12h-CIT的肺在任何给定时间之间的肺损伤评分或湿干比没有差异。IL-1β和IL6在再灌注期间均呈上升趋势。TNF-α在再灌注30min内达到峰值。几乎检测不到IFN-γ。Caspase-3免疫表达通过染色细胞的百分比进行半定量分级。再灌注后30min观察到20%的凋亡细胞。
结论:我们观察到6和12h的CIT在微观肺损伤方面是等效的,LT猪模型中的炎症谱和细胞凋亡。用显微肺损伤评分测量肺损伤程度,促炎细胞因子和caspase-3测定轻度。