不同胆道灌洗液对肝移植供体胆道冷保存损伤的影响

Effect of different bile duct flush solutions on biliary tract cold preservation injury of donated liver for liver transplantation

目的:冷保存/再灌注损伤是导致非外科性移植物胆管树损害最重要的始动因素。观察不同胆道灌洗液对大鼠肝移植供肝胆道保存的影响,以筛选出一种较为理想的胆道灌洗液,使胆道在冷保存期间获得器官水平上的最佳保护。方法:实验于2006-08/12在中国医科大学附属盛京医院动物实验中心完成。①实验设计及分组:实验因素包括胆道灌洗液与冷保存时间,不同的灌洗液分别为胆道灌洗液a(生理盐水、HCA液),胆道灌洗液b(HTK液、UW液);冷保存时间为4h,8h,12h。采用正交设计方法,选用[L9(34)]正交表对实验因素随机分组,共分为9组。②实验方法:选择雄性Wistar大鼠45只,分为9组,每组5只。制作大鼠取供肝模型,按照实验设计分别选择不同的胆道灌洗液进行胆道灌洗,在相应的冷保存时间取胆管组织进行检测。③实验评估:采用原位末端标记法进行胆管上皮细胞凋亡检测,计算凋亡指数;在电镜下观察胆管细胞的病理形态变化,并利用图像分析系统求出每张照片中所有胆管上皮细胞的线粒体平均体积和数密度。结果:45只大鼠全部进入结果分析。通过正交设计的方差分析与直观分析得出下列结果。①胆管上皮细胞凋亡指数的正交设计结果:胆道灌洗液a、胆道灌洗液b及冷保存时间是决定大鼠供肝胆道冷保存损伤的主要因素(P均<0.01);其中HCA液、HTK液及4h冷保存时间较为理想,且三者结合效果更好。②胆道上皮细胞线粒体平均体积及数密度的正交设计结果:胆道灌洗液a、胆道灌洗液b及冷保存时间是决定大鼠供肝胆道冷保存损伤的主要因素(P均<0.01);其中HCA液、HTK液及4h冷保存时间较为理想,且三者结合效果更好。结论:①正确选择胆道灌洗液可以大幅度减轻肝移植供肝胆道的冷保存损伤,保护胆管上皮细胞。②胆道灌洗液以选择HCA液与HTK液结合灌洗的方式较为理想,冷保存时间越短越理想。

AIM： Cold preservation/reperfusion injury is the most important initial factor of inducing the biliary ductal tree. This study is designed to study the effect of different bile duct flush solutions on biliary tract preservation of rat donated liver, find out one ideal bile duct flush solution, so as to provide the best preservation in the level of ＂organs＂ for biliary tract during the cold preservation. METHODS： The expenment was performed at the Expenmental Animal Center of Shengjing Hospital of China Medical University from August to December in 2006. （1）The factors of the study included bile duct flush solution and cold preservation time. The bile duct flush solutions were divided into solution a [including normal saline and hypertonic citrate adenine （HCA）], and solution b [including University of Wisconsin solution and histidine tryptophan ketoglutarate （HTK） solution]. The cold preservation times were defined as 4, 8, 12 hours. The factors were divided into 9 groups at random with the [L9（3^4）] orthogonal table.（2）Forty-flve healthy male Wistar rats were selected and randomly divider into 9 groups, there were 5 rats in each group. After the rat donor liver model was built, according to orthogonal design, different bile duct flush solutions were selected to flush bile ducts, then biliary tract tissues were taken to be examined at the corresponding cold preservation time. （3）Apoptotic indexes of endothelial calls of biliary tract were taken by TUNEL way. Pathomorphological changes of bile duct calls were observed under electron microscope. The average volumes and number density of mitochondna in all the endothelial cells of biliary tract were calculated by image analysis system. RESULTS： All the 45 rats were involved in the result analysis. The results were obtained by analysis of vananca and direct analysis of orthogonal design. （1）The apoptotic indexes of endothelial calls of biliary tract： The cold preservation injury of biliary tract of donated liver was related to bile duct flush solutions a, b and cold preservation time （P〈 0.01）, furthermore factors HCA, HTK and 4-hour cold preservation were all ideal. Moreover, it was the best for the combination of the above three factors. （2）The average volumes and number density of mitochondda endothelial cells in biliary tract presented the identical trend as the apoptotic indexes of endothelial calls. CONCLUSION： （1）The cold preservation injury of biliary tract in the donated liver can be relieved greatly by selecting the suitable bile duct solutions, and the endothelial cells of biliary tract can be protected. （2）It is the ideal to adopt the combination of HCA and HTK as the bile duct flush solutions. And it is as shorter as more ideal for the cold preservation time.