Mueller Thomas H J, Kienle Klaus, Beham Alexander, Geissler Edward K, Jauch Karl W, Rentsch Markus
Department of Surgery, University of Regensburg, 93053 Regensburg, Germany.
Transplantation. 2004 Nov 15;78(9):1267-73. doi: 10.1097/01.tp.0000141095.06273.10.
Apoptosis plays a crucial role after ischemia-reperfusion in organ transplantation. It is executed by caspases and influenced by the rheostat of pro- and anti-apoptotic proteins of the bcl-2 family. This study investigated the effect of specific inhibition of caspases 3 and 7 on graft function, survival, and hepatic bcl-2 levels after liver transplantation.
Lewis rats underwent syngeneic orthotopic liver transplantation after 16 hr of cold graft storage (in University of Wisconsin solution). Livers of donor animals treated with D(OMe)E(OMe)VD(OMe)-fluoromethylketone (specific inhibitor of apoptosis executor caspases 3 and 7), and appropriate control groups, were investigated. Early graft injury was quantified by measurement of bile flow and determination of microvascular graft injury by using in vivo fluorescence microscopy. Apoptosis and its regulation were examined by terminal deoxynucleotide transferase-mediated dUTP nick-end labeling staining and Western blot analysis of cell death effectors, respectively.
After specific in vivo caspase inhibition, Western blot analysis revealed inhibition of caspase-induced cleavage of poly-ADP-ribose-polymerase. Inhibition of caspases 3 and 7 resulted in a significantly decreased number of apoptotic endothelial cells and improved microvascular perfusion. A cell protective effect was also suggested by an increase of bcl-2 levels at 7 days. Most important, specific caspase blockade resulted in improved rat survival after liver transplantation.
Specific inhibition of apoptosis executor caspases effectively reduces graft ischemia-reperfusion injury and improves survival in liver transplantation. Better tissue preservation after caspase inhibition correlates with reduced apoptosis execution, improved microvascular perfusion, and bcl-2 up-regulation. Therefore, specific caspase inhibition represents a promising regimen for clinical use in liver transplantation.
凋亡在器官移植的缺血再灌注后起着关键作用。它由半胱天冬酶执行，并受bcl-2家族促凋亡和抗凋亡蛋白的调节。本研究调查了特异性抑制半胱天冬酶3和7对肝移植后移植物功能、存活及肝脏bcl-2水平的影响。
Lewis大鼠在冷保存16小时（在威斯康星大学溶液中）后进行同基因原位肝移植。对用D(OMe)E(OMe)VD(OMe)-氟甲基酮（凋亡执行半胱天冬酶3和7的特异性抑制剂）处理的供体动物肝脏以及适当的对照组进行研究。通过测量胆汁流量对早期移植物损伤进行定量，并使用体内荧光显微镜确定微血管移植物损伤。分别通过末端脱氧核苷酸转移酶介导的dUTP缺口末端标记染色和细胞死亡效应器的蛋白质印迹分析来检测凋亡及其调节。
在体内特异性抑制半胱天冬酶后，蛋白质印迹分析显示半胱天冬酶诱导的聚ADP-核糖聚合酶裂解受到抑制。抑制半胱天冬酶3和7导致凋亡内皮细胞数量显著减少，微血管灌注改善。7天时bcl-2水平升高也提示了细胞保护作用。最重要的是，特异性半胱天冬酶阻断导致肝移植后大鼠存活率提高。
特异性抑制凋亡执行半胱天冬酶可有效减轻移植物缺血再灌注损伤，提高肝移植的存活率。半胱天冬酶抑制后更好的组织保存与凋亡执行减少、微血管灌注改善和bcl-2上调相关。因此，特异性半胱天冬酶抑制是一种有前景的肝移植临床应用方案。
