肝移植大鼠肝窦内皮细胞细胞凋亡与移植肝肝细胞损害的关系

Relationship between sinusoidal endothelial cell apoptosis and hepatocyte injury after their transplantation into rats

目的探讨冷保存肝移植大鼠肝窦内皮细胞(SEC)细胞凋亡与移植肝肝细胞损害的关系。方法雄性SD大鼠随机分为假手术组(n=6)、UW1h肝移植组(n=48)、UW12h肝移植组(n=48)。参照 Kamada的方法行原位肝移植(OLT)。观察大鼠168h存活率。分别于术后不同时相点采取血液及组织标本, 检测血清丙氨酸氨基转移酶(ALT)及透明质酸(HA)水平；TUNEL法检测SEC凋亡,透射电镜观察细胞凋亡的形态学改变。结果 UW12h组168h存活率为50％,显著低于UW1h组(F=6.39,P＜0.05)。 UW12h组肝移植后血清ALT、HA水平明显高于UW1h组(F=3.99,P＜0.05；F=12.43,P＜0.05),两组大鼠ALT水平均于术后6h达高峰。UW12h组SEC凋亡指数(AI)明显高于UW1h组和假手术组(F值分别为63.58和86.58,P值均＜0.01),两组大鼠SEC的AI也于术后6h达高峰,与血中丙氨酸氨基转移酶(ALT) 的高峰时相点一致。且两组大鼠SEC的AI均与ALT水平呈显著正相关(r值分别为1.0和0.962,P＜0.05)。结论 SEC凋亡程度与移植肝肝细胞损害呈显著正相关,SEC凋亡是冷保存再灌注损伤的关键环节。

Objective To investigate the relationship between the apoptosis of sinusoidal endothelial cells （SEC） and the hepatocyte injury of the livers after their transplantation into rats. Methods Male SD rats were divided into three groups randomly： sham group, UWIh group and UW12 h group. Orthotopic liver transplantations were performed using the technique described by Kamada with a modification. A survival rate curve was made using the Kaplan-Meier method. Liver tissue specimens and blood samples were collected at different time points after the surgeries. Six animals were sacrificed at each timepoint. The liver injury was evaluated by serum ALT and HA levels. The incidence of apoptosis in SECs was measured using the Tunel method. Additionally, the typical morphology indication of apoptosis was observed by transmission electron microscopy. Results The survival rate at 168 h in the UWI 2 h group was significantly lower than that in the UWlh group （F = 6.39, P 〈 0.05）. The levels of serum ALT and HA were significantly higher in UW12h group than those in UWlh group （F = 3.99, P 〈 0.05; F = 12.43, P 〈 0.05）. The serum ALT level reached its peak at 6 h after transplantation in both groups. The AI of SEC was significantly higher in the UW12 h group than those in the UWlh group and sham group （F = 63.58, P 〈 0.01; F = 86.58, P 〈 0.01）. The apoptosis index （AI） in the UWIh group and in the Uw12 h group both reached their peak at 6 h postopera- tively and the AI of SEC of each group was positively correlated with their serum ALT levels significantly （r = 1.0, P 〈 0.01 ; r = 0.962, P 〈 0.05）. Conclusion The increase of apoptosis of SEC is significantly correlated with the dysfunction of the livers after transplantation in rats, and the dysfunction was mainly caused by the process of cold preservation/reperfusion injury.