解偶联蛋白2过度表达对LO2肝细胞缺血再灌注损伤的影响  

Effect of UCP-2 Over-expression on Ischemia-Reperfusion-induced Injury of Human Hepatocyte Strain LO2

作　　者：万赤丹[1] 王宏博[1] 冯贤松[1] 刘涛[1] 程锐[1] 勾善淼[1] 

机构地区：[1]华中科技大学同济医学院附属协和医院普外科,武汉430022

出　　处：《华中科技大学学报（医学版）》2008年第4期469-472,475,558,共6页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

基　　金：国家自然科学基金资助项目(No30571764)

摘　　要：目的探讨解偶联蛋白-2(uncoupling protein-2,UCP-2)的过度表达对人LO2肝细胞系缺血再灌注损伤的影响及可能机制。方法构建UCP-2真核表达质粒,转染LO2肝细胞并筛选出阳性克隆进行鉴定。将转染重组质粒组和对照组肝细胞进行缺血再灌注处理后,采用原位Hoechest/PI染色,荧光显微镜观察细胞损伤情况,AnnexinⅤ/PI双标流式细胞仪检测细胞坏死率、凋亡率和存活率,并检测细胞内三磷酸腺苷(ATP)、活性氧(ROS)、丙二醛(MDA)水平。结果转染重组质粒组细胞在缺血再灌注后遭受的损伤显著重于对照组,转染重组质粒组细胞内ATP水平显著低于对照组,细胞坏死率、凋亡率、存活率与对照组细胞比较差异均有显著性意义(P<0.05)。结论肝细胞内UCP-2的过度表达加剧了缺血再灌注后细胞内ATP的耗竭,导致细胞遭受更为严重的损伤。Objective To observe the effect of UCP-2 over-expression on ischemia-reperfusion-induced injury of human hepatocyte strain LO2 and discuss the possible mechanism.Methods A eukaryotic expression plasmid of UCP-2 was constructed and transfected into LO2 cells. The positive clones were screened out and identified. The cells in the transfection group and control group were subjected to ischemia-reperfusion treatment. By using in situ Hoechest/PI staining and fluorescent microscopy, the cell injury was observed. Annexin Ⅴ/PI double-labeled flow cytometry was used to assay the rate of cell necrosis, apoptosis and survival. The contents of ATP, ROS and MDA in cells were determined.Results After ischemia-reperfusion, the injury in transfection group was more severe than in control group. The intracellular contents of ATP in transfecttion group were significantly lower than in control group （P〈0.05）. There was significant difference in the rate of cell necrosis, apoptosis and survival between two groups （P〈0.05）.Conclusion UCP-2 over-expression in hepatocytes aggravates the exhaustion of intracellular ATP after ischemia-reperfusion, leading to more severe injury of cells.

关 键 词：解偶联蛋白-2 缺血再灌注损伤 肝细胞 

分 类 号：R657.3[医药卫生—外科学]