过表达解偶联蛋白-2基因加重原代肝细胞急性损伤  

Overexpression of Uncoupling Protein 2 Aggravates Acute Primary Hepatocyte Damage

作　　者：万赤丹[1] 程锐[1] 王春友[1] 王宏博[1] 王帅[1] 刘涛[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）》2009年第2期150-152,157,共4页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

基　　金：国家自然科学基金资助项目(No.30571764)

摘　　要：目的研究解偶联蛋白-2(uncoupling protein-2,UCP-2)基因在肝细胞急性损伤过程中的作用。方法构建小鼠UCP-2基因过表达慢病毒载体;二步胶原酶灌注法分离培养小鼠原代肝细胞;慢病毒感染肝细胞,使肝细胞中UCP-2基因表达上调;荧光显微镜镜检确定感染效率,Real ti me PCR检测UCP-2表达;肿瘤坏死因子-α(TNF-α)作用于病毒感染肝细胞24 h后,检测细胞上清液中谷丙转氨酶(ALT)和乳酸脱氢酶(LDH)水平;碘化丙啶(PI)染色流式细胞仪检测细胞凋亡,Western blot检测凋亡因子Caspase 3的活化。结果Real ti me PCR检测证实目的细胞中UCP-2表达上调;TNF-α作用后病毒感染的实验组上清液中ALT和LDH水平、细胞凋亡率、凋亡因子Caspase 3活化程度均高于对照组(均P<0.05)。结论UCP-2基因过度表达可加重肝细胞急性损伤。Objective To investigate the role of uncoupling protein-2（UCP-2） in acute liver injury. Methods Lentivirus overexpression vector to mouse ucp-2 gene was constructed,and mouse primary hepatocytes were isolated by two-step liver perfusion with collagenase type IV and cultured. After infection with the lentivirus,hepatocytes were observed by fluorescence microscope,and real time PCR and Western blot were used to detect the expression of ucp-2 gene. After stimulation with TNF-α for 24 h,ALT and LDH in the cell culture supernatant were determined, and apoptosis was analyzed by flow cytometry using propidium iodide（PI） staining. Activation of caspase 3 was assayed by Western blot. Results Real time PCR revealed that the expression of ucp-2 gene was up-regulated effectively. The levels of ALT and LDH, apoptosis rate, and the activation of Caspase 3 were significantly increased in experimental group as compared with control group（all P〈0.05）. Conclusion Overexpression of ucp-2 aggravates acute liver damage.

关 键 词：解偶联蛋白-2 慢病毒 肝细胞损伤 

分 类 号：R657.3[医药卫生—外科学]