抑制解偶联蛋白-2基因表达减轻急性脂肪肝细胞损伤  

Inhibiting the expression of uncoupling protein-2 attenuates acute damage to fatty liver cells

作　　者：程锐[1] 王春友[1] 刘涛[1] 王宏博[1] 王帅[1] 万赤丹[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：《中华器官移植杂志》2008年第9期540-542,共3页Chinese Journal of Organ Transplantation

基　　金：国家自然科学基金（30571764）

摘　　要：目的探讨下调解偶联蛋白2（UCP-2）基因的表达对脂肪肝细胞损伤的影响，为存在脂肪变性的供肝提供治疗靶点。方法采用二步胶原酶灌注法分离C57BL／6 J—ob／ob转基因肥胖小鼠的原代脂肪肝细胞，用靶向小鼠UCP-2基因的RNA干扰慢病毒载体感染所获得的脂肪肝细胞，实现对UCP2基因的特异性敲减（实验组），另以空载体感染的脂肪肝细胞为对照。荧光显微镜镜检确定感染效率，实时聚合酶链反应鉴定敲减效果。以肿瘤坏死因子-α（TNF-α）作用于感染后的肝细胞，24h后以碘化丙啶染色。流式细胞仪检测细胞凋亡情况；Western印迹法检测凋亡蛋白酶-3（Caspase-3）的活化。结果UCP2基因表达被有效抑制，UCP-2基因的敲减率约为75％。实验组的肝细胞凋亡率为（4．97±0．25）％，明显低于对照组的（21．13±1．28）%（P〈0．05），其Caspase-3的活化程度也低于对照组。结论抑制UCP-2基因表达能减轻脂肪肝细胞的损伤。Objective To investigate the effects of down regulating uncoupling protein-2 （UCP-2） expression on acute damage to fatty liver cells and explore a new target for the donor liver with steatosis. Methods Primary fatty liver cells were isolated from C57BL/6J-ob/ob transgenic mice by two-step collagenase perfusion method. RNAi lentivirus vector targeting mouse UCP-2 gene was used to knock down the UCP-2 gene in the steatosis hepatoeytes （the experimental group）. Empty lentivirus vector was transfected into the steatosis hepatocytes cells as the control group. Under the fluorescence microscopy, the transfection efficiency was tested. Real time PCR was used to determine the effect of RNAi. After the transfected cells were treated with TNF-α for 24 h, apoptosis was analyzed by flow cytometry using PI staining. Activation of caspase3 was detected by Western blot. Results The expression of UCP-2 gene was inhibited effectively, and the knockdown rate of UCP-2 gene was 75 %. The apoptosis rate in the experimental group was （4. 97 ± 0. 25） %, significantly lower than in the control group [（21. 13 ±1.28 ）%, P 〈 0. 05]. Activation of caspase.3 in the experimental group was also weaker than in the control group. Conclusion Inhibiting the expression of UCP 2 can attenuate the injury of fatty liver cells.

关 键 词：解偶联蛋白-2 基因表达 脂肪肝 细胞死亡 

分 类 号：R575.5[医药卫生—消化系统]