PE P-1-血红素加氧酶-1融合蛋白预处理对L02肝细胞缺氧复氧损伤的影响  

Effects of PEP-1-heme oxygenase-1 fusion protein pretreatment on hypoxia-reoxygenation injury in L02 hepatocytes

作　　者：曾天才[1] 蔡庆和[1] 徐阳[2] 张林菲[1] 周晋航[1] 涂华华[1] 陈先祥[1] 

机构地区：[1] 湖北医药学院附属人民医院肝胆外科,十堰442000 [2] 湖北医药学院医务室

出　　处：《临床外科杂志》2014年第9期663-665,共3页Journal of Clinical Surgery

基　　金：湖北省教育厅科学技术研究计划重点项目（编号：D20082405）

摘　　要：目的：评价细胞穿透肽PEP-1介导血红素加氧酶-1（HO-1）预处理对L02肝细胞缺氧复氧损伤的影响。方法利用基因工程手段表达和纯化融合蛋白PEP-1-HO-1（PEP-1-heme oxy-genase-1，PEP-1-HO-1）。使用人肝细胞株（HL-7702）建立培养人肝细胞缺氧复氧模型。按实验需要以10％新生牛血清的DMEM液静置培养L02肝细胞，将细胞进行随机分为7组：A组，正常对照组（常规培养）；B组，缺氧复氧组（缺氧复氧组细胞缺氧18 h，复氧6 h）；C组，PEP-1-HO-1预处理组，按PEP-1-HO-1浓度分为5亚组（C1，0．125μmol/L；C2，0．25μmol/L ；C3，0．5μmol/L；C4，1．0μmol/L；C5，2．0μmol/L，缺氧前以PEP-1-HO-1融合蛋白预处理2 h，缺氧18 h，复氧6 h）。复氧结束后，收集各组细胞及培养液上清，检测MDA、LDH、AST、ALT含量及SOD活性。结果缺氧复氧组较正常对照组相比，MDA、LDH、AST及ALT水平明显升高（P＜0．05），而 SOD 水平下降（P＜0．05）；PEP-1-HO-1预处理组与缺氧复氧组相比，预处理组能明显降低MDA、LDH、AST及ALT水平（P＜0．05），使得SOD水平上升（P＜0．05），且在一定浓度下与剂量呈正相关。结论 PEP-1-HO-1融合蛋白预处理可减轻细胞缺氧复氧损伤。Objective To evaluate the pretreatment effects of heme oxygenase-1 mediated by cell-penetrating peptide PEP-1 on cultured human hepatocytes（L02）against hypoxia-reoxygenation injury. Methods Genetic engineering techniques were used to express and purify PEP-1-heme oxygenase-1 （PEP-1-HO-1）fusion protein.Human hepatic cell line（HL-7702）was used to establish the model of he-patic hypoxia-reoxygenation injury.According to the experimental requirement,L02 cells were fed with Dulbecco＆#39;s modified Eagle Medium supplemented with the 10%fetal bovine serum.The cultured L02 cells were divided into 7 groups randomly.Group A（CTL）received no anoxia and served as control and Group B （H/R）received 18 hours of anoxia,followed by reoxygenation for 6 hours.According to the concentration of PEP-1-HO-1 pretreatment,Group C were divided into 5 subgroups,including Group C1 （0.125 μmol/L）,Group C2（0.25 μmol/L）,Group C3（0.5 μmol/L）,Group C4（1.0 μmol/L）and Group C5（2.0μmol/L）.PEP-1-HO-1 fusion protein was pretreated for 2 hours before 18 hours of hypoxia and 6 hours of reoxygenation.The contents of ALT,AST,LDH,MDA and SOD viability in the culture medium were col-lected and measured at the end of the experiment.Results Compared with the control group,the level of MDA,LDH,AST and ALT increased and the SOD activity of L02 cells decreased in the H/R group（P〈0.05）.Compared with the H/R group,the level of MDA,LDH,AST and ALT decreased in the pretreat-ment groups and the SOD activity of L02 cells increased and positively correlated with pretreatment dosage under a certain concentration（P〈0.05）.Conclusion PEP-1-HO-1 fusion protein pretreatment can pro-tect L02 hepatocytes against hypoxia-rexygenation injury.

关 键 词：PEP-1肽 血红素加氧酶-1 肝细胞 缺氧复氧 保护 HEME Oxygenase-1 

分 类 号：R657.3[医药卫生—外科学]