CAJ | 학술논문

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。
목적：평개세포천투태PEP-1개도혈홍소가양매-1（HO-1）예처리대L02간세포결양복양손상적영향。방법이용기인공정수단표체화순화융합단백PEP-1-HO-1（PEP-1-heme oxy-genase-1，PEP-1-HO-1）。사용인간세포주（HL-7702）건립배양인간세포결양복양모형。안실험수요이10％신생우혈청적DMEM액정치배양L02간세포，장세포진행수궤분위7조：A조，정상대조조（상규배양）；B조，결양복양조（결양복양조세포결양18 h，복양6 h）；C조，PEP-1-HO-1예처리조，안PEP-1-HO-1농도분위5아조（C1，0．125μmol/L；C2，0．25μmol/L ；C3，0．5μmol/L；C4，1．0μmol/L；C5，2．0μmol/L，결양전이PEP-1-HO-1융합단백예처리2 h，결양18 h，복양6 h）。복양결속후，수집각조세포급배양액상청，검측MDA、LDH、AST、ALT함량급SOD활성。결과결양복양조교정상대조조상비，MDA、LDH、AST급ALT수평명현승고（P＜0．05），이 SOD 수평하강（P＜0．05）；PEP-1-HO-1예처리조여결양복양조상비，예처리조능명현강저MDA、LDH、AST급ALT수평（P＜0．05），사득SOD수평상승（P＜0．05），차재일정농도하여제량정정상관。결론 PEP-1-HO-1융합단백예처리가감경세포결양복양손상。
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'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.