两种细胞建立肝细胞氧化损伤模型比较

Comparative study on utilization of two cell lines in liver cell oxidative damage model

目的比较过氧化氢诱导人肝癌细胞株（Hep G2）与正常肝细胞株（Chang liver）建立的肝细胞氧化应激损伤模型。方法用不同浓度过氧化氢诱导Hep G2细胞和Chang liver细胞氧化应激损伤,采用噻唑蓝法检测细胞生长抑制率;分光光度法测定培养液中乳酸脱氢酶（LDH）、谷丙转氨酶（ALT）、谷草转氨酶（AST）活性,以及肝细胞中超氧化物歧化酶（SOD）活性、还原型谷胱甘肽（GSH）及丙二醛（MDA）含量。结果作用时间在0.5~4 h时,在75~600μmol/L浓度范围内,过氧化氢可浓度和时间依赖性地抑制2种肝细胞增殖,促使细胞内AST、ALT和LDH向培养液中释放;细胞中SOD和GSH活性明显降低,MDA含量明显升高,表明2种肝细胞氧化损伤模型构建成功;选择300μmol/L H2O2作用4 h为致Hep G2和Chang liver细胞氧化应激损伤的最佳条件,结果显示,Hep G2和Chang liver细胞的生长抑制率分别为62%和76%,培养液中ALT、AST、LDH活性分别为（18.2±0.2）、（34.2±4.6）、（544.2±26.8）和（19.1±0.1）、（30.3±2.5）、（536.8±22.3）U/L,细胞中MDA含量分别为（7.8±0.9）和（8.6±1.1）nmol/mgprot。结论Hep G2与Chang liver细胞均可用于氧化应激细胞损伤模型的制备。

Objective To compare the utilization of Hep G2 and Chang liver cell lines in hydrogen peroxide（ H2O2）-induced hepatotoxicity cell models in vitro. Methods Methylthiazolyldiphenyl-tetrazolium bromide（ MTT）assay was used to evaluate the inhibitory effect of H2O2 on cell proliferation. Lactate dehydrogenase（ LDH）,alanine amino-transferase（ ALT）,and aspartate aminotransferase（ AST） in culture solution and malondialdelyde（ MDA）,superoxide dismutase（ SOD）,and reduced glutathione（ GSH） in the cells were detected with spectrophotometric method. Results H2O2 at the concentrations of 75- 600 μmol / L and with the treatment time of 0. 5 to 4 hours inhibited the proliferation of the two cell lines and resulted in the leakage of cellular LDH,ALT and AST into culture solution in a concentrationand time-dependent manner; furthermore,H2O2 increased MDA formation and reduced GSH level in the cells of the two cell lines. The results suggested that H2O2 could induce cellular oxidative injury in both Hep G2 and Chang liver cell line.With the optimal H2O2 concentration of 300 μmol / L and treatment time of 4 hours,the proliferation inhibitory ratio,the activities of ALT,AST and LDH in the culture solution,and the concentration of MDA in the cells were 62%,18. 2 ± 0. 2U / L,34. 2 ± 4. 6 U / L and 544. 2 ± 26. 8 U / L,and 8. 6 ± 1. 1 nmol / mg prot for the model utilizing Hep G2 cell line,and those indicators were 76%,19. 1 ± 0. 1 U / L,30. 3 ± 2. 5 U / L and 536. 8 ± 22. 3U / L,and 7. 8 ± 0. 9 nmol / mg prot for the model utilizing Chang liver cell line. Conclusion The two cell lines could be utilized in the construction of H2O2-induced cellular oxidative damage.