BACKGROUND: Low-dose active oxygen can promote the proliferation and differentiation of normal cells and tumor cells, but the relationship between the signal pathway and cell sensitive phase involved in this process is unclear at present.OBJECTIVE: To observe the relationship between HepG2 cell proliferations induced by hydrogen peroxide(HP), and NF-κB signal pathway and sensitive phase through the blockage of intracellular NF-κB signal pathway and the induction of difference cell phase to provide theoretical gist for inhibiting and preventing the development of tumor cell and improving prognosis of the patients.DESIGN: A complete randomized controlled study.SETTING: Staff Room of Toxicology, Department of Preventive Medicine,Fourth Military Medical University of Chinese PLA.PARTICIPANTS: The study was conducted in the Staff Room of Toxicology, Department of Preventive Medicine, Fourth Military Medical University of Chinese PLA from April to August 2004. The material was human liver cancer cell strain HepG2 cell, which was a present from the Department of Pathology of the Faculty of Basic Medicine, Fourth Military Medical University of Chinese PLA.METHODS: HP was directly acted on the cultured HepG2 cell in different dosages for the screening of concentration of HP, which could adequately promote the proliferation of HepG2 cell. 50 μmol/L of HP was used to manage HepG2 cell pre-managed by NF-κB inhibitor(pyrrolidine dithiocarbamate, PDTC) or the HepG2 cells in G1, G2/M or S phase obtained through the synchronization by thymidine separately.MAIN OUTCOME MEASURES: Absorbency of the cell in each group was detected by MTT colorimetric method after 1, 24 and 48 hours of the management by HP for the comparison of cellular proliferation activity.RESULTS: After 24 hours of the reaction by 50 μmol/L of HP, the absorbency of HepG2 cell in MTT assay was significantly higher than that of the control group( t =0. 001, P ＜ 0.01) . There was no significant difference of absorbency between HepG2 cell acted with 50 μmol/L of HP after 2 hours of pre-management by 20 μ mol/L of PDTC( t =0. 211, P ＞ 0. 05) . The absorbency of HepG2 cell in G1 phase was significant higher than that of the control group after reacting with 50 μmol/L of HP for 24 hours( t = 0. 031,P ＜0.05).CONCLUSION: Low-dose HP can induce the proliferation of HepG2 cell and this process might include the dependence on the participation of NF-κB signal pathway. The induction of HP on the proliferation of HepG2 cell has different sensitive phase in different cell cycle, of which it mainly induces cell proliferation of G1 phase. This conclusion will provide laboratorial data for tumor intervention and the improvement of prognosis.%背景:低剂量活性氧可以促进正常细胞和肿瘤细胞增殖、分化,但目前对此过程中所涉及的信号途径及其与细胞敏感时相的关系尚认识不足.目的:通过阻断细胞内NF-κB途径和诱导不同细胞时相,观察过氧化氢促进HepG2细胞增殖与NF-κB途径和细胞敏感时相的关系,为抑制、干预肿瘤细胞的生长及改善患者预后功能提供理论依据.设计:完全随机实验研究.单位:解放军第四军医大学预防医学系毒理学教研室.对象:实验于2004-04/2004-08在解放军第四军医大学预防医学系毒理学教研室完成,对象为人肝癌细胞株HepG2细胞,由解放军第四军医大学基础部病理学教研室惠赠.方法:以不同剂量过氧化氢直接作用于培养的HepG2细胞,筛选能够促进HepG2细胞增殖的过氧化氢浓度.用50μmol/L的过氧化氢分别处理经NF-κB抑制剂(PDTC)预处理的HepG2细胞和用胸腺嘧啶核苷同步化得到的G1,G2/M,S期HepG2细胞.主要观察指标:细胞分别经过氧化氢处理1,24,48 h后,通过四甲基偶氮唑蓝(MTT)比色法检测各组细胞吸光度值,比较细胞增殖活性.结果:50 μmol/L的过氧化氢作用HepG2细胞24 h后,MTT实验所测吸光度值与对照组相比显著升高(t=0.001,P＜0.01).20 μmol/L PDTC预处理HepG2细胞2 h后,给予50μmol/L过氧化氢作用细胞,MTT实验所测吸光度值与对照组相比差异无显著性意义(t=0.211,P＞0.05).50 μmol/L的过氧化氢作用G期HepG2细胞24 h后,MTT实验所测吸光度值与对照组相比显著升高(t=0.031,P＜0.05).结论:低剂量过氧化氢可诱导HepG2细胞增殖,该过程可能包含依赖NF-κB的信号途径的参与;过氧化氢诱导HepG2细胞增殖在细胞周期的不同时相效应敏感性不同,主要诱导G1期细胞增殖.此结论将为肿瘤干预和改善其预后提供实验学数据.
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