Analysis of damage to human genes for estimating the carcinogenicity of environmental chemicals

分析人类基因损伤以评估环境化学物质的致癌性

• 批准号: 06557028
• 负责人: KAWANISHI Shosuke
• 金额: $ 7.3万
• 依托单位: KYOTO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1995
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-06557028/
• 关键词: human gene; Ame's test; carcinogen; active oxygen species; DNA damage; NADH; Mn; transition metal; Ames試験; 環境因子; 金属イオン; トリプトファン代謝物; ペンタクロロフェノール; 過酸化水素

Pulsed-field gel electrophoresis (PFGE) has emerged as a powerful tool for the study of high-molecular weight DNA.PFGE is generally used for detection of cellular DNA double-strand breaks. In addition, we have designed an experimental protocol which allows the detection of DNA single-strand breaks plus alkali-labile sites by PFGE.In this study, we have investigated damage to cellular and isolated DNA by Ames-test negative carcinogens in the presence of metal ions using both PFGE and DNA sequencing techNique. Active species causing DNA damage were investigated by the ESR-spin trapping method. We have shown that in the presence of Cu (II), Ames-test negative carcinogens (benzene metabolites, o-phenylphenol metabolites, caffeic acid, pentachlorophenol metabolites, tryptophan metabolites) caused damage to isolated DNA through H2O2 formation. PFGE showed that Ames-test negative carcinogens induced DNA strand breaks in cultured human cells in the presence of Mn (II). With alkali treatment, DNA single-strand breaks were observed. The strand breakage was increased by 3-aminotriazol (a catalase inhibitor) and decreased by catalase, indicating the involvement of H2O2. The DNA damage was decreased by o-phenanthroline, indicating the involvement of transition metal ion. These results suggest that in the presence of Mn (II) or Cu (II), Ames-test negative carcinogens produce H2O2, which is activated by transition metals to cause damage to DNA in vitro and probably in cultured cells.

脉冲场凝胶电泳（PFGE）已成为研究高分子量DNA的有力工具。PFGE一般用于检测细胞DNA双链断裂。此外，我们还设计了一个实验方案，允许通过PFGE检测DNA单链断裂和碱不稳定位点。在这项研究中，我们使用PFGE和DNA测序技术研究了金属离子存在下ames测试阴性致癌物对细胞和分离DNA的损伤。用esr自旋捕获法研究了引起DNA损伤的活性物质。我们已经证明，在Cu （II）存在下，ames测试阴性致癌物（苯代谢物、邻苯酚代谢物、咖啡酸、五氯酚代谢物、色氨酸代谢物）通过H2O2形成对分离的DNA造成损伤。PFGE显示，在Mn （II）存在下，ames测试阴性致癌物诱导培养的人细胞DNA链断裂。碱液处理后，DNA单链断裂。3-氨基三唑（一种过氧化氢酶抑制剂）增加了链断裂，过氧化氢酶减少了链断裂，表明H2O2参与了链断裂。邻菲罗啉降低了DNA的损伤，表明过渡金属离子的参与。这些结果表明，在Mn （II）或Cu （II）存在下，ames测试阴性的致癌物产生H2O2， H2O2被过渡金属激活，在体外和可能在培养细胞中对DNA造成损伤。

项目成果

川西 正祐: "金属化合物の発がん性と発がん機構" 環境保全. 9. 39-47 (1994)

川西正介：“金属化合物的致癌性和致癌机制”环境保护。 9. 39-47 (1994)

Y. Hiraku and S. Kawanishi: "Mechanism of Oxidative DNA Damage Induced by δ-Aminolevulinic Acid in the Presence of Copper lons." Cancer Res.(1996)

Y. Hiraku 和 S. Kawanishi：“铜离子存在下 δ-氨基乙酰丙酸诱导氧化 DNA 损伤的机制。”(1996)

S.Oikawa, and S.Kawanishi: "Site-specific DNA Damage Induced by NADH in the Presence of Copper(II) : Role of Active Oxygen Species." Biochemistry. (in press). (1996)

S.Oikawa 和 S.Kawanishi：“铜 (II) 存在下 NADH 诱导的位点特异性 DNA 损伤：活性氧的作用。”

Y.Hiraku and S.Kawanishi: "Mechanism of Oxidative DNA Damage Induced by Aminolevulinic Acid in the Presence of Copper Ions." Cancer Res.(in press). (1996)

Y.Hiraku 和 S.Kawanishi：“铜离子存在下氨基乙酰丙酸诱导氧化 DNA 损伤的机制”。

I.Saito, M.Takayama, and S.Kawanishi: "Photoactivatable DNA-Cleaving Amino Acids : Highly Sequence-Selective DNA Photocleavage by Novel-Lysine Derivatives." J.Amer.Chem.Soc.117. 5590-5591 (1995)

I.Saito、M.Takayama 和 S.Kawanishi：“光活化 DNA 裂解氨基酸：新型赖氨酸衍生物的高度序列选择性 DNA 光裂解”。