Analysis of the human DNA damageby antioxidants for evaluation of safety in cancer chemoprevention

作为一种评估癌症化学预防安全性的方法，我们利用人类培养细胞和32p标记的人类肿瘤抑制基因p53和c- ha -ras- 1的分离DNA片段，研究了抗氧化剂是否具有人类DNA损伤的能力。用于检测celulardn。用抗氧化剂处理人体培养的损伤细胞，用脉冲场get电泳法检测。结果是。通过维生素A、视网膜、α -生育酚和槲皮素处理检测细胞dna损伤。此外。我们用HPLC-ECD定量测量了8-羟基脱氧鸟苷（8-OH-dG）的形成，这是DNA氧化损伤的一个指标。在维生素A、视网膜和n -乙酰半胱氨酸处理的细胞中，细胞内ar8- oh - dg的形成显著增加。用于检测对分离DN的损害。A，抗氧化剂与32p标记的DNA片段在金属离子存在下孵育，获得自射线图。α -生育酚、维生素A、视网膜、槲皮素和n -乙酰半胱氨酸在Cm存在下引起位点特异性DN，A损伤（II）。过氧化氢酶和根茎碱。一种Cu(I)特异性螯合剂。抑制Co（II）介导的DNA损伤，提示H_2O_2和Curl的参与。在Cu（II）存在下，胸腺嘧啶和胞嘧啶残基上的DNA切割频率较高。所谓的“抗氧化剂”在某些情况下起到抗氧化剂的作用，但在其他情况下也起到促氧化剂的作用。换句话说，抗氧化剂在某些情况下具有破坏DNA的能力，尽管抗氧化剂在其他情况下保护免受氧化应激。这些DNA氧化损伤可能导致癌变的起始和/或肿瘤的促进。在建议使用抗氧化剂进行癌症化学预防之前，应评估其安全性和有效性。

As a method to evaluatesafety in cancer chemoprevention, we have investigated whether antioxidants have the ability of the human DNA damage, using human cultured cells and 32P-labeledisolated DNA fragnrents of the human p53 tumor suppressor gene and c-Ha-ras-l prctooncogene. For detection of ce1lularDN.A damage, human cultured celles were treated with antioxidants, and examined by the pulsed-field get electrophoresis method. As the result. celularDNA damage was detected by the treatment of vitamin A, retinal, alpha-tocopherol and quercetin. Furthermore. we measured 8-hydroxy deoxyguanosine (8-OH-dG) formation, which was an index to an oxidative DNA lesion, quaruitativelyby HPLC-ECD.lntracellu1ar8-OH-dG formation significantly increased in cells treated with vitamin A, retinal and N-acetylcysteine. For detection of damage to isolated DN.A, antioxidants were incubated with 32P-labled DNA fragment in the presence of metalions, and the autoradiogram was obtained. alpha-Tocopherol, vitamin A, retinal, quercetin and N-acetylcysceine caused site-specific DN,A-damage in the presence of Cm(II). Catalase and bathocuproine. a Cu(I)-specific chelaror. inhibited the Co(II)-mediated DNA damage, suggesting the involvement of H_2O_2 and Curl).The DNA cleavage was observed frequency at thymine and cytosine residues in the presence of Cu(II).So-called "antioxidants" act as antioxidants in some circumstances, but also act as prooxidants in other circumstances. In another word, antioxidants become to have ability of damaging DNA in some cases, although antioxidants protect from oxidative stress in other cases. These oxidative DNA damage could be responsible for initiation and/or tumor promotion the multi-stage of carcinogenesis. In is requested chat safety and efficacy should be estimated before recommending use of antioxidants for cancer chemopreventton.