MUTAGENICITY OF BENZENE METABOLITES

描述：(改编自研究人员的摘要)苯是 少数几个明确被确定为 引发人类癌症。这是一个重要的公共卫生问题，因为 事实上，几乎每个人都暴露在不同水平的苯中。在……里面 与它在生物学上的重要性相比，人们对它的了解相对较少 苯诱导其遗传毒性效应的分子机制。至 为了解决这个问题，他们提出了开发一种人类细胞系，这种细胞系可以 用于研究苯的代谢物的致突变性和 相关的环境代理人。他们提出：(1)分离HL-60细胞 含有稳定整合的lambda/laci穿梭载体拷贝的线。 该细胞系将通过转染HL-60细胞获得 Lambda/LacI穿梭载体和编码新霉素抗性的质粒。 为本研究选定的克隆将具有以下属性：(A)a 对Lambda/Laci穿梭载体救援效率高，(B)包含 显著提高过氧化物酶活性并激活对苯二酚形成 DNA加合物，和(C)显示低自发和显著增加 LacI在处理细胞中的突变频率。(2)测量频率 不同剂量处理HL-60/LACI细胞诱导的LACI突变 对苯二酚或对苯二酚的浓度。他们将决定 这些代谢产物对HL-60/LACI细胞的作用显著 与溶剂处理相比，提高了LacI突变频率 独自一人。DNA加合物在处理细胞中的形成将通过以下方式确定 32P-后标记；允许我们估计形成的DNA加合物的水平 在所用条件下进行LacI突变分析。(3)按顺序排列 来自对照组和治疗组的单个LacI突变体。这 分析将使他们能够确定是否存在突变谱 治疗组与对照组的突变谱不同 一群人。他们将确定观察到的突变谱是否 对苯二酚和氢醌治疗组符合 来源于DNA加合物的N_2-(4-羟基苯基)脱氧鸟苷 由这两种代谢物形成。这些研究将提供 对这些苯代谢物诱导突变的本质的洞察。 虽然这项建议使用既定的程序，但它对 原因如下：(1)它将使用细胞过氧化物酶 代谢产物的激活，(2)它将允许比较突变 加合物诱导；(3)所建立的细胞系可应用 对大范围环境的诱变潜力的分析， 职业和医药制剂。

DESCRIPTION: (Adapted from the investigator's abstract) Benzene is one of the few environmental agents which is unequivocally established as initiating huma cancer. This is a important public health issue due to the fact that virtually everyone is exposed to varying levels of benzene. In contrast to its biologica importance, relatively little is known about the molecular mechanisms by which benzene induces its genotoxic effects. To address this, they proposed the development of a human cell line which can be used to investigate the mutageni properties of benzene's metabolites and related environmental agents. They proposed: (1) To isolate HL-60 cell line containing stably integrated copies o a lambda/lacI shuttle vector. This cell line will be developed by transfection of HL-60 cells with a lambda/lacI shuttle vector and a plasmid encoding resistance to neomycin. The clone selected for this study will have the following properties: (a) a high rescue efficiency for lambda/lacI shuttle vector, (b) contain significant levels of peroxidase enzyme and activate hydroquinone to form DNA adducts, and (c) shows a low spontaneous and a significant increase in lacI mutation frequency in treated cells. (2) To measure the frequency of lacI mutations induced by treatment of HL-60/lacI cells with varying concentrations of either p-benzouione or hydroquinone. They will determine if treatment of HL-60/lacI cells with these metabolites significantly increases the lacI mutation frequency compared to treatment with solvent alone. The formation of DNA adducts in treated cells will be determine by 32P-postlabeling; allowing us to estimate the levels of DNA adducts formed under the conditions used for analysis of lacI mutations. (3) To sequence individual lacI mutants from the control and the treatment groups. This analysis will allow them to determine if the mutation spectrum in the treatmen groups are different than the mutation spectrum in the control group. They wil determine if the spectrum of mutations observed in the para-benzoquione and hydroquione treatment groups are consistent with being derived from N2-(4-hydroxy phenyl) deoxyguanosine the DNA adduct identified as being formed by these two metabolites. These studies will provide insights as to the nature of mutations induced by these benzene metabolites. Although this proposal uses established procedures, it is innovative for the following reasons: (1) it wil use cellular peroxidase enzymes for activation of the metabolites, (2) it will allow for comparison of mutation induction with adduct formation, (3) the cell line developed can be applied to the analysis of the mutagenic potential of a wide range of environmental, occupational and medicinal agents.