Stierum R H, van Herwijnen M H, Pasman P C, Hageman G J, Kleinjans J C, van Agen B
Department of Health Risk Analysis and Toxicology, University of Limburg, Maastricht, The Netherlands.
Carcinogenesis. 1995 Nov;16(11):2765-71. doi: 10.1093/carcin/16.11.2765.
In response to DNA damage, in particular DNA strand breaks, the proposed roles for normal tumour suppressor protein p53 are to increase the period of time available for DNA repair prior to replication, or to direct damaged cells into programmed cell-death. Since treatment of mammalian cells with (+/-)-anti-benzo[a]pyrene diolepoxide [(+/-)-anti-BPDE] --a mixture of metabolites comprising the most reactive (+)-anti-enantiomer of the full environmental carcinogen benzo[a]pyrene--has been shown to result in induction of DNA repair processes and consequently in DNA strand break formation, the aim of the present study was to investigate whether p53 accumulation is induced in (+/-)-anti-BPDE-treated phytohaemagglutinin-stimulated human peripheral blood lymphocytes (PBLs). Both immunocytochemical and immunoblot analysis indicated that treatment of PBLs with (+/-)-anti-BPDE results in p53 accumulation. Optimal accumulation was observed at 2.5 microM, while no increase of p53 levels was observed at concentrations < 2.5 microM and > 10 microM. Further, (+/-)-anti-BPDE-induced p53 accumulation in PBLs was found to be time-dependent with accumulation up to 24 h after the onset of treatment. Treatment of PBLs with 2.5 microM of (+/-)-anti-BPDE and 1 mM of 3-aminobenzamide, an inhibitor of the DNA strand break-dependent enzyme poly(ADP-ribose) polymerase, resulted in increased p53 levels, in comparison to cells treated with (+/-)-anti-BPDE alone. This combination also potentiated the frequency of (+/-)-anti-BPDE-induced micronuclei. These findings suggest that (+/-)-anti-BPDE-induced DNA strand break formation is responsible for the observed p53 accumulation. It is unlikely that poly(ADP-ribose) polymer formation is a prerequisite in the process of p53 accumulation, as triggered by DNA strand-break inducing agents like (+/-)-anti-BPDE. It is hypothesized that p53-dependent pathways may be activated in phytohaemagglutinin-stimulated human peripheral blood lymphocytes exposed ex vivo to (+/-)-anti-BPDE.
为应对DNA损伤，尤其是DNA链断裂，正常肿瘤抑制蛋白p53的作用是延长复制前DNA修复的时间，或引导受损细胞进入程序性细胞死亡。由于用（±）-反式苯并[a]芘二氢二醇环氧化物[（±）-反式-BPDE]（一种包含完全环境致癌物苯并[a]芘最具反应性的（+）-反式对映体的代谢物混合物）处理哺乳动物细胞已被证明会导致DNA修复过程的诱导，并因此导致DNA链断裂的形成，本研究的目的是调查在（±）-反式-BPDE处理的植物血凝素刺激的人外周血淋巴细胞（PBL）中是否诱导了p53积累。免疫细胞化学和免疫印迹分析均表明，用（±）-反式-BPDE处理PBL会导致p53积累。在2.5 microM时观察到最佳积累，而在浓度<2.5 microM和>10 microM时未观察到p53水平的增加。此外，发现（±）-反式-BPDE诱导PBL中的p53积累是时间依赖性的，在处理开始后长达24小时积累。用2.5 microM的（±）-反式-BPDE和1 mM的3-氨基苯甲酰胺（一种DNA链断裂依赖性酶聚（ADP-核糖）聚合酶的抑制剂）处理PBL，与仅用（±）-反式-BPDE处理的细胞相比，导致p53水平升高。这种组合还增强了（±）-反式-BPDE诱导的微核频率。这些发现表明，（±）-反式-BPDE诱导的DNA链断裂形成是观察到的p53积累的原因。聚（ADP-核糖）聚合物的形成不太可能是由（±）-反式-BPDE等DNA链断裂诱导剂引发的p53积累过程中的先决条件。据推测，在体外暴露于（±）-反式-BPDE的植物血凝素刺激的人外周血淋巴细胞中，p53依赖性途径可能被激活。
