Lee E W, Johnson J T, Garner C D
Biomedical Science Department, General Motors Research Laboratories, Warren, Michigan 48090-9055.
J Toxicol Environ Health. 1989;26(3):277-91. doi: 10.1080/15287398909531254.
Effects of endogenously produced and exogenously added benzene metabolites on the nuclear DNA synthetic activity were investigated using a culture system of mouse bone marrow cells. Effects of the metabolites were evaluated by a 30-min incorporation of [3H]thymidine into DNA following a 30-min interaction with the cells in McCoy's 5a medium with 10% fetal calf serum. Phenol and muconic acid did not inhibit nuclear DNA synthesis. However, catechol, 1,2,4-benzenetriol, hydroquinone, and p-benzoquinone were able to inhibit 52, 64, 79, and 98% of the nuclear DNA synthetic activity, respectively, at 24 microM. In a cell-free DNA synthetic system, catechol and hydroquinone did not inhibit the incorporation of [3H]thymidine triphosphate into DNA up to 24 microM but 1,2,4-benzenetriol and p-benzoquinone did. The effect of the latter two benzene metabolites was completely blocked in the presence of 1,4-dithiothreitol (1 mM) in the cell-free assay system. Furthermore, when DNA polymerase alpha, which requires a sulfhydryl (SH) group as an active site, was replaced by DNA polymerase I, which does not require an SH group for its catalytic activity, p-benzoquinone and 1,2,4-benzenetriol were unable to inhibit DNA synthesis. Thus, the data imply that p-benzoquinone and 1,2,4-benzenetriol inhibited DNA polymerase alpha, consequently resulting in inhibition of DNA synthesis in both cellular and cell-free DNA synthetic systems. The present study identifies catechol, hydroquinone, p-benzoquinone, and 1,2,4-benzenetriol as toxic benzene metabolites in bone marrow cells and also suggests that their inhibitory action on DNA synthesis is mediated by mechanism(s) other than that involving DNA damage as a primary cause.
利用小鼠骨髓细胞培养系统，研究了内源性产生和外源性添加的苯代谢产物对核DNA合成活性的影响。在含有10%胎牛血清的 McCoy's 5a培养基中，代谢产物与细胞相互作用30分钟后，通过30分钟内[3H]胸苷掺入DNA来评估代谢产物的作用。苯酚和粘康酸不抑制核DNA合成。然而，儿茶酚、1,2,4-苯三酚、对苯二酚和对苯醌在24 microM时分别能够抑制52%、64%、79%和98%的核DNA合成活性。在无细胞DNA合成系统中，儿茶酚和对苯二酚在浓度高达24 microM时不抑制[3H]三磷酸胸苷掺入DNA，但1,2,4-苯三酚和对苯醌会抑制。在无细胞检测系统中，当存在1 mM的1,4-二硫苏糖醇时，后两种苯代谢产物的作用被完全阻断。此外，当需要巯基（SH）作为活性位点的DNA聚合酶α被催化活性不需要SH基团的DNA聚合酶I取代时，对苯醌和1,2,4-苯三酚无法抑制DNA合成。因此，数据表明对苯醌和1,2,4-苯三酚抑制了DNA聚合酶α，从而导致细胞和无细胞DNA合成系统中的DNA合成受到抑制。本研究确定儿茶酚、对苯二酚、对苯醌和1,2,4-苯三酚为骨髓细胞中的有毒苯代谢产物，并表明它们对DNA合成的抑制作用是由除涉及DNA损伤作为主要原因之外的其他机制介导的。
