Department of Pathology, University of Texas Medical Branch, Galveston, TX 77555, USA.
Toxicol Appl Pharmacol. 2013 Mar 15;267(3):276-83. doi: 10.1016/j.taap.2013.01.005. Epub 2013 Jan 23.
Mechanisms by which aniline exposure elicits splenotoxicity, especially a tumorigenic response, are not well-understood. Earlier, we have shown that aniline exposure leads to oxidative DNA damage and up-regulation of OGG1 and NEIL1/2 DNA glycosylases in rat spleen. However, the contribution of endonuclease III homolog 1 (NTH1) and apurinic/apyrimidinic endonuclease 1 (APE1) in the repair of aniline-induced oxidative DNA damage in the spleen is not known. This study was, therefore, focused on examining whether NTH1 and APE1 contribute to the repair of oxidative DNA lesions in the spleen, in an experimental condition preceding tumorigenesis. To achieve this, male SD rats were subchronically exposed to aniline (0.5 mmol/kg/day via drinking water for 30 days), while controls received drinking water only. By quantitating the cleavage products, the activities of NTH1 and APE1 were assayed using substrates containing thymine glycol (Tg) and tetrahydrofuran, respectively. Aniline treatment led to significant increases in NTH1- and APE1-mediated BER activity in the nuclear extracts of spleen of aniline-treated rats compared to the controls. NTH1 and APE1 mRNA expression in the spleen showed 2.9- and 3.2-fold increases, respectively, in aniline-treated rats compared to the controls. Likewise, Western blot analysis showed that protein expression of NTH1 and APE1 in the nuclear extracts of spleen from aniline-treated rats was 1.9- and 2.7-fold higher than the controls, respectively. Immunohistochemistry indicated that aniline treatment also led to stronger immunoreactivity for both NTH1 and APE1 in the spleens, confined to the red pulp areas. These results, thus, show that aniline exposure is associated with induction of NTH1 and APE1 in the spleen. The increased repair activity of NTH1 and APE1 could be an important mechanism for the removal of oxidative DNA lesions. These findings thus identify a novel mechanism through which NTH1 and APE1 may regulate the repair of oxidative DNA damage in aniline-induced splenic toxicity.
苯胺暴露引发脾毒性，尤其是致癌反应的机制尚不清楚。我们之前已经表明，苯胺暴露会导致大鼠脾脏中的氧化 DNA 损伤和 OGG1 和 NEIL1/2 DNA 糖苷酶的上调。然而，endonuclease III homolog 1 (NTH1) 和 apurinic/apyrimidinic endonuclease 1 (APE1) 在修复苯胺诱导的脾脏氧化 DNA 损伤中的作用尚不清楚。因此，本研究的重点是研究在致癌发生之前的实验条件下，NTH1 和 APE1 是否有助于修复脾脏中的氧化 DNA 损伤。为此，雄性 SD 大鼠通过饮用水（每天 0.5 mmol/kg，持续 30 天）亚慢性暴露于苯胺，而对照组仅接受饮用水。通过定量分析切割产物，使用含有胸腺嘧啶二醇（Tg）和四氢呋喃的底物分别测定 NTH1 和 APE1 的活性。与对照组相比，苯胺处理导致苯胺处理大鼠脾脏核提取物中 NTH1 和 APE1 介导的 BER 活性显著增加。与对照组相比，苯胺处理大鼠脾脏中 NTH1 和 APE1 的 mRNA 表达分别增加了 2.9 倍和 3.2 倍。同样，Western blot 分析表明，苯胺处理大鼠脾脏核提取物中 NTH1 和 APE1 的蛋白表达分别比对照组高 1.9 倍和 2.7 倍。免疫组织化学分析表明，苯胺处理也导致 NTH1 和 APE1 在脾脏中的免疫反应性增强，仅限于红髓区域。因此，这些结果表明，苯胺暴露与脾脏中 NTH1 和 APE1 的诱导有关。NTH1 和 APE1 修复活性的增加可能是清除氧化 DNA 损伤的重要机制。这些发现确定了一种新的机制，通过该机制，NTH1 和 APE1 可能调节苯胺诱导的脾毒性中氧化 DNA 损伤的修复。
