摘要
Objective To determine if DNA excision repair enzymes oxoguanine glycosylase 1 （OGG1） and xeroderma pigmentosum group F protein （XPF） are involved in the pathogenesis of Parkinson＇s disease （PD） in a cell model. Methods PC12 cells were treated with 1-Methyl-4-phenylpyridine ion （MPP＋） for various periods of time to induce oxidative DNA damage. MTT assay was used to determine cell viability. Immunocytochemistry with antibody against 8-hydroxy-2＇- deoxyguanosine （8-oxodG） was used to evaluate oxidative DNA damage. Immunoblotting was used to detect the protein levels of OGG1 and XPF. Results MPP＋ treatment （1 mmol/L） for 18 h and 24 h reduced cell viability to 78.6% and 70.3% of the control, respectively, in a time-dependent way. MPP＋ increased the immunoreactivity of 8-oxodG in the cytoplasm at 3 h and in the nucleus at 24 h of treatment. With the treatment of MPP＋, the expression of OGG1 was significantly increased at 1 h, reaching a peak at 3 h, and then it was decreased at 24 h, as compared to that with vehicle treatment. The same effect was exerted on XPF level, except that the XPF level reached a peak at 18 h of MPP＋ treatment. Moreover, the maximally-increased protein level of OGG1 by MPP＋ was approximately 2-fold higher than that of XPF. Conclusion MPP＋ treatment could time- dependently induce increases in OGG1 and XPF expressions in PC12 cells. Also, this study indicates that the base and nucleotide excision repair pathways may be compensatorily activated in the early stage of pathogenesis in the cells after MPP＋ treatment.
目的探讨离体条件下DNA剪切修复相关蛋白酶氧基-鸟嘌呤DNA糖苷酶 (OGG1) 和着色性干皮病蛋白F(XPF) 是否参与帕金森病 (PD) 的病理过程。方法用1 mmol/L 1-甲基-4-苯基吡啶 (MPP+) 处理PC12细胞不同时间,诱导细胞产生DNA氧化损伤,建立PD细胞模型。在此模型上,采用MTT法检测细胞活力;采用8-氧基-7,8- 双氢脱氧鸟嘌呤 (8-oxodG) 免疫细胞化学技术检测细胞DNA氧化损伤;用Western blot技术检测细胞内OGG1和XPF的蛋白表达水平。结果MPP+ 时程依赖性地降低细胞活力,18 h 和24 h 后细胞活力分别降至对照组的78.6% 和70.3%。MPP+ 显著增加了 8-oxodG 免疫阳性反应,在 3 h 时 8-oxodG 主要分布于细胞质,在 24 h 时其主要位于细胞核内。此外,MPP+ 显著改变了 OGG1 和 XPF 的蛋白表达水平。在 MPP+ 处理 1 h 后,OGG1 和 XPF 蛋白表达水平均显著上调,分别于3 h和18 h达到顶峰,随后依次下降,且OGG1的峰值几乎是XPF的两倍。结论MPP+时程依赖性地增加了PC12细胞内XPF和OGG1的表达水平。本研究提示,在MPP+致PD的病理模型早期,碱基切除修复和核苷酸剪切修复可能被代偿性地激活。
基金
supported by the National Natural Science Foundation of China (No. 30770660,J0730860)
