研究报告
梁冯,王泽夏,晏彪.NF-κB通路参与邻苯二甲酸二丁酯诱导U251细胞毒性的作用研究[J].环境科学学报,2019,39(2):640-648
NF-κB通路参与邻苯二甲酸二丁酯诱导U251细胞毒性的作用研究
- Role of the NF-κB pathway in dibutyl phthalate mediated effects in U251 cells
- 基金项目：湖北省卫生计生委面上项目（No.WJ2017M166）；湖北省教育厅科学技术研究项目（No.B2018172）
- 作者
- 单位
- 梁冯
- 湖北中医药大学检验学院, 武汉 430065
- 王泽夏
- 湖北科技学院基础医学研究中心, 咸宁 437100
- 晏彪
- 湖北科技学院基础医学研究中心, 咸宁 437100
- 摘要：为探究nuclear factor-κB（NF-κB）通路及其分子事件相关蛋白参与邻苯二甲酸二丁酯（dibutyl-phthalate，DBP）诱导神经毒性的作用机制，以人胶质瘤细胞U251为对象，设置6个实验组：Control组，Tween 80组（阳性对照组）、25 μmol·L-1 DBP组、100 μmol·L-1 DBP组、20 μmol·L-1维生素E（vitamin E，VE）组、100 μmol·L-1 DBP+20 μmol·L-1 VE组.U251细胞经不同处理组分别暴露12 h及24 h后，观察细胞形态学结晶紫染色结果，检测细胞活性（CCK-8）、活性氧（reactive oxygen species，ROS）、丙二醛（malondialdehyde，MDA）、氧化应激转录因子NF-κB，以及NF-κB通路相关蛋白钙调神经磷酸酶（calcineurin，CaN）、泛素特异性蛋白酶14（ubiquitin-specific protease 14，USP14）、胶质细胞源性神经营养因子（glial cell line-derived neurotrophic factor，GDNF）的表达水平.与对照组相比，100 μmol·L-1 DBP组U251细胞的数量降低，细胞损伤，CCK-8降低，ROS、MDA含量显著升高，NF-κB水平显著升高，CaN、USP14表达均升高，GDNF表达降低，差异均有统计学意义（p<0.05，p<0.01）；加入抗氧化剂VE处理后，与100 μmol·L-1 DBP组相比，100 μmol·L-1 DBP+20 μmol·L-1 M VE组U251细胞的数量增多，细胞损伤得到缓解，CCK-8上升，ROS、MDA含量显著降低，NF-κB水平显著降低，CaN、USP14表达均降低，GDNF表达增加，差异均有统计学意义（p<0.05，p<0.01）.NF-κB和NF-κB通路相关蛋白的水平或含量变化表明，NF-κB作为氧化应激转录因子参与DBP诱导的U251细胞毒性作用；VE作为抗氧化剂可阻断ROS的生成从而抑制NF-κB的活化，提示VE在体外对DBP介导的神经毒性具有一定的保护作用.
- Abstract：To investigate the role of nuclear factor-κB (NF-κB) pathway and its related proteins involved in dibutyl-phthalate (DBP) induced neurotoxicity in U251 glioma cells. Six experimental groups were treated as follows:control group, Tween 80 group, 25 μmol·L-1 DBP group, 100 μmol·L-1 DBP, 20 μmol·L-1 vitamin E (VE), and 100 μmol·L-1 DBP combined with after-treatment of VE group. U251 cells were exposed to different treatment groups for 12 h and 24 h, respectively. The morphological changes of U251 cells were observed by crystal violet staining. The expression levels of CCK-8, reactive oxygen species (ROS), malondialdehyde (MDA), NF-κB (an oxidative stress transcription factor), calcineurin (CaN), ubiquitin-specific protease 14 (USP14) and glial cell line-derived neurotrophic factor (GDNF) were detected. Compared with the control group, the number of U251 cells in 100 μmol·L-1 DBP group was decreased, the contents of ROS and MDA were significantly increased (p<0.05, p<0.01), the expression of CaN and USP14 were significantly increased (p<0.05, p<0.01), and the expression of GDNF was decreased (p<0.05, p<0.01), respectively. After the antioxidant VE was added to the treatment, compared with the100 μmol·L-1 DBP group, the number of U251 cells in 100 μmol·L-1 DBP + 20 μmol·L-1 VE group was increased, the cell damage was alleviated, the content of ROS and MDA were significantly decreased (p<0.05, p<0.01), and the expression of CaN and USP14 were decreased significantly (p<0.05, p<0.01), and the expression of GDNF was increased (p<0.05, p<0.01). The changes of NF-κB and NF-κB pathway related proteins suggest that NF-κB, as an oxidative stress transcription factor, is associated with the cytotoxicity of U251 cells induced by DBP. And VE can inhibit the activation of NF-κB by blocking the production of ROS, suggesting that VE has a protective effect on neurotoxicity mediated by DBP in vitro.
摘要点击次数： 742 全文下载次数： 901
