PC12细胞中氧化应激对神经黑色素合成代谢的影响

Effect of Oxidative Stress on the Synthesis of Neuromelanin in PC12 Cells

目的研究PC12细胞中氧化应激对神经黑色素(neuromelanin,NM)合成量的影响及其可能机制。方法将PC12细胞置于含50μmol/L左旋多巴(L-dopa)的培养液中培养,采用改良的MTT法观察不同浓度(0、10、20、40 mU/mL)的葡萄糖氧化酶(glucose oxidase,GO)对PC12细胞活力的影响,对照组含同等浓度GO但不含L-dopa,共聚焦显微镜检测GO作用4 h后含L-dopa不同组之间的活性氧(ROS)水平,采用SP法检测含L-dopa不同组之间酪氨酸羟化酶(TH)表达量的变化,使用透射电镜观察PC12细胞超微结构变化。结果随着GO浓度增加,细胞内ROS明显增加,TH表达量逐渐增高,在第20天观察到细胞内出现NM,且随GO浓度增加而增加,对照组中未见NM出现。结论氧化应激可以促进NM的合成,其机制可能与提高TH活性有关。

Objective To study the effect of oxidative stress on the synthesis of neuromelanin in PC12 cells and its possible mechanism.Methods PC12 cells were incubated in 50 μmol/L levodopa（L-dopa）with different concentrations（0,10,20,40 mU/mL）of glucose oxidase（GO）.The modified MTT assay was used to evaluate the cell viabilities.In the control groups with the same concentrations of GO in the absence of L-dopa,the level of ROS was determined by a laser scanning confocal microscope 4 h later.The activity of tyroxine hydroxylase in PC12 cells was detected by immunohistochemistry（SP method）.The morphological changes of PC12 cells were observed under the transmisssion electron microscopy.Results With the increase in concentration of GO,the level of ROS was increased significantly,and TH activity was gradually increased.At the 20th day,there appeared neuromelanin in PC12 cells,which was increased with the increased concentration of GO,while there was no neuromelanin in the control groups.Conclusion Oxidative stress can promote the synthesis of neuromelanin,which may be related to the increased TH activity.