摘要
目的研究茶多酚在甲基苯丙胺(MPP^(+))诱导的PC12细胞神经元损伤中的作用机制。方法研究分为对照组、模型组和低、高剂量实验组。对照组在RPMI 1640培养基中培养,模型组在对照组的基础上用MPP^(+)(1 mmol·L^(-1))处理细胞24 h。低、高剂量实验组在添加MPP^(+)之前分别用10μmol·L^(-1)和20μmol·L^(-1)的茶多酚预处理1 h。观察各组细胞形态变化,以细胞计数法-8(CCK-8)检测细胞活力,用流式细胞术检测细胞凋亡情况,检测活性氧(ROS)表达水平,用Comet assay分析各组细胞DNA损伤情况,用蛋白质印迹法检测DNA修复相关蛋白表达水平。结果通过CCK-8实验和细胞形态学观察,发现模型组的细胞增殖率为0.36±0.02,而低、高剂量实验组的细胞增殖率为0.55±0.02、0.67±0.03,表明茶多酚可显著减轻MPP^(+)对PC12细胞的细胞毒性和形态损伤。进一步的实验表明,茶多酚可以降低MPP^(+)诱导的氧化应激程度,增强细胞内抗氧化能力,对照组、模型组和低、高剂量实验组的ROS水平分别为(14.36±1.34)%、(32.64±3.72)%、(21.64±2.08)%和(18.45±1.66)%,结果表明与对照组相比,模型组的ROS水平显著升高(P<0.05),而低、高剂量实验组组的ROS水平显著低于模型组(P<0.05),从而减轻氧化应激对细胞的损伤。此外,茶多酚还能够促进细胞DNA修复功能的活化,对照组、模型组和低、高剂量实验组的γ-H2AX水平分别为1.00±0.08、1.82±0.13、1.35±0.09和1.22±0.07;Ku70水平分别为1.00±0.10、0.49±0.06、0.82±0.08和0.98±0.10;XRCC1水平分别为1.00±0.11、0.51±0.06、0.90±0.09和1.15±0.12;与对照组相比,模型组γ-H2AX表达显著增加,Ku70和XRCC1的表达降低。低、高剂量实验组Ku70和XRCC1的表达显著增高,从而改善MPP^(+)对细胞DNA的损伤。结论茶多酚可以通过减少氧化应激和促进DNA修复来保护PC12细胞免受MPP^(+)诱导的神经毒性。
Objective To investigate the mechanism of tea polyphenols in methamphetamine(MPP^(+))-induced neuronal injury in PC12 cells.Methods This study was divided into control group,model group,low dose experimental group,high dose experimental group.The control group was cultured in RPMI 1640 medium,model group was treated with MPP^(+)(1 mmol·L^(-1))for 24 h on the basis of control group.The low dose and high dose experimental groups were pretreated with 10μmol·L^(-1) and 20μmol·L^(-1) tea polyphenols for 1 h before adding MPP^(+).The morphological changes of cells in each group were obs erv ed,the cell viability was detected by cell counting method-8(CCK-8),the apoptosis of cells was detected by flow cytometry,the expression level of reactive oxygen species(ROS)was detected,and the DNA damage of cells in each group was analyzed by Comet assay.Western blotting was used to detect the expression levels of DNA repair-related proteins.Results Through CCK-8 experiment and cell morphology observation,it was found that the cell proliferation rate of the model group were 0.36±0.02,while those in low dose experimental group,high dose experimental group were 0.55±0.02 and 0.67±0.03,indicating that tea polyphenols can significantly alleviate cytotoxicity and morphological damage of PC12 cells by MPP^(+).Further experiments showed that tea polyphenols could reduce the degree of oxidative stress induced by MPP^(+)and enhance intracellular antioxidant capacity.The ROS levels of control group,model group,low dose experimental group,high dose experimental group were(14.36±1.34)%,(32.64±3.72)%,(21.64±2.08)%and(18.45±1.66)%,the results showed that compared with the control group,the ROS level of model group was significantly increased(P<0.05),while the ROS level of low dose and high dose experimental groups was significantly lower than that of model group(P<0.05),thereby reducing the damage of oxidative stress to cells.In addition,tea polyphenols can also promote the activation of cellular DNA repair function.The levels ofγ-
作者
周妮娜
邓明珠
王灿
ZHOU Ni-na;DENG Ming-zhu;WANG Can(Department of Neurology,Hunan Brain Hospital,Changsha 410007,Hunan Province,China)
出处
《中国临床药理学杂志》
CAS
CSCD
北大核心
2023年第21期3097-3101,共5页
The Chinese Journal of Clinical Pharmacology
基金
湖南省卫生健康委科研计划基金资助项目(20201642)。
