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目的探讨茶多酚中主要活性成分没食子儿茶素没食子酸酯(EGCG)拮抗百草枯诱导人神经母细胞瘤细胞株SK-N-SH细胞凋亡的作用机制。方法培养SK-N-SH细胞并给予400μmol.L-1百草枯作用72 h诱导细胞凋亡,建立细胞凋亡模型。实验分为6组:空白对照组、百草枯模型组、维生素E(10μmol.L-1)组和EGCG(10、5、1μmol.L-1)3个剂量组。药物处理细胞2 h后,加入百草枯,72 h后检测细胞内超氧化物歧化酶(SOD)活性及丙二醛(MDA)、活性氧(ROS)的含量,罗丹明123染色及流式细胞术测线粒体膜电位,RT-PCR检测细胞bcl-2 mRNA与bax mRNA表达情况。结果与空白对照组比较,百草枯模型组细胞SOD活性降低,MDA和ROS含量增加,线粒体膜电位下降,bcl-2 mRNA表达减弱,bax mRNA表达增强,bcl-2/bax比值降低(均P<0.01)。对百草枯损伤的SK-N-SH细胞,维生素E和EGCG可明显提高SOD活性,降低MDA和ROS的含量,恢复线粒体膜电位,增强bcl-2 mRNA表达,减弱bax mRNA表达,升高bcl-2/bax比值(P<0.05或P<0.01)。结论 EGCG拮抗百草枯诱导SK-N-SH细胞凋亡的机制可能与提高细胞抗氧化能力、保护线粒体结构和功能的完整性、抑制线粒体凋亡基因过度
Objective To investigate the mechanism of action of epigallocatechin gallate (EGCG), a main active component of tea polyphenols, on paraquat-induced apoptosis in human neuroblastoma SK-N-SH cells. Methods SK-N-SH cells were cultured and paracetamol (400μmol·L-1) was added for 72 h to induce apoptosis. Apoptosis model was established. The experiment was divided into 6 groups: blank control group, paraquat model group, vitamin E (10μmol.L-1) group and EGCG (10,5,1μmol.L-1) three dose groups. Paraquat was added to the cells for 2 h after the drug treatment, and the activity of superoxide dismutase (SOD) and the contents of malondialdehyde (MDA) and reactive oxygen species (ROS) in the cells were measured 72 h later, and rhodamine 123 staining and flow cytometry The mitochondrial membrane potential was measured and the expression of bcl-2 mRNA and bax mRNA was detected by RT-PCR. Results Compared with the blank control group, the activity of SOD, the content of MDA and ROS in paraquat model group decreased, the expression of bcl-2 mRNA and bcl-2 / bax decreased, the mitochondrial membrane potential decreased, 0.01). Vitamin E and EGCG could significantly increase the activity of SOD, decrease the contents of MDA and ROS, restore the mitochondrial membrane potential, increase the expression of bcl-2 mRNA, decrease the expression of bax mRNA and increase the expression of bcl-2 in paraquat-damaged SK-N-SH cells. 2 / bax ratio (P <0.05 or P <0.01). Conclusion The mechanism of EGCG antagonizing the paraquat-induced apoptosis in SK-N-SH cells may be related to enhancing cell antioxidant capacity, protecting the integrity of mitochondrial structure and function, inhibiting mitochondrial apoptotic gene over-expression