摘要
目的探究京尼平苷(geniposide,GP)通过AMP依赖的蛋白激酶[Adenosine 5′-monophosphate(AMP)-activated protein kinase,AMPK]/哺乳动物雷帕霉素靶点(mammalian target of rapamycin,mTOR)信号通路对1-甲基4-苯基-四氢吡啶离子(1-methyl-4-phenyl 1,2,3,6-tetrahydropyridine,MPP+)诱导人神经母细胞瘤株SH-SY5Y细胞获得的帕金森细胞模型细胞凋亡及氧化应激损伤的影响。方法以SH-SY5Y细胞为研究对象,按照处理方法将其分为Control组(正常培养)、MPP+组(在Control组基础上添加1 mmol/L MPP+)、GP组(在Control组基础上添加100μmol/L GP)、MPP++GP组(在MPP+组基础上添加100μmol/L GP)。使用丙二醛(malondialdehyde,MDA)、乳酸脱氢酶(lactate dehydrogenase,LDH)及超氧化物歧化酶(superoxide dismutase,SOD)的酶联免疫吸附测定试剂盒、流式细胞术及Western blot法检测各组细胞氧化应激损伤、细胞凋亡及AMPK、mTOR、LC3蛋白相关表达水平的差异。应用AMPK激活剂、mTOR抑制剂处理后,再次使用Western blot检测MPP+组及MPP++GP组细胞上述蛋白表达水平的变化。结果相较于Control组,MPP+组细胞凋亡率、LDH活性、MAD含量、AMPK磷酸化水平及LC3-Ⅱ/LC3-Ⅰ比值明显增加,SOD活性及mTOR磷酸化水平明显降低(P<0.01)。相较与MPP+组,MPP++GP组细胞凋亡率、LDH活性、MAD含量、AMPK磷酸化水平及LC3-Ⅱ/LC3-Ⅰ比值明显降低,SOD活性及mTOR磷酸化水平明显增加(P<0.01)。而添加AMPK激活剂或mTOR抑制剂后,MPP+组及MPP++GP组细胞AMPK磷酸化水平及LC3-Ⅱ/LC3-Ⅰ比值明显升高,mTOR磷酸化水平明显降低(P<0.05)。结论GP部分通过调控AMPK/mTOR信号通路抑制MPP+诱导的SH-SY5Y细胞凋亡、氧化应激损伤及自噬。
Objective To investigate the effect of geniposide(GP)on the apoptosis and oxidative stress injury of the Parkinson′s disease model obtained by 1-methyl-4-phenyl 1,2,3,6-tetrahydropyridine(MPP+)induced human neuroblastoma SH-SY5Y cells through Adenosine 5′-monophosphate(AMP)-activated protein kinase/mammalian target of rapamycin(AMPK/mTOR)pathway.Methods The SH-SY5Y cells were selected as the research subject,and according to different treatment methods,they were divided into Control group(normal culture),MPP+group(supplemented with 1 mmol/L MPP+on the basis of the Control group),GP group(supplemented with 100μmol/L GP on the basis of the Control group),and MPP++GP group(supplemented with 100μmol/L GP on the basis of the MPP+group).Malondialdehyde(MDA),lactate dehydrogenase(LDH)and superoxide dismutase(SOD)kits,flow cytometry and Western blot were used to detect the changes in oxidative stress injury of cells,cell apoptosis and related proteins expression levels of AMPK,mTOR and LC3 in each group.The changes in the related expression levels of the above proteins in MPP+group and MPP++GP group were detected by Western blot after treatment with AMPK activator or mTOR inhibitor.Results Compared with the Control group,the cell apoptosis,LDH activity,MDA content,AMPK phosphorylation level,LC3-Ⅱ/LC3-Ⅰratio were increased,while SOD activity and mTOR phosphorylation level were reduced in the MPP+group(P<0.01).Compared with the MPP+group,the cell apoptosis,LDH activity,MDA content,AMPK phosphorylation level,and LC3-Ⅱ/LC3-Ⅰratio were reduced,while SOD activity and mTOR phosphorylation level were increased in the MPP++GP group(P<0.01).After AMPK activator or mTOR inhibitor was added,the AMPK phosphorylation level and LC3-Ⅱ/LC3-Ⅰratio were increased,while mTOR phosphorylation level was reduced in the MPP+group and MPP++GP group.Conclusion GP can inhibit MPP+induced apoptosis of SH-SY5Y cells,oxidative stress injury and autophagy partly by AMPK/mTOR pathway.
作者
朱宁
马晓珊
陈春丽
黎国仙
杨秀娟
谢东
ZHU Ning;MA Xiao-shan;CHEN Chun-li;LI Guo-xian;YANG Xiu-juan;XIE Dong(Department of Pharmacy, Nanhai Hospital of Southern Medical University, Guangdong Province, Foshan 528244, China;Department of Pharmacy, Zhujiang Hospital, Southern Medical University, Guangdong Province, Guangzhou 510282, China)
出处
《河北医科大学学报》
CAS
2021年第9期1008-1015,共8页
Journal of Hebei Medical University
基金
国家自然科学基金(82073805)
广东省自然科学基金(2018030310289)
广东省医学科学技术研究基金项目(A2020114)。
关键词
帕金森病
细胞凋亡
京尼平苷
Parkinson disease
cell apoptosis
geniposide
