摘要
研究三七总皂苷对6-羟基多巴胺诱导SH-SY5Y细胞损伤的保护作用并探讨可能机制。采用MTT法检测SH-SY5Y细胞存活率;试剂盒测定乳酸脱氢酶(LDH)漏出率、活性氧(ROS)水平、丙二醛(MDA)含量以及抗氧化酶超氧化物歧化酶(SOD)、过氧化氢酶(CAT)和谷胱甘肽过氧化物酶(GSH-Px)活力;TUNEL染色检测细胞凋亡;JC-1染色观察线粒体膜电位;caspase-3活性检测试剂盒测定caspase-3活性;蛋白印迹(Western blot)检测细胞核中转录因子NF-F2相关因子(Nrf2)和胞浆中血红素加氧酶(HO-1)蛋白表达。结果显示,三七总皂苷能明显提高SH-SY5Y细胞存活率,抑制LDH漏出,减少ROS和MDA产生,增加SOD、CAT和GSH-Px活力,稳定线粒体膜电位,抑制caspase-3的活化。Western blot结果显示,三七总皂苷能够上调SH-SY5Y细胞核Nrf2和HO-1的蛋白表达,同时HO-1抑制剂锡原卟啉(Sn PP)能够抑制三七总皂苷的细胞保护作用。结果表明,三七总皂苷对6-OHDA诱导的SH-SY5Y细胞损伤具有一定的保护作用。该保护作用可能与激活Nrf2、上调HO-1表达、抑制6-OHDA诱导SH-SY5Y细胞氧化应激和凋亡有关。
The aim of this study is to investigate the protective effects of Panax notoginseng saponins（PNS） against 6-hydroxydopamine（6-OHDA）-induced apoptosis in SH-SY5Y cells and the possible underlying mechanisms. Cell viability was examined by MTT assay. The levels of lactate dehydrogenase（LDH）, reactive oxygen species（ROS）, malondialdehyde（MDA） and the activities of superoxide dismutase（SOD）, catalase（CAT）, glutathione peroxidase（GSH-Px） were measured using the respective assay kits. Apoptosis was measured by TUNEL kit, JC-1 and ROS was measured by staining with fluorescent dyes. The activation of caspase-3 was measured with the caspase-3 assay kit. The expression of nuclear protein Nrf2 and HO-1 were determined by Western blot. PNS had significant protective effects against 6-OHDA-induced apoptosis in SH-SY5Y cells in a time- and dose-dependent manner. PNS could attenuate 6-OHDA-induced suppression of SOD, GAT, GSH-Px（P 0.01）. PNS reduced the level of LDH, decreased the levels of ROS, MDA and increased cell viability and the mitochondrial membrane potential（P 0.01）. PNS also inhibited DNA fragmentation, mitochondrial response and the activation of caspase-3（P 0.01）. Moreover, PNS pretreatment increased the expression of the nuclear Nrf2 and up-regulate HO-1. The protective effects of PNS could be inhibited by HO-1 inhibitor Sn PP. In conclusion,PNS has significant protective effects against 6-OHDA- induced apoptosis in SH-SY5Y cells. The possible mechanisms of PNS are due to PNS-mediated activation of Nrf2, up-regulation of HO-1 and inhibition of oxidative stress.
出处
《药学学报》
CAS
CSCD
北大核心
2016年第6期898-906,共9页
Acta Pharmaceutica Sinica
基金
国家自然科学基金资助项目(81503290)
中医药行业科研专项(201507004)
国家重大创新药创制专项(2012ZX09501001-004)
关键词
帕金森病
三七总皂苷
氧化应激
凋亡
NRF2
Parkinson＇s disease
Panax notoginseng saponin
oxidative stress
apoptosis
Nrf2
