矢车菊素-3-O-葡萄糖苷保护RAW264.7细胞免受过氧化氢诱导的氧化损伤  

Cyanidin-3-O-Glucoside Protects RAW264.7 Cells against Hydrogen Peroxide-Induced Oxidative Damage

作　　者：薛宏坤 谭佳琪 李倩[1] 唐劲天[1] XUE Hongkun;TAN Jiaqi;LI Qian;TANG Jintian(Key Laboratory of Particle&Radiation Imaging,Ministry of Education,Department of Engineering Physics,Tsinghua University,Beijing 100084,China;Academy for Advanced Interdisciplinary Studies,Peking University,Beijing 100080,China)

机构地区：[1]清华大学工程物理系,粒子与辐射成像教育部重点实验室,北京100084 [2]北京大学前沿交叉学科研究院,北京100080

出　　处：《食品科学》2021年第13期103-113,共11页Food Science

基　　金：国家自然科学基金面上项目(31560396)。

摘　　要：本研究旨在探究矢车菊素-3-O-葡萄糖苷(cyanidin-3-O-glucoside,C3G)对H2O2诱导RAW264.7细胞氧化损伤的保护作用及其机制。通过噻唑蓝法测定C3G和过氧化氢分别对RAW264.7细胞存活率的影响;采用酶联免疫吸附测定法检测细胞内超氧化物歧化酶(superoxide dismutase,SOD)和谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)活力以及一氧化氮(nitric oxide,NO)和丙二醛(malondialdehyde,MDA)水平;利用2’,7’-二氯荧光素二乙酸酯活性氧荧光探针检测细胞内活性氧(reactive oxygen species,ROS)水平;通过逆转录定量聚合酶链式反应和Western blot法分别测定相关mRNA和蛋白表达水平。结果表明,C3G(6.25~25.00μmol/L)能显著抑制H2O2诱导RAW264.7细胞活性降低(P<0.05)。C3G显著降低H2O2诱导RAW264.7细胞内ROS过表达、MDA水平和NO释放(P<0.05),显著增加SOD和GSH-Px活力(P<0.05)。此外,与空白对照相比,400μmol/L H2O2处理组中,蛋白激酶Mst1/2和Kelch样环氧氯丙烷相关蛋白Keap1的mRNA及蛋白相对表达水平显著上调(P<0.05),而细胞中核因子E2相关因子和下游抗氧化酶HO-1的mRNA及蛋白相对表达水平显著下调(P<0.05);而采用C3G干预RAW264.7细胞,上述相关mRNA及蛋白的相对表达水平被逆转。结论:C3G对H2O2诱导RAW264.7细胞氧化损伤保护作用,可能与激活Mst/Nrf2信号通路和提高抗氧化酶活性有关。The aim of this study is to evaluate the cytoprotection and potential molecular mechanisms of cyanidin-3-Oglucoside(C3G)on hydrogen peroxide(H2O2)-induced oxidative damage in RAW264.7 cells.The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)assay was conducted to determine the viability of RAW264.7 cells exposure to H2O2 or C3G.Meanwhile,we measured the antioxidant properties of C3G by determining the activities of superoxide dismutase(SOD)and glutathione peroxidase(GSH-Px),nitric oxide(NO)release and malondialdehyde(MDA)levels by enzyme-linked immunosorbent assay(ELISA).2’,7’-dichlorofluorescin diacetate(DCFH-DA)was employed to evaluate the production of intracellular reactive oxygen species(ROS).Finally,the expression levels of related mRNA/protein were evaluated by reverse transcription polymerase chain reaction(RT-PCR)and Western blot analysis,respectively.The results showed that the H2O2-induced decrease in the cell viability of RAW264.7 cells was remarkably suppressed C3G(6.25–25.00μmol/L).C3G significantly inhibited the H2O2-induced of overproduction of intracellular ROS,NO release and MDA levels,but increased the activities of intracellular SOD and GSH-Px(P<0.05).In addition,the relative mRNA and protein expression levels of Mst1,Mst2 and Keap1 were up-regulated,while the mRNA and protein relative expression levels of Nrf2 and HO-1 were down-regulated in the 400μmol/L H2O2-treated group when compared to the vehicle-treated group.However,the above changes were reversed by intervention with C3G.C3G could exert a cytoprotective effect possibly by activating the Mst/Nrf2 signaling pathway and improving the activities of antioxidant enzymes.

关 键 词：矢车菊素-3-O-葡萄糖苷 RAW264.7细胞 氧化损伤 Mst/Nrf2信号通路 

分 类 号：TS201.4[轻工技术与工程—食品科学]