摘要
目的 分析枸杞多糖(LBP)干预对氧化损伤的人肾小管上皮细胞(HK-2)内的核因子E2相关因子2(Nrf2)/血红素氧合酶-1(HO-1)通路蛋白表达的变化,探索LBP拮抗HK-2细胞氧化损伤的分子机制.方法 采用过氧化氢诱导HK-2细胞制备氧化损伤细胞模型,HK-2被分成4组:正常组、LBP组、氧化损伤组及LBP干预组.观察4组细胞的长势和形态变化;细胞活力测定法比较每组细胞的存活率;比色法分析氧化产物丙二醛(malondialde-hyde,MDA)含量以及超氧化物歧化酶(superoxide dismutase,SOD)和谷胱甘肽过氧化酶(GSH-Px)的水平;免疫印迹技术检测细胞内Nrf2/HO-1通路蛋白Nrf2、HO-1的相对水平.结果 正常组和LBP组相比,两组的细胞长势、形态和存活率,细胞产生MDA的量、SOD和GSH-Px的水平,通路蛋白Nrf2、HO-1的相对水平均无明显差异(均P＞0.05);氧化损伤组和正常组相比,细胞皱缩变圆并脱落、贴壁细胞变少,存活率减少、MDA量增加、SOD和GSH-Px降低,通路蛋白Nrf2、HO-1的相对值下降(均P＜0.05);LBP干预组和氧化损伤组相比,细胞长势、形状恢复、贴壁细胞较多,存活率增加、MDA量减少、SOD和GSH-Px升高,通路蛋白Nrf2、HO-1的相对值升高(均P＜0.05).结论 枸杞多糖能通过激活Nrf2/HO-1通路提高HK-2细胞的抗氧化酶活性达到减轻细胞氧化损伤的目的.
Abstract
Objective To analyze the expression changes of nuclear factor erythroid-2 related factor 2(Nrf2)/heme oxygenase-1(HO-1)pathway protein in oxidative damaged human renal tubular epithelial cells(HK-2)after interven-tion with lycium barbarum polysaccharides(LBP),and explore the molecular mechanism of LBP antagonizing oxidative damage in HK-2 cells.Methods A oxidative damage cell model was prepared by inducing HK-2 cells with hydrogen peroxide.The cells were arranged into 4 groups:normal group,LBP group,oxidative damage group,and LBP interven-tion group.The 4 groups have the same duration of action.Observe the growth and morphological changes of 4 groups of cells;Compare the survival rate of each group cells using cell viability assay;Colorimetric test box was used to ana-lyze the content of malondialdehyde(MDA),superoxide dismutase(SOD)and glutathione peroxidase(GSH-Px);West-ern blotting was used to detect the relative levels of Nrf2/HO-1 pathway proteins:Nrf2 and HO-1.Results In contrast to normal group,LBP group had not obvious differences in cell growth,morphology,survival rate,MDA,SOD and GSH-Px levels,relative levels of pathway proteins Nrf2,HO-1(all P＞0.05);The oxidative damage group showed re-duced cell shrinkage and detachment,fewer adherent cells,survival rate reduced,MDA increased,SOD and GSH-Px de-creased,and relative values of Nrf2 and HO-1 increased(all P＜0.05);In contrast to the oxidative damage group,the LBP intervention group showed more cell growth,shape recovery,more adherent cells,survival rate increased,MDA de-creased,SOD and GSH-Px increased,and relative values of Nrf2 and HO-1(all P＜0.05).Conclusion Lycium barba-rum polysaccharides can enhance the antioxidant enzyme activity of HK-2 cells by activating the Nrf2/HO-1 pathway,achieving the goal of reducing cellular oxidative damage.
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