虾肽对过氧化氢诱导RAW 264.7细胞氧化损伤的保护作用

Protective effects of shrimp peptides on hydrogen peroxide-induced oxidative damage in RAW264.7 Cells

为研究虾肽对H_(2)O_(2)诱导的RAW 264.7细胞氧化损伤的保护作用,利用CCK-8法检测不同剂量虾肽对RAW 264.7细胞的存活率的影响,通过酶联免疫试剂盒和荧光实时定量PCR检测细胞因子和mRNA的表达水平;Western Blotting检测细胞中核因子E2相关因子2(nuclear factor E2-related factor 2,Nrf2)、Kelch样环氧氯丙烷相关蛋白1(Kelch like-ECH-associated protein 1,Keap1)蛋白含量的变化。结果表明,提前给予虾肽干预的细胞的存活率明显增加,虾肽能提高细胞内超氧化物歧化酶(superoxide dismutase,SOD)、谷胱甘肽过氧化酶和过氧化氢酶的活力,而且细胞内SOD 1、谷胱甘肽过氧化物酶1、Nrf2、醌氧化还原酶1、血红素加氧酶-1 mRNA的表达量显著升高(P<0.05)。此外,细胞内Keap1蛋白相对表达量显著降低,而Nrf2蛋白相对表达量显著升高(P<0.05),说明提前给予虾肽干预对H_(2)O_(2)诱导RAW 264.7细胞氧化应激具有预防保护作用,其作用机制与激活Nrf2/Keap1通路有关。

The aim of the study was to investigate the protective effects of shrimp peptides against hydrogen peroxide(H_(2)O_(2))induced oxidative damage in RAW264.7 cells by CCK-8 assay,enzyme linked immunosorbent assay(ELISA),realtime-PCR and Western Blotting.The results from CCK-8 assay showed that shrimp peptides could significantly increase viability of cells compared with RAW 264.7 cells exposed to H_(2)O_(2)(P<0.05).Moreover,shrimp peptides significantly increase the levels of superoxide dismutase(SOD),glutathione reductase(GSH-Px)and catalase(CAT)in RAW264.7 cells.RT-PCR showed that shrimp peptides could increase the gene expression levels of SOD1,GPX1,Nrf2,NQO1 and HO-1(P<0.05).In addition,shrimp peptides could also observably increase the protein expression level of Nrf2 and decrease the protein expression level of Keap1 with Western Blotting.Collectively,this study suggested that shrimp peptides could effectively protect RAW264.7 cells against H 2O 2-induced oxidative damage.This protection mechanism may be related to activation of the Nrf2/Keap1 signaling pathway.