番茄红素对人肝L02细胞氧化损伤的保护作用及其机制
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摘要
目的探索番茄红素(Lyc)对过氧化氢(H_2O_2)诱导的人肝L02细胞氧化损伤的保护作用及其机制。方法以H_2O_2诱导的L02细胞氧化损伤为模型,通过测定不同浓度番茄红素预处理后L02细胞的生存率以确定最佳处理浓度;通过测定细胞活性氧(ROS)水平、细胞内超氧化物歧化酶(SOD)与谷胱甘肽过氧化物酶(GSH-Px)活性、细胞内丙二醛(MDA)水平,培养液上清中的丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)及乳酸脱氢酶(LDH)活性等指标观察番茄红素对细胞氧化损伤的保护作用;通过检测细胞核蛋白中的核因子E2相关因子2(Nrf2)蛋白表达量及其靶基因血红素加氧酶1(HO-1)和辅酶Ⅰ醌类氧化还原酶1(NQO1)的mRNA表达水平等指标,观察番茄红素对Nrf2的核转位及其下游靶基因的激活作用。结果 10μmol/L番茄红素可显著提高H_2O_2培养条件下L02细胞的生存率,提高细胞内SOD、GSH-Px酶活性,降低胞内MDA含量及培养液中的ALT、AST、LDH酶活性(P<0.05),并能提高细胞核蛋白的Nrf2含量及Nrf2靶基因HO-1和NQO1表达水平(P<0.05)。结论番茄红素可通过促进Nrf2的核转位、增强其下游抗氧化基因的表达从而对H_2O_2诱导的L02细胞氧化损伤起保护作用。
Objective To explore lycopene 's protective effect on H_2O_2 induced oxidative damage of L02 cells and its mechanism. Methods L02 cells were cultured by H_2O_2 to build the model of cellular oxidative damage. Different dose of lycopene was used to pretreat the cells, and cell survival rate was detected to verify the appropriate concentration. Then cellular ROS level,activity of cellular SOD and GSH-Px,cellular MDA content,and activity of ALT,AST and LDH in the culture medium were detected to observe lycopene 's effect on cellular oxidant damage. Finally,to observe lycopene 's activating effect on nuclear-translocation of Nrf2,L02 cells' nuclear protein was extracted to detect Nrf2 protein content,and also,mRNA expression level of Nrf2 target genes HO-1 and NQO1 was assayed to verify this mechanism. Results Pretreatment of 10 μmol/Llycopene raised cellular viability of L02 cells on H_2O_2 culturing condition,reduced cellular ROS,enhanced enzymatic activity of cellular SOD and GSH-Px,reduced cellular MDA content,and depressed the activity of ALT,AST and LDH in culture medium. Lycopene also increased nuclear Nrf2 protein content and enhanced the expression of its target genes HO-1 and NQO1. Conclusion Lycopene could protect L02 cells from H_2O_2 induced oxidative damage,probably by promoting nuclear-translocation of Nrf2 and activating expression of its target antioxidant genes.
Objective To explore lycopene 's protective effect on H_2O_2 induced oxidative damage of L02 cells and its mechanism. Methods L02 cells were cultured by H_2O_2 to build the model of cellular oxidative damage. Different dose of lycopene was used to pretreat the cells, and cell survival rate was detected to verify the appropriate concentration. Then cellular ROS level,activity of cellular SOD and GSH-Px,cellular MDA content,and activity of ALT,AST and LDH in the culture medium were detected to observe lycopene 's effect on cellular oxidant damage. Finally,to observe lycopene 's activating effect on nuclear-translocation of Nrf2,L02 cells' nuclear protein was extracted to detect Nrf2 protein content,and also,mRNA expression level of Nrf2 target genes HO-1 and NQO1 was assayed to verify this mechanism. Results Pretreatment of 10 μmol/Llycopene raised cellular viability of L02 cells on H_2O_2 culturing condition,reduced cellular ROS,enhanced enzymatic activity of cellular SOD and GSH-Px,reduced cellular MDA content,and depressed the activity of ALT,AST and LDH in culture medium. Lycopene also increased nuclear Nrf2 protein content and enhanced the expression of its target genes HO-1 and NQO1. Conclusion Lycopene could protect L02 cells from H_2O_2 induced oxidative damage,probably by promoting nuclear-translocation of Nrf2 and activating expression of its target antioxidant genes.
引文
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[14]LI S,TAN H Y,WANG N,et al.The role of oxidative stress and antioxidants in liver diseases[J].Int J Mol Sci,2015,16(11):26087-26124.
[15]YANG J J,TAO H,HUANG C,et al.Nuclear erythroid 2-related factor 2:a novel potential therapeutic target for liver fibrosis[J].Food Chem Toxicol,2013,59(8):421-427.
[16]ZHANG H,DAVIES K J,FORMAN H J.Oxidative stress response and Nrf2 signaling in aging[J].Free Radic Biol Med,2015,88(Pt B):314-336.
[2]SERVIDDIO G,BELLANTI F,VENDEMIALE G.Free radical biology for medicine:learning from nonalcoholic fatty liver disease[J].Free Radic Biol Med,2013,65(6):952-968.
[3]SNCHEZ-VALLE V,CHVEZ-TAPIA N C,URIBE M,et al.Role of oxidative stress and molecular changes in liver fibrosis:a review[J].Curr Med Chem,2012,19(28):4850-4860.
[4]SHIN S M,YANG J H,KI S H.Role of the Nrf2-ARE pathway in liver diseases[J].Oxid Med Cell Longev,2013,2013(6):763257.
[5]KLAASSEN C D,REISMAN S A.Nrf2 the rescue:effects of the antioxidative/electrophilic response on the liver[J].Toxicol Appl Pharmacol,2010,244(1):57-65.
[6]SHIN S M,YANG J H,KI S H.Role of Nrf2 in chronic liver disease[J].Crit Rev Food Sci Nutr,2015,55(7):919-929.
[7]VITAGLIONE P,MORISCO F,CAPORASO N,et al.Dietary antioxidant compounds and liver health[J].Crit Rev Food Sci Nutr,2004,44(7-8):575-586.
[8]SEREN S,MUTCHNICK M,HUTCHINSON D,et al.Potential role of lycopene in the treatment of hepatitis C and prevention of hepatocellular carcinoma[J].Nutr Cancer,2008,60(6):729-735.
[9]ABASS M A,ELKHATEEB S A,ABD EL-BASET S A,et al.Lycopene ameliorates atrazine-induced oxidative damage in adrenal cortex of male rats by activation of the Nrf2/HO-1 pathway[J].Environ Sci Pollut Res Int,2016,23(15):15262-15274.
[10]YANG P M,WU Z Z,ZHANG Y Q,et al.Lycopene inhibits ICAM-1 expression and NF-κB activation by Nrf2-regulated cell redox state in human retinal pigment epithelial cells[J].Life Sci,2016,155:94-101.
[11]DAI C,TANG S,DENG S,et al.Lycopene attenuates colistin-induced nephrotoxicity in mice via activation of the Nrf2/HO-1 pathway[J].Antimicrob Agents Chemother,2015,59(1):579-585.
[12]BEN-DOR A,STEINER M,GHEBER L,et al.Carotenoids activate the antioxidant response element transcription system[J].Mol Cancer Ther,2005,4(1):177-186.
[13]CICHOZ·-LACH H,MICHALAK A.Oxidative stress as a crucial factor in liver diseases[J].World J Gastroenterol,2014,20(25):8082-8091.
[14]LI S,TAN H Y,WANG N,et al.The role of oxidative stress and antioxidants in liver diseases[J].Int J Mol Sci,2015,16(11):26087-26124.
[15]YANG J J,TAO H,HUANG C,et al.Nuclear erythroid 2-related factor 2:a novel potential therapeutic target for liver fibrosis[J].Food Chem Toxicol,2013,59(8):421-427.
[16]ZHANG H,DAVIES K J,FORMAN H J.Oxidative stress response and Nrf2 signaling in aging[J].Free Radic Biol Med,2015,88(Pt B):314-336.