中华心血管病杂志
中華心血管病雜誌
중화심혈관병잡지
Chinese Journal of Cardiology
2015年
4期
341-346
,共6页
刘源%唐其柱%胡哲夫%邓伟%李利娜
劉源%唐其柱%鬍哲伕%鄧偉%李利娜
류원%당기주%호철부%산위%리리나
心肌%氧化性应激%番茄红素
心肌%氧化性應激%番茄紅素
심기%양화성응격%번가홍소
Myocardium%Oxidative stress%Lycopene
目的 观察番茄红素(lycopene,Lyc)对血管紧张素Ⅱ(angiotensionⅡ,AngⅡ)诱导的H9c2细胞氧化应激的影响,并探讨其机制.方法 本研究为细胞实验,采用的是大鼠H9c2细胞系.实验将H9c2细胞分为6组,分别为空白对照组(对照组)、单加AngⅡ组(1.μmol/L)(AngⅡ组)、AngⅡ(1μmol/L) +Lyc (3.125 nmol/L)组(AngⅡ+低剂量Lyc组)、AngⅡ(1μmol/L)+Lyc(6.25nmol/L)组(AngⅡ+中剂量Lyc组)、AngⅡ(1μmol/L)+Lyc(12.5 nmol/L)组(AngⅡ+高剂量Lyc组)和单加Lyc(12.5 nnmol/L)组(Lyc组).采用不同因素干预H9c2细胞12h后,细胞毒性检测试剂盒(CCK-8)检测不同浓度Lyc和(或)AngⅡ对H9c2活性的影响,酶标仪及荧光显微镜检测Lyc对AngⅡ诱导细胞产生活性氧(ROS)的影响,实时荧光定量聚合酶链反应检测Lyc对AngⅡ诱导的细胞内还原型辅酶Ⅱ(NADPH)氧化酶2(NOX2)、NADPH氧化酶p47 phox亚基(p47phox)、超氧化物歧化酶1(SODl)及超氧化物歧化酶2(SOD2)基因表达的影响,细胞丙二醛(MDA)检测试剂盒检测MDA的变化.结果 (1)各组H9c2细胞活力的检测结果:AngⅡ组细胞生存率明显低于对照组(P＜0.01).而AngⅡ+低剂量Lyc组、AngⅡ+中剂量Lyc组和AngⅡ+高剂量Lyc组H9c2细胞生存率均明显高于AngⅡ组(P均＜0.01),且呈浓度依赖性.(2)各组H9c2细胞中ROS表达水平的检测结果:AngⅡ组H9c2细胞中ROS表达水平明显高于对照组(P＜0.01).而AngⅡ+低剂量Lyc组、AngⅡ+中剂量Lyc组和AngⅡ+高剂量Lyc组H9c2细胞中ROS表达水平均明显低于AngⅡ组(P均＜0.01),且呈浓度依赖性.(3)各组H9c2细胞中SOD1、SOD2 mRNA表达水平及MDA含量的检测结果:AngⅡ组H9c2细胞中SOD1、SOD2 mRNA水平均明显低于对照组,而MDA含量明显高于对照组(P均＜0.01).而AngⅡ+低剂量Lyc组、AngⅡ+中剂量Lyc组和AngⅡ+高剂量Lyc组H9c2细胞中SOD1、SOD2 mRNA水平均明显高于AngⅡ组,MDA含量则均低于AngⅡ组(P均＜0.01),且均呈浓度依赖性.(4)各组H9c2细胞中NOX2蛋白及其亚单位p47phox mRNA表达水平的检测结果:AngⅡ组H9c2细胞中NOX2蛋白表达水平及其亚单位p47 phox的mRNA水平均明显高于对照组(P均＜0.01),而AngⅡ+低剂量Lyc组、AngⅡ+中剂量Lyc组和AngⅡ+高剂量Lyc组二者的表达水平均明显低于AngⅡ组(P均＜0.01),且呈浓度依赖性.结论 Lyc可改善AngⅡ诱导的H9c2细胞氧化应激,其机制与Lyc可抑制AngⅡ诱导的H9c2细胞中ROS的生成,同时可促进细胞ROS的清除能力有关.
目的 觀察番茄紅素(lycopene,Lyc)對血管緊張素Ⅱ(angiotensionⅡ,AngⅡ)誘導的H9c2細胞氧化應激的影響,併探討其機製.方法 本研究為細胞實驗,採用的是大鼠H9c2細胞繫.實驗將H9c2細胞分為6組,分彆為空白對照組(對照組)、單加AngⅡ組(1.μmol/L)(AngⅡ組)、AngⅡ(1μmol/L) +Lyc (3.125 nmol/L)組(AngⅡ+低劑量Lyc組)、AngⅡ(1μmol/L)+Lyc(6.25nmol/L)組(AngⅡ+中劑量Lyc組)、AngⅡ(1μmol/L)+Lyc(12.5 nmol/L)組(AngⅡ+高劑量Lyc組)和單加Lyc(12.5 nnmol/L)組(Lyc組).採用不同因素榦預H9c2細胞12h後,細胞毒性檢測試劑盒(CCK-8)檢測不同濃度Lyc和(或)AngⅡ對H9c2活性的影響,酶標儀及熒光顯微鏡檢測Lyc對AngⅡ誘導細胞產生活性氧(ROS)的影響,實時熒光定量聚閤酶鏈反應檢測Lyc對AngⅡ誘導的細胞內還原型輔酶Ⅱ(NADPH)氧化酶2(NOX2)、NADPH氧化酶p47 phox亞基(p47phox)、超氧化物歧化酶1(SODl)及超氧化物歧化酶2(SOD2)基因錶達的影響,細胞丙二醛(MDA)檢測試劑盒檢測MDA的變化.結果 (1)各組H9c2細胞活力的檢測結果:AngⅡ組細胞生存率明顯低于對照組(P＜0.01).而AngⅡ+低劑量Lyc組、AngⅡ+中劑量Lyc組和AngⅡ+高劑量Lyc組H9c2細胞生存率均明顯高于AngⅡ組(P均＜0.01),且呈濃度依賴性.(2)各組H9c2細胞中ROS錶達水平的檢測結果:AngⅡ組H9c2細胞中ROS錶達水平明顯高于對照組(P＜0.01).而AngⅡ+低劑量Lyc組、AngⅡ+中劑量Lyc組和AngⅡ+高劑量Lyc組H9c2細胞中ROS錶達水平均明顯低于AngⅡ組(P均＜0.01),且呈濃度依賴性.(3)各組H9c2細胞中SOD1、SOD2 mRNA錶達水平及MDA含量的檢測結果:AngⅡ組H9c2細胞中SOD1、SOD2 mRNA水平均明顯低于對照組,而MDA含量明顯高于對照組(P均＜0.01).而AngⅡ+低劑量Lyc組、AngⅡ+中劑量Lyc組和AngⅡ+高劑量Lyc組H9c2細胞中SOD1、SOD2 mRNA水平均明顯高于AngⅡ組,MDA含量則均低于AngⅡ組(P均＜0.01),且均呈濃度依賴性.(4)各組H9c2細胞中NOX2蛋白及其亞單位p47phox mRNA錶達水平的檢測結果:AngⅡ組H9c2細胞中NOX2蛋白錶達水平及其亞單位p47 phox的mRNA水平均明顯高于對照組(P均＜0.01),而AngⅡ+低劑量Lyc組、AngⅡ+中劑量Lyc組和AngⅡ+高劑量Lyc組二者的錶達水平均明顯低于AngⅡ組(P均＜0.01),且呈濃度依賴性.結論 Lyc可改善AngⅡ誘導的H9c2細胞氧化應激,其機製與Lyc可抑製AngⅡ誘導的H9c2細胞中ROS的生成,同時可促進細胞ROS的清除能力有關.
목적 관찰번가홍소(lycopene,Lyc)대혈관긴장소Ⅱ(angiotensionⅡ,AngⅡ)유도적H9c2세포양화응격적영향,병탐토기궤제.방법 본연구위세포실험,채용적시대서H9c2세포계.실험장H9c2세포분위6조,분별위공백대조조(대조조)、단가AngⅡ조(1.μmol/L)(AngⅡ조)、AngⅡ(1μmol/L) +Lyc (3.125 nmol/L)조(AngⅡ+저제량Lyc조)、AngⅡ(1μmol/L)+Lyc(6.25nmol/L)조(AngⅡ+중제량Lyc조)、AngⅡ(1μmol/L)+Lyc(12.5 nmol/L)조(AngⅡ+고제량Lyc조)화단가Lyc(12.5 nnmol/L)조(Lyc조).채용불동인소간예H9c2세포12h후,세포독성검측시제합(CCK-8)검측불동농도Lyc화(혹)AngⅡ대H9c2활성적영향,매표의급형광현미경검측Lyc대AngⅡ유도세포산생활성양(ROS)적영향,실시형광정량취합매련반응검측Lyc대AngⅡ유도적세포내환원형보매Ⅱ(NADPH)양화매2(NOX2)、NADPH양화매p47 phox아기(p47phox)、초양화물기화매1(SODl)급초양화물기화매2(SOD2)기인표체적영향,세포병이철(MDA)검측시제합검측MDA적변화.결과 (1)각조H9c2세포활력적검측결과:AngⅡ조세포생존솔명현저우대조조(P＜0.01).이AngⅡ+저제량Lyc조、AngⅡ+중제량Lyc조화AngⅡ+고제량Lyc조H9c2세포생존솔균명현고우AngⅡ조(P균＜0.01),차정농도의뢰성.(2)각조H9c2세포중ROS표체수평적검측결과:AngⅡ조H9c2세포중ROS표체수평명현고우대조조(P＜0.01).이AngⅡ+저제량Lyc조、AngⅡ+중제량Lyc조화AngⅡ+고제량Lyc조H9c2세포중ROS표체수평균명현저우AngⅡ조(P균＜0.01),차정농도의뢰성.(3)각조H9c2세포중SOD1、SOD2 mRNA표체수평급MDA함량적검측결과:AngⅡ조H9c2세포중SOD1、SOD2 mRNA수평균명현저우대조조,이MDA함량명현고우대조조(P균＜0.01).이AngⅡ+저제량Lyc조、AngⅡ+중제량Lyc조화AngⅡ+고제량Lyc조H9c2세포중SOD1、SOD2 mRNA수평균명현고우AngⅡ조,MDA함량칙균저우AngⅡ조(P균＜0.01),차균정농도의뢰성.(4)각조H9c2세포중NOX2단백급기아단위p47phox mRNA표체수평적검측결과:AngⅡ조H9c2세포중NOX2단백표체수평급기아단위p47 phox적mRNA수평균명현고우대조조(P균＜0.01),이AngⅡ+저제량Lyc조、AngⅡ+중제량Lyc조화AngⅡ+고제량Lyc조이자적표체수평균명현저우AngⅡ조(P균＜0.01),차정농도의뢰성.결론 Lyc가개선AngⅡ유도적H9c2세포양화응격,기궤제여Lyc가억제AngⅡ유도적H9c2세포중ROS적생성,동시가촉진세포ROS적청제능력유관.
Objective To investigate the effect of Lycopene (Lyc) on Ang Ⅱ induced oxidative stress in H9c2 cell line derived from rat cardiac tissue,and to explore related mechanisms.Methods H9c2 cells were divided into 6 groups:control group,Ang Ⅱ group (1 μmol/L),Ang Ⅱ (1 μmol/L) + low dose Lyc (3.125 nmol/L) group,AngⅡ (1 μmol/L) + moderate dose Lyc(6.25 nmol/L)group and Ang Ⅱ (1 μmol/L) + high dose Lyc(12.5 nmol/L) group and Lyc group (12.5 nnmol/L).Cell growth was determined by CCK8 assay,ROS generation was detected using a Microplate reader and Fluorescence microscopy,the expression of NOX2 was determined by Western blot,mRNA expression of p47phox,SOD1 and SOD2 were determined by Real Time-PCR,MDA was detected by ELISA kit.Results Compared to control group,cell survival was significantly reduced and ROS generation was significantly increased post Ang Ⅱ stimulation,cotreatment with Lyc significantly improved cell survival and reduced ROS generation in a dosedependent manner (all P ＜ 0.01).mRNA expression of SOD1 and SOD2 was significantly downregulated while MDA concentration was significantly increased in Ang Ⅱ treated cells,which could be significantly reversed by cotreatment with Lyc in a dose dependent manner (all P ＜ 0.01).Protein expression of NOX2 and mRNA expression of p47phox were significantly upregulated post Ang Ⅱ and which could be significantly downregulated by cotreatment with Lyc in a dose-dependent manner (all P ＜ 0.01).Conclusion Lyc could attenuate Ang Ⅱ induced oxidative stress and this effect is linked with its capacity of reducing ROS generation and enhancing cellular ROS scavenging ability in H9c2 cells.
