辛伐他汀对氧化型低密度脂蛋白诱导的内皮细胞氧化应激损伤的保护作用及其机制的实验研究
신벌타정대양화형저밀도지단백유도적내피세포양화응격손상적보호작용급기궤제적실험연구
Simvastatin attenuates oxidized low density lipoprotein induced oxidative stress in human umbilical vein endothelial cells via downregulating NADPH oxidase activity
  • 万方数据
    中华心血管病杂志 중화심혈관병잡지
    Chinese Journal of Cardiology
    2014年 1期 43-47 ,共5页

    张磊善%郭锐%崔立宝

    장뢰선%곽예%최립보

    内皮细胞%斯伐他汀%脂蛋白类,LDL%氧化性应激 內皮細胞%斯伐他汀%脂蛋白類,LDL%氧化性應激
    내피세포%사벌타정%지단백류,LDL%양화성응격
    Endothelial cells%Simvastatin%Lipoproteins,LDL%Oxidative stress
    目的 观察辛伐他汀对氧化型低密度脂蛋白(oxidized low-density lipoprotein,ox-LDL)诱导的内皮细胞氧化应激损伤的保护作用,并初步探讨其机制.方法 体外分离培养人脐静脉内皮细胞后分为6组,分别是对照组、ox-LDL处理组(ox-LDL组)、辛伐他汀溶剂+ox-LDL组(溶剂对照组),辛伐他汀0.1 μmol/L+ ox-LDL组、辛伐他汀0.5 μmol/L+ ox-LDL组和辛伐他汀1.0 μmol/L+ox-LDL组.ox-LDL处理组直接给予ox-LDL(120 μg/ml)刺激24 h.辛伐他汀+ox-LDL组分别采用不同浓度的辛伐他汀(0.1、0.5和1.0 μmol/L)预先孵育人脐静脉内皮细胞30 rin,再给予ox-LDL(120 μg/ml)刺激24 h.荧光分光光度法测定胞内活性氧簇(ROS)的水平.化学发光法测定胞内还原型烟酰胺腺嘌呤二核苷酸磷酸氧化酶(NADPH氧化酶)活性.RT-PCR测定NADPH氧化酶亚基p22phox、gp91phox、p47phox和p67phoxmRNA的表达水平.免疫共沉淀以及免疫印迹方法测定p47 phox与p22phox(p47 phox/p22phox)的蛋白结合量.结果 ox-LDL组ROS水平和NADPH氧化酶活性均显著高于对照组[分别为1.507±0.048比0.442±0.021(P＜0.01)和(87.90±10.34) RLU·s-1·mg-1比(38.52±4.20) RLU·s-1·mg-1(P＜0.01)],p22phox、gp91 phox、p47phox和p67phox mRNA水平亦均显著高于对照组(P均＜0.01),p47 phox/p22 phox蛋白结合量亦显著高于对照组[(292±12)％比100％,P＜0.01].辛伐他汀不同浓度+ox-LDL各组ROS水平和NADPH氧化酶活性随着辛伐他汀浓度的增加而降低,组间差异均有统计学意义(P均＜0.05),且均低于ox-LDL组(P＜0.05或0.01).辛伐他汀1.0 μmol/L+ ox-LDL组p22phox、gp91phox、p47phox和p67phoxmRNA水平均显著低于ox-LDL组(P均＜0.05),p47phox/p22phox蛋白结合量亦显著低于ox-LDL组[(117±9)％比(292±l2)％,P＜0.01].结论 辛伐他汀可能通过降低NADPH氧化酶活性的途径保护ox-LDL诱导的内皮细胞氧化应激损伤,其机制可能与下调NADPH氧化酶亚基mRNA表达水平和抑制p47phox/p22phox蛋白结合有关.
    목적 관찰신벌타정대양화형저밀도지단백(oxidized low-density lipoprotein,ox-LDL)유도적내피세포양화응격손상적보호작용,병초보탐토기궤제.방법 체외분리배양인제정맥내피세포후분위6조,분별시대조조、ox-LDL처리조(ox-LDL조)、신벌타정용제+ox-LDL조(용제대조조),신벌타정0.1 μmol/L+ ox-LDL조、신벌타정0.5 μmol/L+ ox-LDL조화신벌타정1.0 μmol/L+ox-LDL조.ox-LDL처리조직접급여ox-LDL(120 μg/ml)자격24 h.신벌타정+ox-LDL조분별채용불동농도적신벌타정(0.1、0.5화1.0 μmol/L)예선부육인제정맥내피세포30 rin,재급여ox-LDL(120 μg/ml)자격24 h.형광분광광도법측정포내활성양족(ROS)적수평.화학발광법측정포내환원형연선알선표령이핵감산린산양화매(NADPH양화매)활성.RT-PCR측정NADPH양화매아기p22phox、gp91phox、p47phox화p67phoxmRNA적표체수평.면역공침정이급면역인적방법측정p47 phox여p22phox(p47 phox/p22phox)적단백결합량.결과 ox-LDL조ROS수평화NADPH양화매활성균현저고우대조조[분별위1.507±0.048비0.442±0.021(P＜0.01)화(87.90±10.34) RLU·s-1·mg-1비(38.52±4.20) RLU·s-1·mg-1(P＜0.01)],p22phox、gp91 phox、p47phox화p67phox mRNA수평역균현저고우대조조(P균＜0.01),p47 phox/p22 phox단백결합량역현저고우대조조[(292±12)％비100％,P＜0.01].신벌타정불동농도+ox-LDL각조ROS수평화NADPH양화매활성수착신벌타정농도적증가이강저,조간차이균유통계학의의(P균＜0.05),차균저우ox-LDL조(P＜0.05혹0.01).신벌타정1.0 μmol/L+ ox-LDL조p22phox、gp91phox、p47phox화p67phoxmRNA수평균현저저우ox-LDL조(P균＜0.05),p47phox/p22phox단백결합량역현저저우ox-LDL조[(117±9)％비(292±l2)％,P＜0.01].결론 신벌타정가능통과강저NADPH양화매활성적도경보호ox-LDL유도적내피세포양화응격손상,기궤제가능여하조NADPH양화매아기mRNA표체수평화억제p47phox/p22phox단백결합유관.
    Objective To investigate the effects and related mechanism of simvastatin on oxidized low density lipoprotein (ox-LDL) induced oxidative stress in human umbilical vein endothelial cells (HUVECs).Methods HUVECs were cultured in 6 different culture media:control,ox-LDL,ox-LDL + vehicle,ox-LDL + 0.1 μmol/L simvastatin,ox-LDL + 0.5 μmol/L simvastatin,ox-LDL + 1.0 μmol/L simvastatin.HUVECs were incubated with ox-LDL (120 μg/ml) for 24 h in the presence or absence of different concentrations of simvastatin (0.1,0.5,1.0 μmol/L).The fluorescence intensity for reactive oxygen species (ROS) in HUVECs was measured by a laser confocal scanning microscopy and a microplate reader.NADPH oxidase activity was measured by lucigenin chemiluminescence,p22phox,gp91 phox,p47phox and p67phox mRNA expression of HUVECs post various treatments was detected by RT-PCR.p22phox immunoprecipitates were immunoblotted for p47phox and total p22phox levels to indentify p47phox/p22phox interaction.Results Simvastatin attenuated ox-LDL induced ROS generation and NADPH oxidase activity in a concentration dependent manner (all P ＜ 0.05).In addition,simvastatin significantly downregulated mRNA expression of p22phox,gp91phox,p47phox and p67phox (all P ＜ 0.05),as well as the interaction of p47phox/p22phox (P ＜ 0.01).Conclusions Simvastatin is an important regulator on NADPH subunits mRNA expressions and p47phox/p22phox interaction.Simvastatin attenuates ox-LDL-induced oxidative stress in HUVECs via reducing NADPH oxidase activity.
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