中华急诊医学杂志
中華急診醫學雜誌
중화급진의학잡지
CHINESE JOURNAL OF EMERGENCY MEDICINE
2008年
5期
491-495
,共5页
丁惠民%邱海波%王连%刘玲%李红鹏
丁惠民%邱海波%王連%劉玲%李紅鵬
정혜민%구해파%왕련%류령%리홍붕
血管紧张素Ⅱ%人肺微血管内皮细胞%核因子-κB
血管緊張素Ⅱ%人肺微血管內皮細胞%覈因子-κB
혈관긴장소Ⅱ%인폐미혈관내피세포%핵인자-κB
Angiotensin Ⅱ%Human pulmonary micmvasctdar endothelial cell%Nuclear factor-κB
目的 探讨血管紧张素Ⅱ(Ang Ⅱ)对肺微血管内皮细胞(HPMEC)核因子-κB(NF-κB)的活性的影响,以及经典途径在Ang Ⅱ介导的NF-κB活化过程中的作用.方法 本实验分为:Ang Ⅱ组:10-6 mol/L AngⅡ分别刺激HPMEC 0、0.5、1、2、4 h;氯沙坦组:10-6mol/L氯沙坦(AngⅡ 1型受体阻滞剂)预先处理HPMEC 1 h,再予相同10-6mol/L AngⅡ刺激细胞2 h,提取胞浆蛋白和胞核蛋白.凝胶电泳迁移率分析实验(EMSA)观察细胞中NF-κB的DNA结合活性.Western印迹检测细胞浆中抑制因子κBα(IκBα)的含量.结果 AngⅡ刺激HPMEC 0.5 h后细胞中NF-κB活性明显上升(144.5±16.1),在2h达到峰值(270.1±27.2),4 h(215.1±17.8)较2 h有所下降,但仍高于0 h水平,以上各时间点与AngⅡ刺激细胞0 h(100.0±25.1)相比,差异有统计学意义(P＜0.05).与Ang Ⅱ刺激HPMEC 0 h IκBα含量(44.4%±2.1%)相比,0.5 h后IκBα含量即开始明显下降(38.9%±3.6%,P＜0.05),2 h后IκBα含量下降最为明显(32.6%±2.3%,P＜0.05),4 h细胞中IκBα含量较2 h有所上升,但仍然明显低于ArgⅡ刺激0 h细胞中IκBα的含量(40.1%±4.7%,P＜0.05).氯沙坦组NF-κB活性(115.4±10.7)和IκBα含量(43.6%±3.7%)与AngⅡ组0 h相比无明显差异.氯沙坦组中NF-κB活性明显低于AngⅡ组2 h,氯沙坦组中iκBα的含量亦明显高于AngⅡ组2 h.结论 AugⅡ能通过AT1介导HPMEC中NF-κB活化.经典途径参与了AagⅡ诱导的HPMEC中NF-κB活化过程.
目的 探討血管緊張素Ⅱ(Ang Ⅱ)對肺微血管內皮細胞(HPMEC)覈因子-κB(NF-κB)的活性的影響,以及經典途徑在Ang Ⅱ介導的NF-κB活化過程中的作用.方法 本實驗分為:Ang Ⅱ組:10-6 mol/L AngⅡ分彆刺激HPMEC 0、0.5、1、2、4 h;氯沙坦組:10-6mol/L氯沙坦(AngⅡ 1型受體阻滯劑)預先處理HPMEC 1 h,再予相同10-6mol/L AngⅡ刺激細胞2 h,提取胞漿蛋白和胞覈蛋白.凝膠電泳遷移率分析實驗(EMSA)觀察細胞中NF-κB的DNA結閤活性.Western印跡檢測細胞漿中抑製因子κBα(IκBα)的含量.結果 AngⅡ刺激HPMEC 0.5 h後細胞中NF-κB活性明顯上升(144.5±16.1),在2h達到峰值(270.1±27.2),4 h(215.1±17.8)較2 h有所下降,但仍高于0 h水平,以上各時間點與AngⅡ刺激細胞0 h(100.0±25.1)相比,差異有統計學意義(P＜0.05).與Ang Ⅱ刺激HPMEC 0 h IκBα含量(44.4%±2.1%)相比,0.5 h後IκBα含量即開始明顯下降(38.9%±3.6%,P＜0.05),2 h後IκBα含量下降最為明顯(32.6%±2.3%,P＜0.05),4 h細胞中IκBα含量較2 h有所上升,但仍然明顯低于ArgⅡ刺激0 h細胞中IκBα的含量(40.1%±4.7%,P＜0.05).氯沙坦組NF-κB活性(115.4±10.7)和IκBα含量(43.6%±3.7%)與AngⅡ組0 h相比無明顯差異.氯沙坦組中NF-κB活性明顯低于AngⅡ組2 h,氯沙坦組中iκBα的含量亦明顯高于AngⅡ組2 h.結論 AugⅡ能通過AT1介導HPMEC中NF-κB活化.經典途徑參與瞭AagⅡ誘導的HPMEC中NF-κB活化過程.
목적 탐토혈관긴장소Ⅱ(Ang Ⅱ)대폐미혈관내피세포(HPMEC)핵인자-κB(NF-κB)적활성적영향,이급경전도경재Ang Ⅱ개도적NF-κB활화과정중적작용.방법 본실험분위:Ang Ⅱ조:10-6 mol/L AngⅡ분별자격HPMEC 0、0.5、1、2、4 h;록사탄조:10-6mol/L록사탄(AngⅡ 1형수체조체제)예선처리HPMEC 1 h,재여상동10-6mol/L AngⅡ자격세포2 h,제취포장단백화포핵단백.응효전영천이솔분석실험(EMSA)관찰세포중NF-κB적DNA결합활성.Western인적검측세포장중억제인자κBα(IκBα)적함량.결과 AngⅡ자격HPMEC 0.5 h후세포중NF-κB활성명현상승(144.5±16.1),재2h체도봉치(270.1±27.2),4 h(215.1±17.8)교2 h유소하강,단잉고우0 h수평,이상각시간점여AngⅡ자격세포0 h(100.0±25.1)상비,차이유통계학의의(P＜0.05).여Ang Ⅱ자격HPMEC 0 h IκBα함량(44.4%±2.1%)상비,0.5 h후IκBα함량즉개시명현하강(38.9%±3.6%,P＜0.05),2 h후IκBα함량하강최위명현(32.6%±2.3%,P＜0.05),4 h세포중IκBα함량교2 h유소상승,단잉연명현저우ArgⅡ자격0 h세포중IκBα적함량(40.1%±4.7%,P＜0.05).록사탄조NF-κB활성(115.4±10.7)화IκBα함량(43.6%±3.7%)여AngⅡ조0 h상비무명현차이.록사탄조중NF-κB활성명현저우AngⅡ조2 h,록사탄조중iκBα적함량역명현고우AngⅡ조2 h.결론 AugⅡ능통과AT1개도HPMEC중NF-κB활화.경전도경삼여료AagⅡ유도적HPMEC중NF-κB활화과정.
Objective To investigate the activation of nuclear factor-κB(NF-κB),which was stimulated by angiotensin-Ⅱ(AngⅡ)through classical pathway in human pulmonary microvascular endothehal cells(HPMEC).Method The experiment was divided into two groups:in Ang Ⅱ group,HPMECS were incubated with 10-6mol/L AngⅡ for 0,0.5,1,2 and 4 hours,respectively;in losartan group,HPMEC was pretreated with 10-6mol/L losartan(inhibitor of AngⅡ type 1 receptor)for 1 hour,and then stimulated with 10-6mol/L AngⅡ for 2 hours,and the nucleax protein and the cell plasma protein were prepared by lysis and centrifugation.Electrophoretic mobility shift assay(EMSA)was used to detect the NF-κB DNA binding activity.The inhibitor of κBa(IκBα)was detected by Western blotting.The data were expressed as(x±s)and analyzed with one way analysis of variance.A P value less than 0.05 indicated significant difference.Results Compared with the activity of NF-κB at 0 h (100.0±25.1)after AngⅡstimalation,the activity increased significantly at 0.5 hour(144.5±16.1,P＜0.05),and reached peak value at 2 hours(270.1±27.2,P＜0.05).The concentration of IκBα at 0 hours was 44.4%±2.1%,decreased markedly at 0.5 hours(38.9%±3.6%,P＜0.05),and to the lowest level at 2hours(32.6%±2.3%,P＜0.05).The activity of NF-κB(115.4±10.7)and the concentration of IκBα(43.6%±3.7%)in losartan group had ilo significant difference with AngⅡ group at 0 h(P＞0.05).The activity of NF-κB and the concentration of IκBα in losartan group had significant difference with AngⅡ group at 2hours.Conclusions NF-κB can be activated through classical pathway,which stimulated by AngⅡ in HPMEC.
