医学论文-碘化N-正丁基氟哌啶醇对缺氧血管平滑肌细胞增殖的影响及其作用机制的研究.docVIP

医学论文-碘化N-正丁基氟哌啶醇对缺氧血管平滑肌细胞增殖的影响及其作用机制的研究 ??????????????? 作者：刘清,石刚刚，陈一村，周燕琼，郭福晓，郑燕珊??? 【摘要】? 目的:研究碘化N-正丁基氟哌啶醇(F2)对缺氧大鼠胸主动脉平滑肌细胞(SMCs)增殖的影响，并探讨其作用机制是否与早期生长反应基因-1(egr-1)有关。方法:取第3～4代培养的大鼠胸主动脉SMCs制作缺氧模型，应用噻唑蓝比色法检测细胞增殖情况,流式细胞术测定细胞周期,蛋白印迹杂交方法检测Egr-1及血小板源生长因子(PDGF-A)蛋白表达情况。结果:与缺氧组比,反义寡核苷酸组及F2组比色吸光度A值、细胞周期S+G2/M期比例均下降(P0.01)，Egr-1及PDGF-A蛋白表达均减少。结论:F2可通过抑制Egr-1蛋白表达来抑制缺氧SMCs的过度增殖。 【关键词】? 碘化N-正丁基氟哌啶醇;血管平滑肌细胞；缺氧;增殖；反义寡核苷酸;早期生长反应基因-1 　　［Abstract］Objective : To investigate the effect and mechanism of N-n-butyl haloperidol iodide(F2)on hypoxia-induced proliferation of rat thoracic aortic smooth muscle cells(SMCs)in vitro， and to indicate the relationship between the effect of F2 and the express of early growth response gene-1(egr-1). Methods:　Passages 3 to 4 of cultured rat thoracic aortic SMCs were used to establish hypoxia models. The viability of VSMC was detected by mono-nuclear cell direct cytotoxicity assay. The cell cycle was analyzed by flow cytometry. The levels of Egr-1 and PDGF-A were assessed by Western blot. Results: Compared with the hypoxia group,　the absorption value,proliferation index and the levels of Egr-1 and PDGF-A of antisense oligodeoxynucleotide group and F2 group were all decreased.　Conclusion: The inhibition of F2 on hypoxia-induced proliferation of rat thoracic aortic SMCs in vitro is mediated by Egr-1. 　　［Key Words］N-n-butyl haloperidol iodide; vascular smooth muscle cell; Hypoxia; proliferation； antisense oligodeoxynucleotides; early growth response gene-1 ??? 碘化N-正丁基氟哌啶醇(F2)是本实验室设计合成的一种新型氟哌啶醇季铵盐衍生物,已有实验表明它对心肌缺血再灌注损伤以及心肌细胞、内皮细胞缺氧复氧损伤具有保护作用，且这种作用与抑制早期生长反应基因-1(egr-1)mRNA及蛋白表达水平的过度升高有关［1～6］。缺氧可致血管平滑肌细胞(VSMCs)的过度增殖［7］，进而导致一系列血管增生性疾病。本研究拟在观察F2对心脏整体、离体以及内皮细胞的作用基础上，进一步观察F2对缺氧诱导的VSMCs增殖的作用及其与Egr-1的关系。 　　1? 材料与方法 　　1.1? 仪器与试剂 ??? 倒置相差显微镜(上海光学仪器厂,中国);超净工作台(苏州净化设备有限公司,中国);CO2培养箱(Thermo Forma公司,美国);凯基MTT细胞增殖检测试剂盒以及细胞周期检测试剂盒(凯基生物科技发展有限公司，中国)；multiskan spectrum酶标仪(Thermo Labsystem公司,芬兰);FACSCalibur流式细胞仪(Becton Dickinson公司，美国)；聚丙烯酰胺凝胶电泳相关材料(Bio-RAD公司,美国);硝酸纤维素膜(Inv