哇巴因对人血管内皮细胞死亡和钠泵α1、β1亚单位表达的影响  

THE ROLE OF OUABAIN IN CELL DEATH OF VASCULAR ENDOTHELIAL CELLS ECV304 AND THE CHANGES OF EXPRESSION OF Na^+,K^+-ATPase α1,β1-SUBUNIT

作　　者：王娜[1] 徐瑞成[1] 陈小义[1] 呼文亮[1] 

机构地区：[1]中国人民武装警察部队医学院细胞生物学教研室,天津300162

出　　处：《中国应用生理学杂志》2008年第2期177-183,I0003,共8页Chinese Journal of Applied Physiology

基　　金：天津市自然科学基金项目(06YFJMJC10400)

摘　　要：目的:研究钠泵抑制剂哇巴因(ouabain)对人血管内皮细胞死亡的影响及其作用机制。方法:以脐静脉内皮细胞系ECV304为靶细胞,应用MTT实验检测哇巴因对细胞生长的作用;采用Hoechst33342/PI双荧光染色、透射电镜和DNA琼脂糖凝胶电泳等分析细胞死亡特征,半定量RT-PCR法检测钠泵α1和β1亚单位mRNA的表达。结果:哇巴因以浓度和作用时间依赖的方式抑制ECV304细胞生长。10μmol/L哇巴因作用24 h,引起细胞坏死;0.1μmol/L哇巴因作用24~48 h,细胞明显脱落,细胞间连接丧失,细胞出现染色质凝集、分布于核膜内缘、DNA裂解等凋亡特征。哇巴因能明显上调ECV304细胞钠泵α1亚单位mRNA的表达,下调β1亚单位mRNA表达,且两者均呈时间依赖性。结论:哇巴因能诱导人血管内皮细胞ECV304死亡,其上调钠泵α1亚单位表达、下调β1亚单位表达,可能与亚单位介导信号传递、降低细胞黏附有关。Aim： To study the effect of Na^＋ , K^＋ -ATPase inhibition by ouabain on growth and death of vascular endothelial cells ECV304 and involved mechanisms. Methods： Growth inhibition of ouabain on ECV304 cells was analyzed using MTT assay. The feature of cell death was studied by Hoechst33342/PI staining, transmission electron microscopy and DNA agarose gel electrophoresis in ECV304 cells treated with ouabain. The mRNA expression of Na^＋ , K^＋ -ATPase al ,β1-subunit was examined by reverse transcription PCR（RT-PCR） .Results： Ouabain inhibited the growth of ECV304 cells in a dose and time-dependent manner. 10 μmol/L ouabain treated for 24 hours could stimulate the necrosis of ECV304 cells; When treated with 0.1μmol/L ouabain for 24-48 hours, the cells showed obviously defluxion, the loss of cell-cell contacts, nuclear chromatin condensation, chromatin margination and DNA fragmentation. Na^＋ , K^＋ -ATPase al-subunit mRNA expression was significantly up-regulated in ECV304 cells treated with ouabain while the β-subunit expression conversely showed a significant decrease. Conclusion： Ouabain could up-regulate Na^＋, K^＋ -ATPase al-Subunit expression and reduce β-Subunit expression which mediated signal transduction and decreased cell-cell adhesions and induced ECV304 cells death.

关 键 词：哇巴因 血管内皮细胞 细胞死亡 细胞连接 钠泵亚单位 

分 类 号：R363[医药卫生—病理学]