锰超氧化物岐化酶模拟化合物诱导K562细胞凋亡的机制研究  

The apoptosis-inducing effect of MnSOD_m on K562 cells and its molecular mechanism

作　　者：范临兰[1] 魏虎来[1] 窦伟[2] 刘伟生[2] 张慧芳[1] 

机构地区：[1]兰州大学医学实验中心,甘肃省新药临床前研究重点实验室,甘肃兰州730000 [2]兰州大学化学化工学院,甘肃兰州730000

出　　处：《中国药理学通报》2009年第10期1363-1366,共4页Chinese Pharmacological Bulletin

基　　金：甘肃省新药临床前研究重点实验室开放基金资助项目(NoGSKFKT-0702)

摘　　要：目的观察锰超氧化物岐化酶模拟化合物（mimics of manganese superoxide dismutase,MnSODm）对人白血病K562细胞凋亡诱导作用,并探讨其分子机制。方法以人白血病K562细胞为靶细胞,四氮唑蓝比色法（MTT法）测定细胞增殖活性;Annexin V/PI双标记和细胞形态学法检测细胞凋亡;RT-PCR检测bcl-2和bax基因mRNA的表达水平;流式细胞术（FCM）测定Bcl-2和Bax蛋白表达水平、线粒体跨膜电位（Δψm）、细胞色素C（Cyt C）释放和Caspase-3活性变化。结果0.5~10mg·mL-1MnSODm明显抑制K562细胞增殖（P〈0.01）,Annexin V/PI染色显示凋亡细胞明显增多,光学显微镜和透射电镜观察呈现典型的凋亡形态改变;bcl-2基因mRNA和蛋白表达下调,bax基因mRNA和蛋白表达上调,线粒体Δψm降低,Cyt C释放增多,Caspase-3活性增强。结论MnSODm调控Bax/Bcl-2表达,通过线粒体途径诱导K562细胞凋亡。Aim To explore the apoptotic effect of mimics of manganese superoxide dismutase （ MnSODm ） on human leukemia cell line K562 in vitro and the possible molecular mechanisms. Methods Human leukemia K562 cells were used as the target cells. The cell proliferating activity was examined by a MTT colorimetrie assay, and the apoptosis of K562 cells was assessed with FITC-Annexin V and propidium iodide （PI） double staining and morphological changes. The expressions of bcl-2 and bax mRNA were detected by reverse transcription polymerase chain reaction （RT- PCR）, and flow eytometry （FCM） was employed to measure the expressions of Bcl-2 and Bax protein,mitoehondrial inner membrane potential （AOm）, Cytochrome C（Cyt C） release and Caspase-3 activity. Results The proliferation of K562 cells was obviously inhibited by 0. 5 - l0 mg · L^-1MnSODm （P 〈 0. 01 ）,and the apoptosis rate of K562 cells detected with Annexin V/PI staining was markedly increased, and the typical apoptotic morphological changes of K562 cells was observed under optical and electronic microscopic morphology. The down-regulation of expressions of bcl- 2 mRNA and protein was accompanied with the up-regulation of bax mRNA and protein. The disruption of mitochondria △ψm and the increased release of Cyt C from mitochondria to cytoplasm were noted, and the activity of Caspase-3 was significantly enhanced. Conclusions MnSODm can induce apoptosis of K562 cells via mitochondrial pathway by regulating the expression of Bax/Bcl-2.

关 键 词：MnSOD模拟化合物 K562细胞 凋亡 Bcl-2 BAX 膜电位 细胞色素C CASPASE-3 

分 类 号：R329.25[医药卫生—人体解剖和组织胚胎学] R733.702.2[医药卫生—基础医学]