哇巴因诱导Jurkat细胞凋亡及胱冬酶-3及bcl-2基因家族关系.docVIP

哇巴因诱导Jurkat细胞凋亡及胱冬酶-3及bcl-2基因家族关系 　 作者：熊安秀，王敏， 金润铭， 白燕， 林雯 【摘要】 本研究探讨哇巴因诱导Jurkat细胞凋亡及其机制。采用四甲基偶氮唑盐（MTT）法观察哇巴因对Jurkat细胞生长的抑制作用，应用流式细胞术、原位末端标记技术（TUNEL）等观察细胞凋亡，应用Western-blot测定胱冬酶-3（caspase-3）的活性亚单位及Bcl-2、Bax蛋白表达水平，用比色法检测caspase-3活性。结果表明：哇巴因能诱导Jurkat细胞凋亡，在细胞凋亡过程中， Bax蛋白表达增加，caspase-3活性明显升高。结论：哇巴因可能通过调节Bcl-2、Bax蛋白表达，从而激活caspase途径诱导Jurkat细胞凋亡。 【关键词】 哇巴因；Jurkat细胞； 细胞凋亡； Bcl-2； Bax； caspase-3 　　Ouabain-induced Apoptosis of Jurkat Cells Correlates with Acti-vation of caspase-3 and Regulation of bcl-2 Gene Family 　　　　Department of Pediatrics, Yichang Central Hospital, The First Clinical Medical College of Sanxia University, Yichang 443003, China; 1Department of Pediatrics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology , Wuhan 430022, China AbstractThe study was aimed to explore the apoptosis effect of ouabain on Jurkat cells and its mechanism. MTT method was used to observe the inhibitory effect of ouabain on Jurkat cell proliferation. Apoptosis was detected by using flow cytometry(FCM) and the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling reaction method (TUNEL). The protein expressions of Bax, Bcl-2 and active subunits of caspase-3 were measured by Western blot. Activities of caspase-3 were determined by colorimetry method. The results showed that ouabain could induce apoptosis of Jurkat cells, the expression of Bax protein in process of cell apoptosis, caspase-3 activity of Jurkat cells were remarkably enhanced after ouabain treatment. It is concluded that ouabain may induce apoptosis of Jurkat cells due to the activation of caspase-3 resulting from regulation of Bax protein and bcl-2 gene expressions. 　　Key wordsouabain; Jurkat cell; apoptosis; Bcl-2; Bax; caspase-3 　　内源性哇巴因（endogenous ouabain,EO）是由肾上腺皮质分泌的一种类固醇激素，为特异的钠钾三磷酸腺苷转移酶(Na+-K+ATPase)抑制剂，具有调节水钠代谢、血管收缩和心肌收缩等重要的生理功能。最近研究表明，哇巴因对细胞具有促进增殖和诱导细胞凋亡的双重效应［1,2］，诱导细胞增殖和凋亡的信号传导［3］，但对哇巴因诱导细胞凋亡的机制目前仍未彻底阐明。据此，我们以白血病细胞株Jurkat为体外模型，探讨了Bcl-2、Bax和