倍半萜烯内酯诱导WEHI-3细胞凋亡的实验  

Experimental on Apoptosis in Leukemia Cells Induced by Thapsigargin

作　　者：易雪[1] 刘芳[2] 邹萍[2] 肖娟[2] 程辉[1] 

机构地区：[1]武汉市中西医结合医院,武汉43002 [2]华中科技大学同济医学院血液病研究所

出　　处：《肿瘤防治研究》2011年第12期1389-1392,共4页Cancer Research on Prevention and Treatment

摘　　要：目的探讨内质网Ca2+-ATP酶抑制剂Thapsigargin的抗白血病作用机制。方法利用An-nexin-Ⅴ/PI双染实验和琼脂糖凝胶电泳、TUNEL染色测定鼠白血病细胞株(WEHI-3)细胞凋亡,Fura-2荧光负荷技术检测细胞内游离钙离子浓度([Ca2+]i),并检测在此过程中Caspase-12的表达。结果 0.5、1、2μmol/L Thapsigargin作用于WEHI-3细胞后,分别用Annexin-V/PI双染实验、琼脂糖凝胶电泳、TUNEL法测定WEHI-3凋亡率结果均较对照组显著升高(如Annexin-V/PI双染实验测定凋亡率分别为(25.3±3.2)%、(46.7±3.9)%、(70.2±2.3)%较对照组(7.6±0.4)%显著升高(F=26.52,P<0.01)),且凋亡率呈剂量依赖性。Thapsigargin作用后[Ca2+]i分别为(156.5±10.3)nmol/L、(180.3±15.6)nmol/L和(210.7±15.3)nmol/L,均较对照组(78.3±11.2)nmol/L显著升高(F=21.26,P<0.01)。Caspase-12 mRNA的表达随着Thapsigargin作用剂量增加而增加。结论 Thapsigargin通过Ca2+超载后诱发内质网应激诱导了WEHI-3细胞凋亡,Ca2+可能成为抗肿瘤药物作用新靶点。Objective To investigate apoptosis in mouse leukemia cell（WEHI-3） induced by Thapsigargin and its mechanism.Methods Apoptosis induced by Thapsigargin was examined by flow cytometry and Agarose gel electrophoresis of DNA and TUNEL staining.Free calcium（i）was determined by Fura-2 fluorescein load techinique.RT-PCR was used to analyze the mRNA of Caspase-12.Results After exposure to Thapsigargin of 0.5,1 and 2 μmol/L,WEHI-3 cells were induced to apoptosis.The apoptotic index was examined by Annexin-V/PI double-stained,Agarose gel electrophoresis of DNA and TUNEL.The results showed that the apoptotic index of the experimental groups was significantly higher than that of the control group（For example：the apoptotic index of the experimental groups determined by Annexin-V/PI double-stained were（25.3±3.2）%,（46.7±3.9）% and（70.2±2.3）%,respectively,which were significantly higher than the apoptotic index of the control group which was（7.6±0.4）%（F=26.52,P0.01）;and the apoptotic index was in dose dependent manner.After exposure to Thapsigargin,that i in the experimental group was significantly higher than that of the control group[（156.5±10.3）nmol/L,（180.3±15.6） nmol/L and（10.7±15.3）nmol/L vs.（78.3±11.2）nmol/L（F=21.26,P0.01）].The mRNA expression of Caspase-12 increased with an increase in Thapsigargin concentration.Conclusion Thapsigargin can induce the WEHI-3 to apoptosis by Endoplasmic reticulum stress of i overload,Ca2＋ may play a promising target for cancer therapy.

关 键 词：倍半萜烯内酯 细胞凋亡 游离钙离子浓度 CASPASE-12 

分 类 号：R733.7[医药卫生—肿瘤]